Abstract
Pre-weaned porcine islets (PPIs) represent an unlimited source for islet transplantation but are functionally immature. We previously showed that necrostatin-1 (Nec-1) immediately after islet isolation enhanced the in vitro development of PPIs. Here, we examined the impact of Nec-1 on the in vivo function of PPIs after transplantation in diabetic mice. PPIs were isolated from pancreata of 8–15-day-old, pre-weaned pigs and cultured in media alone, or supplemented with Nec-1 (100 µM) on day 0 or on day 3 of culture (n = 5 for each group). On day 7, islet recovery, viability, oxygen consumption rate, insulin content, cellular composition, insulin secretion capacity, and transplant outcomes were evaluated. While islet viability and oxygen consumption rate remained high throughout 7-day tissue culture, Nec-1 supplementation on day 3 significantly improved islet recovery, insulin content, endocrine composition, GLUT2 expression, differentiation potential, proliferation capacity of endocrine cells, and insulin secretion. Adding Nec-1 on day 3 of tissue culture enhanced the islet recovery, proportion of delta cells, beta-cell differentiation and proliferation, and stimulation index. In vivo, this leads to shorter times to normoglycemia, better glycemic control, and higher circulating insulin. Our findings identify the novel time-dependent effects of Nec-1 supplementation on porcine islet quantity and quality prior to transplantation.
Highlights
The current standard treatment of Type 1 diabetes mellitus (T1DM) using subcutaneous administration of exogenous insulin is associated with daily stress and increased rate of severe hypoglycemic events due to suboptimal absorption and imprecise dosing [1].To alleviate these adverse effects, islet allotransplantation using the Edmonton protocol has been deemed a propitious therapy to restore normoglycemia and promote long-term insulin independence in T1DM patients [2]
Nec-1 supplementation either immediately after islet isolation or on day 3 did not affect islet viability compared to untreated islets on day 3 and day 7 (Supplementary Figure S1)
Pre-weaned porcine islets (PPIs) in study, the effects that Nec-1 supplementation on day 3 of tissue culture have on PPIs inon vitro vitro vivo after transplantation in STZ-induced diabetic mice wereOur examined
Summary
The current standard treatment of Type 1 diabetes mellitus (T1DM) using subcutaneous administration of exogenous insulin is associated with daily stress and increased rate of severe hypoglycemic events due to suboptimal absorption and imprecise dosing [1]. To alleviate these adverse effects, islet allotransplantation using the Edmonton protocol has been deemed a propitious therapy to restore normoglycemia and promote long-term insulin independence in T1DM patients [2]. Our recent studies have further shown that 100 μM of Nec-1 was the most effective dose to enhance the in vitro maturation of PPIs for up to 7 days during tissue culture [16,17]. The present study compared the effects of supplementing Nec-1 on day 3 of tissue culture to immediately after islet isolation on PPIs in vitro and, for the first time, evaluated the in vivo function of PPIs treated with Nec-1 on day 3 of tissue culture after transplantation in streptozotocin-induced diabetic mice
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