Abstract
One of the key limitations to successful human islet transplantation is loss of islets due to stress responses pre- and post-transplantation. Nutrient deprivation and ER stress have been identified as important mechanisms leading to apoptosis. Glial Cell-line Derived Neurotrophic Factor (GDNF) has recently been found to promote islet survival after isolation. However, whether GDNF could rescue human islets from nutrient deprivation and ER stress-mediated apoptosis is unknown. Herein, by mimicking those conditions in vitro, we have shown that GDNF significantly improved glucose stimulated insulin secretion, reduced apoptosis and proinsulin:insulin ratio in nutrient deprived human islets. Furthermore, GDNF alleviated thapsigargin-induced ER stress evidenced by reduced expressions of IRE1α and BiP and consequently apoptosis. Importantly, this was associated with an increase in phosphorylation of PI3K/AKT and GSK3B signaling pathway. Transplantation of ER stressed human islets pre-treated with GDNF under kidney capsule of diabetic mice resulted in reduced expressions of IRE1α and BiP in human islet grafts with improved grafts function shown by higher levels of human C-peptide post-transplantation. We suggest that GDNF has protective and anti-apoptotic effects on nutrient deprived and ER stress activated human islets and could play a significant role in rescuing human islets from stress responses.
Highlights
Type-1 diabetes (T1D) results from autoimmune destruction of insulin producing pancreatic beta cells in islets of Langerhans, which is largely due to reactive T-cells[1]
In order to investigate the effect of Glial cell-line derived neurotrophic factor (GDNF) on islet function and survival under nutrient deprivation, isolated human islets were cultured for 72 hrs under low concentration of serum (0.5%) with or without GDNF
We have shown that GDNF could reverse the adverse effect of nutrient deprivation and SERCA channel blocker, Tg by improving human islets function and viability through reduction in endoplasmic reticulum (ER) stress induced apoptosis and activation of PI3K/p-AKT/p-GSK3B survival pathway
Summary
GDNF improves function and viability of nutrient deprived human islets. In order to investigate the effect of GDNF on islet function and survival under nutrient deprivation, isolated human islets were cultured for 72 hrs under low concentration of serum (0.5%) with or without GDNF. We performed viability staining (FDA/PI) of intact islets and showed enhanced PI staining in the Tg treated islets compared to Tg+GDNF (Fig. 2h) Taken together, these results suggest that GDNF protects human islets from ER stress and ER stress induced apoptosis. Co-treatment of human islets with Tg+GDNF significantly recovered the levels of p-PI3K, p-AKT, p-GSK3B compared to Tg alone (Fig. 3), suggesting that GDNF protects human islets from ER-stress via activation of the PI3K/AKT signaling pathways. We found a significantly decrease in protein expression of IRE1α (Fig. 4f) and BiP (Fig. 4g) in grafts containing islets pre-treated with Tg+GDNF compared to Tg alone (2.0 and 3.0 fold reduction, respectively) Taken together, these results suggest that GDNF protects human islets from ER stress and further contributes to improve islet grafts function post transplantation
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