RESUMEN
For patients with type 1 diabetes mellitus complicated by severe hypoglycemia, clinical islet transplantation is an efficacious alternative to whole pancreas transplantation. While islet transplantation has improved over the last few years, there remain questions regarding its cost-effectiveness and donor allosensitization, which is exacerbated when islets from more than one donor are required. Understanding the features of a pancreas that would provide viable islets prior to isolation may lead to development of an accurate assay that could identify suitable pancreases and provide significant cost savings to a clinical islet transplantation program. In this pilot study, solid-state high-resolution magic angle spinning (HRMAS) nuclear magnetic resonance (NMR) spectroscopy was used to assess samples of convenience of human pancreatic tissue taken prior to islet isolation both before and after incubation using the two-layer perfluorocarbon (PFC)/University of Wisconsin (UW) solution cold-storage method. We observed that, prior to incubation, human pancreatic tissue exhibited evidence of hypoxia with decreased peak integrals associated with glucose and increased peak integrals corresponding to lactate and free fatty acids. After incubation, we observed a reversal of the hypoxia-induced damage, as integrals corresponding to glucose increased, and those corresponding to lactate and free fatty acid resonances decreased. Interestingly, a significant correlation between the ratio of the glucose integral (at 3.0-4.5 ppm) to the sum of the fatty acid (at 0.9 ppm) and lactate + fatty acid (at 1.3 ppm) integrals and glucose responsiveness, a measure of islet viability, of the isolated islets, was observed after incubation in PFC/UW solution for pancreases that responded to PFC/UW solution incubation (p = 0.02). Notably, pancreases with little or no change in the integral ratio after PFC/UW solution incubation had poor recovery. These results suggest that tissue recovery is a key feature for determining islet cell viability, and further that HRMAS NMR may be a practical method to quickly assess human donor pancreatic tissue prior to islet isolation for clinical transplantation.
RESUMEN
Pancreatic islet transplantation is proposed as a cure for type 1 diabetes mellitus (T1D). Despite its success in optimal regulation of glucose levels, limitations in longevity of islet grafts still require innovative solutions. Inflammatory stress post-transplantation and loss of extracellular matrix attribute to the limited ß-cell survival. Pancreatic stellate cells (PSCs), identified as pancreatic-specific stromal cells, have the potential to play a crucial role in preserving islet survival. Our study aimed to determine the effects of PSCs co-cultured with human CM ß-cells and human islets under inflammatory stress induced by a cytokine cocktail of IFN-γ, TNF-α and IL-1ß. Transwell culture inserts were utilized to assess the paracrine impact of PSCs on ß-cells, alongside co-cultures enabling direct interaction between PSCs and human islets. We found that co-culturing PSCs with human CM ß-cells and human cadaveric islets had rescuing effects on cytokine-induced stress. Effects were different under normoglycemic and hyperglycemic conditions. PSCs were associated with upregulation of ß-cell mitochondrial activity and suppression of inflammatory gene expression. The rescuing effects exist both in indirect and direct co-culture methods. Furthermore, we tested whether PSCs have rescuing effects on human islets in conventional alginate-based microcapsules and in composite microcapsules composed of alginate-pectin collagen type IV, laminin sequence RGD, Nec-1, and amino acid. PSCs partially prevented cytokine-induced stress in both systems, but beneficial effects were stronger in composite capsules. Our findings show novel effects of PSCs on islet health. Islets and PSCs coculturing or co-transplantation might mitigate the inflammation stress and improve islet transplantation outcomes.