• 中國醫(yī)科大學附屬第一醫(yī)院普通外科教研室器官移植科(沈陽 110001);

目的 　探討在細胞因子與大鼠胰島細胞共同培養(yǎng)過程中，誘導型一氧化氮合酶(iNOS)抑制劑氨基胍對胰島細胞功能和存活的影響及其機理。
方法 　分離純化大鼠胰島，進行胰島細胞培養(yǎng)。根據(jù)培養(yǎng)基中是否加入氨基胍或細胞因子IL-1β和TNF-α，按隨機對照原則分為空白對照組(完全培養(yǎng)基)、細胞因子組(加IL-1β和TNF-α)、氨基胍組(加氨基胍)及氨基胍＋細胞因子組(加氨基胍及細胞因子)。檢測指標包括： 培養(yǎng)液中NO水平、胰島組織中iNOS活性、胰島細胞存活情況（丫啶橙/溴乙錠染色）、胰島細胞凋亡情況（TUNEL法）及胰島功能（胰島素釋放試驗）。
結(jié)果 　與空白對照組比較,細胞因子組大鼠胰島組織中iNOS的活性明顯提高，培養(yǎng)液中NO的水平明顯上升，同時胰島細胞的存活率下降，大量細胞凋亡，胰島素分泌明顯減少(P＜0.01)。與細胞因子組比較,氨基胍＋細胞因子組的iNOS的活性〔（3.17±0.51） U/ml比（38.93±4.72） U/ml〕及NO水平〔（50.5±10.4） μmol/L比（313.0±35.4） μmol/L〕明顯下降，胰島細胞存活率活明顯升高〔（72.73±3.14）%比（57.07±5.07）%〕，凋亡率明顯下降〔（20.11±8.48）%比（41.17±6.87）%〕，胰島素分泌指數(shù)明顯升高(3.50±0.27比1.96±0.19),差異均有統(tǒng)計學意義(P＜0.01)。
結(jié)論 　氨基胍通過抑制iNOS活性，控制NO過量產(chǎn)生，從而減輕細胞因子對胰島的損害，改善胰島的存活與功能。

引用本文： 李柏峰,劉永鋒,程穎,張佳林. 氨基胍對大鼠胰島保護作用的實驗研究. 中國普外基礎(chǔ)與臨床雜志, 2009, 16(9): 729-733. doi: 復制

1. Kwon KB, Kim EK, Jeong ES, et al. Cortex cinnamomi extract prevents streptozotocin- and cytokine-induced beta-cell damage by inhibiting NF-kappaB [Ｊ]. World J Gastroenterol， 2006; 12(27): 4331-4337.
2. Mosén H, Salehi A, Henningsson R, et al. Nitric oxide inhibits, and carbon monoxide activates, islet acid alpha-glucoside hydrolase activities in parallel with glucose-stimulated insulin secretion [Ｊ]. J Endocrinol， 2006; 190(3): 681-693.
3. Wahoff DC, Sutherland DE, Hower CD, et al. Free intraperitoneal islet autografts in pancreatectomized dogs-impact of islet purity and posttransplantation exogenous insulin [Ｊ]. Surgery， 1994; 116(4): 742-748.
4. Shapiro AM, Lakey JR, Ryan EA, et al. Islet transplantation in seven patients with type 1 diabetes mellitus using a glucocorticoid-free immunosuppressive regimen [Ｊ]. N Engl J Med， 2000; 343(4): 230-238.
5. Frank A, Deng S, Huang X, et al. Transplantation for type Ⅰ diabetes: comparison of vascularized whole-organ pancreas with isolated pancreatic islets [Ｊ]. Ann Surg， 2004; 240(4): 631-640.
6. 劉方峰, 程穎, 劉永鋒, 等. 激活型Akt1轉(zhuǎn)染大鼠胰島對移植物凋亡和再血管化的影響 [Ｊ]. 中國普外基礎(chǔ)與臨床雜志, 2009; 16(1): 28-31.
7. Ozmen L, Ekdahl KN, Elgue G, et al. Inhibition of thrombin abrogates the instant blood-mediated inflammatory reaction triggered by isolated human islets: possible application of the thrombin inhibitor melagatran in clinical islet transplantation [Ｊ]. Diabetes, 2002; 51(6): 1779-1784.
8. Fotiadis C, Xekouki P, Papalois AE, et al. Effects of mycophenolate mofetil vs cyclosporine administration on graft survival and function after islet allotransplantation in diabetic rats [Ｊ]. World J Gastroenterol, 2005; 11(18): 2733-2778.
9. Fontaine MJ, Fan W. Islet cell transplantation as a cure for insulin dependent diabetes: current improvements in preserving islet cell mass and function [Ｊ]. Hepatobiliary Pancreat Dis Int, 2003; 2(2): 170-179.
10. Moberg L. The role of the innate immunity in islet transplantation [Ｊ]. Ups J Med Sci, 2005; 110(1): 17-55.
11. Lewis EC, Shapiro L, Bowers OJ, et al. α1-Antitrypsin mo-notherapy prolongs islet allograft survival in mice [Ｊ]. Proc Natl Acad Sci USA, 2005; 102(34): 12153-12158.
12. Cattan P, Berney T, Schena S, et al. Early assessment of apoptosis in isolated islets of Langerhans [Ｊ]. Transplantation, 2001; 71(7): 857-862.
13. Matsuda T, Omori K, Vuong T, et al. Inhibition of p38 pathway suppresses human islet production of pro-inflammatory cytokines and improves islet graft function [Ｊ]. Am J Transplant, 2005; 5(3): 484-493.
14. Mi QS, Ly D, Zucker P, et al. Interleukin-4 but not interleukin-10 protects against spontaneous and recurrent type 1 diabetes by activated CD1d-restricted invariant natural killer T-cells [Ｊ]. Diabetes, 2004; 53(5): 1303-1310.
15. Choi SE, Choi KM, Yoon IH, et al. IL-6 protects pancreatic islet beta cells from pro-inflammatory cytokines-induced cell death and functional impairment in vitro and in vivo [Ｊ]. Transpl Immunol, 2004; 13(1): 43-53.
16. Sung HH, Juang JH, Lin YC, et al. Transgenic expression of decoy receptor 3 protects islets from spontaneous and chemical-induced autoimmune destruction in nonobese diabetic mice [Ｊ]. J Exp Med, 2004; 199(8): 1143-1151.
17. Grey ST, Lock J, Bach FH, et al. Adenovirus-mediated gene transfer of A20 in murine islets inhibits Fas-induced apoptosis [Ｊ]. Transplant Proc, 2001; 33(1-2): 577-578.
18. Shapiro AM, Ricordi C, Hering BJ, et al. International trial of the Edmonton protocol for islet transplantation [Ｊ]. N Engl J Med, 2006; 355(13): 1318-1330.
19. Ryan EA, Paty BW, Senior PA, et al. Five-year follow-up after clinical islet transplantation [Ｊ]. Diabetes, 2005; 54(7): 2060-2069.
20. Mandrup-Poulsen T. Apoptotic signal transduction pathways in diabetes [Ｊ]. Biochem Pharmacol, 2003; 66(8): 1433-1440.
21. Donath MY, Strling J, Maedler K, et al. Inflammatory mediators and islet beta-cell failure: a link between type 1 and type 2 diabetes [Ｊ]. J Mol Med, 2003; 81(8): 455-470.
22. Saldeen J. Cytokines induce both necrosis and apoptosis via a common Bcl-2-inhibitable pathway in rat insulin-producing cells [Ｊ]. Endocrinology, 2000; 141(6): 2003-2010.
23. Stickings P, Cunningham JM. Interleukin-1beta-induced nitric oxide production and inhibition of insulin secretion in rat islets of langerhans is dependent upon the nitric oxide synthase cofactor tetrahidrobiopterin [Ｊ]. Cytokine, 2002; 18(2): 81-85.
24. 宋文哲， 宋燕， 譚巖, 等. 氨基胍對內(nèi)毒素休克大鼠肝損傷的保護作用研究 [Ｊ]. 中國普外基礎(chǔ)與臨床雜志, 2003; 10（4）: 347-350.
25. Twigg SM, Cao Z, MCLennan SV, et al. Renal connective tissue growth factor induction in experimental diabetes is prevented by aminoguanidine [Ｊ]. Endocrinology, 2002; 143(12): 4907-4915.
26. Osicka TM, Yu Y, Lee V, et al. Aminoguanidine and rami-pril prevent diabetes-induced increases in protein kinase C activity in glomeruli, retina and mesenteric artery [Ｊ]. Clin Sci (Lond), 2001; 100(3): 249-257.
27. Jerums G, Panagiotopoulos S, Forbes J, et al. Evolving concepts in advanced glycation, diabetic nephropathy, and diabetic vascular disease [Ｊ]. Arch Biochem Biophys, 2003; 419(1): 55-62.
  1. 1. Kwon KB, Kim EK, Jeong ES, et al. Cortex cinnamomi extract prevents streptozotocin- and cytokine-induced beta-cell damage by inhibiting NF-kappaB [Ｊ]. World J Gastroenterol， 2006; 12(27): 4331-4337.
  2. 2. Mosén H, Salehi A, Henningsson R, et al. Nitric oxide inhibits, and carbon monoxide activates, islet acid alpha-glucoside hydrolase activities in parallel with glucose-stimulated insulin secretion [Ｊ]. J Endocrinol， 2006; 190(3): 681-693.
  3. 3. Wahoff DC, Sutherland DE, Hower CD, et al. Free intraperitoneal islet autografts in pancreatectomized dogs-impact of islet purity and posttransplantation exogenous insulin [Ｊ]. Surgery， 1994; 116(4): 742-748.
  4. 4. Shapiro AM, Lakey JR, Ryan EA, et al. Islet transplantation in seven patients with type 1 diabetes mellitus using a glucocorticoid-free immunosuppressive regimen [Ｊ]. N Engl J Med， 2000; 343(4): 230-238.
  5. 5. Frank A, Deng S, Huang X, et al. Transplantation for type Ⅰ diabetes: comparison of vascularized whole-organ pancreas with isolated pancreatic islets [Ｊ]. Ann Surg， 2004; 240(4): 631-640.
  6. 6. 劉方峰, 程穎, 劉永鋒, 等. 激活型Akt1轉(zhuǎn)染大鼠胰島對移植物凋亡和再血管化的影響 [Ｊ]. 中國普外基礎(chǔ)與臨床雜志, 2009; 16(1): 28-31.
  7. 7. Ozmen L, Ekdahl KN, Elgue G, et al. Inhibition of thrombin abrogates the instant blood-mediated inflammatory reaction triggered by isolated human islets: possible application of the thrombin inhibitor melagatran in clinical islet transplantation [Ｊ]. Diabetes, 2002; 51(6): 1779-1784.
  8. 8. Fotiadis C, Xekouki P, Papalois AE, et al. Effects of mycophenolate mofetil vs cyclosporine administration on graft survival and function after islet allotransplantation in diabetic rats [Ｊ]. World J Gastroenterol, 2005; 11(18): 2733-2778.
  9. 9. Fontaine MJ, Fan W. Islet cell transplantation as a cure for insulin dependent diabetes: current improvements in preserving islet cell mass and function [Ｊ]. Hepatobiliary Pancreat Dis Int, 2003; 2(2): 170-179.
  10. 10. Moberg L. The role of the innate immunity in islet transplantation [Ｊ]. Ups J Med Sci, 2005; 110(1): 17-55.
  11. 11. Lewis EC, Shapiro L, Bowers OJ, et al. α1-Antitrypsin mo-notherapy prolongs islet allograft survival in mice [Ｊ]. Proc Natl Acad Sci USA, 2005; 102(34): 12153-12158.
  12. 12. Cattan P, Berney T, Schena S, et al. Early assessment of apoptosis in isolated islets of Langerhans [Ｊ]. Transplantation, 2001; 71(7): 857-862.
  13. 13. Matsuda T, Omori K, Vuong T, et al. Inhibition of p38 pathway suppresses human islet production of pro-inflammatory cytokines and improves islet graft function [Ｊ]. Am J Transplant, 2005; 5(3): 484-493.
  14. 14. Mi QS, Ly D, Zucker P, et al. Interleukin-4 but not interleukin-10 protects against spontaneous and recurrent type 1 diabetes by activated CD1d-restricted invariant natural killer T-cells [Ｊ]. Diabetes, 2004; 53(5): 1303-1310.
  15. 15. Choi SE, Choi KM, Yoon IH, et al. IL-6 protects pancreatic islet beta cells from pro-inflammatory cytokines-induced cell death and functional impairment in vitro and in vivo [Ｊ]. Transpl Immunol, 2004; 13(1): 43-53.
  16. 16. Sung HH, Juang JH, Lin YC, et al. Transgenic expression of decoy receptor 3 protects islets from spontaneous and chemical-induced autoimmune destruction in nonobese diabetic mice [Ｊ]. J Exp Med, 2004; 199(8): 1143-1151.
  17. 17. Grey ST, Lock J, Bach FH, et al. Adenovirus-mediated gene transfer of A20 in murine islets inhibits Fas-induced apoptosis [Ｊ]. Transplant Proc, 2001; 33(1-2): 577-578.
  18. 18. Shapiro AM, Ricordi C, Hering BJ, et al. International trial of the Edmonton protocol for islet transplantation [Ｊ]. N Engl J Med, 2006; 355(13): 1318-1330.
  19. 19. Ryan EA, Paty BW, Senior PA, et al. Five-year follow-up after clinical islet transplantation [Ｊ]. Diabetes, 2005; 54(7): 2060-2069.
  20. 20. Mandrup-Poulsen T. Apoptotic signal transduction pathways in diabetes [Ｊ]. Biochem Pharmacol, 2003; 66(8): 1433-1440.
  21. 21. Donath MY, Strling J, Maedler K, et al. Inflammatory mediators and islet beta-cell failure: a link between type 1 and type 2 diabetes [Ｊ]. J Mol Med, 2003; 81(8): 455-470.
  22. 22. Saldeen J. Cytokines induce both necrosis and apoptosis via a common Bcl-2-inhibitable pathway in rat insulin-producing cells [Ｊ]. Endocrinology, 2000; 141(6): 2003-2010.
  23. 23. Stickings P, Cunningham JM. Interleukin-1beta-induced nitric oxide production and inhibition of insulin secretion in rat islets of langerhans is dependent upon the nitric oxide synthase cofactor tetrahidrobiopterin [Ｊ]. Cytokine, 2002; 18(2): 81-85.
  24. 24. 宋文哲， 宋燕， 譚巖, 等. 氨基胍對內(nèi)毒素休克大鼠肝損傷的保護作用研究 [Ｊ]. 中國普外基礎(chǔ)與臨床雜志, 2003; 10（4）: 347-350.
  25. 25. Twigg SM, Cao Z, MCLennan SV, et al. Renal connective tissue growth factor induction in experimental diabetes is prevented by aminoguanidine [Ｊ]. Endocrinology, 2002; 143(12): 4907-4915.
  26. 26. Osicka TM, Yu Y, Lee V, et al. Aminoguanidine and rami-pril prevent diabetes-induced increases in protein kinase C activity in glomeruli, retina and mesenteric artery [Ｊ]. Clin Sci (Lond), 2001; 100(3): 249-257.
  27. 27. Jerums G, Panagiotopoulos S, Forbes J, et al. Evolving concepts in advanced glycation, diabetic nephropathy, and diabetic vascular disease [Ｊ]. Arch Biochem Biophys, 2003; 419(1): 55-62.