Genetic Engineering of a Suboptimal Islet Graft with A20 Preserves β Cell Mass and Function

Abstract

Transplantation of an excessive number of islets of Langerhans (two to four pancreata per recipient) into patients with type I diabetes is required to restore euglycemia. Hypoxia, nutrient deprivation, local inflammation, and the β cell inflammatory response (up-regulation of NF-κB-dependent genes such as inos) result in β cell destruction in the early post-transplantation period. Genetic engineering of islets with anti-inflammatory and antiapoptotic genes may prevent β cell loss and primary nonfunction. We have shown in vitro that A20 inhibits NF-κB activation in islets and protects from cytokine- and death receptor-mediated apoptosis. In vivo, protection of newly transplanted islets would reduce the number of islets required for successful transplantation. Transplantation of 500 B6/AF1 mouse islets into syngeneic, diabetic recipients resulted in a cure rate of 100% within 5 days. Transplantation of 250 islets resulted in a cure rate of only 20%. Transplantation of 250 islets overexpressing A20 resulted in a cure rate of 75% with a mean time to cure of 5.2 days, comparable to that achieved with 500 islets. A20-expressing islets preserve functional β cell mass and are protected from cell death. These data demonstrate that A20 is an ideal cytoprotective gene therapy candidate for islet transplantation.

Suppressor of cytokine signalling (SOCS)-3 protects beta cells against IL-1?-mediated toxicity through inhibition of multiple nuclear factor-?B-regulated proapoptotic pathways

“…The possibility of intervening in this process by overexpressing protective proteins in islets transplanted to diabetic animals has recently been demonstrated. Overexpression of A20, a zinc finger protein inhibiting NF-κB activation and subsequent cytokine-or Fas-mediated apoptosis, protected transplanted islets and reduced the number of islets required to obtain normoglycaemia by 50% in a rat model of type 1 diabetes [11]. Furthermore, the potential of pharmacological modulation of protective processes was substantiated by the finding that a vitamin D analogue inhibited cytokine-induced beta cell apoptosis through induction of A20 expression [56].…”

“…the free radical scavengers manganese superoxide dismutase (Mn-SOD), glutathione peroxidase and catalase [6,7], and elements with anti-apoptotic properties, e.g. thioredoxin, calbindin, bcl-2 and A20 [8,9,10,11]. In general, however, the ability of the beta cell to activate a sufficient protective response is impaired, which could, in part, explain their sensitivity to the toxic effect of cytokines [12].…”

Adenoviral overproduction of interleukin-1 receptor antagonist increases beta cell replication and mass in syngeneically transplanted islets, and improves metabolic outcome

“…Adenoviral vectors have been used previously to achieve transient transgene expression in islet grafts aiming to increase the survival or the growth of transplanted islets [38][39][40][41][42][43][44]. In islet grafts, overexpression of the antiapoptotic gene Bcl2 [40] and the cytoprotective gene A20 (also known as Tnfaip3) [41], and overproduction of the antioxidant metallothionein [42], the mitogenic and survival factor hepatocyte growth factor [38,43] or the key protein kinase B/Akt [44], have successfully improved the metabolic outcome in experimental islet transplantation.…”

“…In islet grafts, overexpression of the antiapoptotic gene Bcl2 [40] and the cytoprotective gene A20 (also known as Tnfaip3) [41], and overproduction of the antioxidant metallothionein [42], the mitogenic and survival factor hepatocyte growth factor [38,43] or the key protein kinase B/Akt [44], have successfully improved the metabolic outcome in experimental islet transplantation. However, the effects of transgene overexpression on apoptosis, growth or beta cell mass of transplanted cells were not quantified, and the confounding effects of the different blood glucose concentrations achieved in experimental groups were not taken into account, thereby limiting overall understanding of the events that took place in the graft.…”

Co-encapsulation of bioengineered IGF-II-producing cells and pancreatic islets: effect on beta-cell survival

“…Other research groups have experimented gene transfer with the aim of controlling the autoimmune reaction against islets. [21][22][23][24][25][26][27][28] Considering the limited duration of insulin independence in recipients of islet transplantation, an important area of research is to design methods to promote islet survival. The mere immobilization of islets in alginate beads has been shown to improve islet survival both in vitro and in vivo.…”