Long-term correction of diabetes in rats after lentiviral hepatic insulin gene therapy

Abstract

Aims/hypothesis Type 1 diabetes results from the autoimmune destruction of pancreatic beta cells. Exogenous insulin therapy cannot achieve precise physiological control of blood glucose concentrations, and debilitating complications develop. Lentiviral vectors are promising tools for liver-directed gene therapy. However, to date, transduction rates in vivo remain low in hepatocytes, without the induction of cell cycling. We investigated long-term transgene expression in quiescent hepatocytes in vitro and determined whether the lentiviral delivery of furin-cleavable insulin to the liver could reverse diabetes in rats. Materials and methods To improve transduction efficiency in vitro, we optimised hepatocyte isolation and maintenance protocols and, using an improved surgical delivery method, delivered furin-cleavable insulin alone or empty vector to the livers of streptozotocin-induced diabetic rats by means of a lentiviral vector. Rats were monitored for changes in body weight and blood glucose, and intravenous glucose tolerance tests were performed. Expression of insulin was determined by RT-PCR, immunohistochemistry and electron microscopy. Results We achieved long-term transgene expression in quiescent hepatocytes in vitro (87±1.2% transduction efficiency), with up to 60±3.2% transduction in vivo. We normalised blood glucose for 500 days-a significantly longer period than previously reported-making this the first successful study using a lentiviral vector. This procedure resulted in the expression of genes encoding several beta cell transcription factors, some pancreatic endocrine transdifferentiation, hepatic insulin storage in granules, and restoration of glucose tolerance. Liver function tests remained normal. Importantly, pancreatic exocrine transdifferentiation did not occur. Conclusions/interpretation Our data suggest that this regimen may ultimately be employed for the treatment of type 1 diabetes.

Diabetes therapy by lentiviral hepatic insulin gene expression without transformation of liver

“…1). In contrast to Ren et al [1], in our experiments we used furin-cleavable insulin and not the furin-cleavable insulin analogue B10. In this insulin analogue the histidine in position 10 of the B chain of the insulin molecule is replaced by aspartic acid [4].…”

“…To the editor: We read with interest the paper by Ren et al [1] published in Diabetologia. In this study the liver of diabetic rats was transduced by a lentiviral vector system with the furincleavable insulin analogue B10.…”

“…In this insulin analogue the histidine in position 10 of the B chain of the insulin molecule is replaced by aspartic acid [4]. The mutation of this insulin analogue has been shown to induce mitogenic effects [5][6][7], and according to the authors' [1] conclusion, could also be responsible for the observed liver transdifferentiation. Using the furin-cleavable insulin for the lentiviral transductions we detected a decrease in blood glucose concentration of STZ-diabetic rats from 22.3±3.4 to 15.6±2.1 mmol/l after 3 days and to 15.0±2.8 mmol/l after 30 days.…”

Őssejtterápia, β-sejt- és Langerhans-sziget-neogenezis: a jövő lehetséges terápiái 1-es típusú diabetesben?

“…Az így kezelt állatok májsejtjeiben számos, a β-sejtekre jellemző transzkripciós faktor génje is expresszálódott. Mindez nem váltott ki gyulladásos vagy egyéb reakciót, a májenzimek szintje nem emelkedett [15,16]. Hasonló experimentális eredményeket közöltek Elsner és mtsai, valamint Gerace és mtsai is [17,18].…”

Diabetes therapy by lentiviral hepatic insulin gene expression without transformation of liver. Reply to Elsner M, Jörns A, Lenzen S [letter]

“…Ren B, O'Brien BA, Swan MA et al (2007) Long-term correction of diabetes in rats after lentiviral hepatic insulin gene therapy. …”