A Physiological Pattern of Oxygenation Using Perfluorocarbon-Based Culture Devices Maximizes Pancreatic Islet Viability and Enhances β-Cell Function

Abstract

Conventional culture vessels are not designed for physiological oxygen (O 2 ) delivery. Both hyperoxia and hypoxia-commonly observed when culturing cells in regular plasticware-have been linked to reduced cellular function and death. Pancreatic islets, used for the clinical treatment of diabetes, are especially sensitive to sub-and supraphysiological O 2 concentrations. A result of current culture standards is that a high percentage of islet preparations are never transplanted because of cell death and loss of function in the 24-48 h postisolation. Here, we describe a new culture system designed to provide quasiphysiological oxygenation to islets in culture. The use of dishes where islets rest atop a perfluorocarbon (PFC)-based membrane, coupled with a careful adjustment of environmental O 2 concentration to target the islet physiological pO 2 range, resulted in dramatic gains in viability and function. These observations underline the importance of approximating culture conditions as closely as possible to those of the native microenvironment, and fill a widely acknowledged gap in our ability to preserve islet functionality in vitro. As stem cell-derived insulin-producing cells are likely to suffer from the same limitations as those observed in real islets, our findings are especially timely in the context of current efforts to define renewable sources for transplantation.

Mitigating hypoxic stress on pancreatic islets via in situ oxygen generating biomaterial

“…This HIF-1α mediated cascade is activated from the start of islet procurement, as islets suffer from fluctuations in ambient oxygen tension during pancreas processing due to the stripping of their native vasculature. Cultured islet spheroids continue to experience unfavorable oxygen gradients, despite the practice of low cell density culture [10, 11]. These detrimental oxygen conditions are intensified following intrahepatic transplantation, as grafts become hypoxic due to inadequate site oxygenation [12, 13].…”

Hyperoxia reduces insulin release and induces mitochondrial dysfunction with possible implications for hyperoxic treatment of neonates

“…Using rat islets, we did not detect any positive effects of culture at 30 or 40% oxygen (unpublished observations). In another study, culture of human islets below atmospheric oxygen was reported to induce optimal functionality (Fraker et al 2013). A discrepancy between studies could be due to differences in oxygen concentrations between studies as well as other factors; here we tested for effects after 24 h, whereas the published report (Komatsu et al 2016) tested after 7 days of oxygen exposure.…”

“…In another study, culture of human islets below atmospheric oxygen was reported to induce optimal functionality (Fraker et al. ). A discrepancy between studies could be due to differences in oxygen concentrations between studies as well as other factors; here we tested for effects after 24 h, whereas the published report (Komatsu et al.…”

Beta-cell replacement therapy

“…A short culture period to cull nonviable cells and to decrease tissue factor expression and IBMIR is also an important improvement in alloislet transplantation [57]. Importantly, recent advances in islet isolation techniques and characterization have really augmented islet yield and function and may drive further improvements in outcomes after single donor islet allotransplantation by way of an increased functional b-cell mass [53,[58][59][60][61][62]63 && ]. However, unlike islet autotransplants, the function of allotransplants is not as durable over time [10].…”