Coexpression of vascular endothelial growth factor and interleukin-1 receptor antagonist for improved human islet survival and function.
Ex vivo gene therapy approaches can improve the outcome of islet transplantation for treating type I diabetes. We have recently shown improvement in islet survival and function following ex vivo infection of islets with a mixture of adenoviral vectors encoding human vascular endothelial growth factor (Adv-hVEGF) and human interleukin-1 receptor antagonist (Adv-hIL-1Ra). In this study, we constructed a bicistronic vector encoding these two genes (phVEGF-hIL-1Ra) by cloning hIL-1Ra under the cytomegalovirus (CMV) promoter and hVEGF under the elongation factor-1alpha (EF-1 alpha) promoter in pBudCE4.1 vector. There was dose and time dependent expression of hVEGF and hIL-1Ra at both mRNA and protein levels after transfection with human islets. Transfected islets were viable, as evidenced by insulin release upon glucose challenge. Coexpression of hVEGF and hIL-1Ra suppressed nitric oxide production, total caspases, apoptosis, and necrosis in the presence of inflammatory cytokine cocktail consisting of IL-1beta, TNFalpha, and IFNgamma. In conclusion, our results indicated that coexpression of growth factor and antiapoptic genes can improve islet survival and function.
Jia
Xiangxu
X
Department of Pharmaceutical Sciences, University of Tennessee Health Science Center, Memphis, TN 38163, USA.
Cheng
Kun
K
Mahato
Ram I
RI
D020169:Caspases / Q000378:metabolism; D016923:Cell Death; D002478:Cells, Cultured; D016207:Cytokines / Q000494:pharmacology; D005786:Gene Expression Regulation; D005822:Genetic Vectors / Q000235:genetics*; D006801:Humans; D053590:Interleukin 1 Receptor Antagonist Protein / Q000378:metabolism*; D007515:Islets of Langerhans / Q000502:physiology*; D009569:Nitric Oxide / Q000378:metabolism; D013997:Time Factors; D014162:Transfection; D042461:Vascular Endothelial Growth Factor A / Q000378:metabolism*
D016207:Cytokines; D053590:Interleukin 1 Receptor Antagonist Protein; D042461:Vascular Endothelial Growth Factor A; D009569:Nitric Oxide; D020169:Caspases