Scaffold-Based Cell Delivery for Cardiac Repair

C H A P T E R

26 Scaffold-Based Cell Delivery for Cardiac Repair Lisle Blackbourn1, Eric G. Schmuck1, and Amish N. Raval1,2 1

Division of Cardiovascular Medicine, Department of Medicine, University of Wisconsin School of Medicine and Public Health, Madison, WI, USA; 2Department of Biomedical Engineering, University of Wisconsin, Madison, WI, USA

INTRODUCTION

THE OPTIMAL CELL DELIVERY SCAFFOLD FOR CARDIAC REPAIR

Cellular therapy promises to be a revolutionary technology for the treatment of advanced heart disease. Despite remarkable advances in this field, the optimal cell type and delivery approach are widely debated. Cell delivery methods that have been adopted in clinical trials include intravenous infusion, catheter-directed coronary artery or coronary sinus infusion, and intramuscular injection. Each method offers certain advantages, yet all are limited by poor cell retention. Explanations include inadequate tissue homing signals, an inhospitable microenvironment within the injured tissue, cell escape from the vascular circulation and, in the case of intramuscular injection, egress of cells immediately from the needle tract or outward into the lymph/capillary circulation. It is now widely accepted that certain somatic cells offer therapeutic benefit through paracrine mechanisms and not through local transdifferentiation and engraftment. Even so, sustained retention of viable cells within the myocardium is likely a prerequisite for a durable therapeutic effect. The concept of a cell-seeded biologic scaffold has emerged as an alternative approach to deliver a large number of cells directly to the injured area of the heart. The cellular arrangement, protein composition, pore size, geometric and mechanical characteristics of the scaffold may be engineered to create a highly deliverable and favorable microenvironment for enhanced cell retention and a more long-lasting therapeutic result. Herein, we discuss features of an optimal cell delivery scaffold for cardiac repair and review the experience to date in this field. We will focus this discussion on the scaffolds, as the properties of therapeutic cells will be discussed in other chapters.

Stem Cell and Gene Therapy for Cardiovascular Disease DOI: http://dx.doi.org/10.1016/B978-0-12-801888-0.00026-6

The ideal cell delivery scaffolds for cardiac repair should possess several properties (Table 26.1). The scaffold should permit predictable, timedependent dissociation of cells from the scaffold to the site of myocardial injury. The ideal scaffold should ideally have an interconnected pore system with high porosity ensuring cellular penetration and adequate diffusion of nutrients to cells. The pore size should be large enough to allow cells to migrate into the scaffold, yet small enough to establish a sufficiently high specific surface area to allow adhesion. The scaffold should be nonimmunogenic and biodegradable with nontoxic degradation byproducts. The scaffold should be manufactured ideally without chemical fixation. If chemical fixation is required, then cytotoxic chemical fixatives should be thoroughly removed prior to cell seeding and transplant. Chemical fixation could also have an effect on the immune response. Furthermore, the ideal scaffold material should inherently adhere to the epicardial surface, without sutures or glue. If sutures or glues are required, then these adhesives should not cause constriction or impede ventricular filling. Finally, the scaffold should have sufficient tensile strength to permit physical handling during surgical implantation.

CLINICAL REGULATORY CONSIDERATIONS The United States Food and Drug Administration (FDA) considers cell delivery scaffolds to be an Investigational Cellular and Gene Therapy Product.

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26. SCAFFOLD-BASED CELL DELIVERY FOR CARDIAC REPAIR

TABLE 26.1 Ideal Features of Scaffolds Designed to Deliver Cells to Treat Heart Disease Ideal scaffold Promotes adherence of cells Allows dissociation of cells Interconnected pore system, high porosity, proper pore size Nonimmunogenic Biodegradable with nontoxic degradation byproducts Sufficiently compliant Low cost Easy to replicate Easy to store/long shelf-life

The FDA Guidance for Industry recommends preclinical assessment standards and advises that the cell delivery scaffold must not alter the relevant biological characteristics of cells or tissues. These attributes should be demonstrated in vitro and in vivo in appropriate animal models. For example, an isolated “minimally manipulated” cell product may lose this distinction if there are cell-scaffold interactions that significantly alter the cell properties. The final cell and scaffold therapeutic product must meet final product chemical composition, manufacturing and storage specifications prior to being delivered into humans.

are derived from tissues, whereas synthetic scaffolds are derived using inanimate materials. Natural scaffolds may offer intrinsic bioactivity, but reproducibility and large-scale production may be challenging. In contrast, synthetic scaffolds are more readily prepared with stringent control over the biochemical and mechanical properties, to create a well-defined and reproducible therapeutic product. A systematic review of the published literature was conducted using PubMed, Wiley Online Library, Science Direct, and OVID from June 2014 to January 2015. The search was limited to those published after 1990. Keyword search terms were scaffold, bioscaffold, tissue scaffold, extracellular matrix (ECM), patch, stem cell transplantation, stem cell delivery, stem cells, mesenchymal stem cells, cellularized, bioengineered, synthetic, natural, regenerate, cardiovascular, heart, and cardiac. Certain papers were excluded because they (i) discussed “cell sheets” which are an alternative to scaffolds for cell delivery, (ii) discussed a scaffold free method of cell delivery, (iii) did not include cells in the scaffolding, or (iv) did not pertain to treating cardiac disease. Results of this search yielded 473 papers. Forty-three scaffold papers met the search criteria and are discussed herein. We have summarized the experience to date for natural and synthetic bioscaffolds developed for cardiac repair (Table 26.2).

Natural DELIVERY APPROACHES There are three approaches of how cells can be combined with scaffolds for therapeutic delivery (Figure 26.1): 1. Administering a scaffold first and then cells directly within the heart (A1), or administering cells first and then scaffolds (A2) 2. Combining cells with scaffolds out of the body and then immediately placing the cell-scaffold product onto the heart (B) 3. Combining cells with scaffolds at a manufacturing facility in advance of the procedure, storing it, shipping it to the site for therapeutic administration (C) [1]. Each of these approaches has unique advantages and disadvantages but approach (3) may be clinically simplest and most practical.

TYPES OF CELL DELIVERY SCAFFOLDS FOR CARDIAC REPAIR There are two broad categories of engineered scaffolds: (i) natural and (ii) synthetic [2]. Natural scaffolds

There are a several natural materials that have been well described in the published literature. In general, these scaffolds are immunoprivileged and interact favorably with host tissue depending on the degree of chemical cross-linking [2]. These materials can contain native ECM proteins including collagen (I, II, III, IV, V), elastin, laminin, and fibronectin. In addition to structural proteins, natural materials may contain cytokines and growth factors. Selected materials are discussed later. Myocardium ECM derived from decellularized adult myocardium is an attractive, tissue-specific (homologous) device for therapeutic cell delivery [8,40,41]. Human adult myocardial ECM may be isolated from donor hearts that are not suitable for transplant. An advantage of decellularized adult myocardial ECM is that it is tissue specific and considered homologous to the intended diseased heart of the recipient. However, there are several disadvantages as well. Adult ECM consists of collagen (I, II, III, IV, V), elastin, laminin, and fibronectin [2]. Collagen degradation products may have certain toxicities in high concentrations such

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FIGURE 26.1 Approaches for scaffoldbased therapeutic cell delivery. (A) The scaffold is administered first and cells are injected directly into the myocardium through the scaffold (A1) or cells are administered first and then scaffolds placed over the site of injection (A2). (B) Cells are seeded into scaffolds ex vivo just prior to placement on the heart. (C) Cells are preseeded onto scaffolds at a manufacturing facility then shipping to the site for therapeutic administration.

TABLE 26.2

Preclinical Studies of Scaffold-Based Cell Delivery for Cardiac Repair

Scaffold type

Cells delivered

Model

Species

Attachment

Endpoint

References

Bioengineered cardiac matrix (fibronectin)

MSC

MI

Rat

Self-adherent Feasibility

[3]

B-ECM

Neonatal cardiomyocytes

HF

Rat

Suture

Increased contractility

[4]

Small intestine submucosa (pig)

MSC

MI

Rabbit

Suture

Increased contractility

[5]

Pericardium (bovine)

MSC

MI

Rat

Suture

Increased contractility

[6]

Pericardium (human)

Cardiac progenitor cells

Healthy

Rat

Suture

Feasibility

[7]

Myocardium (human) and fibrin hydrogel

MSC

MI

Rat

Suture

Increased contractility

[8]

Fibrin

hESC

MI

Mouse and pig

Fibrin glue

Increased contractility

[9]

Fibrin

ADSC

MI

Rat

Injection

Increased contractility

[10]

Fibrin

iPSC

MI

Pig

Injection

Increased contractility, reduced infarct size

[1]

Fibrin glue

ADSC

MI

Rat

Fibrin glue

Increased vascularity

[11]

Collagen

MSC

MI

Rodent

Fibrin glue

Increased contractility

[12]

Collagen/matrigel

Skeletal muscle cells

MI

Rat

Fibrin glue

Increased contractility

[13]

Collagen

ADSC

MI

Rat and pig Suture

Increased contractility, increased vascularity

[14]

NATURAL

(Continued)

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26. SCAFFOLD-BASED CELL DELIVERY FOR CARDIAC REPAIR

(Continued)

Scaffold type

Cells delivered

Model

Species

Attachment

Endpoint

References

Collagen

MSC

MI

Rat

Suture

Increased contractility, increased infarct thickness

[15]

Collagen/glycosaminoglycan

MSC

MI

Rat

Suture

Increased vascularity

[16]

Collagen/vitronectin

EPC

MI

Rat

Suture

Increased vascularity

[17]

Collagen (bovine)

MSC

MI

Rat

Injection

Failure to improve cardiac function

[18]

Collagen

ADSC

MI

Rat

Injection

Increased contractility

[10]

Bovine serum albumin

MSC

MI

Rat

Fibrin glue

Increased vascularity, decreased infarct size

[19]

Alginate

Neonatal cardiomyocytes

MI

Rat

Suture

Increased vascularity

[20]

Alginate/chitosan

MSC

MI

Rat

Suture

Increased contractility, improved neovascularization

[21]

Collagen/TissueMend

MSC

MI

Mouse

Suture

Feasibility

[22]

Chitosan

ADSC

MI

Rat

Injection

Feasibility

[23]

Silk/hyaluronic acid

MSC

MI

Rat

Fibrin glue

Increased vascularity, increased infarct thickness

[24]

Poly(glycolide-co-caprolactone)

BMMNC

MI

Rat

Suture

Increased contractility

[25]

Poly(lactide-co-ε-caprolactone)

MSC

MI

Rat

Suture

Increased contractility, decreased infarct size

[26]

Poly-glycolic acid

BMMNC

MI

Rat

Suture

Increased contractility

[27]

Poly(lactic-co-glycolic acid)

ADSC

MI

Rat

Injection

Increased contractility

[28,29]

PLGA

MSC

MI

Rat

Suture

Increased contractility

[30]

Polyurethane

Skeletal myoblasts

MI

Rat

Suture

Increased contractility

[13,31]

Poly(ethylene glycol)

iPSC

MI

Mouse

Suture

Increased contractility

[32]

Silanized hydroxypropyl methylcellulose

MSC

MI

Rat

Injection

Increased contractility, decreased infarct size

[33]

Synthetic polysaccharide-based scaffold

MSC

MI

Rat

Glue

Reduced ventricular dimension

[34]

Copolymer P(NIPAMmPEGMA-MDO-MATA)

MSC

MI

Rat

Injection

Feasibility

[35]

Oligo(poly(ethylene glycol) fumarate)

ESC

MI

Rat

Injection

Reduced infarct size

[36]

Poly(ε-caprolactone)

MSC

MI

Rat

Glue

Increased contractility

[37]

PCL/fibronectin

MSC

MI

Rat

Fibrin glue

Increased contractility

[38]

PCL/gelatin

MSC

MI

Rat

Suture

Reduced infarct size, reduced ventricular dimension

[39]

SYNTHETIC

as hydroxyl proline [42]. Furthermore, the relative proportion of collagens changes in disease conditions such as acute myocardial infarction (MI) [43]. Therefore, the variability in the structural protein composition may restrict the suitability of donor hearts to only those

that fall within a narrow collagen composition specification. Xenogeneic sources of myocardium may be an alternative; however, functional similarity and immunologic concerns remain [41]. Another disadvantage is that adult ECM does not naturally adhere to surfaces

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TABLE 26.3 Pros and Cons of Using Myocardium as a Source for Scaffold Delivery of Cells

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neonatal dermal fibroblast-derived ECM for efficient delivery of cardiomyocytes (Theregen, San Francisco, CA) in a rat MI model [4].

Pros

Cons

Tissue specific

Potential toxic degradation

Mucosal Tissues

Homologous

Collagen remodeling in diseased heart

PERICARDIUM

Human source available

Does not naturally adhere to surface

Protein composition varies

Pore structure varies

Both bovine and human pericardium have been described as a cell delivery platform for cardiac disease treatment [6,7,45 47]. The protein composition and pore structure of bovine pericardium is highly consistent [48]. In addition, this material is biocompatible and a potential off-the-shelf product. Typically, bovine pericardium is fixed in glutaraldehyde (GA), which increases the strength and stability of the material [47,48]. Tissues stabilized with 1% or more GA are unsuitable as a cell delivery scaffold because they are resistant to cell infiltration [47]. Pericardium scaffolds also require sutures or glue to affix it to the surface of the heart. Although bovine pericardium is well studied, it is unknown how it will react in the human body, and human pericardium has not been studied for cell delivery scaffolding for cardiac treatment in any cardiac disease animal model. Sung and co-workers have fabricated and documented a bovine pericardium scaffold that delivers MSCs into a rat MI model [6,45,46,49]. Research has shown better ventricular function, prevention of enlargement of the left ventricle (LV) cavity, improvement in LV end-systolic (ES) and end-diastolic (ED) pressures [6]. Tissue regeneration has also been reported in one study [46]. CardioCel (Admedus, Australia) is FDA approved for cardiac indications and is composed of bovine pericardium making it an attractive cell delivery scaffold, but its use as a cell delivery scaffold has not been tested in vivo [47].

and therefore requires sutures or glue for epicardial placement. Finally, pore architecture of decellularized matrix varies depending on the direction of sectioning. Apex-to-base and radial sections exhibit low porosity and closed pores, whereas circumferential sections contain large interconnected pores with smooth channels (Table 26.3). The experience of using decellularized human myocardium as a cell delivery scaffold is limited. GodierFurnemont et al. delivered a cell matrix composite consisting of a fibrin hydrogel, human mesenchymal precursor cells, and decellularized human myocardium obtained from patients with both ischemic and nonischemic cardiomyopathies onto the epicardial surface in a rat MI model [8]. The fibrin hydrogel was required to attach the decellularized myocardium with sutures to the epicardial surface. The composite therapy increased local vascular networks and enhanced cardiac functional recovery. Bioengineered Cardiac Matrix Fibroblasts when cultured at high-density form a unique bioactive, homologous ECM, or in this case, a cardiac ECM [3,4,44]. This B-ECM (bioengineered cardiac matrix) is high in fibronectin (80%) with lesser amounts of collagen types I, II, and III (20%) [3]. The structural protein composition of a B-ECM scaffold is similar to that of ECM observed in the healing phase post-MI and in developing myocardium. B-ECM can be fabricated without chemical cross-linking, which allows the maintenance of the porosity and avoids toxic chemical residue. B-ECM is easily delivered and self-adheres to the epicardium without sutures or adhesives. Finally, B-ECM contains over 18 bioactive molecules with potent angiogenic, immune-modulatory, and proregenerative properties (Figure 26.2) [3]. Mesenchymal stem cells (WiCell, Madison, WI) were delivered by a cardiac fibroblast-derived B-ECM scaffold onto the epicardial surface in a rat MI model [3]. At 48 h, no toxicity was observed and viable transplanted MSCs were observed to be highly retained in the endo-, mid-, and epicardial layers of the myocardium suggesting MSCs are efficiently released off of the scaffold in vivo. Lancaster et al. described a similar

SEROSAL INTESTINAL SUBMUCOSA

Serosal intestinal submucosa (SIS) has been derived from porcine jejunum, but can be derived from other vertebrate animals (Figure 26.3) [5,51,52]. The submucosa is isolated from the layer of smooth muscle and the mucosa and the tissue decellularized. The resulting material has a 3D microstructure and natural porosity allowing for cell, nutrient, and waste movement [51,52]. The protein composition of SIS includes collagen type I, collagen type IV, fibronectin, and laminin [52]. The protein growth factors that are seen in SIS promote angiogenesis, which could be valuable in the treatment of ischemic heart disease [5]. SIS is biodegradable and has been shown to have low immunogenicity. However, SIS is nonadherent and requires suture or glue to affix it to the epicardial surface of the heart. SIS is similar to collagen-based constructs

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26. SCAFFOLD-BASED CELL DELIVERY FOR CARDIAC REPAIR

FIGURE 26.2 (A) Bioengineered cardiac matrix. (B) B-ECM (yellow dotted line) attached to the infarcted mouse heart 48 h posttransplantation. The suture visible in the image was used to ligate the left anterior descending artery. No sutures or glue were used to secure the B-ECM to the epicardial surface. (C) Bioactive molecules present in B-ECM. (D) Breakdown of ECM proteins found in B-EMC ([3], University of Wisconsin-Madison).

FIGURE 26.3 Electron micrograph of mucosal side of decellularized SIS displaying the transformation from (A) untreated (fresh) SIS to (B) decellularized SIS (no treatment) to (C) treated decellularized SIS [50].

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TYPES OF CELL DELIVERY SCAFFOLDS FOR CARDIAC REPAIR

FIGURE 26.4 Formation of fibrin by the protease activity of thrombin on fibrinogen. Thrombin cleaves fibrinopeptides from fibrinogen allowing fibrin monomers to self-assemble, resulting in the polymerization of fibrin.

MI 65 60

(B) 25

MI+P

20

MI+P+C P<0.05

55 P<0.05

50 45 40 35 Baseline

1 week

4 week

Thickening fraction (TF, %)

Ejection fraction (EF, %)

(A)

MI MI+P MI+P+C

P<0.05

15 10

P<0.05

5 0 –5 –10

Infarct zone

Border zone

–15

FIGURE 26.5 Temporal change of LV contractile function in terms of ejection fraction (A) and thickening fraction (B). The cell transplanted group (MI 1 P 1 C) showed significant LV functional improvement as compared to the control groups (MI, MI 1 P) [9].

in that the pore architecture varies geometrically depending on the direction of sectioning [52]. It has been shown that sectioning in the serosal to mucosal direction was found to have porosity four times greater than the porosity in the opposite direction. Tan et al. applied porcine SIS seeded with mesenchymal stem cells in a rabbit MI model [5]. An increase in vascular density and improvement of LV contractile function were reported, however, these effects were not sustained long-term. Although there was no immunologic rejection, some mild inflammation was observed with the presence of lymphocytes and small macrophages in the graft area. SIS has many intriguing properties and continues to be investigated as a potential cell delivery platform. Fibrin Fibrin, also called Factor Ia, plays an important role in blood clotting, but more recently has been investigated as potential cell delivery platform [1,8,10,11,53]. Fibrin is formed by the protease activity of thrombin

on fibrinogen, which results in the polymerization of fibrinogen into fibrin (Figure 26.4) [54]. Fibrin has many advantageous characteristics for a cell delivery platform. It is naturally adhesive, does not illicit an immune response, promotes cell adhesion and local signaling [8,55]. Commercially available fibrin products are available for use as surgical glue [56,57]. In porcine and mouse MI models, Xiong et al. fabricated a fibrin scaffold to deliver hESC-derived endothelial cells and smooth muscle cells [9]. The authors report improved ejection fraction and an increase of neovascularization. Results also showed lower systolic LV wall stresses (Figure 26.5). The significance of replicating mouse model results in swine model results is huge due to the difference in heart rate between mouse and humans. A mouse heart beats  600 times per minute compared to  70 beats per minute for a human heart. There are also arrhythmia concerns with human cells engrafted in a mouse heart.

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A large animal study, such as a pig model, that is closer to how a human heart works that displays these promising results brings this field closer to clinical application. The Transplantation of Human Embryonic Stem Cell-derived Progenitors in Severe Heart Failure (ESCORT) trial is currently underway that combines a fibrin scaffold with human embryonic stem cellderived cardiac progenitors for patients with advanced ischemic cardiomyopathy. Collagen

through the material [63]. There is concern that one of collagen’s breakdown products, hydroxyl proline, may be toxic to cells in high concentrations [42]. Furthermore, collagen-based scaffolds under development today require sutures or glues for epicardial fixation [58]. Arana et al. used a bovine collagen-based type I scaffold to deliver adipose-derived mesenchymal stem cells in rat and pig MI models [14]. The adipose MSCseeded collagen improved cardiac function, decreased fibrosis, and increased vasculogenesis (Table 26.4).

Collagen has been widely investigated as a cell delivery platform for cardiac applications [10,12 18,53,58]. Collagen exists in several distinct subtypes, types I, II, III, IV, and V with collagen type 1 being the most widely abundant [59]. Each collagen subtype has unique physical properties ranging from large fibrillar proteins (types I, II, III) to small stabilizing strut proteins (types IV, V). Collagen is commercially available, inexpensive, and FDA approved for certain applications such as medicated sponges (i.e., TachoSil) and nerve conduits for nerve gap repair (i.e., NeuraGen) [53,56,58,60]. Scaffolds made from collagen can be easily fabricated with tight control over the density of collagen (Figure 26.6) [62]. Collagen scaffolds are nonimmunogenic and biodegradable [53,58]. Collagen also promotes cellular interactions in terms of adhesion and mobility, or the ability of a cell to travel

FIGURE 26.6 (A) Collagen scaffold used in the MAGNUM clinical trial. (B) Scanning electron micrograph of the collagen scaffold used in the MAGNUM clinical trial [61].

TABLE 26.4 Left Ventricle Ejection Fractions (LVEF), End-Systolic Diameters (ESD), End-Diastolic Diameters (EDD), End-Systolic Volumes (ESV), and End-Diastolic Volumes (EDV) in the Rat Model of MI were Recorded and Compared Co

CS

ADSC

CS-ADSC

Preimplant

35.13 6 2.51

36.96 6 1.44

34.55 6 1.87

31.26 6 2.41

Postimplant

38.83 6 3.15

37.37 6 3.13

39.50 6 3.71

47.61 6 3.86

Preimplant

0.77 6 0.02

0.72 6 0.02

0.72 6 0.02

0.71 6 0.02

Postimplant

0.76 6 0.04

0.72 6 0.03

0.74 6 0.04

0.67 6 0.04

Preimplant

0.90 6 0.02

0.85 6 0.02

0.84 6 0.02

0.81 6 0.02

Postimplant

0.89 6 0.03

0.86 6 0.03

0.89 6 0.03

0.85 6 0.03

Preimplant

1.02 6 0.07

0.85 6 0.06

0.84 6 0.06

0.82 6 0.06

Postimplant

1.02 6 0.14

0.89 6 0.09

0.96 6 0.12

0.77 6 0.13

Preimplant

1.56 6 0.10

1.33 6 0.08

1.29 6 0.09

1.19 6 0.07

Postimplant

1.56 6 0.16

1.39 6 0.11

1.54 6 0.14

1.38 6 0.13

LVEF (%)

ESD (CM)

EDD (CM)

ESV (ML)

EDV (ML)

Statistically significant differences are noted by collagen scaffold seeded with ADSC [14].



(P , 0.05) and



(P , 0.01) in terms of mean 6 SEM. Abbreviations: Co, control; CS, collagen scaffold; CS-ADSC,

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Other Natural Materials Other natural polymers considered for scaffold delivery of stem cells include bovine serum albumin (BSA), alginate, TissueMend, chitosan, silk, and hyaluronic acid, among others [19 24,64]. These materials have similar features to other natural materials such as favorable biocompatibility and defined porous structures, usually in a uniform distribution [20,65]. They are all biodegradable.

Synthetic Synthetic materials are able to vary widely in their characteristics for better control over the material [2]. Compared to natural materials, synthetic materials are more consistent in their composition between batches. Their mechanical properties (i.e., stiffness, elasticity, and porosity) have the ability to be precisely controlled, unlike natural materials. Their biocompatibility and biodegradability, however, depend on the material used. Selected materials are discussed later. Poly-glycolic Acid and Poly-lactic Acid Many scaffolds have been developed for cardiac treatment based on poly-glycolic acid (PGA), polylactic acid (PLA), or a combination of both [25 30,66]. PGA and PLA have many favorable properties. First, these materials are elastic and suitable for cardiac delivery. Second, the mechanical properties are easily manipulated by modifying cross-linking [25,26]. Third, these materials have also been shown to have favorable cellular interactions [25,26,28,29,53]. PGA- and PLAbased materials have defined porous structures, which is advantageous for cell and nutrient transport [65]. These materials are biodegradable with nontoxic byproducts [26,29,53]. By changing the ratio of lactic and glycolic acids, the degradation rate can be modified [29,53]. Certain compounds that consist of these materials are FDA approved for drug delivery approaches. However, potential toxicity in the form of a foreign body reaction pattern has been described [67]. Piao et al. developed a poly(glycolide-co-caprolactone) (PGCL) scaffold to support the delivery of bone marrow-derived mononuclear cells (BMMNCs) in a rat MI model [25]. Treated rats showed improved fractional shortening and reduced left ventricular end-diastolic pressure, end-diastolic and end-systolic diameters. Polyurethane Polyurethane (PU) has favorable mechanical properties and is reproducibly manufactured [68]. PU is biocompatible and can be fabricated into a highly porous material depending on the solvent used to create the

335

pores [68,69]. Pores are randomly distributed throughout the scaffold in a nonuniform manner [68]. The best solvent for the preparation of PU scaffolds appears to be dimethylformamide (DMF) [69]. Other solvents can give numerous, but closed pores, which do not allow proper nutrient and cell transport. PU scaffolds are degraded by hydrolysis [68]. However, the time frame for PU degradation may be as long as 5 years, which may not be appealing for cardiac delivery due to infection risk. Giraud et al. developed a PU (Artelon) scaffold seeded with myoblasts that was highly porous but rigid in structure [13,31]. In a rat MI model, early improvements in contractile function and ejection fraction were observed with PU compared to controls. However, in a subsequent longer term study, PU offered no effects on structural remodeling. Although a delay in the progression toward heart failure was observed, this effect disappeared after 12 months [70]. Poly(Ethylene Glycol) Poly(ethylene glycol) (PEG) has been manufactured and tested for therapeutic delivery for cardiac repair [71]. PEG degrades rapidly as a hydrogel when cross-linked. Usually, a natural material is cross-linked with PEG to confer bioactivity to the seeded cells which manufactured PEG cannot normally give [72]. For example, a PEG-fibrinogen cross-linked hydrogel scaffold has malleable mechanical properties and is biodegradable [32]. It is believed that such a cross-link can curb cardiac remodeling via simple ratio modifications to the scaffold material composition [72]. However, chemical crosslinking may also have unintended negative effects depending on the degree of cross-linking [2]. Bearzi et al. made a composite hydrogel scaffold of PEG-fibrinogen seeded with both iPS cells engineered to secrete MMP9 and iPS cells engineered to secrete PlGF [32]. After optimization of its stiffness, the scaffold was implanted in a mouse MI model. Results showed that the velocity/time integral (VTI), which reflects the velocity of blood flow in the left ventricular outflow tract, was nearly restored to physiologic values. Other Synthetic Materials Other synthetic polymers considered for scaffold delivery of stem cells include poly(e-caprolactone) (PCL), polyamide (PA), silanized hydroxypropyl methylcellulose (Si-HPMC), poly(N-isopropylacrylamide) (PNIPAAM), polysaccharide-based scaffold (pullulan and dextran), oligo(poly(ethylene glycol) fumarate) (OPF), and poly(ether)urethane-polydimethylsiloxane (PEtU-PDMS) [33 39,66,73]. These materials have been shown to induce a classical foreign body reaction pattern when implanted on the epicardial surface of healthy rats [67]. Cross-linking or coating these

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scaffolds with other materials may resolve this issue. For example, a MSC-seeded plasma-coated PCL scaffold stabilized cardiac function in one study [37].

Scaffold Delivery Methods Scaffold delivery should be simple and easily implemented into the clinical workflow (Figure 26.7). For example, direct application of a cell-scaffold construct onto the epicardial surface through a sternotomy requires no specialized imaging or delivery tools [74]. However, this approach is invasive and regions of the heart such as the interventricular septum are inaccessible. This approach may also require the use of either suture or glue to affix the material to the heart [3,6,9,12,13,45,49,58,75 84]. Suturing the scaffold may increase damage to the heart and risk perforation. Glues may reduce the permeation of homing signals to the cells loaded onto the materials. This may prevent cell homing to the injured tissue. Thoracoscopicguided cell-scaffold administration is less invasive but increases the number of inaccessible locations such as the posterior wall. Schmuck et al. [3] described a selfadhering scaffold that does not require sutures or glues and may be promising implementation of an epicardial patch.

An alternative delivery method currently under investigation consists of fragmenting the cell-scaffold construct into a suspension thus allowing for direct intramyocardial injection [74,85]. Once in suspension, the cell-scaffold construct can be delivered via epicardial injection or transendocardial injection using a steerable catheter. The latter method is perhaps the most appealing, as it is minimally invasive and allows for access to the whole ventricle, including the interventricular septum and posterior wall. Precision targeting of transendocardial injections may be performed with a multimodality image coregistration system. Alternatively, electro-anatomic mapping may be used for interventional imaging guidance and is discussed in detail elsewhere.

Human Trials of Cell-Seeded Scaffolds for Cardiac Repair The Myocardial Assistance by Grafting New Bioartificial Upgraded Myocardium (MAGNUM) trial administered collagen I matrix seeded with bone marrow mononuclear cells (Figure 26.8A) in patients with ischemic cardiomyopathy with prior MI [61,86]. Bone marrow mononuclear cells were injected into scar followed by epicardial application of collagen I matrix seeded with bone marrow mononuclear cells. The matrix-cell graft was sutured to the infarct location. Comparisons were made to patients who received just an injection of human bone marrow mononuclear cells (n 5 10 for both groups) (Table 26.5). Improvements in ejection fraction, scar thickness area, and end-diastolic volume were seen in the scaffold-cell group. Heart failure functional class also improved in patients who received the scaffold-cell treatment. As previously mentioned, the Transplantation of Human Embryonic Stem Cell-derived Progenitors in Severe Heart Failure (ESCORT) clinical trial is currently underway in Europe testing the combination of embryonic stem cellderived cardiac precursor cells with a fibrin scaffold.

CONCLUSION

FIGURE 26.7 Schematic showing clinical workflow for scaffoldbased cell delivery.

Scaffold-based cell delivery has great potential for treating heart disease. There are a number of natural and synthetic scaffolds under development and each have unique properties. Porosity, compliance, and tissue adherence are among the properties that need to be considered when engineering an “ideal” biomaterial for cell delivery. Preclinical studies to date have shown tremendous promise, yet human experience using this approach remains limited.

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FIGURE 26.8 (A) Photograph of a collagen matrix seed with bone marrow mononuclear cells sutured onto the epicardial surface of a human heart [86]. (B) Photograph of a cardiac fibroblast-derived B-ECM loaded with mesenchymal stem cells that is self-adhered onto the epicardial surface of a pig heart. (Schmuck et al., University of Wisconsin-Madison). TABLE 26.5 Echocardiogram and Single Photon Emission Computed Tomography (SPECT) Radioisotropic Analysis of Myocardial Geometry, Function and Viability in the MAGNUM Trial Endpoint

CS 1 BMC (n 5 10)

BMC (n 5 10)

P value

Change in EF% (echo)

6.7 6 5.3

7.4 6 7.5

NS

Change in LVEDV (echo)

229.5 6 12.3 9.8 6 12.1

Change in LVESV (echo)

240.1 6 10.1 214.4 6 14.9 ,0.01

% of cardiac segments with improved viability and contractility (SPECT)

62% 6 5%

58% 6 9%

,0.01

0.06

CS, collagen scaffold; BMC, bone marrow cell; EF, ejection fraction; LVEDV, left ventricular end-diastolic volume; LVESV, left ventricular end-systolic volume (adapted from Ref. [86]).

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