Abstract
Islet transplantation is limited by islet graft failure due to poor revascularization, host immune rejection and nonspecific inflammatory response. Delivery of human vascular endothelial growth factor (hVEGF) gene to the islets is likely to promote islet revascularization and survival. We used a bicistronic adenoviral vector encoding hVEGF and CpG-free allele of green fluorescent protein (Adv-GFP-hVEGF) and introduced into human pancreatic islets by transfection. We found that transfection efficiency and apoptosis were dependent on the multiplicity of infection (MOI). Compared to Adv-GFP transfected and nontransfected islets, the levels of hVEGF secreted from Adv-GFP-hVEGF transfected islets were higher and exhibit a linear relationship between hVEGF expression and MOI (10–5000). Persistent, but low level expression of hVEGF from nontransfected islets was also observed. This may be due to expression of the endogenous hVEGF gene under hypoxic conditions. The levels of DNA fragmentation determined by ELISA of islet lysates were dependent on the MOI of Adv-GFP-hVEGF. On glucose challenge, insulin release from transfected islets was comparable to nontransfected islets. Immunohistochemical staining for hVEGF was very high in Adv-GFP-hVEGF transfected islets. Weak staining was also observed for hCD31 in both transfected and nontransfected islets. These findings suggest that Adv-GFP-hVEGF is a potential candidate for promoting islet revascularization.
This is a preview of subscription content, access via your institution
Access options
Subscribe to this journal
Receive 12 print issues and online access
$259.00 per year
only $21.58 per issue
Rent or buy this article
Prices vary by article type
from$1.95
to$39.95
Prices may be subject to local taxes which are calculated during checkout
Similar content being viewed by others
References
Garcia-Ocana A et al. Adenovirus-mediated hepatocyte growth factor expression in mouse islets improves pancreatic islet transplant performance and reduces beta cell death. J Biol Chem 2003; 278: 343–351.
Ryan EA et al. Successful islet transplantation: continued insulin reserve provides long-term glycemic control. Diabetes 2002; 51: 2148–2157.
Embury J et al. Proteins linked to a protein transduction domain efficiently transduce pancreatic islets. Diabetes 2001; 50: 1706–1713.
Hering BaR C . Results, research priorities and reason for optimism: islet transplantation for patients with Type I diabetes. Graft 2000; 2: 12–27.
Gaber AO et al. Improved in vivo pancreatic islet function after prolonged in vitro islet culture. Transplantation 2001; 72: 1730–1736.
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: 1060–1066.
Contreras JL et al. Cytoprotection of pancreatic islets before and soon after transplantation by gene transfer of the anti-apoptotic Bcl-2 gene. Transplantation 2001; 71: 1015–1023.
Giannoukakis N et al. Adenoviral gene transfer of the interleukin-1 receptor antagonist protein to human islets prevents IL-1beta-induced beta-cell impairment and activation of islet cell apoptosis in vitro. Diabetes 1999; 48: 1730–1736.
Gorden DL et al. Vascular endothelial growth factor is increased in devascularized rat islets of Langerhans in vitro. Transplantation 1997; 63: 436–443.
Goudy K et al. Adeno-associated virus vector-mediated IL-10 gene delivery prevents type 1 diabetes in NOD mice. Proc Natl Acad Sci USA 2001; 98: 13913–13918.
Ko KS, Lee M, Koh JJ, Kim SW . Combined administration of plasmids encoding IL-4 and IL-10 prevents the development of autoimmune diabetes in nonobese diabetic mice. Mol Ther 2001; 4: 313–316.
Reiser J, Lai Z, Zhang XY, Brady RO . Development of multigene and regulated lentivirus vectors. J Virol 2000; 74: 10589–10599.
Kapturczak M et al. Transduction of human and mouse pancreatic islet cells using a bicistronic recombinant adeno-associated viral vector. Mol Ther 2002; 5: 154–160.
Menger MD, Vajkoczy P, Beger C, Messmer K . Orientation of microvascular blood flow in pancreatic islet isografts. J Clin Invest 1994; 93: 2280–2285.
Lukinius A, Jansson L, Korsgren O . Ultrastructural evidence for blood microvessels devoid of an endothelial cell lining in transplanted pancreatic islets. Am J Pathol 1995; 146: 429–435.
Vasir B et al. Hypoxia induces vascular endothelial growth factor gene and protein expression in cultured rat islet cells. Diabetes 1998; 47: 1894–1903.
Gerber HP et al. Vascular endothelial growth factor regulates endothelial cell survival through the phosphatidylinositol 3′-kinase/Akt signal transduction pathway. Requirement for Flk-1/KDR activation. J Biol Chem 1998; 273: 30336–30343.
Ferrara N, Gerber HP, LeCouter J . The biology of VEGF and its receptors. Nat Med 2003; 9: 669–676.
Mahato RI et al. Cationic lipid and polymer-based gene delivery to human pancreatic islets. Mol Ther 2003; 7: 89–100.
Narang AS et al. Vascular endothelial growth factor gene delivery to human islets for neoangiogenesis after transplantation. Pharm Res 2004; 21: 15–25.
Lakey JR et al. Nonviral transfection of intact pancreatic islets. Cell Transplant 2001; 10: 697–708.
Benhamou PY et al. Standardization of procedure for efficient ex vivo gene transfer into porcine pancreatic islets with cationic liposomes. Transplantation 1997; 63: 1798–1803.
Saldeen J et al. Efficient gene transfer to dispersed human pancreatic islet cells in vitro using adenovirus-polylysine/DNA complexes or polycationic liposomes. Diabetes 1996; 45: 1197–1203.
Csete ME et al. Efficient gene transfer to pancreatic islets mediated by adenoviral vectors. Transplantation 1995; 59: 263–268.
Muruve DA, Manfro RC, Strom TB, Libermann TA . Ex vivo adenovirus-mediated gene delivery leads to long-term expression in pancreatic islet transplants. Transplantation 1997; 64: 542–546.
Dobson T et al. Human pancreatic islets transfected to produce an inhibitor of TNF are protected against destruction by human leukocytes. Cell Transplant 2000; 9: 857–865.
Weber M et al. Adenoviral transfection of isolated pancreatic islets: a study of programmed cell death (apoptosis) and islet function. J Surg Res 1997; 69: 23–32.
Rivard A et al. Rescue of diabetes-related impairment of angiogenesis by intramuscular gene therapy with adeno-VEGF. Am J Pathol 1999; 154: 355–363.
Vasir B et al. Effects of diabetes and hypoxia on gene markers of angiogenesis (HGF, cMET, uPA and uPAR, TGF-alpha, TGF-beta, bFGF and Vimentin) in cultured and transplanted rat islets. Diabetologia 2000; 43: 763–772.
Ajioka I, Akaike T, Watanabe Y . Expression of vascular endothelial growth factor promotes colonization, vascularization, and growth of transplanted hepatic tissues in the mouse. Hepatology 1999; 29: 396–402.
Linn T et al. Angiogenic capacity of endothelial cells in islets of Langerhans. Faseb J 2003; 17: 881–883.
Linn T et al. Polymers for induction of revascularization in the rat fascial flap: application of vascular endothelial growth factor and pancreatic islet cells. Cell Transplant 2003; 12: 769–778.
Wilson JM . Adenoviruses as gene-delivery vehicles. N Engl J Med 1996; 334: 1185–1187.
Johnson LG et al. In vitro assessment of variables affecting the efficiency and efficacy of adenovirus-mediated gene transfer to cystic fibrosis airway epithelia. Hum Gene Ther 1996; 7: 51–59.
Teramoto S, Ito H, Ouchi Y . Variables affecting the transduction efficiency of adenovirus vectors in bovine aortic endothelial cells. Thromb Res 1999; 93: 35–42.
Mendola JF et al. Immunocytochemical study of pancreatic islet revascularization in islet isograft. Effect of hyperglycemia of the recipient and of in vitro culture of islets. Transplantation 1994; 57: 725–730.
Schmittgen TD, Zakrajsek BA . Effect of experimental treatment on housekeeping gene expression: validation by real-time, quantitative RT–PCR. J Biochem Biophys Methods 2000; 46: 69–81.
Simpson DA, Feeney S, Boyle C, Stitt AW . Retinal VEGF mRNA measured by SYBR green I fluorescence: a versatile approach to quantitative PCR. Mol Vision 2000; 6: 178–183.
Muruve DA et al. Adenovirus-mediated expression of Fas ligand induces hepatic apoptosis after systemic administration and apoptosis of ex vivo-infected pancreatic islet allografts and isografts. Hum Gene Ther 1997; 8: 955–963.
Wyllie AH . Cell death: A new classification separating apoptosis from necrosis. In: Bowen IDaL RA (ed). Cell Death in Biology and Pathology. Chapman & Hall: New York, 1981: 9–34.
Quintana FJ, Rotem A, Carmi P, Cohen IR . Vaccination with empty plasmid DNA or CpG oligonucleotide inhibits diabetes in nonobese diabetic mice: modulation of spontaneous 60-kDa heat shock protein autoimmunity. J Immunol 2000; 165: 6148–6155.
Campbell IL, Colman PG, Harrison LC . Adult human pancreatic islet cells in tissue culture: function and immunoreactivity. J Clin Endocrinol Metab 1985; 61: 681–685.
Redick SD, Bautch VL . Developmental platelet endothelial cell adhesion molecule expression suggests multiple roles for a vascular adhesion molecule. Am J Pathol 1999; 154: 1137–1147.
Vajkoczy P et al. Histogenesis and ultrastructure of pancreatic islet graft microvasculature. Evidence for graft revascularization by endothelial cells of host origin. Am J Pathol 1995; 146: 1397–1405.
Furuya H et al. Revascularization and function of pancreatic islet isografts in diabetic rats following transplantation. Cell Transplant 2003; 12: 537–544.
Maheshwari A, Han S, Mahato RI, Kim SW . Biodegradable polymer-based interleukin-12 gene delivery: role of induced cytokines, tumor infiltrating cells and nitric oxide in anti-tumor activity. Gene Therapy 2002; 9: 1075–1084.
Sigrist S et al. Induction of angiogenesis in omentum with vascular endothelial growth factor: influence on the viability of encapsulated rat pancreatic islets during transplantation. J Vasc Res 2003; 40: 359–367.
Arnush M et al. IL-1 produced and released endogenously within human islets inhibits beta cell function. J Clin Invest 1998; 102: 516–526.
Acknowledgements
We acknowledge the University of Tennessee Memphis start-up funds to Mahato's lab, and USPHS/NIH DK57700, USPHS/NCRR RR16602 and Juvenile Diabetes Research Foundation 1-2000-416 grants to Gaber's lab.
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Cheng, K., Fraga, D., Zhang, C. et al. Adenovirus-based vascular endothelial growth factor gene delivery to human pancreatic islets. Gene Ther 11, 1105–1116 (2004). https://doi.org/10.1038/sj.gt.3302267
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1038/sj.gt.3302267
Keywords
This article is cited by
-
Engineered VEGF-releasing PEG–MAL hydrogel for pancreatic islet vascularization
Drug Delivery and Translational Research (2015)
-
PEG–Maleimide Hydrogels for Protein and Cell Delivery in Regenerative Medicine
Annals of Biomedical Engineering (2014)
-
Optimization of pancreatic islet spheroid using various concave patterned-films
Macromolecular Research (2012)
-
Functional clustering of pancreatic islet cells using concave microwell array
Macromolecular Research (2011)
-
Islet surface PEGylation attenuate the instant blood-mediated inflammatory reaction in intrahepatic islet transplantation
Macromolecular Research (2011)