Autoimmune destruction of beta cells results in Type 1 diabetes, and these individuals become dependent on exogenous insulin administration for survival. Unfortunately, exogenous insulin administration lacks the precision of that which the beta cells are capable, so these individuals suffer from complications associated with hyperglycemic and hypoglycemic episodes. Therefore, islet transplantation and subsequent restoration of fully functional beta cells, holds much promise in addressing this specific problem. Preliminary work in islet transplantation demonstrates that though islet transplantation is successful in liberating these individuals from exogenous insulin administration, this effect is not sustained. This is mainly due to both pre- and post- transplantation loss of beta cell mass and poor or failed islet revascularization. In the pancreas, connective tissue growth factor (CTGF), a secreted protein in the CCN family, is expressed in embryonic beta cells and pancreatic vasculature and has been shown to be important in beta cell mass and proliferation. Specifically, the Gannon lab has shown that heterozygous and homozygous CTGF null mutant mouse embryos have reduced beta cell area and proliferation. Studies of bone development in CTGF homozygous null mutant mouse embryos have shown reduced angiogenesis at the growth plate. Based on these results, a bigenic mouse model was generated to control CTGF over-expression specifically in the beta cells. Transgenic mice with tetracycline operator-driven CTGF cDNA expression (TetO- CTGF) were interbred with mice in which the rat insulin promoter (RIP) drives the expression of the reverse tetracycline transcriptional activator (rtTA). In the presence of doxycycline, these bigenic mice over-express CTGF in the beta cells. Unpublished studies in the Gannon lab have shown that when doxycycline is given to pregnant females, the bigenic offspring show increased endocrine area in the pancreas, specifically in the number of alpha and beta cells, when compared to their RIP-rtTA littermates. We hypothesize that over-expression of CTGF in mature beta cells will increase beta cell mass, proliferation, and angiogenesis in adult bigenic mice as well as in transplanted bigenic islets. This will be investigated by administering doxycycline to adult bigenic mice and analyzing islet morphology using immunohistochemistry (insulin, glucagon, proliferation markers, apoptosis markers) and islet function using perifusion and intraperitoneal glucose tolerance tests. To test the effect of CTGF on islet transplantation, bigenic islets will be transplanted under the kidney capsule of control littermates who are administered doxycycline. The islets will be explanted and examined for beta cell mass, proliferation, and angiogenesis using immunohistochemistry. Additionally, since CTGF is a secreted factor, control islets will be treated with recombinant CTGF protein to see if there is an increase in beta cell mass and angiogenesis. These studies will determine whether CTGF could be part of a solution to improve islet transplantation survival and sustainability.

Public Health Relevance

Type 1 diabetes portends increased morbidity and mortality from hypoglycemia and hyperglycemia due to the imprecision of exogenous insulin administration. Islet transplantation is a promising solution to this problem, but these transplantations tend to fail within a three-year post-transplantation period. This proposal will test whether connective tissue growth factor (CTGF) could be used to increase pre-transplantation beta cell survival by increasing beta cell mass and proliferation as well as improving post-transplantation islet survival by also increasing beta cell mass, proliferation, and revascularization.

National Institute of Health (NIH)
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Postdoctoral Individual National Research Service Award (F32)
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Special Emphasis Panel (ZDK1-GRB-9 (J1))
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Castle, Arthur
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Vanderbilt University Medical Center
Internal Medicine/Medicine
Schools of Medicine
United States
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Gunasekaran, Uma; Hudgens, Courtney W; Wright, Brian T et al. (2012) Differential regulation of embryonic and adult ýý cell replication. Cell Cycle 11:2431-42