Pancreas and islet preservation

C H A P T E R

42 Pancreas and islet preservation Klearchos K. Papas, Hector De Leon University of Arizona, Department of Surgery, Institute of Cellular Transplantation, Tucson, AZ, United States

O U T L I N E Introduction

503

Key determinants of islet yield, function and viability during preservation 504 Impact of ischemia time during pancreas procurement, storage and transportation on transplantation outcomes 504 Ischemia/reperfusion injury 505 Effects of hypoxia during islet isolation, culture and distribution 506 Pancreas preservation prior to islet isolation Static methods/pancreas immersion Dynamic methods/pancreas perfusion

507 507 511

Introduction Autoimmune destruction of insulin-producing βcells in the endocrine pancreas leads to type 1 diabetes (T1D). T1D patients are currently treated with insulin therapy through infusion pumps or daily injections, whereas severe cases are considered for whole pancreas transplantation.1,2 Human pancreas transplantation represents a procedure that may attain normoglycemia without the need for exogenous insulin; nonetheless, it is a major surgical procedure that requires systemic immunosuppression for the life of the graft. Simultaneous pancreas and kidney transplant (SPK) is the most common transplant category3; however, pancreas transplants are also conducted as single grafts in diabetic patients.4 Although graft survival rates have improved for all transplant categories one

Transplantation, Bioengineering, and Regeneration of the Endocrine Pancreas, Volume 1 https://doi.org/10.1016/B978-0-12-814833-4.00042-3

Islet preservation during isolation and purification Isolation of human islets Pancreas distension, tissue dissociation, and islet collection Islet purification

513 514 515 515

Islet preservation during culture and distribution Culture ware and culture methods Islet culture supplements Cryopreservation

516 516 517 518

Outlook

518

References

519

year after transplantation (79%–90%),4 the need for major surgery, immunosuppression regimens for the life of the graft, and graft loss a few years after transplantation are major limitations to the use of whole pancreas transplantation as a therapeutic option for the majority of T1D patients.4–6 Results from studies comparing pancreas transplant outcomes of organs obtained from donors after cardiorespiratory death (DCD) to those obtained from donors after brain death (DBD) show comparable results3,7,8 encouraging the inclusion of less than optimal donors in clinical practice. The transplantation of isolated human islets and ultimately porcine islets or human stem cell-derived beta cells in the long-term represent alternative therapeutic approaches for T1D currently pursued by several research groups to reduce the need for whole pancreas transplantation.9–17

503

© 2020 Elsevier Inc. All rights reserved.

504

42.  Pancreas and islet preservation

Versions of the Edmonton Protocol, an intrahepatic allogeneic islet transplantation procedure,18 are currently implemented globally for a selected number of qualified T1D patients,19,20 including those with a high surgical risk.21 Islet transplantation has the advantage of being a minimally invasive procedure with intra-­ portal islet infusion conducted under radiological guidance. Follow up studies evaluating long-term outcomes of islet transplantation with the Edmonton protocol indicate that only about 10% of patients were insulin-­ independent after five years, with a median duration of insulin independence of 15 months; both parameters were significantly worse than those of a single vascularized whole pancreas transplant.22 Over the past decade, islet transplantation has continued to evolve with substantial improvements in outcomes.19,23 However, islets from more than one donor pancreas are still required to achieve the same short and long-term (5  year) outcomes achieved with a single whole pancreas transplant (donor to recipient ratio of 1:1).20,22–28 The need of islets from multiple donors and the high cost associated to a complex and laborious islet isolation process are major roadblocks to expand the number of diabetic patients that can benefit from islet transplantation. The high attrition rate of islets during pancreas preservation, islet isolation, culture and engraftment is the primary reason behind the need for islets from multiple donors for a single recipient. Islets are richly vascularized, complex multicellular structures, highly vulnerable to the harmful effects of ischemia and hypoxia.29–35 Only a fraction (30%–50%) of human islets can be isolated using current isolation protocols,36,37 a variable islet fraction dies during islet culture and clinical-grade preparations are obtained only in approximately 50% of isolations.38 Additionally, more than 50% of isolated islets do not engraft after transplantation and this is at least in part associated with hypoxia-induced damage prior to islet revascularization.39 Using diffusion-­ reaction

in silico models, it has been shown that hypoxia and hypoxia-­ related events may be at the center of islet loss of function after transplantation.40 Multiple experimental studies have demonstrated that islet size and oxygen microenvironment are key determinants of islet survival and clinical outcomes lending further support to predictions generated by mathematical models.39–41 Therefore, preservation measures must be implemented at the organ, tissue and cellular level before and after islet isolation and purification to increase beta cell viability, preserve cell function and improve clinical transplantation outcomes.

Key determinants of islet yield, function and viability during preservation Impact of ischemia time during pancreas procurement, storage and transportation on transplantation outcomes DBD and DCD are critically important organ donor categories for physicians, surgeons, health organizations and decision-making bodies within transplant programs because current transplantation conventions consider organs from DCD donors to be of inferior quality due to exposure to warm ischemia. Warm ischemia is used to describe two distinct periods: (1) ischemia during organ retrieval from a donor’s body—donor warm ischemia time (DWIT)—and (2) ischemia during organ implantation. A DBD organ donor exhibiting normal heart rate and cardiac contractility ensures that organs retrieved from the body after withdrawal of life support undergo the shortest possible DWIT. For DBD donors, DWIT is the interval between withdrawal of life support—onset of asystole for DCD donors—to the start of organ flushing with cold solutions. DWIT is followed by graft/­organ cold ischemia time (CIT), which ends with

FIG. 1  Steps involved in human islet preservation prior to ITx. These steps involve pancreas procurement and preservation, islet isolation and purification as well as islet culture and distribution as necessary. DWIT is minimal in DBD donors. OQA and IQA can be performed right after procurement. Preservation measures (e.g., oxygenation, additives) are usually taken at all steps during islet isolation, culture and distribution before ITx. CIT, cold ischemia time; DBD, donation after brain death; DCD, donation after circulatory death; IQA, islet quality assessment; ITx, islet transplantation; OQA, organ quality assessment; DWIT, donor warm ischemia time.

B. Islet allo-transplantation



Key determinants of islet yield, function and viability during preservation

retrieval of the graft from cold storage at ~4°C (Fig. 1). The time it takes from organ retrieval from cold storage to organ reperfusion in the recipient’s body is referred to as graft warm ischemia time (WIT).42,43 The length of DWIT is dependent upon type of organ donor—DBD donors have shorter or no DWIT compared to DCD donors (Fig. 1), whereas CIT varies primarily as a function of organ storage and transportation times. Since DWIT starts during the phase of cardiorespiratory collapse that precedes the onset of asystole in controlled DCD donors, it has been suggested that a better measure of injury during DWIT— also called functional DWIT—starts when the donor’s arterial pressure drops below 50 mmHg, the arterial oxygen saturation decreases below 70%, or both, and ends with cold perfusion.44 Islets isolated from DCD or DBD donors have been reported to give similar yields, purity and viability, as well as glucose stimulated insulin secretion (GSIS)45 suggesting that quality of islets from both donor types are comparable. However, the sample size of the DCD group in this study was disproportionately smaller compared to the DBD group. DCD represented less than 5% of all processed pancreata. Other reports indicate that islets from DCD pancreata are associated with reduced long-term survival.46 Although pancreata from DCD donors seem suitable for islet transplantation,47,48 more studies to confirm these findings are warranted.

Ischemia/reperfusion injury Hypoxia/cold ischemia The cellular pathobiology underlying warm and cold ischemia is related to ischemia reperfusion (I/R) injury. Donor(s) risk factors, organ retrieval time and technique, as well as organ and islet preservation methods are all terms in an equation that outputs I/R injury, a highly impactful component of transplantation outcomes.49 Regardless of temperature and donor-recipient considerations, interruption of blood flow to an organ jeopardizes its function and may cause irreversible cell and tissue damage in a matter of minutes due to hypoxia, the lack of oxygen and other nutrients and accumulation of toxic metabolic byproducts. As an acceptor of electrons in cellular respiration, oxygen is an indispensable component to generate energy in the form of ATP for cellular function. Hypoxia prevents oxidative phosphorylation and depletes ATP.50 The chain of events that lead to cell dysfunction and death during hypoxia starts with an inability to generate enough ATP to meet the demands of living tissue. Cell depolarization and an unregulated rise of intracellular Ca++ lead to enzymatic activation (e.g., proteases), cell swelling and eventually to cell apoptosis and/or necrosis. Increased cytosolic Ca++ activates calcium-dependent hydrolases including phospholipase

505

A2 and other proteases such as calpain.51–54 The protein degradation that ensues in membranes and intracellular compartments is considered a key event that eventually leads to cell death/necrosis. To minimize cell and tissue injury due to hypoxia, organs are stored in cold solutions (static cold storage (SCS)) or stored and perfused with cold liquid solutions (hypothermic machine perfusion (HMP)) or humidified oxygen gas (persufflation, (PSF))—in experimental settings. Hypothermia (2–4°C) slows down cellular metabolic rate, ATP use and production, oxygen consumption rate (OCR), and Na+/K+ ATPase activity. Depending on the preservation method and solutions used (e.g., oxygenated vs nonoxygenated solutions) organs are exposed to varying degrees of hypoxia and even to complete anoxia in a time span that varies greatly between organ procurement and restoration of blood flow upon transplantation completion. Ionic disturbances To a large extent, functional living cells are defined by the energy-dependent maintenance of electrochemical potentials across cellular membranes, which enable a tightly regulated molecular traffic across them. During hypoxia, the demands on anaerobic metabolism to provide energy (ATP) from glucose or glycogen to sustain electrical potentials—ion gradients—across biological membranes are markedly increased, and yet the energy needs to sustain functional membranes are unmet. Consequently, ions across cell membranes begin drifting towards their thermodynamic equilibrium. Maintenance of physiological ionic cell and tissue gradients—the cornerstone of cellular homeostasis—during hypothermia/ hypoxia requires preventing ion leakage and redistribution of ions that already leaked to restore concentrations across membranes compatible with cellular excitability and function.55 ATP-dependent mechanisms and molecules, including but not limited to, Na+/K+-ATPases are largely responsible for maintaining and restoring an ionic homeostasis compatible with living cells. Alterations of ion pumps and transporters contributing to regulating pH including the Na+/H+ exchanger, the Na + / HCO 3 − symporter, and the Na+-K+-2Cl cotransporter have been extensively documented in liver cells and cell lines.56–63 Of note, ionic dysregulation occurs not only in cell membranes but also in membranes of intracellular organelles (e.g., mitochondria, lysosomes). Thus, alterations of cellular ion homeostasis are extensive and at the center of hypothermic injury. ROS-mediated injury The paradoxical observation that reoxygenation enhances cell and tissue injury has perplexed investigators for over a century. Disorders characterized by chronic and acute reductions in organ blood flow and I/R i­ njury

B. Islet allo-transplantation

506

42.  Pancreas and islet preservation

include myocardial infarction and stroke.64 The I/R undergone by transplanted organs shares with those disorders the abrupt halting of blood flow after procurement and the reperfusion that occurs when blood flow is restored to the graft in the recipient’s body. According to one premise, the reoxygenation injury is fully dependent on the biochemical events and cellular alterations that occur during the ischemic/hypoxic period, whereas another school of thought argues that blood reperfusion inflicts tissue injury due to release of reactive oxygen species (ROS), dysregulation of Ca++ storage and microvascular dysfunction.65 Reoxygenation of a dysfunctional respiratory chain that leaks electrons translates into the formation of superoxide anion radical (O 2 −). Through a series of reactions, two O 2 − can form molecular oxygen (O2) and hydrogen peroxide (H2O2), which in turn may generate hydroxyl radicals (−OH) that react with other molecules and render them nonfunctional.66 ROS can induce direct damage of cellular proteins and molecules and activate signaling pathways involved in inflammation including cytokines and cell adhesion molecules.67 Disturbances of mitochondrial membrane potential and mitochondrial permeability may also lead to release of proapoptotic molecules and further production of ROS, which are themselves able to initiate cell death programs.68 Numerous comprehensive reviews have been published on the subject of I/R injury,64,69 and on the specific role of ROS in hypoxia-reoxygenation injury.65,68 The reader is referred to those reviews for indepth information regarding the cellular mechanisms of I/R injury, the intracellular sources of ROS (e.g., xanthine dehydrogenase oxidase, NADPH oxidase), as well as the antioxidant defense mechanisms that inhibit reoxygenation injury (e.g., glutathione). The pancreas is particularly sensitive to ischemia as reflected by the high incidence of pancreatitis during hemorrhagic shock and cardiac surgery.70,71 Significant correlations between pancreatic cellular injury after cardiac surgery and a battery of biomarkers including pancreatic enzymes and alpha2-macroglobulin have been reported.72 Microcirculatory failure is considered by some investigators the distinctive pathophysiological feature of pancreatic I/R injury.49 Using continuous tissue oximetry, impaired pancreatic microcirculation has been reported in the early reperfusion period after pancreas transplantation.73 Intravital microscopy studies with direct visualization of the capillary network and interactions between leukocytes and endothelial cells have described impairment of capillary perfusion and tissue oxygenation during reperfusion.74–76 To varying degrees of success, therapeutic approaches for I/R injury in experimental pancreas transplantation have included nitric oxide (NO)77,78 and tetrahydrobiopterin (BH4)79 supplementation, anticoagulation with antithrombin III,80 and antiinflammatory molecules—

monoclonal antiICAM-1 antibodies81 and antiadhesive P-selectin antagonists.82

Effects of hypoxia during islet isolation, culture and distribution Current isolation and purification protocols expose islets to a fluctuating range of temperatures from ~4°C to ~37°C. Moreover, islet preparations may be repeatedly exposed to hypoxia and reoxygenation, heat or cold shock during the various steps involved in islet isolation, purification and culture. Various studies have examined the effects of a hypoxic environment created by high seeding densities and temperature changes that islets undergo during isolation and culture. Some investigators have examined the effects of culturing islets at low temperatures to mitigate the effects of a hypoxic environment.83,84 It is important to emphasize that the exposure to hypoxia before organ procurement and islet isolation such as anemia33 or exposure of living animals to intermittent hypoxia85 may have a different mechanistic basis for its effect compared to the hypoxic environment and mechanical stress that islets are exposed to during islet isolation from distended and minced pancreata. The gene expression signatures of isolated islets exposed to hypoxic conditions include upregulation of the classical hypoxia-induced factor (HIF)-related pathways and increased expression of proinflammatory genes.34,35,86–89 Moderate hypoxia of MIN6 beta cells also exhibited downregulation in the expression of other genes relevant to beta cell pathways and function including Mafa, Pdx1, Slc2a2, Ndufa5, Kcnj11, Ins1, Wfs1, Foxa2, and Neurod1. Hypoxia also induced apoptosis and abnormal insulin secretion.90 Genes involved in endoplasmic reticulum homeostasis were also downregulated in MIN6 cells.91 It remains to be elucidated whether the gene expression signature of islets exposed to hypoxia during isolation is similar to the one reported for MIN6 beta cells. The mammalian pancreas has high concentrations of zinc (Zn++), a key ion required for insulin processing, storage and secretion.92 A disturbed zinc metabolism has been linked to insulin deregulation. Hypoxia-driven decreased levels of zinc transporters (e.g., ZIP8) in beta cells leads to decreased Zn++ concentration in the cytosol that results in hypoinsulinemia.93 Islets undergo considerable vascular and structural damage during isolation, a multistep process that encompasses distention of the pancreatic parenchyma with collagenase solutions, enzymatic disaggregation of minced pancreatic tissue, islet purification by density gradients and islet culture. The common element during a typical islet isolation procedure is hypoxia. Thus, the less severe the enzymatic conditions used, and the better oxygenated islets are during isolation, the higher

B. Islet allo-transplantation



Pancreas preservation prior to islet isolation

the islet yield and viability. In the following sections we examine the preservation steps taken to protect islets prior to and during the isolation and culture procedures before transplantation. We also highlight the areas that may have the largest impact to protect islet function and translate into significant benefits to islet transplantation recipients.

Pancreas preservation prior to islet isolation Conventional organ preservation methods may be efficacious to preserve pancreata from DBD donors with relatively short CIT, however, islet isolation from pancreata from DCD donors and extended preservation times may require further optimization of preservation techniques. In the sections below, we will review current

507

static and dynamic methods being tested clinically and experimentally to preserve whole pancreata prior to islet isolation and purification.

Static methods/pancreas immersion Several organ preservation procedures involving immersion of the whole organ in cold solution/medium have been developed over the last few decades. To minimize the effects of I/R injury, cytoprotective agents may be added to preservation solutions (Fig. 2). We briefly review the use of preservation solutions and additives. We refer the reader to previously published comprehensive reviews on the specific subject.94–98 Efforts to minimize temperature fluctuations are being considered by a number of investigators to improve islet yield and quality, and transplantation outcomes.

FIG. 2  Interventions aimed at improving islet preservation prior to ITx. ITx emphasis on a 1:1 donor to recipient strategy requires implementation of novel methods to reduce islet attrition rates during organ procurement and storage as well as islet isolation, purification, culture and distribution. Cytoprotective agents include a wide range of molecules to protect islets from hypoxia, apoptosis and inflammation including PFCs, AAT, glutathione, NO, CO, H2S, and ATP. The 1:1 donor to recipient strategy also requires implementation of a set of validated organ and islet quality assessment tests—OQA and IQA, respectively—at key “check points” along the process from organ procurement to ITx and islet engraftment, as well as development of appropriate “isletware” to enable oxygenation of islet preparations. AAT, alpha 1-antitrypsin; ATP, adenosine triphosphate; CO, carbon monoxide; ECM, extracellular matrix; H2, hydrogen; HMP, hypothermic machine perfusion; H2S, hydrogen sulfide; IQA, islet quality assessment; ITx, islet transplantation; NHP, nonhuman primate; NMP, normothermic machine perfusion; NO, nitric oxide; OQA, organ quality assessment; PFCs: perfluorocarbons, PSF, persufflation; PGE1, prostaglandin E1; SCS, static cold storage; TLM, two-layer method.

B. Islet allo-transplantation

508

42.  Pancreas and islet preservation

Static cold storage SCS is the most extensively used preservation method regardless of whether the pancreas would be used for whole organ transplantation or islet isolation, and it remains as the most extensively utilized method for pancreas preservation during retrieval and transport. Tissues under hypothermic conditions (2–4°C) reduce drastically their metabolic activity and their ATP levels.99,100 Maintenance of glucose oxidation rate to CO2 during hypothermia has been used as a surrogate measure of organ function. Although short-term storage and transport are currently accepted indications of SCS, it is clearly an insufficient method for “marginal” organs from expanded criteria donors (ECDs) that exhibit high risk of disfunction after transplantation and long-term storage.95 For example, prolonged CIT is associated with primary nonfunction in the liver101 and delayed graft function (DGF) in renal transplantation.102 Long periods of cold ischemic stress (16 h) of adult porcine pancreata have also been shown to have a detrimental effect on islet yield and quality.103 A significant drop in metabolism during SCS of large organs (e.g., liver, pancreas, kidney) does not occur fast enough to prevent ischemic injury; tissue is not appropriately oxygenated under such conditions even while immersed in fully oxygenated solutions—saturated with 100% oxygen gas. Organ oxygenation during SCS is heavily dependent on tissue thickness and OCR. Preserving an organ by immersion in a cold solution equilibrated with pure oxygen (pO2 = 760 mmHg at the solution/organ interphase), as performed in SCS, results in a maximum oxygen penetration depth of approximately 1 mm.104 Therefore, using the same immersion solution and keeping all other factors constant (e.g., temperature, time), relative oxygenated organ volumes from small species (e.g., mice, rats) will always be greater compared to those from large species such as pigs and humans.105 Additionally, cellular edema occurs during SCS and, because of anaerobic metabolism, ROS accumulate and further damage tissues upon reperfusion after transplantation (see I/R injury section above). Due to these limitations and the inability to sample biomarkers of quality and function from organs kept in cold medium, the use of SCS has begun to decrease as more effective alternative preservation methods including HMP and PSF have emerged. Preservation solutions Preservation solutions contain impermeant agents, osmotic molecules that prevent cell swelling, purine nucleotide precursors (adenine, phosphates, ribose) to sustain a low metabolic rate, antioxidants (glutathione, vitamin E), enzyme inhibitors, buffers (e.g., HCO3) and electrolytes mimicking intracellular (e.g., UW solution) or extracellular (e.g., IGL-1 solution) fluids. In short,

preservation solutions are aimed at sustaining metabolic rates, prevent edema, acidosis and ROS formation during hypothermia.106 Preservation solutions for SCS of abdominal organs include the Euro-Collins (EC), University of Wisconsin (UW), histidine-tryptophan-ketoglutarate (HTK), Celsior and Institut Georges Lopez-1 (IGL-1) solutions.94 The preservation rationale behind the EC and UW solutions resides in protecting the intracellular space from ischemic injury, as reflected by their high potassium and low sodium content. EC contains glucose as an osmotic protective barrier, whereas UW replaced glucose with more efficient osmotically active components such as lactobionate and raffinose to aid in preventing edema. The EC solution was introduced by Collins and his colleagues in 1969 and it was extensively used for organ preservation for two decades.98,107 The shortage of solid organs has forced transplantation experts to utilize organs from DCD and ECD donors and develop preservation solutions that are better at limiting the greater risk of tissue damage posed by organs from DCD and ECD donors. Introduced in 1989, the UW solution has become the standard for static cold preservation of liver, kidney, pancreas and intestine.94 UW is a highly viscous solution because of the presence of osmotically active colloid hydroxyethyl starch, raffinose, and lactobionate (Table 1). Developed for pancreas preservation, UW remains the preferred SCS solution.108 Disadvantages of using UW solution include resistance to flushing due to its high viscosity, the need to filter it due to formation of adenosine crystals and risk of inducing hyperkalemia since UW has a high potassium concentration (125 nmol/L). EC, the predecessor of UW solution, contains glucose, chloride and larger concentrations of phosphate compared to UW. HTK contains osmotically active mannitol, high concentration of histidine (198 mmol/L) as a buffer, tryptophan (1 mmol/L), an aromatic amino acid that stabilizes cell membranes, and ketoglutarate (2 mmol/L) as an energy substrate. HTK is not as viscous as UW and may therefore be used for organ flushing and faster cooling of organs from living donors when short preservation times are required.109 HTK and UW appear to be clinically comparable for organ preservation in pancreas transplantation.110–112 However, retrospective studies have reported a higher incidence of postoperative complications including graft loss, pancreatitis and thrombosis with HTK solution.113,114 Further prospective, randomized clinical trials are needed to determine whether a causal relationship exists. HTK and UW appear to be equally effective in the preservation of human pancreata intended for islet isolation.115,116 Like HTK, Celsior has a higher buffer capacity than UW solution. Both HTK and Celsior contain mannitol, but Celsior also contains lactobionate,

B. Islet allo-transplantation



509

Pancreas preservation prior to islet isolation

TABLE 1  Composition of common preservation solutions Euro-Collins UW (Viaspan)

HTK (Custodiol)

HTK-Ti-Protec

Celsior

IGL-1

 Sodium

10

30

15

16

100

120

 Potassium

115

125

9

93

15

30

 Chloride

15

–

50

103

28

20

 Calcium

–

–

0.02

0.05

0.25

–

 Magnesium

–

5

4

6

13

5

 Phosphate

57.5

25

–

–

–

25

 Sulfate

–

5

–

–

–

5

 Bicarbonate

–

10

–

–

–

–

  Lactobionate (mmol/L)

–

100

–

–

80

100

  HES (g/L)

–

50

–

–

–

–

  Raffinose (mmol/L)

–

30

–

–

–

30

  Mannitol (mmol/L)

–

–

30

–

60

–

  Ketoglutarate (mmol/L)

–

–

1

–

–

–

 Polyethylene glycol (35 kDa) (mmol/L)

–

–

–

–

–

0.03

  Glucose (mmol/L)

126

–

–

10

–

–

  Sucrose (mmol/L)

–

–

–

37

–

–

 Histidine

–

–

198

198

30

–

 Tryptophan

–

–

2

2

–

–

 Glutamate

–

–

–

20

–

 Glycine

–

–

–

5

–

–

  Insulin (U/L)

–

100

–

–

–

–

  Penicillin (U/L)

–

200

–

–

–

–

  Adenosine (mmol/L)

–

5

–

–

–

5

  Allopurinol (mmol/L)

–

1

–

–

–

1

  Dexamethasone (mg/L)

–

8

–

–

–

–

  Glutathione (mmol/L)

–

3

–

–

3

3

  N-Acetyl histidine c (mmol/L)

–

–

–

30

–

–

 Defreroxamine/L20 iron chelator

–

–

–

0.5/0.02

–

–

 pH

–

7.4

7.2

7

7.3

7.4

  Osmolality (mOs/L)

375

320

310

305

320

320

Electrolytes (MMOL/L)

Osmotic agents/colloids

  Buffers/AA (mmol/L)

Pharmacological agents

AA, amino acids; HES, hydroxyethyl starch; HTK, histidine-tryptophan-ketoglutarate; IGL-1, Institut Georges Lopez-1; UW, University of Wisconsin. Adapted from Parsons RF, Guarrera JV. Preservation solutions for static cold storage of abdominal allografts: which is best? Curr Opin Organ Transplant. 2014;19(2):100–107 and Fuller B, Froghi F, Davidson B. Organ preservation solutions: linking pharmacology to survival for the donor organ pathway. Curr Opin Organ Transplant. 2018;23(3):361–368.

B. Islet allo-transplantation

510

42.  Pancreas and islet preservation

which provides better osmotic properties to limit cellular edema. Although two small randomized clinical trials have shown comparable outcomes between Celsior and UW solution after pancreas transplantation,117,118 larger randomized, controlled clinical studies are needed to ensure that Celsior is as safe as UW solution. A preclinical pancreas transplantation study in pigs (n = 8) reported that IGL-1 was as safe as UW solution after 16 h of CIT.119 A small case series (n = 5) study of human pancreas transplantation reported exogeneous insulin independence, no DGF, no graft losses and no thrombosis in all 5 recipients of pancreata preserved with IGL-1.120 A retrospective analysis of several hundred islet isolations from pancreata flushed and transported with IGL1, UW or Celsior showed that the three solutions were equivalent when islet function was evaluated at 1 and 6-month posttransplantation follow-up.121 Therefore, HTK, Celsior and IGL-1, three alternative solutions to UW exhibit similar clinical outcomes and seem to be as safe as UW particularly in the setting of short cold ischemic periods (<10 h). Determining the extent to which preservation solutions prevent ionic and metabolic disturbances induced by hypoxia during hypothermia is a challenging undertaking in the clinical setting because of the costly, properly powered, complex clinical trials required to compare different formulations.122 Perfluorocarbons (PFCs) PFCs are hydrocarbon molecules in which hydrogen atoms are replaced by fluorine. PFC-containing solutions can dissolve 20–25 times more oxygen than plasma at room temperature95 and 10 times as much as water at 4°C.123 A low oxygen binding constant allows PFCs to release oxygen more effectively than hemoglobin.124 PFCs have been used as oxygen carriers to preserve pancreatic tissue by the two-layer method (TLM).123,124 Pancreatic grafts are placed in chambers containing PFCs and UW solution or other preservation solutions. Substrates are added to the UW solution prior to adding PFCs. The hydrophobicity and higher density of PFCs (lower layer) compared to UW solution results in the physical separation of both fluids. The pancreas floats at the interface between the layers. The TLM may presumably help provide ATP by direct phosphorylation of adenosine provided in the solution.125 Although PFCs have great potential as oxygen carriers126–128 and blood substitutes,129 their use in organ preservation solutions for organ immersion suffer from the same physical constraints as SCS using UW solution or any other medium alone (e.g., Euro-Collins solution). Nonetheless, increased islet recovery and function after transplantation using the TLM has been reported in rodent130 canine125 and human131 pancreata. The oxygenation effectiveness of the procedure is primarily driven by passive diffusion and it is thus highly dependent on the organ thickness.105 The

predicted oxygenated volume fraction of rat pancreas (thickness range: 1–4 mm) would be 60%–100% with the thinner segments of the head region (1–2 mm) reaching 100% oxygenation. In contrast, the oxygenated volume fraction of a 25-mm thick human pancreas would be less than 20% during similar preservation periods.104 The improved oxygenation and larger islet yields observed with the TLM method may be accounted for by a PFCmediated temporospatial decrease in oxygen gradients that facilitates oxygen diffusion and penetration. A one-layer method (OLM) whereby a PFC constitutes the preservation layer has been compared to the TLM in their ability to successfully achieve long-term preservation of porcine pancreata.132,133 In those studies, no differences were found regarding islet purity, viability and ATP content, and islet transplantation into diabetic nude mice demonstrated graft function in both groups (OLM vs TLM). To assess the effects of long-term storage (clinically relevant CIT) in PFC solutions, pig pancreata were immersed in preoxygenated perfluorohexyloctan (F6H8) or perfluorodecalin for 8–10 h and followed by pancreas digestion and islet isolation.134 Intrapancreatic pO2 and the ratio of ATP to inorganic phosphate (Pi) was compared by 31P-NMR spectroscopy prior to tissue digestion. Although islet yield was similar in both groups, GSIS, viability and survival were improved in the F6H8 group. Preoxygenated F6H8 pancreata exhibited a higher pO2 compared to pancreata stored in preoxygenated perfluorodecalin. These data suggest that in addition to the beneficial effects of PFDs on islet quality, differences between PFCs exist and require a more extensive assessment of other hyperoxygen carriers within the family of PFCs. The sole intraperitoneal administration of a PFC (perfluorodecalin) before intraportal delivery of islets in Lewis rats resulted in an increased pO2 in the portal vein and a significant increase in islet function postgrafting.135 Alternative novel formulations including perfluorodecalin-filled poly(n-butyl-cyanoacrylate)127 and albumin-derived PFC-based nanoparticles128 are being currently explored as artificial oxygen carriers. Additives Efforts to develop solutions containing cytoprotective additives and supplements that minimize the impact of postmortem ischemia on islet quality are an active research area. A large number of molecules have been tested over the past two decades and the subject has been reviewed extensively elsewhere.122,136 Preservation additives, supplements and pharmacological agents can have highly specific actions at the cellular and mitochondrial level (e.g., oxygen, NO), they can modify the expression of genes through gene therapy approaches (e.g., antiapoptotic genes) or affect critical functions and entire pathways (e.g., inflammation, apoptosis, coagulation/thrombosis) (Fig.  2). The large array of biological pathways affected by DWIT, CIT, and WIT provides

B. Islet allo-transplantation



Pancreas preservation prior to islet isolation

multiple opportunities for modulation via additives and pharmacological agents. Nonetheless, precise molecular and pathway signatures driven by I/R injury of changes that occur over time from organ procurement to engraftment are missing. Abdominal and thoracic organs used for transplantation have remarkable differences in function and metabolic needs (e.g., kidney vs liver vs heart vs pancreas), therefore, comparing performance and viability in the presence of different preservation solutions is typically performed for each organ to make decisions based on objective measurements.110–112,137–139 Running such comparisons in the presence and absence of additives and pharmacological agents are challenging and costly studies to design. Thus, decisions by the transplantation team on the use of additives are primarily based on the limited mechanistic evidence available in the setting of transplantation for the large number of agents. IGL-1, a preservation solution that has been recently introduced for clinical use, contains PEG (Table  1), which may reduce inflammation in the setting of I/R injury.140,141 Oxidative stress has been targeted by adding iron chelators (e.g., deferoxamine), n-acetyl histidine, and l-arginine as a NO substrate supply to HTK-TiProtec solution (Table  1). Addition of a caspase inhibitor, quinoline-Val-Asp-difluorophenoxymethylketone (Q-VD-OPH), and alpha tocopherol to Unisol and Belzer’s machine perfusion solutions showed a reduction in apoptosis.142 Intraductal l-glutamine administration combined with SCS in UW solution supplemented with l-glutamine resulted in improved posttransplant islet function in diabetic nude mice.143 Thus, additives to preservation solutions protect islet function during tissue disaggregation and islet isolation (see islet isolation section below). Addition of n-acetyl-cysteine, l-arginine, nitroglycerin, prostaglandin E1 (PGE1), and alpha-­ ketoglutarate to KPS-1 base solution generated Vasosol, which has been used with HMP.144,145 Using ­animal models of I/R injury, gene therapy approaches using siRNAs that have targeted the expression of caspase 3, receptors for endothelin—a potent vasoconstrictor—and p53, have successfully improved renal transplantation outcomes.146–150 Knocking down the expression of TNF-alpha, C3 and Fas genes with small interference RNAs (siRNAs) added to UW solution has been shown to protect cardiac function in a model of I/R injury.151 This and other delivery systems of siRNAs and micro-RNAs against other inflammatory molecules (e.g., TLR4, ICAM-1) that have shown to be protective against I/R injury in other organs152–154 should also be assayed in pancreatic models. The signaling functions of carbon monoxide (CO), NO, hydrogen sulfide (H2S) and their derived species have been investigated in a variety of biological settings.155–158 CO, a product of heme oxygenase-1 (HO-1)

511

has antiapoptotic and vasodilating properties. It reduces the levels of O2- and increase the generation of glutathione. Adding gaseous CO to preservation solutions has been shown to inhibit apoptosis and to protect against I/R injury.159,160 Increasing the production of CO by induction of HO-1 has been demonstrated to have cytoprotective effects in rodent lung, liver, heart, and intestine models of transplantation.161 NO is a potent vasodilator and antiinflammatory molecule that requires BH4 as a cofactor. Administration of either NO77,78 or BH479 has protective effects in pancreatic I/R injury. Pancreatic grafts missing the neuronal form of the nitric oxide synthase (nNOS) were shown to confer protection from disruption of the capillary network in the transplanted organ.162 Retrograde PSF supplemented with NO in a warm ischemia model of liver injury reduced oxidative stress and the extent of ROS-mediated damage during preservation with HTK solution.163 These results highlight the therapeutic potential of NO in pancreas and islet transplantation and the role of nNOS as a new therapeutic target to preserve vascular homeostasis during transplantation. H2S, a messenger generated by cystathionine gamma-lyase (CGL), cystathionine beta-synthase (CBS) and 3-mercaptopyruvate sulfurtransferase (3MST), has multiple cellular effectors; it possesses ROS scavenging properties and thus reduces oxidative stress. H2S also activates Nrf-2, a molecular switch that results in increased levels of glutathione, one of the most abundant antioxidants. Just as NO induces S-nitrosylation, H2S induces S-sulfhydration of intracellular proteins leading to a range of effects including vasodilation and antiinflammatory actions.155,157 The short half-life of these molecules (CO, NO, and H2S) and their potential toxicity at high concentrations could be circumvented by generating controlled release compounds with long half-lives that can be added to preservation solutions. Delivery of therapeutic gases, CO, NO, and H2S in addition to oxygen via controlled release agents would accelerate the translation of these gaseous cytoprotectants from the research laboratory to the clinical setting.

Dynamic methods/pancreas perfusion HMP, normothermic machine perfusion (NMP), and PSF comprise dynamic preservation approaches to organ preservation developed in response, at least partially, to the need for an increase in allowable organ preservation times, expansion of the donor pool by including “marginal” organs from DCDs and ECDs. Dynamic methods can be also categorized as liquid (HMP, NMP) and gaseous (PSF) perfusion procedures. Any of these methods and their variations are costlier and technically more demanding because surgical and perfusion procedures must be carefully performed before and after organ retrieval from the body. Perfusate composition, perfusion

B. Islet allo-transplantation

512

42.  Pancreas and islet preservation

pressure and flow rate must be cautiously determined. Nonetheless, the rewards of improved human clinical outcomes using dynamic methods over conventional static procedures —longer survival rates and improved quality of life—far exceed the technical challenges that the transplantation team must overcome. Just as static preservation procedures, dynamic perfusion methods are used to better maintain organ function for whole organ transplantation and to enhance islet yield, quality and function during isolation and purification for transplantation of islets in either hepatic or extra-hepatic sites. Liquid perfusion Hypothermic machine perfusion During HMP, cold preservation solution is recirculated through the organ’s main vessels in a continuous or pulsatile manner. Toxic metabolites (e.g., ROS) that accumulate during SCS are removed during HMP by a continuous flushing of the entire organ’s circulatory bed. The system provides the means to administer pharmacological agents as needed and HMP with oxygenated solutions supports ATP synthesis, promotes functional recovery and reduces enzyme leakage upon reperfusion.164 Compared to SCS, HMP preservation reduced to a greater degree the risk of DGF and improved 1- and 3-year graft survival in DBD and ECD kidneys.165–167 However, no improvements in the incidence of DGF between kidneys from HMP or SCS have also been reported.168 Development of HMP methods for pancreas transplantation remains a challenging undertaking.169 Intravascular whole organ perfusion with fluids mimicking physiological or supraphysiological levels of oxygen confers the advantage of a faster and more uniform distribution of oxygen within the organ. Early experimental data in dogs showed that pancreaticoduodenal grafts remained viable after 24 h of pulsatile HMP170 and more recent data confirmed the feasibility of using pulsatile HMP to preserve porcine pancreata.171 A small study where human pancreata were preserved by HMP and compared to SCS (n = 4 DCD + 4 DBD pancreata per group) showed increased levels of ATP in the HMP group.172 Increased levels of LDL, amylase and lipase in the preservation fluid was attributed to a potential “washing out” effect of degrading enzymes. The roadblocks ahead to develop specific HMP protocols for human pancreas seem to reside on the fragile pancreatic vasculature, the organ’s low blood flow and its proneness to edema when subjected to perfusion,173 which heightens the risk of thrombosis and graft failure.174 UW solution (KPS-1, Organ Recovery Systems, Chicago, Illinois) is the preferred solution used for HMP preservation for organ transplantation. Despite the moderate edema induced by 24 h of HMP with KPS-1 solution, yield and purity of islets was greater compared with pancreas subjected

to 24 h of SCS.175 Further exploration of HMP protocols are warranted to fully discern its usefulness in pancreas preservation and graft quality improvement. Normothermic machine perfusion Intuitively, preservation by cooling down of organs (~4°C) and perfusing them with liquid (HMP) or gaseous (PSF) solutions may seem more effective than perfusion at body temperature (37°C). However, normothermic (37°C) perfusion (NMP) with oxygenated autologous blood or an acellular perfusate confers the benefit of restoring oxidative metabolism and avoiding cold ischemia injury.96,176 The realization that the pancreas was particularly sensitive to blood perfusion changes was reported by Babkin and Starling in 1926 in their description of a canine pancreas perfusion model to study the effects of exogenous secretin on pancreatic blood flow.177 Using a perfusion apparatus and an oxygenator, Eckhauser et al. (1981), perfused canine pancreata with an autologous blood-perfusate mixture to assess pancreatic exocrine function. They concluded that oxygen consumption and vascular resistance were the most useful criteria to assess organ viability.178 Pancreata undergoing NMP have been used to assess graft viability,179 insulin secretion dynamics,180,181 and interspecies differences in insulin secretion.182 An ex vivo NMP has been successfully evaluated in human heart transplantation and represents a viable option for organs not only from DBDs but also from DCD donors.183 The clinical use of NMP to reduce ischemic injury has also been documented for kidney,176,184–186 liver,187 pancreas,185 and lung188 transplantation. Perfusion pressure and flow are critical elements of HMP and NMP in pancreas preservation. Because of the lower organ metabolic needs at low temperature, HMP studies report the use of much lower pressures compared to NMP (10–30 mmHg vs 75–110 mmHg, respecti vely).170,171,175,178,180,189–192 Pressures of 75–110 mmHg in pancreata undergoing NMP resulted in normal exocrine (e.g., amylase, lipase secretion) and endocrine (e.g., GSIS) function. Nonetheless, morphological evidence indicated moderate levels of edema after several hours of perfusion.178,193 The delicate nature of the pancreatic vasculature may require further NMP research to establish optimal ex vivo perfusate pressures and flow rates in different species. ex  vivo perfusion models in pigs have been proposed as reliable models to investigate machine perfusion preservation methods.194 One such model of NMP using autologous blood has been developed recently.195 The authors demonstrated exocrine (e.g., amylase and lipase secretion) and endocrine (e.g., GSIS) function in pancreata perfused at low pressure (20 mmHg). Low perfusion pressures may better recreate the physiological pressures that the pancreas is exposed to in  vivo and prevent endothelial

B. Islet allo-transplantation



Islet preservation during isolation and purification

and ­parenchymal damage.195 The use of autologous blood or red blood cells as the oxygen carrier is common in NMP. Packed red blood cells are sometimes combined with albumin—as an osmotic constituent to prevent edema formation—and electrolyte solutions.179,193,196–198 In short, Although NMP has been touted as a superior preservation method compared to SCS, the renewed interest in NMP warrants further studies to expand comparisons with other dynamic methods of pancreas preservation to objectively assess whether NMP is a better preservation method to protect the pancreas’ endocrine and exocrine functions for whole organ or islet transplantation. Gaseous perfusion Persufflation PSF is an organ preservation method initially explored in a series of cardiopulmonary bypass and neurobiology experiments in the mid and late 1950s.199–201 These and other studies encouraged investigators to study PSF as a preservation method for other organs including liver, kidney and pancreas.202 The simplicity of PSF as an oxygen delivery system and the experimental evidence indicating that PSF not only preserves organ function but may also rescue organs otherwise unusable for transplantation, has received renewed attention in recent years.202–207 PSF can be delivered through pancreatic arteries (anterograde PSF) or through the venous system (retrograde PSF). Although retrograde PSF in kidneys has been performed and reported to have shorter recovery times compared to anterograde PSF,208,209 no studies comparing both approaches have been conducted in pancreas. Nonetheless, using a rat model of DCD pancreas, Reddy et  al found that retrograde portal venous oxygen PSF was superior to SCS and HMP in islet yield, viability and function (GSIS) of isolated islets.207 A study in porcine pancreata preserved by either TLM or PSF for 24 h showed that persufflated pancreata exhibited improved tissue quality compared to TLM-treated pancreata,210 highlighting again the potential for PSF to increase islet yields isolated from human pancreata. Using a similar experimental design (TLM vs PSF), the bioenergetic status of pancreatic tissue was determined by measuring the ratio of adenosine triphosphate to inorganic phosphate (ATP:Pi) by 31P-magnetic resonance spectroscopy (31P-MRS). ATP levels were dramatically decreased in the TLM group, whereas the PSF group showed ATP levels similar to those observed in rat pancreata.206 Preservation of a metabolic status where higher levels of ATP are available may enable the use of marginal human donors and thus expand an important donor pool. Experiments conducted in rat livers showed that adding pulsatility to PSF improved parenchymal integrity, expression of vasoprotective genes (e.g., Kruppel-like factor 2) and increased levels of NO, all factors that may

513

promote early engraftment.204 Physiological circulatory patterns and levels of sheer stress provided by pulsatile PSF may ameliorate I/R-mediated damage to endothelial cells and protect pancreatic tissue and islets. PSF is a highly promising strategy for pancreas and islet preservation in the clinical setting. Compared to SCS, PSF of human pancreata has been recently shown to promote reduced expression levels of inflammation-related pathways, greater transcriptional responses of metabolic genes (e.g., pancreatic secretion, Na+/Cl+ transporters) and improved beta cell function (GSIS) in isolated islets in a cohort matched for total preservation time (Fig.  3, panels A–C).211 To better understand the differences between anterograde and retrograde PSF, additional studies may be needed to examine gene and protein expression of molecular biomarkers and signaling pathways of inflammation and oxidative stress in response to anterograde vs retrograde PSF.

Islet preservation during isolation and purification Microdissection was the first islet isolation technique essayed in pancreas from hyperglycemic mice.212 The low yield of hand-picking isolation was soon realized as a serious scalability limitation that would prevent translation to the clinical setting. Bacterial collagenase to separate exocrine from endocrine tissue was introduced by Moskalewski.213 Further improvements in the islet isolation technique included the intra-ductal injection of cold saline buffer to distend the pancreas followed by tissue mincing and enzymatic digestion.214,215 Intraductal delivery of collagenase instead of buffer facilitated tissue disaggregation while also distending the organ.216 Novel purification methods including islet separation by density gradients (Ficoll, Percoll albumin,) were developed.217–219 Investigators soon realized that for islet transplantation to succeed, isolation and purification should be considered two related yet independent sets of steps of a procedure aimed at maintaining cellular function.220 Improvements of experimental diabetes (partial and full reversal) were initially reported in the early 70s.221,222 However, human islet transplants faced setbacks when partially purified pancreatic preparations yielded poor clinical results in the first human trials.223,224 Focus on development of better islet isolation methods shifted toward identification of the ideal transplantation site after a report by Kemp in 1973 showed that islet transplantation into the liver was superior to subcutaneous or peritoneal delivery.225 A few years later, development of improved islet isolation methodologies refocused on mechanical (e.g., syringe passages, nylon mesh, stainless steel filters) and enzymatic (collagenase) tissue dissociation that resulted in higher islet recovery yields.215,226–228

B. Islet allo-transplantation

514

42.  Pancreas and islet preservation

FIG. 3  Functional measurements from islets isolated from human pancreata preserved on SCS and SCS+PSF. Data from panels (A–C) were

obtained in islets isolated from a cohort matched for total preservation time. (A) Total preservation time is depicted for SCS (n = 6; blue area = SCS) and SCS+PSF (n = 8; blue area = SCS, red area = PSF) groups. (B) GSIS rate was determined in islets isolated from SCS and SCS+PSF pancreata. GSIS rate was higher in SCS+PSF than in SCS islets. (C) First and second phase area under the curve of insulin secretion were higher for SCS+PSF compared to SCS. Data from islets isolated from a cohort matched for duration of SCS is presented in panels (D–F) (n = 6 for SCS islets, blue area = SCS; n = 5 for SCS+PSF islets, red area = PSF). No differences were observed in GSIS rate (E) or first and second phase area under the curve of insulin secretion between SCS and SCS+PSF in the cohort matched for SCS duration (F). ⁎ = P<.05, # = P<.05 indicates difference between total preservation time in cohort matched for duration of SCS (D). Abbreviations as in Fig. 2. Data from Kelly AC, Smith KE, Purvis WG, et al. Oxygen perfusion (persufflation) of human pancreata enhances insulin secretion and attenuates islet proinflammatory signaling. Transplantation 103 (1), 2019, 160–167.

Isolation of human islets The success of the Edmonton protocol, islet transplantation in conjunction with a steroid-free immunosuppressive regimen, triggered renewed interest in developing more efficient islet isolation methods for diabetes research and clinical use.229,230 Current knowledge of islet biology has primarily been derived from studies conducted mostly in murine islets.231–234 The interest in porcine islets resides primarily in pigs being a theoretically unlimited source of islets for xenotransplantation in T1D patients. Various factors including temperature, quality and concentration of dissociating enzymes, duration of pancreas dissociation, total isolation time, postdissociation purification methods and culture conditions, are all involved in obtaining a pure preparation of islets that is also viable and functional to be either transplanted to humans or used in animal or in vitro studies.215,220,234,235 Islet isolation methods from mammalian pancreatic

t­issue are relatively similar among a variety of species. However, differences in islet cytoarchitecture from one species to another may contribute to variations in islet yield and quality.31,234,236,237 Identification of the primary factors affecting yield, purity and viability of human islet preparations should aid in developing standardized methods aimed at improving quality and reducing variability between laboratories and transplantation centers around the world. Unlike rodent islets,236 human and nonhuman primate islets exhibit alpha (glucagon-producing), beta (insulin-producing) and delta (somatostatin-producing) cells randomly distributed throughout the islet microstructure and along the islet vasculature.31,237–239 Human islets contain fewer beta cells and more alpha cells compared to rodent islets.237 The relative proportion of beta cells within each islet, as well as regional differences in the number of islets and endocrine cells across the human pancreas should be taken into consideration during tissue procurement and islet isolation procedures as they

B. Islet allo-transplantation



Islet preservation during isolation and purification

may have implications for inadvertent enrichment of undesired cell types. Due to its embryological development from two distinct primordia, the human pancreas exhibits an uneven distribution of glucagon (alpha) and pancreatic polypeptide (PP) containing cells. PP cells are concentrated in the posterior region of the pancreatic head, which originates from the ventral primordium and also exhibits a low number of alpha cells.240

Pancreas distension, tissue dissociation, and islet collection Common steps to isolation procedures in all species include in situ/ex vivo distension and digestion of the pancreas via the common bile duct using an injection of a collagenase solution followed by ex vivo enzymatic digestion at 37°C once the pancreas is retrieved.216,231–233,241,242 Additional factors to consider during the enzymatic digestion and cell dissociation steps are the strength and concentration of the collagenase or the combination of enzymes used, the species, age and strain,243,244 as well as whether the incubation was static or dynamic. Dynamic incubation has been shown to improve porcine islet yield compared to a manual method.245 Better islet yields have also been reported when collagenase was administered in UW solution compared to Hanks balanced salt solution.246 Morphological screening of the pancreas before isolation, WIT <10 min and low endotoxin content (<30 EU/mg) in commercial batches of dissociating enzymes have all shown to influence porcine islet yield and viability.247 The most common enzyme used to dissociate pancreatic tissue is bacterial collagenase Clostridium histolyticum introduced in the mid 1960s to isolate guinea pig islets.213 Crude collagenase blends are composed of six collagenases, neutral proteases and other enzymes used to isolate rodent and human islets.248,249 Nonetheless, enzyme selection should be ideally guided by knowledge of the pancreatic, especially the peri-insular, composition of the extracellular matrix in the species studied.250 Collagenase II (collagenase H) has been shown to be critical to isolate rat islets,251 whereas combining collagenase I (collagenase G) and II resulted in faster tissue dissociation and higher islet yields.252–254 Unlike rat islet isolation, collagenase I seems to be essential in human islet isolation.255 In this same study, Barnett et al reported variation in enzyme potency, a critical factor in clinical islet transplantation within lots of commercial collagenase blends (Liberase HI). However, no differences in islet yields and purity were observed when human islets were isolated using a crude Sigma V collagenase batch and high-purity Serva NB1 collagenase.249 The discrepancies reported in the islet isolation literature warrant further and systematic analysis of the effectiveness of different commercially available enzyme blends

515

(e.g., Liberase MTF, SERVA, NEM)256 and purity grades (e.g., endotoxin content) in isolating islets from different species, particularly human islet isolation. It is also important to note that during intraductal and in  vitro digestion, pancreatic tissue is not being oxygenated and islets experience considerable hypoxic stress. Distention of the pancreas is achieved by hand distention or using a mechanical perfusion device. Separation of islets from acinar and connective tissue after pancreas distention and tissue mincing is aided by the Ricordi chamber, a tissue dissociation instrument introduced in 1988 that consists of stainless steel ball bearings, a closed circulation tubing system and a 500 μm mesh on the chamber’s lid.257 The chamber is operated at 37°C and manual or mechanical shaking can be performed to facilitate dislodging of acinar and insular tissue. Monitoring of islet release is important to avoid over digestion and islet fragmentation. Sampling and staining of islet’s zinc granules with dithizone aids in gauging the degree of separation of islets from acinar tissue.242,258,259 To stop the tissue digestion/dissociation process, diluting the preparation with fresh, cold solutions drastically reduces enzymatic activity and has the added benefit of also reducing islet’s metabolism. Following washing, the preparation is usually placed in a preservation solution (e.g., UW) and supplemented with albumin and heparin.234 Organ distention and connective tissue digestion are typically performed at temperatures gradients that range from 25°C (room temperature) to 37°C in the absence of external sources of oxygen, that is, in hypoxic conditions. Given the adverse role that hypoxia plays at impairing islet function and reducing islet viability,260 modified methods and improved technologies and devices designed to deliver oxygen during pancreatic enzymatic distention and islet disaggregation and collection may increase islet yield and quality.

Islet purification Although many density gradient variations and preservation solutions to isolate pancreatic islets have been used and evolved in the past three decades, currently utilized separation media revolve around solutions of high molecular weight (1 × 103 − 4 × 107 Da) sucrose polymers (e.g., Ficoll, Dextran, Biocoll).261–263 In addition to optimization of separation media, a significant advance in islet purification has been the development of semiautomated purification protocols using the IBM 2991 COBE cell separator.264–268 Even though no oxygenation is provided during separation, controlled low temperatures are maintained throughout the purification/­ centrifugation process in modified COBE separators264 to ensure a hypothermic environment and minimize temperature-induced stress and hypoxic damage to

B. Islet allo-transplantation

516

42.  Pancreas and islet preservation

i­slets. A clean cold room also contributes to the same goal. Purified fractions coming out of the COBE cell separator are sampled and stained with dithizone to assess islet yield and purity before islets are placed in culture or transplanted directly in hepatic or extra-hepatic sites.

exerted during pancreatic tissue dissociation and islet purification have been difficult to identify. Despite the evidence supporting transplantation of fresh islets, reports of long-term culture of islets indicate that expression of antigens in beta cells may be abrogated and may contribute to preventing islet rejection.283,284 A key advantage of islet culture is the selective depletion of acinar tissue and Islet preservation during culture and distribution passenger leukocytes. The development of islet culture methods to keep less immunogenic, more viable and Up to this point, islets have been isolated and puri- functional islets for several days/weeks before shipment fied by a process that involves highly trained personnel is of paramount importance for islet banking. Several to execute a series of rigorous steps in a clean room to studies have reported the ability of human and porcine provide high quality islets in sufficient numbers to im- islets in culture to reverse hyperglycemia in mice.285–288 prove the likelihood of the recipient attaining insulin-­ Culturing islets at low temperatures (22–24°C) reindependence shortly after transplantation. Many of the duces metabolism and may partially alleviate the deletechallenges of culturing and distributing a viable islet rious effects of hypoxia during culture at 37°C. Although product—a product that exhibits comparable quality most transplantation centers culture islets at 37°C for assessment scores before and after distribution are very 24–48 h before transplantation, it has been suggested similar. Islets should remain structurally and function- that human islets cultured at low temperatures exhibit ally intact after a standard culture period, during ship- prolonged survival in mouse xenograft transplantation ment and, upon arrival to destination, and right before models.84,286,289 Preservation of islet architecture and retransplantation. Hypoxia should be minimized by using duction of hypoxia-driven central necrosis have been bags and containers that enable oxygenation at high reported in human and rat islets cultured at 24°C for 1 islet seeding densities.269,270 A comprehensive analysis and 4 weeks.83,290 Nonetheless, low-temperature culture of clinical grade islet isolations performed in centers of pig islets has also been reported to damage islet moraround the globe showed that IEQs transplanted in the phology and decrease islet function and viability.277 As most recent year interval examined (2007–10) increased a result of a lower metabolic rate, islet culture at low over previous intervals (1999–2002, 2003–06) and it was temperatures also results in reduced insulin secretion independently associated with insulin independence af- in response to glucose,277,291 which may be restored to ter transplantation.271 Islet culture, banking and distri- physiological levels by re-setting temperature to 37°C. bution are three closely related areas that would benefit Although culture at low temperatures may not seem to from technological innovation in either area aimed at have an advantage over culture at 37°C as evaluated by avoiding hypoxic conditions and provide constant phys- islet yield, OCR and mRNA expression of biomarkers of iological levels of oxygen. stress (e.g., lymphokines, chemokines),292 some clinical transplant groups are trying to take advantage of the potential benefits provided by using both temperatures.293 Culture ware and culture methods Human islet culture at high seeding densities promotes Standard cell culture conditions in conventional plas- a hypoxic and proinflammatory microenvironment that tic ware (e.g., plastic T-flasks) are not optimal for pan- results in increased expression of proinflammatory cytocreatic islets—multicellular, spheroidal structures with kines, chemokines and hypoxia-response signaling.272,273 a complex 3D cytoarchitecture. High islet seeding den- Experimental data on central necrosis of large islets is sities in culture exacerbate hypoxic conditions, reduce in good agreement with computational models using a islet function and viability, and trigger inflammatory finite element method to simulate islet viability in static responses.272,273 Islet culture may constitute an ideal step and moving culture media.294 To minimize the damagfor islets to recover from damage inflicted during isola- ing effects of hypoxia, enhance survival and preserve tion. Survival of beta, alpha and endothelial cells within ­function, islets are cultured at low densities in convenislets could also be stimulated via oxygenation and using tional dishes and flasks (<1000 IEQ/mL culture mea variety of agents while in culture. However, culturing dia). This poses logistic obstacles for product handling, islets is a source of concern because some culture studies transportation and shipment. The use of commercially have reported reduced islet recovery and function, DNA available gas-permeable cell culture bags has practical damage, and cell death.274–279 Better engraftment and limitations (e.g., contamination, propensity to puncture) transplantation outcomes have also been reported with and do not provide enough oxygenation to islets.269 fresh islets compared to cultured ones in several spe- Alternative islet culture flasks with oxygen-permeable cies.280–282 The requirements for islets to survive in culture silicon rubber-based membranes able to deliver oxygen and recover from the mechanical and enzymatic damage simultaneously to islets seeded at 10–20-fold higher

B. Islet allo-transplantation



Islet preservation during culture and distribution

densities compared to conventional flasks has been considered as an alternative approach to prevent hypoxia in cultured and shipped islets.270 Islet culture bag systems295 and rotational cell culture systems296 have also been shown to increase cell viability and GSIS compared to conventional cell culture systems. Strategies aimed at preserving islet viability and function using high seeding density culture systems would enable transplantation centers to safely transport and ship human islets from one institution to another regardless of global location. Even though pancreatic islets and their rich network of capillaries undergo significant damage during isolation,277 there is limited information on co-culture studies with islets and endothelial cells to examine islet revascularization in vitro, islet engraftment and transplantation outcomes. Co-transplantation of endothelial progenitor cells and islets has been reported to induce normoglycemia in diabetic rats, to enhance graft neovascularization up to 6  months after transplantation,297 and to improve glycemic control of transplanted islets in a murine model.298 Advancements in 3D cell culture methods and knowledge on the role of extracellular matrix (ECM) proteins in the 3D structure of islets from different species31,236,237,239,299,300 may provide additional cues to develop and test biological scaffolds and a proangiogenic milieu to partially restore the native islet cytoarchitecture and repair its damaged capillary network.

517

Nacher et al have recently shown that HS confers higher in  vitro islet survival, viability and function compared to HSA. In the latter study, the in vitro benefits did not translate to the in  vivo setting when similar HSA- and HS-cultured islets were transplanted in diabetic nude mice.306 Insulin-like growth factors (IGF-1 and IGF-II), pancreatic duct conditioned media, transferrin and selenium have all been tested to improve islet recovery and viability.279,303 Isolated islets have been cultured in CMRL-1066 media (Mediatech, Hendon, VA), supplemented with albumin, insulin-transferrin-selenium (ITS) ciprofloxacin, HEPES and NaHCO3. Selenoproteins expressed by endothelial cells (e.g., glutathione peroxidases, thioredoxin reductases)307 in pancreatic islets or beta cells308 may prevent ROS-mediated injury to islets in culture. Media supplemented with ITS was found to exhibit improved viability and functional capacity compared to CMRL-1066 supplemented with FBS.278 The effect of other additives to ameliorate oxidative stress and cellular damage during islet culture has been tested. Among the most common ones are vitamin E (alpha-­ tocopherol), deferoxamine and nicotinamide.309–314 In combination with serum-free media, pyruvate, a product of intermediate metabolism and a mitochondrial substrate, improved survival of islet beta cells in culture.315 Pyruvate stimulates glucagon secretion. An elegant study has shown that the activation of beta cells by pyruvate suppresses the mitochondrial rise in Ca++ and glucagon secretion in alpha cells, an example of the Islet culture supplements paracrine activity of beta cells on alpha cells in response Research efforts have been dedicated to identifying the to a single agent.316 Stored in beta cell granules, Zn++, a right combination of islet culture medium, supplements key component of insulin synthesis, storage and release and oxygenation strategies to implement alternative may be also involved in pyruvate-mediated secretion of islet culture protocols that stimulate and preserve islet glucagon after stimulation of beta cells316 and in reducmass and function for longer periods during storage and ing apoptosis of islet cells and oxidative stress.317 As the transportation prior to transplantation.262,274,277,278,301,302 precursor of glutathione, a key reducing endogenous Development, validation and implementation of op- molecule, glutamine has been demonstrated to improve timized islet culture ware and the right combination viability of porcine islets in culture.318 Intraductal gluof culture media and supplements has the potential to tamine administration also reduced oxidative damage, drastically improve islet yield and function for days/ improved islet yield, viability, increased endogenous weeks while in culture. glutathione levels and improved outcomes in islet-­ The use of a serum-free, hormone- and factor-­ transplanted mice.319,320 Free fatty acids involvement in supplemented, defined medium has been shown to modulating and amplifying insulin production and sesupport cell culture and cultures of beta cells and cell cretion via increases in cytosolic free Ca++ has been well lines.303,304 Clinical use of human serum albumin (HSA) documented.321–324 Delivery of lipids and ATP to INS-1 is currently the accepted standard that replaced the use of beta cells using a fibronectin-mimetic peptide to facilfetal bovine serum (FBS) in islet research or human serum itate binding and internalization of liposomes showed (HS) in clinical transplantation to avoid the introduction increased cell survival.325 In sum, pyruvate, Zn++, glutaof xenogeneic material or pathogens, respectively. Since mine and FFAs may be essential elements in islet media islet preparations cultured in FBS exhibit higher viability culture to sustain islet function while in culture and afcompared to islets cultured in media supplemented with ter transplantation. Delivery of ATP, lipids and perhaps HSA,302 the quest for a modified culture media of supe- other key nutrients to cell in islets cultured in a hypoxic rior quality to HS and FBS to support clinical islet trans- environment may have an impact in the short- and longplantation continues. HSA has been reported to be more term to improve islet mass recovery and to preserve islet effective than HS to preserve human islets,305 however, function during isolation and after transplantation.

B. Islet allo-transplantation

518

42.  Pancreas and islet preservation

Cryopreservation Preserving the structural integrity and function of islets that endured the stresses of isolation and purification is central to islet transplantation. Cryopreserving high quality islets for extended periods would facilitate conducting quality control tests, pooling islets from different donors, and transport and shipping to remote locations to benefit a larger population of T1D diabetic patients. However, maintaining islet preparations at −80°C or lower temperatures and preserving functionality upon thawing also requires the use of cryoprotectants and precisely controlled freezing and thawing rates to prevent intracellular crystal formation and irreversible cellular damage.326 Furthermore, the 3D native structure of multicellular structures with 1000–10,000 cells and an average diameter of 150 μm poses specific biophysical heat and mass transport challenges to achieve uniform temperature changes across the islet’s spherical geometry.327 A thought-provoking recent study showed that dissociated and reaggregated islets remained viable and able to reverse diabetes after transplantation in rats.328 The authors dissociated islets into single cells329 followed by cryopreservation (at −80°C for 15 hours and storage at −196°C) and reaggregation into spheroids after thawing.330 Furthermore, the reaggregated human spheroids were more metabolically active than fresh cells and exhibited lower percentages of death cells than native islets. The study by Rawal et  al.328 is proof of the remarkable plasticity of islet cells to self-organize after disaggregation and form functional islet-like structures. However, the efficiency of the process needs to be considered given the potential for significant cell losses during the disaggregation/reaggregation process. Dimethyl sulfoxide (DMSO) was one of the first permeating cryoprotectants used for living cells and rat and human pancreatic islets.331–334 The first attempts at cryopreserving rat and porcine islets were partially successful but limited to small sample sizes (1–12).190,335,336 Providing islets enough time to equilibrate with DMSO at low temperatures (0–25°C) has been shown to be more relevant than the cooling rate.337–339 Over 50% of islets cooled at rates spanning over three orders of magnitude (0.3–1000°C/min) and warmed at the same rate (50°C/ min) survived and retained functionality.339 Vitrification, the glass-like phase where cryoprotectants permeate cells shielding the intracellular space while water freezes outside the cell, is also been explored to cryoprotect islets.340 Thawing vitrified or frozen islets is conducted at high rates (150–200°C/min) to prevent recrystallization. Permeating cryoprotectants (e.g., DMSO, ethylene glycol) are not devoid of toxic effects and must be replaced upon thawing islets with nonpermeating cryoprotectants (e.g., trehalose, raffinose, sucrose, dextran)341; typically, a 0.75 M solution of sucrose at low temperature is used.342 Other

s­ upplements and additives that have shown to have beneficial effects on islet survival are added either to the cryopreservation or thawing solutions and include metformin, gamma aminobutyric acid (GABA), docosahexanoic acid (DHA), and eicosapentaenoic acid (EPA).343–345 Optimization and validation of cryopreservation protocols would be invaluable to transplantation programs. Additional studies with whole islets and single cells from disaggregated islets are warranted to better document the recovery efficiency and quality of islets rendered by current and new methods in different species. Protocols able to cryopreserve islets for weeks or months are highly desirable as they provide a much-needed flexibility to: (1) islet banking, (2) pooling islets from different donors if needed, and (3) running a complete battery of quality assessment tests before transplantation.328

Outlook Although remarkable improvements have been made in islet isolation procedures over the past two decades,271 lack of uniform standards and high variability in isolation, enzymatic digestion and purification approaches, as well as limited implementation of quality assessment methods that better predict clinical outcomes, persist and remain as current and future challenges for researchers, clinicians and regulators alike. With nearly a million islets per human pancreas but up to 50% loss in islet mass and function during isolation,36–38,346,347 current strategies to achieve a 1:1 donor to recipient ratio in islet transplantation should emphasize the development of methods to reduce islet attrition rates during isolation, purification, culture and storage. Survival and preservation of alpha and beta cell function within islets separated from their native environment encompasses a complex procedural continuum that starts with procurement of a donor’s pancreas and ends with transplanting purified pancreatic islets into a recipient’s body. Along that continuum, discrete steps allow for multiple interventions to improve yield and quality of the final islet product to extend the recipient’s benefits; from improving donor selection, pancreas procurement, preservation and transportation methods to enhancing in vitro technologies for islet isolation, physiological culture conditions, and islet banking. Emphasis should be placed on developing alternative, less disruptive methods to enzymatic digestion and distention, as structural damage to the islet vasculature and 3D cytoarchitecture occurs mainly during these steps. The transplantation field would also vastly benefit from adopting current and novel methodologies for oxygenation of whole pancreata prior to isolation and islets during isolation, culture and shipment. The use of a validated set of organ and islet quality assessments (OQA, IQA, Figs. 1 and 2) executed

B. Islet allo-transplantation



References

during whole pancreas preservation, after islet isolation and culture, and before islet distribution would be a great step toward standardization to generate more accurate and reproducible data within and across institutions. The islet transplantation field is accelerating the transition from a therapeutic approach that is currently focused on a subpopulation of few selected T1D patients and involves a high donor to recipient ratio to treating larger typical T1D patient populations using islets from single donors. The generation of human stem-cell derived islets that can be expanded in the lab in unlimited quantities, and the development of approaches (such as cell encapsulation) that eliminate the need for immunosuppression may in the near future result in a truly effective and scalable treatment for insulin dependent patients including children.40

References 1. Golden  SH, Sapir  T. Methods for insulin delivery and glucose monitoring in diabetes: summary of a comparative effectiveness review. J Manag Care Pharm. 2012;18(6 Suppl):S1–17. 2. Yardley JE, Iscoe KE, Sigal RJ, Kenny GP, Perkins BA, Riddell MC. Insulin pump therapy is associated with less post-exercise hyperglycemia than multiple daily injections: an observational study of physically active type 1 diabetes patients. Diabetes Technol Ther. 2013;15(1):84–88. 3. Qureshi MS, Callaghan CJ, Bradley JA, Watson CJ, Pettigrew GJ. Outcomes of simultaneous pancreas-kidney transplantation from brain-dead and controlled circulatory death donors. Br J Surg. 2012;99(6):831–838. 4. Gruessner AC, Gruessner RWG. Pancreas transplantation of US and Non-US cases from 2005 to 2014 as reported to the United Network for Organ Sharing (UNOS) and the International Pancreas Transplant Registry (IPTR). Rev Diabet Stud. 2016;13(1):35–38. 5. Gruessner RW, Sutherland DE, Kandaswamy R, Gruessner AC. Over 500 solitary pancreas transplants in nonuremic patients with brittle diabetes mellitus. Transplantation. 2008;85(1):42–47. 6. Stites  E, Kennealey  P, Wiseman  AC. Current status of pancreas transplantation. Curr Opin Nephrol Hypertens. 2016;25(6):563–569. 7. Salvalaggio  PR, Davies  DB, Fernandez  LA, Kaufman  DB. Outcomes of pancreas transplantation in the United States using cardiac-death donors. Am J Transplant. 2006;6(5 Pt 1):1059–1065. 8. Fernandez  LA, Di Carlo  A, Odorico  JS, et  al. Simultaneous ­pancreas-kidney transplantation from donation after cardiac death: successful long-term outcomes. Ann Surg. 2005;242(5):716–723. 9. D’Amour  KA, Bang  AG, Eliazer  S, et  al. Production of pancreatic hormone-expressing endocrine cells from human embryonic stem cells. Nat Biotechnol. 2006;24(11):1392–1401. 10. Matveyenko AV, Georgia S, Bhushan A, Butler PC. Inconsistent formation and nonfunction of insulin-positive cells from pancreatic endoderm derived from human embryonic stem cells in athymic nude rats. Am J Physiol Endocrinol Metab. 2010;299(5):E713–E720. 11. Rezania A, Bruin JE, Riedel MJ, et al. Maturation of human embryonic stem cell-derived pancreatic progenitors into functional islets capable of treating pre-existing diabetes in mice. Diabetes. 2012;61(8):2016–2029. 12. Rezania  A, Bruin  JE, Arora  P, et  al. Reversal of diabetes with ­insulin-producing cells derived in vitro from human pluripotent stem cells. Nat Biotechnol. 2014;32(11):1121–1133. 13. Bruin JE, Rezania A, Xu J, et al. Maturation and function of human embryonic stem cell-derived pancreatic progenitors in

519

­ acroencapsulation devices following transplant into mice. m Diabetologia. 2013;56(9):1987–1998. 14. Kirk K, Hao E, Lahmy R, Itkin-Ansari P. Human embryonic stem cell derived islet progenitors mature inside an encapsulation device without evidence of increased biomass or cell escape. Stem Cell Res. 2014;12(3):807–814. 15. Motte  E, Szepessy  E, Suenens  K, et  al. Composition and function of macroencapsulated human embryonic stem cell-derived implants: comparison with clinical human islet cell grafts. Am J Physiol Endocrinol Metab. 2014;307(9):E838–E846. 16. Pepper AR, Pawlick R, Bruni A, et al. Transplantation of human pancreatic endoderm cells reverses diabetes post transplantation in a prevascularized subcutaneous site. Stem Cell Reports. 2017;8(6):1689–1700. 17. Pagliuca FW, Millman JR, Gurtler M, et al. Generation of functional human pancreatic beta cells in vitro. Cell. 2014;159(2):428–439. 18. Shapiro  AM, Lakey  JR, Ryan  EA, et  al. Islet transplantation in seven patients with type 1 diabetes mellitus using a ­glucocorticoid-free immunosuppressive regimen. N Engl J Med. 2000;343(4):230–238. 19. Rickels MR, Stock PG, de Koning EJP, et al. Defining outcomes for beta-cell replacement therapy in the treatment of diabetes: a consensus report on the Igls criteria from the IPITA/EPITA opinion leaders Workshop. Transplantation. 2018;102:1479–1486. 20. Foster ED, Bridges ND, Feurer ID, et al. Improved health-­related quality of life in a phase 3 islet transplantation trial in type 1 diabetes complicated by severe hypoglycemia. Diabetes Care. 2018;41(5):1001–1008. 21. Maffi  P, Scavini  M, Socci  C, et  al. Risks and benefits of transplantation in the cure of type 1 diabetes: whole pancreas versus islet transplantation. A single center study. Rev Diabet Stud. 2011;8(1):44–50. 22. Ryan  EA, Paty  BW, Senior  PA, et  al. Five-year follow-up after clinical islet transplantation. Diabetes. 2005;54(7):2060–2069. 23. Rickels MR, Liu C, Shlansky-Goldberg RD, et al. Improvement in beta-cell secretory capacity after human islet transplantation according to the CIT07 protocol. Diabetes. 2013;62(8):2890–2897. 24. Lehmann  R, Weber  M, Berthold  P, et  al. Successful simultaneous islet-kidney transplantation using a steroid-free immunosuppression: two-year follow-up. Am J Transplant. 2004;4(7):1117–1123. 25. Barton FB, Rickels MR, Alejandro R, et al. Improvement in outcomes of clinical islet transplantation: 1999-2010. Diabetes Care. 2012;35(7):1436–1445. 26. Bellin  MD, Barton  FB, Heitman  A, et  al. Potent induction immunotherapy promotes long-term insulin independence after islet transplantation in type 1 diabetes. Am J Transplant. 2012;12(6):1576–1583. 27. Rickels  MR, Peleckis  AJ, Markmann  E, et  al. Long-term improvement in glucose control and counterregulation by islet transplantation for type 1 diabetes. J Clin Endocrinol Metab. 2016;101(11):4421–4430. 28. Hering BJ, Clarke WR, Bridges ND, et al. Phase 3 trial of transplantation of human islets in type 1 diabetes complicated by severe hypoglycemia. Diabetes Care. 2016;39(7):1230–1240. 29. Bonner-Weir  S, Orci  L. New perspectives on the microvasculature of the islets of Langerhans in the rat. Diabetes. 1982;31(10):883–889. 30. Jansson L, Hellerstrom C. Stimulation by glucose of the blood flow to the pancreatic islets of the rat. Diabetologia. 1983;25(1):45–50. 31. Arrojo e Drigo  R, Ali  Y, Diez  J, Srinivasan  DK, Berggren  PO, Boehm  BO. New insights into the architecture of the islet of Langerhans: a focused cross-species assessment. Diabetologia. 2015;58(10):2218–2228.

B. Islet allo-transplantation

520

42.  Pancreas and islet preservation

32. Elayat  AA, el-Naggar  MM, Tahir  M. An immunocytochemical and morphometric study of the rat pancreatic islets. J Anat. 1995;186(Pt 3):629–637. 33. Benjamin JS, Culpepper CB, Brown LD, et al. Chronic anemic hypoxemia attenuates glucose-stimulated insulin secretion in fetal sheep. Am J Physiol Regul Integr Comp Physiol. 2017;312(4):R492–R500. 34. Cantley  J, Grey  ST, Maxwell  PH, Withers  DJ. The hypoxia response pathway and beta-cell function. Diabetes Obes Metab. 2010;12(Suppl 2):159–167. 35. Cheng K, Ho K, Stokes R, et al. Hypoxia-inducible factor-1alpha regulates beta cell function in mouse and human islets. J Clin Invest. 2010;120(6):2171–2183. 36. Sabek  OM, Fraga  DW, Minoru  O, McClaren  JL, Gaber  AO. Assessment of human islet viability using various mouse models. Transplant Proc. 2005;37(8):3415–3416. 37. Plesner A, Verchere CB. Advances and challenges in islet transplantation: islet procurement rates and lessons learned from suboptimal islet transplantation. J Transplant. 2011;2011:979527. 38. Smith RM, Gale EA. Survival of the fittest? Natural selection in islet transplantation. Transplantation. 2005;79(10):1301–1303. 39. Komatsu H, Cook C, Wang CH, et al. Oxygen environment and islet size are the primary limiting factors of isolated pancreatic islet survival. PLoS One. 2017;12(8):e0183780. 40. Papas  KK, Avgoustiniatos  ES, Suszynski  TM. Effect of oxygen supply on the size of implantable islet-containing encapsulation devices. Panminerva Med. 2016;58(1):72–77. 41. Lehmann  R, Zuellig  RA, Kugelmeier  P, et  al. Superiority of small islets in human islet transplantation. Diabetes. 2007;56(3):594–603. 42. Hong  JC, Yersiz  H, Kositamongkol  P, et  al. Liver transplantation using organ donation after cardiac death: a clinical predictive index for graft failure-free survival. Arch Surg. 2011;146(9):1017–1023. 43. Halazun KJ, Al-Mukhtar A, Aldouri A, Willis S, Ahmad N. Warm ischemia in transplantation: search for a consensus definition. Transplant Proc. 2007;39(5):1329–1331. 44. Manara AR, Murphy PG, O’Callaghan G. Donation after circulatory death. Br J Anaesth. 2012;108(suppl_1):i108–i121. 45. Andres A, Kin T, O’Gorman D, et al. Clinical islet isolation and transplantation outcomes with deceased cardiac death donors are similar to neurological determination of death donors. Transpl Int. 2016;29(1):34–40. 46. Anazawa T, Saito T, Goto M, et al. Long-term outcomes of clinical transplantation of pancreatic islets with uncontrolled donors after cardiac death: a multicenter experience in Japan. Transplant Proc. 2014;46(6):1980–1984. 47. Zhao M, Muiesan P, Amiel SA, et al. Human islets derived from donors after cardiac death are fully biofunctional. Am J Transplant. 2007;7(10):2318–2325. 48. Markmann  JF, Deng  S, Desai  NM, et  al. The use of non-heartbeating donors for isolated pancreatic islet transplantation. Transplantation. 2003;75(9):1423–1429. 49. Maglione M, Ploeg RJ, Friend PJ. Donor risk factors, retrieval technique, preservation and ischemia/reperfusion injury in pancreas transplantation. Curr Opin Organ Transplant. 2013;18(1):83–88. 50. Hochachka  PW. Defense strategies against hypoxia and hypothermia. Science. 1986;231(4735):234–241. 51. Padanilam  BJ. Cell death induced by acute renal injury: a perspective on the contributions of apoptosis and necrosis. Am J Physiol Renal Physiol. 2003;284(4):F608–F627. 52. Bilzer M, Gerbes AL. Preservation injury of the liver: mechanisms and novel therapeutic strategies. J Hepatol. 2000;32(3):508–515. 53. Kohli V, Madden JF, Bentley RC, Clavien PA. Calpain mediates ischemic injury of the liver through modulation of apoptosis and necrosis. Gastroenterology. 1999;116(1):168–178.

54. Kohli V, Gao W, Camargo Jr CA, Clavien PA. Calpain is a mediator of preservation-reperfusion injury in rat liver transplantation. Proc Natl Acad Sci U S A. 1997;94(17):9354–9359. 55. Boutilier RG. Mechanisms of cell survival in hypoxia and hypothermia. J Exp Biol. 2001;204(Pt 18):3171–3181. 56. Belzer FO, Southard JH. Principles of solid-organ preservation by cold storage. Transplantation. 1988;45(4):673–676. 57. Kribben A, Wieder ED, Wetzels JF, et al. Evidence for role of cytosolic free calcium in hypoxia-induced proximal tubule injury. J Clin Invest. 1994;93(5):1922–1929. 58. Paffett ML, Walker BR. Vascular adaptations to hypoxia: molecular and cellular mechanisms regulating vascular tone. Essays Biochem. 2007;43:105–119. 59. Piper HM, Garcia-Dorado D. Prime causes of rapid cardiomyocyte death during reperfusion. Ann Thorac Surg. 1999;68(5):1913–1919. 60. Rosser BG, Gores GJ. Liver cell necrosis: cellular mechanisms and clinical implications. Gastroenterology. 1995;108(1):252–275. 61. Frank A, Rauen U, de Groot H. Protection by glycine against hypoxic injury of rat hepatocytes: inhibition of ion fluxes through nonspecific leaks. J Hepatol. 2000;32(1):58–66. 62. Fiegen  RJ, Rauen  U, Hartmann  M, Decking  UK, de Groot  H. Decrease of ischemic injury to the isolated perfused rat liver by loop diuretics. Hepatology. 1997;25(6):1425–1431. 63. Carini R, Bellomo G, Benedetti A, et al. Alteration of Na+ homeostasis as a critical step in the development of irreversible hepatocyte injury after adenosine triphosphate depletion. Hepatology. 1995;21(4):1089–1098. 64. Kalogeris T, Baines CP, Krenz M, Korthuis RJ. Cell biology of ischemia/reperfusion injury. Int Rev Cell Mol Biol. 2012;298:229–317. 65. Gross  GJ, Auchampach  JA. Reperfusion injury: does it exist? J Mol Cell Cardiol. 2007;42(1):12–18. 66. Halliwell  B, Gutteridge  JM. Role of free radicals and catalytic metal ions in human disease: an overview. Methods Enzymol. 1990;186:1–85. 67. Francis  A, Baynosa  R. Ischaemia-reperfusion injury and hyperbaric oxygen pathways: a review of cellular mechanisms. Diving Hyperb Med. 2017;47(2):110–117. 68. Li  C, Jackson  RM. Reactive species mechanisms of cellular hypoxia-reoxygenation injury. Am J Physiol Cell Physiol. 2002;282(2):C227–C241. 69. Rauen  U, de Groot  H. New insights into the cellular and molecular mechanisms of cold storage injury. J Invest Med. 2004;52(5):299–309. 70. Fernandez-del Castillo C, Harringer W, Warshaw AL, et al. Risk factors for pancreatic cellular injury after cardiopulmonary bypass. N Engl J Med. 1991;325(6):382–387. 71. Warshaw AL, O’Hara PJ. Susceptibility of the pancreas to ischemic injury in shock. Ann Surg. 1978;188(2):197–201. 72. Nys  M, Venneman  I, Deby-Dupont  G, et  al. Pancreatic cellular injury after cardiac surgery with cardiopulmonary bypass: frequency, time course and risk factors. Shock. 2007;27(5):474–481. 73. Benz S, Bergt S, Obermaier R, et al. Impairment of microcirculation in the early reperfusion period predicts the degree of graft pancreatitis in clinical pancreas transplantation. Transplantation. 2001;71(6):759–763. 74. Menger  MD, Bonkhoff  H, Vollmar  B. Ischemia-reperfusioninduced pancreatic microvascular injury. An intravital fluorescence microscopic study in rats. Dig Dis Sci. 1996;41(5):823–830. 75. Menger  MD. Microcirculatory disturbances secondary to ­ischemia-reperfusion. Transplant Proc. 1995;27(5):2863–2865. 76. Hoffmann  TF, Leiderer  R, Waldner  H, Arbogast  S, Messmer  K. Ischemia reperfusion of the pancreas: a new in  vivo model for acute pancreatitis in rats. Res Exp Med. 1995;195(3):125–144. 77. Benz  S, Obermaier  R, Wiessner  R, et  al. Effect of nitric oxide in ischemia/reperfusion of the pancreas. J Surg Res. 2002;106(1):46–53.

B. Islet allo-transplantation



References

78. Hegyi  P, Rakonczay Jr Z. The role of nitric oxide in the physiology and pathophysiology of the exocrine pancreas. Antioxid Redox Signal. 2011;15(10):2723–2741. 79. Maglione  M, Oberhuber  R, Cardini  B, et  al. Donor pretreatment with tetrahydrobiopterin saves pancreatic isografts from ischemia reperfusion injury in a mouse model. Am J Transplant. 2010;10(10):2231–2240. 80. Hackert  T, Werner  J, Uhl  W, Gebhard  MM, Buchler  MW, Schmidt  J. Reduction of ischemia/reperfusion injury by antithrombin III after experimental pancreas transplantation. Am J Surg. 2005;189(1):92–97. 81. Preissler  G, Eichhorn  M, Waldner  H, Winter  H, Kleespies  A, Massberg S. Intercellular adhesion molecule-1 blockade attenuates inflammatory response and improves microvascular perfusion in rat pancreas grafts. Pancreas. 2012;41(7):1112–1118. 82. Gaber AO, Mulgaonkar S, Kahan BD, et al. YSPSL (rPSGL-Ig) for improvement of early renal allograft function: a double-blind, placebo-controlled, multi-center Phase IIa study. Clin Transpl. 2011;25(4):523–533. 83. Scharp DW, Lacy PE, Finke E, Olack B. Low-temperature culture of human islets isolated by the distention method and purified with Ficoll or Percoll gradients. Surgery. 1987;102(5):869–879. 84. Ricordi C, Lacy PE, Sterbenz K, Davie JM. Low-temperature culture of human islets or in vivo treatment with L3T4 antibody produces a marked prolongation of islet human-to-mouse xenograft survival. Proc Natl Acad Sci U S A. 1987;84(22):8080–8084. 85. Pae EK, Ahuja B, Kim M, Kim G. Impaired glucose homeostasis after a transient intermittent hypoxic exposure in neonatal rats. Biochem Biophys Res Commun. 2013;441(3):637–642. 86. Maillard E, Juszczak MT, Langlois A, et al. Perfluorocarbon emulsions prevent hypoxia of pancreatic beta-cells. Cell Transplant. 2012;21(4):657–669. 87. Cantley J, Walters SN, Jung MH, et al. A preexistent hypoxic gene signature predicts impaired islet graft function and glucose homeostasis. Cell Transplant. 2013;22(11):2147–2159. 88. Moritz W, Meier F, Stroka DM, et al. Apoptosis in hypoxic human pancreatic islets correlates with HIF-1alpha expression. FASEB J. 2002;16(7):745–747. 89. Marshall D, Sabek O, Fraga D, Kotb M, Gaber AO. Examination of the molecular signature associated with islet dysfunction. Transplant Proc. 2005;37(2):1311–1312. 90. Sato  Y, Inoue  M, Yoshizawa  T, Yamagata  K. Moderate hypoxia induces beta-cell dysfunction with HIF-1-independent gene expression changes. PLoS One. 2014;9(12):e114868. 91. Bensellam M, Maxwell EL, Chan JY, et al. Hypoxia reduces ERto-Golgi protein trafficking and increases cell death by inhibiting the adaptive unfolded protein response in mouse beta cells. Diabetologia. 2016;59(7):1492–1502. 92. Li  YV. Zinc and insulin in pancreatic beta-cells. Endocrine. 2014;45(2):178–189. 93. Pae EK, Kim G. Insulin production hampered by intermittent hypoxia via impaired zinc homeostasis. PLoS One. 2014;9(2):e90192. 94. Parsons  RF, Guarrera  JV. Preservation solutions for static cold storage of abdominal allografts: which is best? Curr Opin Organ Transplant. 2014;19(2):100–107. 95. McLaren AJ, Friend PJ. Trends in organ preservation. Transpl Int. 2003;16(10):701–708. 96. Barlow AD, Hosgood SA, Nicholson ML. Current state of pancreas preservation and implications for DCD pancreas transplantation. Transplantation. 2013;95(12):1419–1424. 97. Dholakia  S, Royston  E, Sharples  EJ, Sankaran  V, Ploeg  RJ, Friend PJ. Preserving and perfusing the allograft pancreas: past, present, and future. Transplant Rev (Orlando). 2018;32(3):127–131. 98. Guibert EE, Petrenko AY, Balaban CL, Somov AY, Rodriguez JV, Fuller BJ. Organ preservation: current concepts and new strategies for the next decade. Transfus Med Hemother. 2011;38(2):125–142.

521

99. Mitchell T, Saba H, Laakman J, Parajuli N, MacMillan-Crow LA. Role of mitochondrial-derived oxidants in renal tubular cell cold-storage injury. Free Radic Biol Med. 2010;49(8):1273–1282. 100. Brinkkoetter PT, Song H, Losel R, et al. Hypothermic injury: the mitochondrial calcium, ATP and ROS love-hate triangle out of balance. Cell Physiol Biochem. 2008;22(1-4):195–204. 101. Porte RJ, Ploeg RJ, Hansen B, et al. Long-term graft survival after liver transplantation in the UW era: late effects of cold ischemia and primary dysfunction. European Multicentre Study Group. Transpl Int. 1998;11(Suppl 1):S164–S167. 102. Morris PJ, Johnson RJ, Fuggle SV, Belger MA, Briggs JD. Analysis of factors that affect outcome of primary cadaveric renal transplantation in the UK. HLA Task Force of the Kidney Advisory Group of the United Kingdom Transplant Support Service Authority (UKTSSA). Lancet. 1999;354(9185):1147–1152. 103. Goto M, Imura T, Inagaki A, et al. The impact of ischemic stress on the quality of isolated pancreatic islets. Transplant Proc. 2010;42(6):2040–2042. 104. Papas  KK, Hering  BJ, Guenther  L, Rappel  MJ, Colton  CK, Avgoustiniatos ES. Pancreas oxygenation is limited during preservation with the two-layer method. Transplant Proc. 2005;37(8):3501–3504. 105. Avgoustiniatos  ES, Hering  BJ, Papas  KK. The rat pancreas is not an appropriate model for testing the preservation of the human pancreas with the two-layer method. Transplantation. 2006;81(10):1471–1472. author reply 1472. 106. Baertschiger  RM, Berney  T, Morel  P. Organ preservation in pancreas and islet transplantation. Curr Opin Organ Transplant. 2008;13(1):59–66. 107. Collins GM, Bravo-Shugarman M, Terasaki PI. Kidney preservation for transportation. Initial perfusion and 30 hours’ ice storage. Lancet. 1969;2(7632):1219–1222. 108. D’Alessandro  AM, Stratta  RJ, Sollinger  HW, Kalayoglu  M, Pirsch JD, Belzer FO. Use of UW solution in pancreas transplantation. Diabetes. 1989;38(Suppl 1):7–9. 109. Pokorny  H, Rasoul-Rockenschaub  S, Langer  F, et  al. ­Histidine-tryptophan-ketoglutarate solution for organ preservation in human liver transplantation—a prospective multi-centre observation study. Transpl Int. 2004;17(5):256–260. 110. Schneeberger  S, Biebl  M, Steurer  W, et  al. A prospective randomized multicenter trial comparing histidine-tryptophane-­ ketoglutarate versus University of Wisconsin perfusion solution in clinical pancreas transplantation. Transpl Int. 2009;22(2):217–224. 111. Englesbe MJ, Moyer A, Kim DY, et al. Early pancreas transplant outcomes with histidine-tryptophan-ketoglutarate preservation: a multicenter study. Transplantation. 2006;82(1):136–139. 112. Fridell  JA, Agarwal  A, Milgrom  ML, Goggins  WC, Murdock  P, Pescovitz MD. Comparison of histidine-tryptophan-­ketoglutarate solution and University of Wisconsin solution for organ preservation in clinical pancreas transplantation. Transplantation. 2004;77(8):1304–1306. 113. Stewart  ZA, Cameron  AM, Singer  AL, Dagher  NN, Montgomery RA, Segev DL. Histidine-tryptophan-ketoglutarate (HTK) is associated with reduced graft survival in pancreas transplantation. Am J Transplant. 2009;9(1):217–221. 114. Alonso D, Dunn TB, Rigley T, et al. Increased pancreatitis in allografts flushed with histidine-tryptophan-ketoglutarate solution: a cautionary tale. Am J Transplant. 2008;8(9):1942–1945. 115. Paushter DH, Qi M, Danielson KK, et al. Histidine-tryptophanketoglutarate and University of Wisconsin solution demonstrate equal effectiveness in the preservation of human pancreata intended for islet isolation: a large-scale, single-center experience. Cell Transplant. 2013;22(7):1113–1121. 116. Salehi P, Hansen MA, Avila JG, et al. Human islet isolation outcomes from pancreata preserved with Histidine-Tryptophan Ketoglutarate versus University of Wisconsin solution. Transplantation. 2006;82(7):983–985.

B. Islet allo-transplantation

522

42.  Pancreas and islet preservation

117. Boggi U, Vistoli F, Del Chiaro M, et al. Pancreas preservation with University of Wisconsin and Celsior solutions: a s­ingle-center, prospective, randomized pilot study. Transplantation. 2004;77(8):1186–1190. 118. Nicoluzzi  J, Macri  M, Fukushima  J, Pereira  A. Celsior versus Wisconsin solution in pancreas transplantation. Transplant Proc. 2008;40(10):3305–3307. 119. Garcia-Gil FA, Fuentes-Broto L, Albendea CD, et al. Evaluation of Institut Georges Lopez-1 preservation solution in pig pancreas transplantation: a pilot study. Transplantation. 2014;97(9):901–907. 120. Chedid  MF, Grezzana-Filho  TJ, Montenegro  RM, et  al. First report of human pancreas transplantation using IGL-1 preservation solution: a case series. Transplantation. 2016;100(9):e46–e47. 121. Niclauss N, Wojtusciszyn A, Morel P, et al. Comparative impact on islet isolation and transplant outcome of the preservation solutions Institut Georges Lopez-1, University of Wisconsin, and Celsior. Transplantation. 2012;93(7):703–708. 122. Saint Yves T, Delpech PO, Giraud S, Thuillier R, Hauet T. Additives to preservation solutions. Prog Urol. 2014;24(Suppl 1):S31–S36. 123. Kuroda  Y, Kawamura  T, Suzuki  Y, Fujiwara  H, Yamamoto  K, Saitoh Y. A new, simple method for cold storage of the pancreas using perfluorochemical. Transplantation. 1988;46(3):457–460. 124. Matsumoto S, Kuroda Y. Perfluorocarbon for organ preservation before transplantation. Transplantation. 2002;74(12):1804–1809. 125. Kuroda Y, Hiraoka K, Tanioka Y, et al. Role of adenosine in preservation by the two-layer method of ischemically damaged canine pancreas. Transplantation. 1994;57(7):1017–1020. 126. Arnaud F, Sanders K, Sieckmann D, Moon-Massat P. In vitro alteration of hematological parameters and blood viscosity by the perfluorocarbon: oxycyte. Int J Hematol. 2016;103(5):584–591. 127. Laudien  J, Gross-Heitfeld  C, Mayer  C, de Groot  H, Kirsch  M, Ferenz  KB. Perfluorodecalin-filled poly(n-butyl-­ cyanoacrylate) nanocapsules as potential artificial oxygen carriers: preclinical safety and biocompatibility. J Nanosci Nanotechnol. 2015;15(8):5637–5648. 128. Wrobeln  A, Schluter  KD, Linders  J, et  al. Functionality of albumin-derived perfluorocarbon-based artificial oxygen car­ riers in the Langendorff-heart (dagger). Artif Cells Nanomed Biotechnol. 2017;45(4):723–730. 129. Waxman  K. Perfluorocarbons as blood substitutes. Ann Emerg Med. 1986;15(12):1423–1424. 130. Tanaka T, Suzuki Y, Tanioka Y, et al. Possibility of islet transplantation from a nonheartbeating donor pancreas resuscitated by the two-layer method. Transplantation. 2005;80(6):738–742. 131. Lakey  JR, Tsujimura  T, Shapiro  AM, Kuroda  Y. Preservation of the human pancreas before islet isolation using a two-layer (UW solution-perfluorochemical) cold storage method. Transplantation. 2002;74(12):1809–1811. 132. Brandhorst D, Iken M, Brendel MD, Bretzel RG, Brandhorst H. Long-term preservation of the pig pancreas by a onelayer method for successful islet isolation. Transplant Proc. 2005;37(1):229–230. 133. Brandhorst  D, Iken  M, Brendel  MD, Bretzel  RG, Brandhorst  H. Successful pancreas preservation by a perfluorocarbon-based onelayer method for subsequent pig islet isolation. Transplantation. 2005;79(4):433–437. 134. Brandhorst H, Iken M, Scott 3rd WE, et al. Quality of isolated pig islets is improved using perfluorohexyloctane for pancreas storage in a split lobe model. Cell Transplant. 2013;22(8):1477–1483. 135. Sakai T, Li S, Kuroda Y, Tanioka Y, Fujino Y, Suzuki Y. Oxygenation of the portal vein by intraperitoneal administration of oxygenated perfluorochemical improves the engraftment and function of intraportally transplanted islets. Pancreas. 2011;40(3):403–409. 136. Fuller  B, Froghi  F, Davidson  B. Organ preservation solutions: linking pharmacology to survival for the donor organ pathway. Curr Opin Organ Transplant. 2018;23(3):361–368.

137. Reichenspurner H, Russ C, Wagner F, et al. Comparison of UW versus HTK solution for myocardial protection in heart transplantation. Transpl Int. 1994;7(Suppl 1):S481–S484. 138. Hendry  PJ, Labow  RS, Keon  WJ. A comparison of intracellular solutions for donor heart preservation. J Thorac Cardiovasc Surg. 1993;105(4):667–673. 139. Arslan  A, Sezgin  A, Gultekin  B, et  al. Low-dose histidine-­ tryptophan-ketoglutarate solution for myocardial protection. Transplant Proc. 2005;37(7):3219–3222. 140. Pasut G, Panisello A, Folch-Puy E, et al. Polyethylene glycols: an effective strategy for limiting liver ischemia reperfusion injury. World J Gastroenterol. 2016;22(28):6501–6508. 141. Hauet  T, Goujon  JM, Baumert  H, et  al. Polyethylene glycol reduces the inflammatory injury due to cold ischemia/reperfusion in autotransplanted pig kidneys. Kidney Int. 2002;62(2):654–667. 142. Campbell  LH, Taylor  MJ, Brockbank  KG. Development of pancreas storage solutions: initial screening of cytoprotective supplements for beta-cell survival and metabolic status after hypothermic storage. Biopreserv Biobanking. 2013;11(1):12–18. 143. Brandhorst  H, Theisinger  B, Guenther  B, Johnson  PR, Brandhorst D. Pancreatic L-glutamine administration protects pig islets from cold ischemic injury and increases resistance toward inflammatory mediators. Cell Transplant. 2016;25(3):531–538. 144. Bae  C, Pichardo  EM, Huang  H, Henry  SD, Guarrera  JV. The benefits of hypothermic machine perfusion are enhanced with Vasosol and alpha-tocopherol in rodent donation after cardiac death livers. Transplant Proc. 2014;46(5):1560–1566. 145. Guarrera JV, Henry SD, Samstein B, et al. Hypothermic machine preservation facilitates successful transplantation of “orphan” extended criteria donor livers. Am J Transplant. 2015;15(1):161–169. 146. Thompson  JD, Kornbrust  DJ, Foy  JW, et  al. Toxicological and pharmacokinetic properties of chemically modified siRNAs targeting p53 RNA following intravenous administration. Nucleic Acid Ther. 2012;22(4):255–264. 147. Jia  Y, Zhao  Z, Xu  M, et  al. Prevention of renal ischemia-­ reperfusion injury by short hairpin RNA of endothelin A receptor in a rat model. Exp Biol Med (Maywood). 2012;237(8):894–902. 148. Yang  C, Jia  Y, Zhao  T, et  al. Naked caspase 3 small interfering RNA is effective in cold preservation but not in autotransplantation of porcine kidneys. J Surg Res. 2013;181(2):342–354. 149. Zhang  ZX, Min  WP, Jevnikar  AM. Use of RNA interference to minimize ischemia reperfusion injury. Transplant Rev (Orlando). 2012;26(2):140–155. 150. Molitoris BA, Dagher PC, Sandoval RM, et al. siRNA targeted to p53 attenuates ischemic and cisplatin-induced acute kidney injury. J Am Soc Nephrol. 2009;20(8):1754–1764. 151. Zheng  X, Lian  D, Wong  A, et  al. Novel small interfering RNA-containing solution protecting donor organs in heart transplantation. Circulation. 2009;120(12):1099–1107. 1091 p following 1107. 152. Weiss  JB, Eisenhardt  SU, Stark  GB, Bode  C, Moser  M, Grundmann S. MicroRNAs in ischemia-reperfusion injury. Am J Cardiovasc Dis. 2012;2(3):237–247. 153. Un  K, Kawakami  S, Yoshida  M, et  al. Efficient suppression of murine intracellular adhesion molecule-1 using ultrasound-­ responsive and mannose-modified lipoplexes inhibits acute hepatic inflammation. Hepatology. 2012;56(1):259–269. 154. Jiang N, Zhang X, Zheng X, et al. Targeted gene silencing of TLR4 using liposomal nanoparticles for preventing liver ischemia reperfusion injury. Am J Transplant. 2011;11(9):1835–1844. 155. Kimura  H. Signaling molecules: hydrogen sulfide and polysulfide. Antioxid Redox Signal. 2015;22(5):362–376. 156. Fukuto JM, Carrington SJ, Tantillo DJ, et al. Small molecule signaling agents: the integrated chemistry and biochemistry of nitrogen oxides, oxides of carbon, dioxygen, hydrogen sulfide, and their derived species. Chem Res Toxicol. 2012;25(4):769–793.

B. Islet allo-transplantation



523

References

157. Mustafa AK, Gadalla MM, Sen N, et al. H2S signals through protein S-sulfhydration. Sci Signal. 2009;2(96):ra72. 158. Mustafa  AK, Gadalla  MM, Snyder  SH. Signaling by gasotransmitters. Sci Signal. 2009;2(68):re2. 159. Ozaki  KS, Kimura  S, Murase  N. Use of carbon monoxide in minimizing ischemia/reperfusion injury in transplantation. Transplant Rev (Orlando). 2012;26(2):125–139. 160. Sandouka A, Fuller BJ, Mann BE, Green CJ, Foresti R, Motterlini R. Treatment with CO-RMs during cold storage improves renal function at reperfusion. Kidney Int. 2006;69(2):239–247. 161. Nakao A, Choi AM, Murase N. Protective effect of carbon monoxide in transplantation. J Cell Mol Med. 2006;10(3):650–671. 162. Cardini  B, Watschinger  K, Hermann  M, et  al. Crucial role for neuronal nitric oxide synthase in early microcirculatory derangement and recipient survival following murine pancreas transplantation. PLoS One. 2014;9(11):e112570. 163. Srinivasan  PK, Yagi  S, Doorschodt  B, et  al. Impact of venous systemic oxygen persufflation supplemented with nitric oxide gas on cold-stored, warm ischemia-damaged experimental liver grafts. Liver Transpl. 2012;18(2):219–225. 164. Luer B, Koetting M, Efferz P, Minor T. Role of oxygen during hypothermic machine perfusion preservation of the liver. Transpl Int. 2010;23(9):944–950. 165. Moers C, Pirenne J, Paul A, Ploeg RJ. Machine preservation trial study G. Machine perfusion or cold storage in deceased-donor kidney transplantation. N Engl J Med. 2012;366(8):770–771. 166. Treckmann J, Moers C, Smits JM, et al. Machine perfusion versus cold storage for preservation of kidneys from expanded criteria donors after brain death. Transpl Int. 2011;24(6):548–554. 167. Cartwright NK. Machine perfusion or cold storage in deceased-­ donor kidney transplantation. N Engl J Med. 2009;360(14):1460– 1461. author reply 1461. 168. Watson CJ, Wells AC, Roberts RJ, et al. Cold machine perfusion versus static cold storage of kidneys donated after cardiac death: a UK multicenter randomized controlled trial. Am J Transplant. 2010;10(9):1991–1999. 169. Balfoussia  D, Yerrakalva  D, Hamaoui  K, Papalois  V. Advances in machine perfusion graft viability assessment in kidney, liver, pancreas, lung, and heart transplant. Exp Clin Transplant. 2012;10(2):87–100. 170. Tersigni  R, Toledo-Pereyra  LH, Pinkham  J, Najarian  JS. Pancreaticoduodenal preservation by hypothermic pulsatile perfusion for twenty-four hours. Ann Surg. 1975;182(6):743–748. 171. Karcz M, Cook HT, Sibbons P, Gray C, Dorling A, Papalois V. An ex-vivo model for hypothermic pulsatile perfusion of porcine pancreata: hemodynamic and morphologic characteristics. Exp Clin Transplant. 2010;8(1):55–60. 172. Leemkuil  M, Engelse  M, Ploeg  R, de Koning  E, Krikke  C, Leuvenink  H. Hypothermic machine perfusion improves the quality of marginal donor pancreata. Transplantation. 2015;99:S231. 173. Hamaoui K, Gowers S, Sandhu B, et al. Development of pancreatic machine perfusion: translational steps from porcine to human models. J Surg Res. 2018;223:263–274. 174. Wright FH, Wright C, Ames SA, Smith JL, Corry RJ. Pancreatic allograft thrombosis: donor and retrieval factors and early postperfusion graft function. Transplant Proc. 1990;22(2):439–441. 175. Taylor MJ, Baicu S, Greene E, Vazquez A, Brassil J. Islet isolation from juvenile porcine pancreas after 24-h hypothermic machine perfusion preservation. Cell Transplant. 2010;19(5):613–628. 176. Brasile  L, Stubenitsky  BM, Booster  MH, et  al. Overcoming severe renal ischemia: the role of ex  vivo warm perfusion. Transplantation. 2002;73(6):897–901. 177. Babkin BP, Starling EH. A method for the study of the perfused pancreas. J Physiol. 1926;61(2):245–247. 178. Eckhauser  F, Knol  JA, Porter-Fink  V, et  al. Ex  vivo normothermic hemoperfusion of the canine pancreas: applications and

179.

180. 181.

182. 183.

184.

185.

186. 187.

188.

189.

190.

191.

192.

193.

194.

195.

196.

197.

198.

199.

limitations of a modified experimental preparation. J Surg Res. 1981;31(1):22–37. Idezuki  Y, Goetz  FC, Kaufman  SE, Lillehei  RC. In  vitro insulin productivity of preserved pancreas: a simple test to assess the viability of pancreatic allografts. Surgery. 1968;64(5):940–947. Curry DL, Bennett LL, Grodsky GM. Dynamics of insulin secretion by the perfused rat pancreas. Endocrinology. 1968;83(3):572–584. Grodsky  GM, Batts  AA, Bennett  LL, Vcella  C, McWilliams  NB, Smith DF. Effects of carbohydrates on secretion of insulin from isolated rat pancreas. Am J Physiol. 1963;205:638–644. Berglund  O. Different dynamics of insulin secretion in the perfused pancreas of mouse and rat. Acta Endocrinol. 1980;93(1):54–60. Macdonald PS, Chew HC, Connellan M, Dhital K. Extracorporeal heart perfusion before heart transplantation: the heart in a box. Curr Opin Organ Transplant. 2016;21(3):336–342. Bagul  A, Hosgood  SA, Kaushik  M, Kay  MD, Waller  HL, Nicholson  ML. Experimental renal preservation by normothermic resuscitation perfusion with autologous blood. Br J Surg. 2008;95(1):111–118. Kuan KG, Wee MN, Chung WY, et al. A study of normothermic hemoperfusion of the porcine pancreas and kidney. Artif Organs. 2017;41(5):490–495. Weissenbacher A, Hunter J. Normothermic machine perfusion of the kidney. Curr Opin Organ Transplant. 2017;22(6):571–576. Angelico  R, Perera  MTPR, Ravikumar  R, et  al. Normothermic machine perfusion of deceased donor liver grafts is associated with improved postreperfusion hemodynamics. Transplant Direct. 2016;2(9):e97. Cypel  M, Yeung  JC, Liu  M, et  al. Normothermic ex  vivo lung perfusion in clinical lung transplantation. N Engl J Med. 2011;364(15):1431–1440. Weegman BP, Taylor MJ, Baicu SC, et al. Hypothermic perfusion preservation of pancreas for islet grafts: validation using a Split Lobe Porcine Model. Cell Med. 2012;2(3):105–110. Kojima  Y, Nakagawara  G, Takeyama  S, Imahori  T, Miyazaki  I. Perfusion preservation of cadaver rat pancreas: II. Culture after perfusion and successful transplantation. Jpn J Surg. 1984;14(1):47–51. Toledo-Pereyra  LH, Valgee  KD, Castellanos  J, Chee  M. Hypothermic pulsatile perfusion: its use in the preservation of pancreases for 24 to 48 hours before islet cell transplantation. Arch Surg. 1980;115(1):95–98. Toledo-Pereyra LH, Valjee KD, Chee M, Lillehei RC. Preservation of the pancreas for transplantation. Surg Gynecol Obstet. 1979;148(1):57–61. Meyer W, Castelfranchi PL, Schulz LS, et al. Physiologic studies during perfusion of the isolated canine pancreas. The endocrine and exocrine behavior. Eur Surg Res. 1973;5(2):105–115. Kumar R, Chung WY, Dennison AR, Garcea G. Ex vivo porcine organ perfusion models as a suitable platform for translational transplant research. Artif Organs. 2017;41(9):E69–E79. Kumar R, Chung WY, Runau F, et al. Ex vivo normothermic porcine pancreas: a physiological model for preservation and transplant study. Int J Surg. 2018;54(Pt A):206–215. Prieto  M, Baron  P, Andreone  PA, et  al. Multiple ex  vivo organ preservation with warm whole blood. J Heart Transplant. 1988;7(3):227–237. O’Malley  VP, Keyes  DM, Postier  RG. The fluosol-perfused isolated canine pancreas: a model for the study of blood component effects in acute pancreatitis. J Surg Res. 1986;40(3):210–215. Eloy MR, Kachelhoffer J, Pousse A, Dauchel J, Grenier JF. Ex vivo vascular perfusion of the isolated canine pancreas. Experimental procedure, haemodynamic data and experimental applications. Eur Surg Res. 1974;6(6):341–353. Bunzl A, Burgen AS, Burns BD, Pedley N, Terroux KG. Methods for studying the reflex activity of the frog’s spinal cord. Br J Pharmacol Chemother. 1954;9(2):229–235.

B. Islet allo-transplantation

524

42.  Pancreas and islet preservation

200. Burns  BD, Robson  JG, Smith  GK. The survival of mammalian tissues perfused with intravascular gas mixtures of oxygen and carbon dioxide. Can J Biochem Physiol. 1958;36(5):499–504. 201. Sabiston Jr DC, Talbert JL, Riley Jr LH, Blalock A. Maintenance of the heart beat by perfusion of the coronary circulation with gaseous oxygen. Ann Surg. 1959;150:361–370. 202. Suszynski  TM, Rizzari  MD, Scott 3rd WE, Tempelman  LA, Taylor  MJ, Papas  KK. Persufflation (or gaseous oxygen perfusion) as a method of organ preservation. Cryobiology. 2012;64(3):125–143. 203. Min CG, Papas KK. Recent developments in persufflation for organ preservation. Curr Opin Organ Transplant. 2018;23(3):330–335. 204. Luer B, Fox M, Efferz P, Minor T. Adding pulsatile vascular stimulation to venous systemic oxygen persufflation of liver grafts. Artif Organs. 2014;38(5):404–410. 205. Suszynski  TM, Rizzari  MD, Scott  WE, et  al. Persufflation (gaseous oxygen perfusion) as a method of heart preservation. J Cardiothorac Surg. 2013;8:105. 206. Scott 3rd WE, Weegman BP, Ferrer-Fabrega J, et al. Pancreas oxygen persufflation increases ATP levels as shown by nuclear magnetic resonance. Transplant Proc. 2010;42(6):2011–2015. 207. Reddy  MS, Carter  N, Cunningham  A, Shaw  J, Talbot  D. Portal venous oxygen persufflation of the donation after cardiac death pancreas in a rat model is superior to static cold storage and hypothermic machine perfusion. Transpl Int. 2014;27(6):634–639. 208. Isselhard  W, Witte  J, Denecke  H, Berger  M, Fischer  JH, Molzberger H. Function and metabolism of canine kidneys after aerobic ischemia by retrograde persufflation with gaseous oxygen. Res Exp Med. 1974;164(1):35–44. 209. Isselhard W, Denecke H, Stelter W, et al. Function and metabolism of canine kidneys after aerobic ischemia by orthograde persufflation with gaseous oxygen. Res Exp Med. 1973;159(4):288–297. 210. Scott 3rd WE, O’Brien  TD, Ferrer-Fabrega  J, et  al. Persufflation improves pancreas preservation when compared with the twolayer method. Transplant Proc. 2010;42(6):2016–2019. 211. Kelly  AC, Smith  KE, Purvis  WG, et  al. Oxygen perfusion (persufflation) of human pancreata enhances insulin secretion and attenuates islet proinflammatory signaling. Transplantation. 2019;103(1):160–167. 212. Hellerstroem C. A method for the microdissection of intact pancreatic islets of mammals. Acta Endocrinol. 1964;45:122–132. 213. Moskalewski S. Isolation and culture of the islets of Langerhans of the guinea pig. Gen Comp Endocrinol. 1965;5:342–353. 214. Lacy PE, Kostianovsky M. Method for the isolation of intact islets of Langerhans from the rat pancreas. Diabetes. 1967;16(1):35–39. 215. Scharp DW, Downing R, Merrell RC, Greider M. Isolating the elusive islet. Diabetes. 1980;29(Suppl 1):19–30. 216. Gotoh  M, Maki  T, Kiyoizumi  T, Satomi  S, Monaco  AP. An improved method for isolation of mouse pancreatic islets. Transplantation. 1985;40(4):437–438. 217. Lindall  A, Steffes  M, Sorenson  R. Immunoassayable insulin content of subcellular fractions of rat islets. Endocrinology. 1969;85(2):218–223. 218. Scharp DW, Kemp CB, Knight MJ, Ballinger WF, Lacy PE. The use of ficoll in the preparation of viable islets of Langerhans from the rat pancreas. Transplantation. 1973;16(6):686–689. 219. Buitrago  A, Gylfe  E, Henriksson  C, Pertoft  H. Rapid isolation of pancreatic islets from collagenase digested pancreas by sedimentation through Percol at unit gravity. Biochem Biophys Res Commun. 1977;79(3):823–828. 220. Brandhorst D, Brandhorst H, Hering BJ, Federlin K, Bretzel RG. Islet isolation from the pancreas of large mammals and humans: 10 years of experience. Exp Clin Endocrinol Diabetes. 1995;103(Suppl 2):3–14. 221. Ballinger WF, Lacy PE. Transplantation of intact pancreatic islets in rats. Surgery. 1972;72(2):175–186.

222. Reckard CR, Ziegler MM, Barker CF. Physiological and immunological consequences of transplanting isolated pancreatic islets. Surgery. 1973;74(1):91–99. 223. Najarian JS, Sutherland DE, Matas AJ, Steffes MW, Simmons RL, Goetz  FC. Human islet transplantation: a preliminary report. Transplant Proc. 1977;9(1):233–236. 224. Sutherland DE, Matas AJ, Goetz FC, Najarian JS. Transplantation of dispersed pancreatic islet tissue in humans: autografts and allografts. Diabetes. 1980;29(Suppl 1):31–44. 225. Kemp CB, Knight MJ, Scharp DW, Ballinger WF, Lacy PE. Effect of transplantation site on the results of pancreatic islet isografts in diabetic rats. Diabetologia. 1973;9(6):486–491. 226. Horaguchi  A, Merrell  RC. Preparation of viable islet cells from dogs by a new method. Diabetes. 1981;30(5):455–458. 227. Noel J, Rabinovitch A, Olson L, Kyriakides G, Miller J, Mintz DH. A method for large-scale, high-yield isolation of canine pancreatic islets of Langerhans. Metabolism. 1982;31(2):184–187. 228. Gray  DW, McShane  P, Grant  A, Morris  PJ. A method for isolation of islets of Langerhans from the human pancreas. Diabetes. 1984;33(11):1055–1061. 229. Kulkarni RN, Stewart AF. Summary of the Keystone islet workshop (April 2014): the increasing demand for human islet availability in diabetes research. Diabetes. 2014;63(12):3979–3981. 230. Chakradhar S. Diabetes researchers fear worsening access to human islets. Nat Med. 2014;20(6):567. 231. Carter JD, Dula SB, Corbin KL, Wu R, Nunemaker CS. A practical guide to rodent islet isolation and assessment. Biol Proced Online. 2009;11:3–31. 232. Zmuda EJ, Powell CA, Hai T. A method for murine islet isolation and subcapsular kidney transplantation. J Vis Exp. 2011;50:e2096. 1–11. 233. Neuman JC, Truchan NA, Joseph JW, Kimple ME. A method for mouse pancreatic islet isolation and intracellular cAMP determination. J Vis Exp. 2014;88:e2096. 1–11. 234. Abdulreda  MH, Rodriguez-Diaz  R, Cabrera  O, Caicedo  A, Berggren PO. The different faces of the pancreatic islet. Adv Exp Med Biol. 2016;938:11–24. 235. Ramirez-Dominguez  M. Isolation of mouse pancreatic islets of Langerhans. Adv Exp Med Biol. 2016;938:25–34. 236. Steiner DJ, Kim A, Miller K, Hara M. Pancreatic islet plasticity: interspecies comparison of islet architecture and composition. Islets. 2010;2(3):135–145. 237. Cabrera  O, Berman  DM, Kenyon  NS, Ricordi  C, Berggren  PO, Caicedo A. The unique cytoarchitecture of human pancreatic islets has implications for islet cell function. Proc Natl Acad Sci U S A. 2006;103(7):2334–2339. 238. Brissova M, Fowler MJ, Nicholson WE, et al. Assessment of human pancreatic islet architecture and composition by laser scanning confocal microscopy. J Histochem Cytochem. 2005;53(9):1087–1097. 239. Kim A, Miller K, Jo J, Kilimnik G, Wojcik P, Hara M. Islet architecture: a comparative study. Islets. 2009;1(2):129–136. 240. Stefan  Y, Grasso  S, Perrelet  A, Orci  L. The pancreatic ­polypeptide-rich lobe of the human pancreas: definitive identification of its derivation from the ventral pancreatic primordium. Diabetologia. 1982;23(2):141–142. 241. Li  DS, Yuan  YH, Tu  HJ, Liang  QL, Dai  LJ. A protocol for islet isolation from mouse pancreas. Nat Protoc. 2009;4(11):1649–1652. 242. Cui W, Gu Y, Miyamoto M, et al. Novel method for isolation of adult porcine pancreatic islets with two-stage digestion procedure. Cell Transplant. 1999;8(4):391–398. 243. Kim HI, Lee SY, Jin SM, et al. Parameters for successful pig islet isolation as determined using 68 specific-pathogen-free miniature pigs. Xenotransplantation. 2009;16(1):11–18. 244. Szot GL, Koudria P, Bluestone JA. Murine pancreatic islet isolation. J Vis Exp. 2007;7:255. 245. Toomey  P, Chadwick  DR, Contractor  H, Bell  PR, James  RF, London  NJ. Porcine islet isolation: prospective comparison of

B. Islet allo-transplantation



246.

247.

248.

249.

250.

251.

252.

253.

254.

255.

256.

257.

258. 259. 260. 261.

262.

263.

264.

265.

525

References

automated and manual methods of pancreatic collagenase digestion. Br J Surg. 1993;80(2):240–243. White SA, Contractor HH, Hughes DP, et al. Influence of different collagenase solvents and timing of their delivery on porcine islet isolation. Br J Surg. 1996;83(10):1350–1355. Dufrane  D, D’Hoore  W, Goebbels  RM, Saliez  A, Guiot  Y, Gianello  P. Parameters favouring successful adult pig islet isolations for xenotransplantation in pig-to-primate models. Xenotransplantation. 2006;13(3):204–214. Wolters  GH, Vos-Scheperkeuter  GH, van Deijnen  JH, van Schilfgaarde  R. An analysis of the role of collagenase and protease in the enzymatic dissociation of the rat pancreas for islet isolation. Diabetologia. 1992;35(8):735–742. Wang Y, Paushter D, Wang S, et al. Highly purified versus filtered crude collagenase: comparable human islet isolation outcomes. Cell Transplant. 2011;20(11–12):1817–1825. van Deijnen JH, Hulstaert CE, Wolters GH, van Schilfgaarde R. Significance of the peri-insular extracellular matrix for islet isolation from the pancreas of rat, dog, pig, and man. Cell Tissue Res. 1992;267(1):139–146. Fujio  A, Murayama  K, Yamagata  Y, et  al. Collagenase H is crucial for isolation of rat pancreatic islets. Cell Transplant. 2014;23(10):1187–1198. Wolters GH, Vos-Scheperkeuter GH, Lin HC, van Schilfgaarde R. Different roles of class I and class II Clostridium histolyticum collagenase in rat pancreatic islet isolation. Diabetes. 1995;44(2):227–233. Brandhorst D, Huettler S, Alt A, et al. Adjustment of the ratio between collagenase class II and I improves islet isolation outcome. Transplant Proc. 2005;37(8):3450–3451. Brandhorst  H, Raemsch-Guenther  N, Raemsch  C, et  al. The ratio between collagenase class I and class II influences the efficient islet release from the rat pancreas. Transplantation. 2008;85(3):456–461. Barnett MJ, Zhai X, LeGatt DF, Cheng SB, Shapiro AM, Lakey JR. Quantitative assessment of collagenase blends for human islet isolation. Transplantation. 2005;80(6):723–728. Balamurugan  AN, Loganathan  G, Bellin  MD, et  al. A new enzyme mixture to increase the yield and transplant rate of autologous and allogeneic human islet products. Transplantation. 2012;93(7):693–702. Ricordi C, Lacy PE, Finke EH, Olack BJ, Scharp DW. Automated method for isolation of human pancreatic islets. Diabetes. 1988;37(4):413–420. Latif ZA, Noel J, Alejandro R. A simple method of staining fresh and cultured islets. Transplantation. 1988;45(4):827–830. Ricordi C, Gray DW, Hering BJ, et al. Islet isolation assessment in man and large animals. Acta Diabetol Lat. 1990;27(3):185–195. Komatsu H, Kandeel F, Mullen Y. Impact of oxygen on pancreatic islet survival. Pancreas. 2018;47(5):533–543. Matsumoto S, Noguchi H, Naziruddin B, et al. Improvement of pancreatic islet cell isolation for transplantation. Proc (Bayl Univ Med Cent). 2007;20(4):357–362. Qi  M, Barbaro  B, Wang  S, Wang  Y, Hansen  M, Oberholzer  J. Human pancreatic islet isolation: part II: purification and culture of human islets. J Vis Exp. 2009;27:1–2. Chadwick DR, Robertson GS, Contractor HH, et al. Storage of pancreatic digest before islet purification. The influence of colloids and the sodium to potassium ratio in University of Wisconsin-based preservation solutions. Transplantation. 1994;58(1):99–104. Adewola  A, Mage  R, Hansen  M, et  al. Comparing cooling systems for the COBE 2991 cell separator used in the purification of human pancreatic islets of Langerhans. Cryo Letters. 2010;31(4):310–317. Eckhard  M, Brandhorst  D, Winter  D, et  al. The role of current product release criteria for identification of human islet prepa-

266.

267.

268.

269.

270.

271.

272.

273.

274.

275.

276. 277.

278.

279.

280.

281.

282.

283.

284.

rations suitable for clinical transplantation. Transplant Proc. 2004;36(5):1528–1531. Friberg AS, Stahle M, Brandhorst H, Korsgren O, Brandhorst D. Human islet separation utilizing a closed automated purification system. Cell Transplant. 2008;17(12):1305–1313. Matsumoto S, Takita M, Chaussabel D, et al. Improving efficacy of clinical islet transplantation with iodixanol-based islet purification, thymoglobulin induction, and blockage of IL-1beta and TNF-alpha. Cell Transplant. 2011;20(10):1641–1647. Miki A, Ricordi C, Yamamoto T, et al. Effect of human islet rescue gradient purification on islet yield and fractional beta cell viability. Transplant Proc. 2008;40(2):360–361. Avgoustiniatos ES, Hering BJ, Rozak PR, et al. Commercially available gas-permeable cell culture bags may not prevent anoxia in cultured or shipped islets. Transplant Proc. 2008;40(2):395–400. Papas KK, Avgoustiniatos ES, Tempelman LA, et al. High-density culture of human islets on top of silicone rubber membranes. Transplant Proc. 2005;37(8):3412–3414. Balamurugan AN, Naziruddin B, Lockridge A, et al. Islet product characteristics and factors related to successful human islet transplantation from the Collaborative Islet Transplant Registry (CITR) 1999-2010. Am J Transplant. 2014;14(11):2595–2606. Brandhorst  D, Brandhorst  H, Mullooly  N, Acreman  S, Johnson  PR. High seeding density induces local hypoxia and triggers a proinflammatory response in isolated human islets. Cell Transplant. 2016;25(8):1539–1546. Smith KE, Kelly AC, Min CG, et al. Acute ischemia induced by high-density culture increases cytokine expression and diminishes the function and viability of highly purified human islets of Langerhans. Transplantation. 2017;101(11):2705–2712. Holmes  MA, Clayton  HA, Chadwick  DR, Bell  PR, London  NJ, James  RF. Functional studies of rat, porcine, and human pancreatic islets cultured in ten commercially available media. Transplantation. 1995;60(8):854–860. Kin  T, Senior  P, O’Gorman  D, Richer  B, Salam  A, Shapiro  AM. Risk factors for islet loss during culture prior to transplantation. Transpl Int. 2008;21(11):1029–1035. Clayton HA, London NJ. Survival and function of islets during culture. Cell Transplant. 1996;5(1):1–12. discussion 13-17, 19. Brandhorst  D, Brandhorst  H, Hering  BJ, Bretzel  RG. Longterm survival, morphology and in  vitro function of isolated pig islets under different culture conditions. Transplantation. 1999;67(12):1533–1541. Fraga DW, Sabek O, Hathaway DK, Gaber AO. A comparison of media supplement methods for the extended culture of human islet tissue. Transplantation. 1998;65(8):1060–1066. Ilieva  A, Yuan  S, Wang  RN, Agapitos  D, Hill  DJ, Rosenberg  L. Pancreatic islet cell survival following islet isolation: the role of cellular interactions in the pancreas. J Endocrinol. 1999;161(3):357–364. Noguchi  H, Naziruddin  B, Jackson  A, et  al. Fresh islets are more effective for islet transplantation than cultured islets. Cell Transplant. 2012;21(2-3):517–523. King A, Lock J, Xu G, Bonner-Weir S, Weir GC. Islet transplantation outcomes in mice are better with fresh islets and exendin-4 treatment. Diabetologia. 2005;48(10):2074–2079. Olsson  R, Carlsson  PO. Better vascular engraftment and function in pancreatic islets transplanted without prior culture. Diabetologia. 2005;48(3):469–476. Kuttler  B, Hartmann  A, Wanka  H. Long-term culture of islets abrogates cytokine-induced or lymphocyte-induced increase of antigen expression on beta cells. Transplantation. 2002;74(4):440–445. Lacy PE, Davie JM, Finke EH. Effect of culture on islet rejection. Diabetes. 1980;29(Suppl 1):93–97.

B. Islet allo-transplantation

526

42.  Pancreas and islet preservation

285. Rijkelijkhuizen  JK, van der Burg  MP, Tons  A, Terpstra  OT, Bouwman  E. Pretransplant culture selects for high-quality porcine islets. Pancreas. 2006;32(4):403–407. 286. Andersson A, Borg H, Groth CG, et al. Survival of isolated human islets of Langerhans maintained in tissue culture. J Clin Invest. 1976;57(5):1295–1301. 287. Gaber  AO, Fraga  DW, Callicutt  CS, Gerling  IC, Sabek  OM, Kotb  MY. Improved in  vivo pancreatic islet function after prolonged in vitro islet culture. Transplantation. 2001;72(11):1730–1736. 288. Jimenez-Vera  E, Davies  S, Phillips  P, O’Connell  PJ, Hawthorne WJ. Long-term cultured neonatal islet cell clusters demonstrate better outcomes for reversal of diabetes: in vivo and molecular profiles. Xenotransplantation. 2015;22(2):114–123. 289. Falqui L, Finke EH, Carel JC, Scharp DW, Lacy PE. Marked prolongation of human islet xenograft survival (human-to-mouse) by low-temperature culture and temporary immunosuppression with human and mouse anti-lymphocyte sera. Transplantation. 1991;51(6):1322–1324. 290. Ono  J, Lacy  PE, Michael  HE, Greider  MH. Studies of the functional and morphologic status of islets maintained at 24 C for four weeks in vitro. Am J Pathol. 1979;97(3):489–503. 291. Escolar JC, Hoo-Paris R, Castex C, Sutter BC. Effect of low temperatures on glucose-induced insulin secretion and glucose metabolism in isolated pancreatic islets of the rat. J Endocrinol. 1990;125(1):45–51. 292. Mueller  KR, Martins  KV, Murtaugh  MP, Schuurman  HJ, Papas KK. Manufacturing porcine islets: culture at 22 degrees C has no advantage above culture at 37 degrees C: a gene expression evaluation. Xenotransplantation. 2013;20(6):418–428. 293. Murdoch TB, McGhee-Wilson D, Shapiro AM, Lakey JR. Methods of human islet culture for transplantation. Cell Transplant. 2004;13(6):605–617. 294. Buchwald  P. FEM-based oxygen consumption and cell viability models for avascular pancreatic islets. Theor Biol Med Model. 2009;6:5. 295. Goto M, Yoshikawa Y, Matsuo K, et al. Optimization of a prominent oxygen-permeable device for pancreatic islets. Transplant Proc. 2008;40(2):411–412. 296. Murray HE, Paget MB, Downing R. Preservation of glucose responsiveness in human islets maintained in a rotational cell culture system. Mol Cell Endocrinol. 2005;238(1-2):39–49. 297. Quaranta P, Antonini S, Spiga S, et al. Co-transplantation of endothelial progenitor cells and pancreatic islets to induce long-­ lasting normoglycemia in streptozotocin-treated diabetic rats. PLoS One. 2014;9(4):e94783. 298. Penko  D, Rojas-Canales  D, Mohanasundaram  D, et  al. Endothelial progenitor cells enhance islet engraftment, influence beta-cell function, and modulate islet connexin 36 expression. Cell Transplant. 2015;24(1):37–48. 299. Arous  C, Wehrle-Haller  B. Role and impact of the extracellular matrix on integrin-mediated pancreatic beta-cell functions. Biol Cell. 2017;109(6):223–237. 300. Christoffersson  G, Walden  T, Sandberg  M, Opdenakker  G, Carlsson PO, Phillipson M. Matrix metalloproteinase-9 is essential for physiological beta cell function and islet vascularization in adult mice. Am J Pathol. 2015;185(4):1094–1103. 301. Kerr-Conte J, Vandewalle B, Moerman E, et al. Upgrading pretransplant human islet culture technology requires human serum combined with media renewal. Transplantation. 2010;89(9):1154–1160. 302. Avgoustiniatos  ES, Scott 3rd WE, Suszynski  TM, et  al. Supplements in human islet culture: human serum albumin is inferior to fetal bovine serum. Cell Transplant. 2012;21(12):2805–2814. 303. Barnes  D, Sato  G. Methods for growth of cultured cells in ­serum-free medium. Anal Biochem. 1980;102(2):255–270. 304. Clark SA, Chick WL. Islet cell culture in defined serum-free medium. Endocrinology. 1990;126(4):1895–1903.

305. Lee  RH, Carter  J, Szot  GL, Posselt  A, Stock  P. Human albumin preserves islet mass and function better than whole serum during pretransplantation islet culture. Transplant Proc. 2008;40(2):384–386. 306. Nacher  M, Estil Les  E, Garcia  A, et  al. Human serum versus human serum albumin supplementation in human islet pretransplantation culture: in  vitro and in  vivo assessment. Cell Transplant. 2016;25(2):343–352. 307. Brigelius-Flohe  R, Banning  A, Schnurr  K. Selenium-dependent enzymes in endothelial cell function. Antioxid Redox Signal. 2003;5(2):205–215. 308. Steinbrenner  H, Hotze  AL, Speckmann  B, et  al. Localization and regulation of pancreatic selenoprotein P. J Mol Endocrinol. 2013;50(1):31–42. 309. Bradley  B, Prowse  SJ, Bauling  P, Lafferty  KJ. Desferrioxamine treatment prevents chronic islet allograft damage. Diabetes. 1986;35(5):550–555. 310. Nomikos IN, Prowse SJ, Carotenuto P, Lafferty KJ. Combined treatment with nicotinamide and desferrioxamine prevents islet allograft destruction in NOD mice. Diabetes. 1986;35(11):1302–1304. 311. Mendola J, Wright Jr JR, Lacy PE. Oxygen free-radical scavengers and immune destruction of murine islets in allograft rejection and multiple low-dose streptozocin-induced insulitis. Diabetes. 1989;38(3):379–385. 312. Eizirik  DL, Sandler  S, Welsh  N, Bendtzen  K, Hellerstrom  C. Nicotinamide decreases nitric oxide production and partially protects human pancreatic islets against the suppressive effects of combinations of cytokines. Autoimmunity. 1994;19(3):193–198. 313. Langlois A, Bietiger W, Mandes K, et al. Overexpression of vascular endothelial growth factor in vitro using deferoxamine: a new drug to increase islet vascularization during transplantation. Transplant Proc. 2008;40(2):473–476. 314. Burkart  V, Gross-Eick  A, Bellmann  K, Radons  J, Kolb  H. Suppression of nitric oxide toxicity in islet cells by alpha-­ tocopherol. FEBS Lett. 1995;364(3):259–263. 315. Bottino R, Inverardi L, Valente U, Ricordi C. Serum-free medium and pyruvate improve survival and glucose responsiveness of islet beta cells in culture. Transplant Proc. 1997;29(4):1978. 316. Ishihara  H, Maechler  P, Gjinovci  A, Herrera  PL, Wollheim  CB. Islet beta-cell secretion determines glucagon release from neighbouring alpha-cells. Nat Cell Biol. 2003;5(4):330–335. 317. Duprez  J, Roma  LP, Close  AF, Jonas  JC. Protective antioxidant and antiapoptotic effects of ZnCl2 in rat pancreatic islets cultured in low and high glucose concentrations. PLoS One. 2012;7(10):e46831. 318. Brandhorst  H, Duan  Y, Iken  M, Bretzel  RG, Brandhorst  D. Effect of stable glutamine compounds on porcine islet culture. Transplant Proc. 2005;37(8):3519–3520. 319. Avila J, Barbaro B, Gangemi A, et al. Intra-ductal glutamine administration reduces oxidative injury during human pancreatic islet isolation. Am J Transplant. 2005;5(12):2830–2837. 320. Avila JG, Tsujimura T, Oberholzer J, et al. Improvement of pancreatic islet isolation outcomes using glutamine perfusion during isolation procedure. Cell Transplant. 2003;12(8):877–881. 321. Schnell  S, Schaefer  M, Schofl  C. Free fatty acids increase cytosolic free calcium and stimulate insulin secretion from beta-cells through activation of GPR40. Mol Cell Endocrinol. ­ 2007;263(1-2):173–180. 322. Wolf  BA, Pasquale  SM, Turk  J. Free fatty acid accumulation in secretagogue-stimulated pancreatic islets and effects of arachidonate on depolarization-induced insulin secretion. Biochemistry. 1991;30(26):6372–6379. 323. Turk J, Gross RW, Ramanadham S. Amplification of insulin secretion by lipid messengers. Diabetes. 1993;42(3):367–374. 324. Dixon  G, Nolan  J, McClenaghan  NH, Flatt  PR, Newsholme  P. Arachidonic acid, palmitic acid and glucose are important for the

B. Islet allo-transplantation



325.

326. 327.

328.

329.

330.

331. 332.

333. 334.

335.

336.

References

modulation of clonal pancreatic beta-cell insulin secretion, growth and functional integrity. Clin Sci (Lond). 2004;106(2):191–199. Atchison N, Swindlehurst G, Papas KK, Tsapatsis M, Kokkoli E. Maintenance of ischemic beta cell viability through delivery of lipids and ATP by targeted liposomes. Biomater Sci. 2014;2(4):548–559. Kojayan GG, Alexander M, Imagawa DK, Lakey JRT. Systematic review of islet cryopreservation. Islets. 2018;10(1):40–49. Benson JD, Benson CT, Critser JK. Mathematical model formulation and validation of water and solute transport in whole hamster pancreatic islets. Math Biosci. 2014;254:64–75. Rawal  S, Harrington  S, Williams  SJ, Ramachandran  K, StehnoBittel L. Long-term cryopreservation of reaggregated pancreatic islets resulting in successful transplantation in rats. Cryobiology. 2017;76:41–50. Williams SJ, Wang Q, Macgregor RR, Siahaan TJ, Stehno-Bittel L, Berkland  C. Adhesion of pancreatic beta cells to biopolymer films. Biopolymers. 2009;91(8):676–685. Ramachandran  K, Williams  SJ, Huang  HH, Novikova  L, Stehno-Bittel  L. Engineering islets for improved performance by optimized reaggregation in a micromold. Tissue Eng Part A. 2013;19(5–6):604–612. Lovelock JE, Bishop MW. Prevention of freezing damage to living cells by dimethyl sulphoxide. Nature. 1959;183(4672):1394–1395. Lakey JR, Rajotte RV, Fedorow CA, Taylor MJ. Islet cryopreservation using intracellular preservation solutions. Cell Transplant. 2001;10(7):583–589. Bank HL. Cryobiology of isolated islets of Langerhans circa 1982. Cryobiology. 1983;20(2):119–128. Bank HL, Davis RF, Emerson D. Cryogenic preservation of isolated rat Islets of Langerhans: effect of cooling and warming rates. Diabetologia. 1979;16(3):195–199. Wise MH, Gordon C, Johnson RW. Intraportal autotransplantation of cryopreserved porcine islets of Langerhans. Cryobiology. 1985;22(4):359–366. Rajotte  RV, Stewart  HL, Voss  WA, Shnitka  TK. Viability studies on frozen—thawed rat islets of Langerhans. Cryobiology. 1977;14(1):116–120.

527

337. Foreman  J, Moriya  H, Taylor  MJ. Effect of cooling rate and its interaction with pre-freeze and post-thaw tissue culture on the in vitro and in vivo function of cryopreserved pancreatic islets. Transpl Int. 1993;6(4):191–200. 338. Ishihara K, Taniguchi H, Hara Y, Ejiri K, Baba S. Effect of cooling rate on insulin release from frozen-thawed dispersed rat islet cells. Diabetes Res Clin Pract. 1989;6(4):243–246. 339. Taylor MJ, Benton MJ. Interaction of cooling rate, warming rate, and extent of permeation of cryoprotectant in determining survival of isolated rat islets of Langerhans during cryopreservation. Diabetes. 1987;36(1):59–65. 340. Taylor  MJ, Baicu  S. Review of vitreous islet cryopreservation: some practical issues and their resolution. Organogenesis. 2009;5(3):155–166. 341. Elliott  TF, Das  GC, Hammond  DK, et  al. Screening and identification of cryopreservative agents for human cellular biotechnology experiments in microgravity. Gravit Space Biol Bull. 2005;18(2):83–84. 342. Rajotte  RV, Warnock  GL, Kneteman  NM, Erickson  C, Ellis  DK. Optimizing cryopreservation of isolated islets. Transplant Proc. 1989;21(1 Pt 3):2638–2640. 343. Tian  J, Dang  H, Chen  Z, et  al. gamma-Aminobutyric acid regulates both the survival and replication of human beta-cells. Diabetes. 2013;62(11):3760–3765. 344. Chandravanshi B, Dhanushkodi A, Bhonde R. High recovery of functional islets stored at low and ultralow temperatures. Rev Diabet Stud. 2014;11(3-4):267–278. 345. Soltani N, Qiu H, Aleksic M, et al. GABA exerts protective and regenerative effects on islet beta cells and reverses diabetes. Proc Natl Acad Sci U S A. 2011;108(28):11692–11697. 346. Al-Adra DP, Gill RS, Imes S, et al. Single-donor islet transplantation and long-term insulin independence in select patients with type 1 diabetes mellitus. Transplantation. 2014;98(9):1007–1012. 347. Colton  CK, Papas  KK, Pisania  A, et  al. Characterization of islet preparations. In: Halberstadt  CR, Emerich  DF, eds. Cellular transplantation: From laboratory to clinic. Oxford: Academic Press; 2007:85–133.

B. Islet allo-transplantation