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Ileal interposition (IT) improves insulin sensitivity in the obese Zucker rat (ZR)
S12
Surgical Forum Abstracts
J Am Coll Surg
suggest that isolation of a critical length of proximal bowel is important to abolish IGS and alter intestinal glucose absorptive capacity. This underscores the need for clinical studies to evaluate the optimal length of BP limb, especially in T2DM patients.
tory has shown that adhesions are reduced by mechanisms that upregulate peritoneal fibrinolysis and antioxidant capacity. NAC is a clinically relevant antioxidant that increases intracellular glutathione levels. Its ability to reduce adhesions is unknown. We hypothesized that NAC would reduce adhesions, increase peritoneal fibrinolysis, and increase antioxidant defenses.
Intestinal alkaline phosphatase targeting of lipopolysaccharide
INTRODUCTION: Intestinal alkaline phosphatase (IAP), an intestinal brush border enzyme, has been shown to protect the host from gut-derived sepsis, but its precise mechanism of action remains unclear. The present studies were designed to identify a specific target for IAP.
METHODS: Abdominal adhesions were induced using our ischemic button model. Male Wistar rats (n ⫽ 40) were randomized to 3 groups: nonoperative controls (NON-OP), operative controls (OP ⫹ saline), or operative treatment with NAC (OP ⫹ NAC). Operated animals were administered either 1 mL normal saline (vehicle) or NAC (150 mg/kg) intraperitoneally BID on preoperative day 1, operative day, and postoperative day 1. Animals were sacrificed at 7 days for adhesion scoring or at 24 hours to evaluate peritoneal fluid for fibrinolytic activity and tissue for tissue plasminogen activator (tPA) and plasminogen activator inhibitor-1 (PAI-1) mRNA by RT-PCR, glutathione (GSH), and the oxidative stress marker, 8-isoprostane, by ELISA.
METHODS: HT29 cells were stably transfected with an IAP expression plasmid, and enzyme activity was confirmed biochemically. Parent and IAP-expressing cells were incubated with Escherichia coli, Salmonella typhimurium, or Listeria monocytogenes. IL-8 secretion was measured by ELISA. In a separate series of experiments, live and heat-killed bacteria were incubated ⫹/– exogenous IAP and used to stimulate HT29 cells.
RESULTS: NAC treatment reduced adhesions by 60% (p⬍0.001) and increased peritoneal fibrinolytic activity by 277% (p⫽0.007). Peritoneal tPA mRNA levels increased by nearly 30% with NAC. Surgery reduced peritoneal adhesion tissue GSH levels by 92%, but NAC reconstituted GSH levels 4-fold to 33% of NON-OP (p ⫽ 0.020) and decreased the oxidative stress marker, 8-isoprostane, by 55% (p⫽0.009).
Kathryn Chen MD, Madhu Malo MD, Skye Zeller BS, Paul Johnson BS, Golam Mostafa MD, Sayeda Alam MD, Sundaram Ramasamy PhD, Elizabeth Hohmann MD, Richard A Hodin MD, FACS Massachusetts General Hospital, Boston, MA
RESULTS: Parent HT-29 cells displayed a dramatic response to bacterial incubation, with elevated IL-8 levels in response to E coli, S typhimurium, and L monocytogenes (⬎10-fold increase; p⬍ 0.005 in all cases). In contrast, IAP conferred protection (⬎50% inhibitions) from the 2 gram-negative strains but had no effect on gram-positive L monocytogenes. IAP did not inhibit the IL-8 response of HT29 cells to live E coli but attenuated the response to heat-killed E coli (⬎50% inhibition). CONCLUSIONS: Intestinal epithelial cells released IL-8 in response to a variety of bacteria, but IAP expression conferred protection against this response in gram-negative bacteria only. There was no appreciable effect of IAP on cytokine response induced by live E coli, but when the same bacteria were heat-killed, thereby exposing LPS, the response was inhibited. These observations suggest that IAP as a gut defense factor does not directly target live bacteria, but rather LPS specifically.
N-acetyl-L-cysteine (NAC) reduces intraabdominal adhesion formation through the upregulation of peritoneal fibrinolytic activity and antioxidant defenses Daniel I Chu MD, Rizal Lim MD, Stanley Heydrick PhD, Karen L Reed PhD, Arthur F Stucchi PhD, James M Becker MD, FACS Boston University Medical Center, Boston, MA INTRODUCTION: Intraabdominal adhesions are a major source of morbidity, and efforts to prevent them remain limited. Our labora-
OP ⴙ Saline
OP ⴙ NAC
Adhesion formation
83.3% ⫾ 5.1%
33.3% ⫾ 4.9%
⬍ .001
Fibrinolytic activityⴱ
594% ⫾ 226%
1646% ⫾ 215%
.007
tPA mRNAⴱ
97.3% ⫾ 11.3%
124% ⫾ 13.3%
.160
PAI-1 mRNAⴱ
918% ⫾ 187%
1123% ⫾ 187%
.456
Reduced GSHⴱ
8.4% ⫾ 6.5%
33.6% ⫾ 6.5%
.020
8-isoprostaneⴱ
405% ⫾ 77%
181% ⫾ 13%
.009
P-Valueⴱⴱ
ⴱExpressed as percentage of NON-OP controls (mean ⫾ SEM). ⴱⴱComparison of OP ⫹ Saline vs OP ⫹ NAC.
CONCLUSIONS: NAC administered intraperitoneally reduces adhesion formation while up-regulating peritoneal fibrinolytic activity and antioxidant defenses. These data suggest a potentially new therapeutic use for NAC in adhesion prevention.
Ileal interposition (IT) improves insulin sensitivity in the obese Zucker rat (ZR) Derek M Culnan MD, Vance Albaugh, Mingjie Sun, Charles Lang, Christopher J Lynch, Robert N Cooney, FACS Pennsylvania State University College of Medicine, Hershey Medical Center, Hershey, PA INTRODUCTION: Type 2 diabetes (T2DM) is caused by insulin resistance and beta-cell dysfunction. Enhanced ileal exposure to nutrients is posited as a mechanism for rapid improvement in T2DM after gastric bypass. Our study examines the effects of IT on glucose tolerance, insulin sensitivity, and tissue glucose uptake in obese ZRs. METHODS: Two groups of obese ZRs were studied: IT and sham surgery fed ad lib (AL). IT rats had a 10-cm segment of neurovascularly intact terminal ileum interposed 10 cm from Treitz. Oral glu-
Vol. 209, No. 3S, September 2009
cose tolerance tests were done 5 weeks postoperatively, and the area under the curve (AUC) was calculated. Body composition was determined via magnetic resonance at 8 weeks postoperatively. Hyperinsulinemic euglycemic clamps were performed week 8 using a maximally stimulating dose of insulin. Data are mean ⫾ SE, n ⫽ 15 (IT); 10 (AL), P ⬍ 0.05 vs AL by Student t test. RESULTS: Food intake, body weight, and body composition were similar between the groups. The IT AUC (8,592 ⫾ 891.4) was significantly lower than the AL AUC (21,440 ⫾ 2,139) 5 weeks postoperatively. Basal hepatic glucose output and fasting blood glucose were similar between the groups, but insulin-stimulated glucose disposal in the IT rats (10.2 ⫾ 2.5) was 3-fold that of the AL group (3.3 ⫾ 0.8). Insulin-stimulated glucose uptake in muscle was increased after IT vs AL, whereas adipose tissues were not improved. CONCLUSIONS: Enhanced nutrient delivery to the ileum following IT improves glucose tolerance, insulin sensitivity, and insulinstimulated muscle glucose uptake in the obese ZR, independent of food intake, body weight, or body composition.
Human dermal tissue allograft use in treating chronic pilonidal sinus Jill Elisabeth Clark MD, James J Matino MD, Saumitra R Banerjee MD, Steven H Brown MD Saint Francis Hospital and Medical Center, Hartford, CT INTRODUCTION: We developed a novel, minimally invasive approach to treating chronic pilonidal sinus using a human dermal tissue allograft with primary closure. METHODS: From January 2007 to September 2008, 54 patients underwent 55 operations for pilonidal disease at our institution. All underwent pore excision, curettage, injection of a human dermal tissue allograft, and primary wound closure. End points at 6 months included narcotic requirement, loss of time from work or school, wound complications, and disease recurrence. RESULTS: Sixty-five percent of patients required no narcotic use postoperatively. Only 10 % required more than 3 narcotic doses postoperatively. Eighty percent of patients returned to work or school on the first postoperative day; 10% of patients missed a week or more of work, although this group admitted that the additional time off was unnecessary in terms of surgical recovery. Thirty-three percent developed minor wound complications that responded to suture removal, drainage, and/or antibiotics. Ten percent suffered major wound failure requiring secondary wound healing. To date, the total disease recurrence rate is 12.5%. CONCLUSIONS: This study is the first to use a human dermal tissue allograft in treating pilonidal disease. The short-term results indicate improved outcome when compared with other methods of treating uncomplicated pilonidal sinus. Patient selection is key. This technique holds promise to become the procedure of choice for patients suffering from uncomplicated sinus disease. Long-term recurrence rates up to at least three years will guide us in improving patient selection and determining the true efficacy of this conservative approach to pilonidal sinus disease.
Surgical Forum Abstracts
S13
Mesoscopic traffic flow analysis of platelet dynamics in chemically induced murine colitis Lino F Miele MD, MS, Grace S Lee MD, Miao Lin MD, Aslihan Turhan PhD, Akira Tsuda PhD, Moritz A Konerding MD, Dennis P Orgill MD, PhD, FACS, Steven J Mentzer MD, FACS Brigham and Women’s Hospital, Harvard Medical School, Boston, MA INTRODUCTION: In the inflammatory colon, intravital microscopy of the mucosal plexus reveals striking perturbations in blood flow, often demonstrating discontinuous and even bidirectional blood flow. Growing evidence that microcirculatory perturbations are involved in the pathogenesis of inflammatory bowel disease suggests the importance of flexible analytical methods to characterize these complex blood flow patterns. METHODS: Colitis was induced in C57BL/6 mice with 3% dextran sodium sulfate (DSS) in drinking water for 6 days or with picryl chloride (TNCB) sensitized 2.5% trinitrobenzene sulfonic acid (TNBS) via rectal installation. After intravascular injection of 6 ⫻ 10e8 500-nm fluorescent tracers, the mucosal plexus was studied using fluorescence intravital videomicroscopy. Tracer flux was quantified using traffic flow theory–inspired space-time plots, tracer flux step curves, and ANOVA. Gene expression of platelet chemokines were analyzed with quantitative RT-PCR. RESULTS: Traffic flow analysis, applied to the flow paths of intravascular tracers within the mucosal plexus, demonstrated comparable total flux in colitis and control mice but significant variability in tracer flux within inflamed mucosal plexus (ANOVA; p⬍.05). The spatial association of platelet aggregates and blood flow exclusion (p⬍.01) suggested a platelet-mediated mechanism. Further, blood flow perturbations were temporally associated with the gene expression of multiple platelet agonists within the mucosal plexus (CCL3, CXCL1, CCL2, CXCL5, CCL7, CCL8, and IL-1b; p⬍.01). CONCLUSIONS: Traffic flow theory facilitated characterization of blood flow heterogeneity within the inflammatory colon mucosal plexus. The traffic analysis provided a spatial and temporal link to platelet aggregation as the primary mechanism of blood flow perturbations in murine chemically induced colitis.
Peptide absorption after massive proximal small bowel resection: Mechanisms of ileal adaptation Hisham G Qandeel MBBS, David J Hernandez, Fernando Alonso, Judith A Duenes BA, Ye Zheng MD, PhD, Michael G Sarr MD, FACS Mayo Clinic College of Medicine, Rochester, MN INTRODUCTION: Protein absorption in the gut occurs primarily as di- and tri-peptides exclusively via the H⫹/peptide cotransporter-1 (PEPT1) and is highly adaptable. Our aim was to identify mechanisms of ileal adaptation for peptide absorption after massive proximal bowel resection. We hypothesized that ileal adaptation in uptake of short peptides is mediated through cellular up-regulation of gene expression of PEPT1.
Surgical Forum Abstracts
J Am Coll Surg
suggest that isolation of a critical length of proximal bowel is important to abolish IGS and alter intestinal glucose absorptive capacity. This underscores the need for clinical studies to evaluate the optimal length of BP limb, especially in T2DM patients.
tory has shown that adhesions are reduced by mechanisms that upregulate peritoneal fibrinolysis and antioxidant capacity. NAC is a clinically relevant antioxidant that increases intracellular glutathione levels. Its ability to reduce adhesions is unknown. We hypothesized that NAC would reduce adhesions, increase peritoneal fibrinolysis, and increase antioxidant defenses.
Intestinal alkaline phosphatase targeting of lipopolysaccharide
INTRODUCTION: Intestinal alkaline phosphatase (IAP), an intestinal brush border enzyme, has been shown to protect the host from gut-derived sepsis, but its precise mechanism of action remains unclear. The present studies were designed to identify a specific target for IAP.
METHODS: Abdominal adhesions were induced using our ischemic button model. Male Wistar rats (n ⫽ 40) were randomized to 3 groups: nonoperative controls (NON-OP), operative controls (OP ⫹ saline), or operative treatment with NAC (OP ⫹ NAC). Operated animals were administered either 1 mL normal saline (vehicle) or NAC (150 mg/kg) intraperitoneally BID on preoperative day 1, operative day, and postoperative day 1. Animals were sacrificed at 7 days for adhesion scoring or at 24 hours to evaluate peritoneal fluid for fibrinolytic activity and tissue for tissue plasminogen activator (tPA) and plasminogen activator inhibitor-1 (PAI-1) mRNA by RT-PCR, glutathione (GSH), and the oxidative stress marker, 8-isoprostane, by ELISA.
METHODS: HT29 cells were stably transfected with an IAP expression plasmid, and enzyme activity was confirmed biochemically. Parent and IAP-expressing cells were incubated with Escherichia coli, Salmonella typhimurium, or Listeria monocytogenes. IL-8 secretion was measured by ELISA. In a separate series of experiments, live and heat-killed bacteria were incubated ⫹/– exogenous IAP and used to stimulate HT29 cells.
RESULTS: NAC treatment reduced adhesions by 60% (p⬍0.001) and increased peritoneal fibrinolytic activity by 277% (p⫽0.007). Peritoneal tPA mRNA levels increased by nearly 30% with NAC. Surgery reduced peritoneal adhesion tissue GSH levels by 92%, but NAC reconstituted GSH levels 4-fold to 33% of NON-OP (p ⫽ 0.020) and decreased the oxidative stress marker, 8-isoprostane, by 55% (p⫽0.009).
Kathryn Chen MD, Madhu Malo MD, Skye Zeller BS, Paul Johnson BS, Golam Mostafa MD, Sayeda Alam MD, Sundaram Ramasamy PhD, Elizabeth Hohmann MD, Richard A Hodin MD, FACS Massachusetts General Hospital, Boston, MA
RESULTS: Parent HT-29 cells displayed a dramatic response to bacterial incubation, with elevated IL-8 levels in response to E coli, S typhimurium, and L monocytogenes (⬎10-fold increase; p⬍ 0.005 in all cases). In contrast, IAP conferred protection (⬎50% inhibitions) from the 2 gram-negative strains but had no effect on gram-positive L monocytogenes. IAP did not inhibit the IL-8 response of HT29 cells to live E coli but attenuated the response to heat-killed E coli (⬎50% inhibition). CONCLUSIONS: Intestinal epithelial cells released IL-8 in response to a variety of bacteria, but IAP expression conferred protection against this response in gram-negative bacteria only. There was no appreciable effect of IAP on cytokine response induced by live E coli, but when the same bacteria were heat-killed, thereby exposing LPS, the response was inhibited. These observations suggest that IAP as a gut defense factor does not directly target live bacteria, but rather LPS specifically.
N-acetyl-L-cysteine (NAC) reduces intraabdominal adhesion formation through the upregulation of peritoneal fibrinolytic activity and antioxidant defenses Daniel I Chu MD, Rizal Lim MD, Stanley Heydrick PhD, Karen L Reed PhD, Arthur F Stucchi PhD, James M Becker MD, FACS Boston University Medical Center, Boston, MA INTRODUCTION: Intraabdominal adhesions are a major source of morbidity, and efforts to prevent them remain limited. Our labora-
OP ⴙ Saline
OP ⴙ NAC
Adhesion formation
83.3% ⫾ 5.1%
33.3% ⫾ 4.9%
⬍ .001
Fibrinolytic activityⴱ
594% ⫾ 226%
1646% ⫾ 215%
.007
tPA mRNAⴱ
97.3% ⫾ 11.3%
124% ⫾ 13.3%
.160
PAI-1 mRNAⴱ
918% ⫾ 187%
1123% ⫾ 187%
.456
Reduced GSHⴱ
8.4% ⫾ 6.5%
33.6% ⫾ 6.5%
.020
8-isoprostaneⴱ
405% ⫾ 77%
181% ⫾ 13%
.009
P-Valueⴱⴱ
ⴱExpressed as percentage of NON-OP controls (mean ⫾ SEM). ⴱⴱComparison of OP ⫹ Saline vs OP ⫹ NAC.
CONCLUSIONS: NAC administered intraperitoneally reduces adhesion formation while up-regulating peritoneal fibrinolytic activity and antioxidant defenses. These data suggest a potentially new therapeutic use for NAC in adhesion prevention.
Ileal interposition (IT) improves insulin sensitivity in the obese Zucker rat (ZR) Derek M Culnan MD, Vance Albaugh, Mingjie Sun, Charles Lang, Christopher J Lynch, Robert N Cooney, FACS Pennsylvania State University College of Medicine, Hershey Medical Center, Hershey, PA INTRODUCTION: Type 2 diabetes (T2DM) is caused by insulin resistance and beta-cell dysfunction. Enhanced ileal exposure to nutrients is posited as a mechanism for rapid improvement in T2DM after gastric bypass. Our study examines the effects of IT on glucose tolerance, insulin sensitivity, and tissue glucose uptake in obese ZRs. METHODS: Two groups of obese ZRs were studied: IT and sham surgery fed ad lib (AL). IT rats had a 10-cm segment of neurovascularly intact terminal ileum interposed 10 cm from Treitz. Oral glu-
Vol. 209, No. 3S, September 2009
cose tolerance tests were done 5 weeks postoperatively, and the area under the curve (AUC) was calculated. Body composition was determined via magnetic resonance at 8 weeks postoperatively. Hyperinsulinemic euglycemic clamps were performed week 8 using a maximally stimulating dose of insulin. Data are mean ⫾ SE, n ⫽ 15 (IT); 10 (AL), P ⬍ 0.05 vs AL by Student t test. RESULTS: Food intake, body weight, and body composition were similar between the groups. The IT AUC (8,592 ⫾ 891.4) was significantly lower than the AL AUC (21,440 ⫾ 2,139) 5 weeks postoperatively. Basal hepatic glucose output and fasting blood glucose were similar between the groups, but insulin-stimulated glucose disposal in the IT rats (10.2 ⫾ 2.5) was 3-fold that of the AL group (3.3 ⫾ 0.8). Insulin-stimulated glucose uptake in muscle was increased after IT vs AL, whereas adipose tissues were not improved. CONCLUSIONS: Enhanced nutrient delivery to the ileum following IT improves glucose tolerance, insulin sensitivity, and insulinstimulated muscle glucose uptake in the obese ZR, independent of food intake, body weight, or body composition.
Human dermal tissue allograft use in treating chronic pilonidal sinus Jill Elisabeth Clark MD, James J Matino MD, Saumitra R Banerjee MD, Steven H Brown MD Saint Francis Hospital and Medical Center, Hartford, CT INTRODUCTION: We developed a novel, minimally invasive approach to treating chronic pilonidal sinus using a human dermal tissue allograft with primary closure. METHODS: From January 2007 to September 2008, 54 patients underwent 55 operations for pilonidal disease at our institution. All underwent pore excision, curettage, injection of a human dermal tissue allograft, and primary wound closure. End points at 6 months included narcotic requirement, loss of time from work or school, wound complications, and disease recurrence. RESULTS: Sixty-five percent of patients required no narcotic use postoperatively. Only 10 % required more than 3 narcotic doses postoperatively. Eighty percent of patients returned to work or school on the first postoperative day; 10% of patients missed a week or more of work, although this group admitted that the additional time off was unnecessary in terms of surgical recovery. Thirty-three percent developed minor wound complications that responded to suture removal, drainage, and/or antibiotics. Ten percent suffered major wound failure requiring secondary wound healing. To date, the total disease recurrence rate is 12.5%. CONCLUSIONS: This study is the first to use a human dermal tissue allograft in treating pilonidal disease. The short-term results indicate improved outcome when compared with other methods of treating uncomplicated pilonidal sinus. Patient selection is key. This technique holds promise to become the procedure of choice for patients suffering from uncomplicated sinus disease. Long-term recurrence rates up to at least three years will guide us in improving patient selection and determining the true efficacy of this conservative approach to pilonidal sinus disease.
Surgical Forum Abstracts
S13
Mesoscopic traffic flow analysis of platelet dynamics in chemically induced murine colitis Lino F Miele MD, MS, Grace S Lee MD, Miao Lin MD, Aslihan Turhan PhD, Akira Tsuda PhD, Moritz A Konerding MD, Dennis P Orgill MD, PhD, FACS, Steven J Mentzer MD, FACS Brigham and Women’s Hospital, Harvard Medical School, Boston, MA INTRODUCTION: In the inflammatory colon, intravital microscopy of the mucosal plexus reveals striking perturbations in blood flow, often demonstrating discontinuous and even bidirectional blood flow. Growing evidence that microcirculatory perturbations are involved in the pathogenesis of inflammatory bowel disease suggests the importance of flexible analytical methods to characterize these complex blood flow patterns. METHODS: Colitis was induced in C57BL/6 mice with 3% dextran sodium sulfate (DSS) in drinking water for 6 days or with picryl chloride (TNCB) sensitized 2.5% trinitrobenzene sulfonic acid (TNBS) via rectal installation. After intravascular injection of 6 ⫻ 10e8 500-nm fluorescent tracers, the mucosal plexus was studied using fluorescence intravital videomicroscopy. Tracer flux was quantified using traffic flow theory–inspired space-time plots, tracer flux step curves, and ANOVA. Gene expression of platelet chemokines were analyzed with quantitative RT-PCR. RESULTS: Traffic flow analysis, applied to the flow paths of intravascular tracers within the mucosal plexus, demonstrated comparable total flux in colitis and control mice but significant variability in tracer flux within inflamed mucosal plexus (ANOVA; p⬍.05). The spatial association of platelet aggregates and blood flow exclusion (p⬍.01) suggested a platelet-mediated mechanism. Further, blood flow perturbations were temporally associated with the gene expression of multiple platelet agonists within the mucosal plexus (CCL3, CXCL1, CCL2, CXCL5, CCL7, CCL8, and IL-1b; p⬍.01). CONCLUSIONS: Traffic flow theory facilitated characterization of blood flow heterogeneity within the inflammatory colon mucosal plexus. The traffic analysis provided a spatial and temporal link to platelet aggregation as the primary mechanism of blood flow perturbations in murine chemically induced colitis.
Peptide absorption after massive proximal small bowel resection: Mechanisms of ileal adaptation Hisham G Qandeel MBBS, David J Hernandez, Fernando Alonso, Judith A Duenes BA, Ye Zheng MD, PhD, Michael G Sarr MD, FACS Mayo Clinic College of Medicine, Rochester, MN INTRODUCTION: Protein absorption in the gut occurs primarily as di- and tri-peptides exclusively via the H⫹/peptide cotransporter-1 (PEPT1) and is highly adaptable. Our aim was to identify mechanisms of ileal adaptation for peptide absorption after massive proximal bowel resection. We hypothesized that ileal adaptation in uptake of short peptides is mediated through cellular up-regulation of gene expression of PEPT1.