腺病毒介导rhbFGF基因转染胰岛后促进小鼠被膜下移植的研究
|
摘要
目的探讨重组人碱性成纤维生长因子(rhbFGF)基因导入胰岛细胞团后经肾被膜下移植,对移植物微血管重建的影响。方法采用携带rhbFGF-flag-GFP基因的腺病毒载体体外转染培养的胰岛细胞团,再移植至同系糖尿病小鼠的肾被膜下,术后30 d内观察糖尿病治疗情况,通过术后14 d糖耐量试验和HE、免疫组化等观察胰岛功能和移植物的微血管重建结果。结果感染复数(MOI)为100的rhbFGF基因转染后胰岛与未处理的胰岛在体外培养60 h后活性无统计学差异(P=0.75);移植200 IEQ转基因胰岛至体内14 d后,胰岛功能良好,微血管重建增加,优于单独400 IEQ胰岛移植组。结论 rhbFGF基因导入胰岛移植有助于提高胰岛的功能,促进移植物微血管重建,并可减少胰岛移植量。
Objective In this paper, we investigate the effect of islets transfected with adenovirus modified recombinant human basic fibroblast growth factor(rhbFGF) gene for angiogenesis after transplantation. Methods The adenoviral vector carrying rhbFGF-flag-GFP gene was transfected into the islets, and then transplanted into the renal capsule of a homologous diabetic mouse. The treatment of diabetes was observed within 30 days and the glucose tolerance test was performed at day 14 after transplantation. HE and immunohist-ochemistry staining were done to determine the islets function and the microvascular remodeling of the graft. Results The adenovirus modified rhbFGF enhanced efficiency of islet transfection, and there was no significant difference in activity between islets infected with a multiplicity of infection of 100 adenovirus and untreated after cultured for 60 h in vitro(P=0.75). The 200 IEQ islets transfected with rhbFGF-flag-GFP significantly increased islets function and accelerated the regeneration of blood vessels in vivo, which was better than the 400 IEQ islets transplantation alone. Conclusion Islets transfected with rhbFGF-flag-GFP gene will improve islets function and promote microvascular remodeling of the grafts, and which can reduce the amount of transplantation needed.
Objective In this paper, we investigate the effect of islets transfected with adenovirus modified recombinant human basic fibroblast growth factor(rhbFGF) gene for angiogenesis after transplantation. Methods The adenoviral vector carrying rhbFGF-flag-GFP gene was transfected into the islets, and then transplanted into the renal capsule of a homologous diabetic mouse. The treatment of diabetes was observed within 30 days and the glucose tolerance test was performed at day 14 after transplantation. HE and immunohist-ochemistry staining were done to determine the islets function and the microvascular remodeling of the graft. Results The adenovirus modified rhbFGF enhanced efficiency of islet transfection, and there was no significant difference in activity between islets infected with a multiplicity of infection of 100 adenovirus and untreated after cultured for 60 h in vitro(P=0.75). The 200 IEQ islets transfected with rhbFGF-flag-GFP significantly increased islets function and accelerated the regeneration of blood vessels in vivo, which was better than the 400 IEQ islets transplantation alone. Conclusion Islets transfected with rhbFGF-flag-GFP gene will improve islets function and promote microvascular remodeling of the grafts, and which can reduce the amount of transplantation needed.
引文
[1] Ballian N, Brunicardi FC. Islet vasculature as a regulator of endocrine pancreas function[J]. World J Surg, 2007, 31(4):705-714.
[2] Rajab A.Islet transplantation:alternative sites[J]. Curr Diab Rep,2010, 10(5):332-337.
[3] Brissova M, Shostak A, Shiota M, et al. Pancreatic islet production of vascular endothelial growth factor-A is essential for islet vascularization, revascularization, and function[J]. Diabetes, 2006, 55(11):2974-2985.
[4] Xiang Q, Xiao J, Zhang H, et al. Preparation and characterisation of bFGF-encapsulated liposomes and evaluation of wound-healing activities in the rat[J]. Burns, 2011, 37(5):886-895.
[5] Stapor PC,Azimi MS,Ahsan T,et al. An angiogenesis model for investigating multicellular interactions across intact microvascular networks[J]. Am J Physiol Heart Circ Physiol, 2013,304(2):235-245.
[6] Laporte LD, Rieux AD, Tuinstra HM, et al. Vascular endothelial growth factor and fibroblast growth factor 2 delivery from spinal cord bridges to enhance angiogenesis following injury[J]. J Biomed Mater Res A,2011,98(3):372-382.
[7] Chen B, Qin J, Wang H, et al. Effects of adenovirus-mediated bFGF, IL-1Ra and IGF-1 gene transfer on human osteoarthritic chondrocytes and osteoarthritis in rabbits[J]. Exp Mol Med,2010,42(10):684-695.
[8] Lee CS,Bishop ES,Zhang R,et al. Adenovirus-mediated gene delivery:potential applications for gene and cell-based therapies in the new era of personalized medicine[J]. Genes Dis, 2017, 4(2):43-63.
[9]叶丛丛,傅红兴,江玲,等.一种快速分离纯化小鼠胰岛的方法[J].肝胆胰外科杂志,2014, 26(5):403-405.
[10]蒋煊,傅红兴,邱凯燕,等.一种小鼠胰岛经门静脉肝内移植方法及疗效评价[J].肝胆胰外科杂志,2016, 28(2):129-132.
[11] Barnett MJ, Mcghee-Wilson D, Shapiro AM, et al. Variation in human islet viability based on diff erent membrane integrity stains[J]. Cell Transplantation, 2004, 13(5):481-488.
[12] Wu H, Yoon AR, Li F, et al. RGD peptide-modified adenovirus expressing hepatocyte growth factor and X-linked inhibitor of apoptosis improves islet transplantation[J]. J Gene Med, 2011, 13(12):658-669.
[13] Lembert N, Weshe J, Petersen P,et al. Areal density measurement is a convenient method for the determination of porcine islet equivalent without counting and sizing individual islets[J]. Cell Transplantation, 2003, 12(1):33-41.
[14] Fu HX, Wu LL, Qiu KY, et al. Construction and evaluation of islet transplantation under the kidney capsule in mice[J]. J Biomater Tissue Eng,2016, 6(3):208-215.
[15]胡倩,傅红兴,蒋煊,等.经脾实质胰岛移植治疗糖尿病小鼠的实验研究[J].肝胆胰外科杂志,2017, 29(6):486-489.
[16] Carlsson PO, Palm F, Andersson A, et al. Markedly decreased oxygen tension in transplanted rat pancreatic islets irrespective of the implantation site[J]. Diabetes, 2001, 50(3):489-495.
[17] Eter WA, Bos D, Frielink C, et al. Graft revascularization is essential for non-invasive monitoring of transplanted islets with radiolabeled exendin[J]. Sci Rep, 2015, 5(1):1-10.
[18] Qiu KY, Fu HX, JIANG X, et al. Sodium hyaluronate gel containing bFGF co-transplanted with islets leading to the promotion of angiogenesis and the reduction of the required grafted islets in diabetic mice[J]. J Biomater Tissue Eng, 2017, 7(4):291-301.
[2] Rajab A.Islet transplantation:alternative sites[J]. Curr Diab Rep,2010, 10(5):332-337.
[3] Brissova M, Shostak A, Shiota M, et al. Pancreatic islet production of vascular endothelial growth factor-A is essential for islet vascularization, revascularization, and function[J]. Diabetes, 2006, 55(11):2974-2985.
[4] Xiang Q, Xiao J, Zhang H, et al. Preparation and characterisation of bFGF-encapsulated liposomes and evaluation of wound-healing activities in the rat[J]. Burns, 2011, 37(5):886-895.
[5] Stapor PC,Azimi MS,Ahsan T,et al. An angiogenesis model for investigating multicellular interactions across intact microvascular networks[J]. Am J Physiol Heart Circ Physiol, 2013,304(2):235-245.
[6] Laporte LD, Rieux AD, Tuinstra HM, et al. Vascular endothelial growth factor and fibroblast growth factor 2 delivery from spinal cord bridges to enhance angiogenesis following injury[J]. J Biomed Mater Res A,2011,98(3):372-382.
[7] Chen B, Qin J, Wang H, et al. Effects of adenovirus-mediated bFGF, IL-1Ra and IGF-1 gene transfer on human osteoarthritic chondrocytes and osteoarthritis in rabbits[J]. Exp Mol Med,2010,42(10):684-695.
[8] Lee CS,Bishop ES,Zhang R,et al. Adenovirus-mediated gene delivery:potential applications for gene and cell-based therapies in the new era of personalized medicine[J]. Genes Dis, 2017, 4(2):43-63.
[9]叶丛丛,傅红兴,江玲,等.一种快速分离纯化小鼠胰岛的方法[J].肝胆胰外科杂志,2014, 26(5):403-405.
[10]蒋煊,傅红兴,邱凯燕,等.一种小鼠胰岛经门静脉肝内移植方法及疗效评价[J].肝胆胰外科杂志,2016, 28(2):129-132.
[11] Barnett MJ, Mcghee-Wilson D, Shapiro AM, et al. Variation in human islet viability based on diff erent membrane integrity stains[J]. Cell Transplantation, 2004, 13(5):481-488.
[12] Wu H, Yoon AR, Li F, et al. RGD peptide-modified adenovirus expressing hepatocyte growth factor and X-linked inhibitor of apoptosis improves islet transplantation[J]. J Gene Med, 2011, 13(12):658-669.
[13] Lembert N, Weshe J, Petersen P,et al. Areal density measurement is a convenient method for the determination of porcine islet equivalent without counting and sizing individual islets[J]. Cell Transplantation, 2003, 12(1):33-41.
[14] Fu HX, Wu LL, Qiu KY, et al. Construction and evaluation of islet transplantation under the kidney capsule in mice[J]. J Biomater Tissue Eng,2016, 6(3):208-215.
[15]胡倩,傅红兴,蒋煊,等.经脾实质胰岛移植治疗糖尿病小鼠的实验研究[J].肝胆胰外科杂志,2017, 29(6):486-489.
[16] Carlsson PO, Palm F, Andersson A, et al. Markedly decreased oxygen tension in transplanted rat pancreatic islets irrespective of the implantation site[J]. Diabetes, 2001, 50(3):489-495.
[17] Eter WA, Bos D, Frielink C, et al. Graft revascularization is essential for non-invasive monitoring of transplanted islets with radiolabeled exendin[J]. Sci Rep, 2015, 5(1):1-10.
[18] Qiu KY, Fu HX, JIANG X, et al. Sodium hyaluronate gel containing bFGF co-transplanted with islets leading to the promotion of angiogenesis and the reduction of the required grafted islets in diabetic mice[J]. J Biomater Tissue Eng, 2017, 7(4):291-301.