Background: In order to facilitate in vitro study of pancreatic islets before the transplant, a microfluidic chip was designed to trap individual islet in an array format which allows us to study physiology and pathophysiology of islets in high-resolution fashion for high-content imaging. METHODS: (1) A microfluidic array was designed based on hydrodynamic principle for pressure drop in a micro channel. (2) Fluid flow simulation performed using COMSOL to verify design principle and minimal shear stress on islet cells. (3) A thin PDMS membrane was integrated as an oxygen controller. RESULTS: (1) Flow simulation verified the design principle, showing maximum velocity in trapping sites with minimum shear stress. (2) The device is capable of trapping up to 200 islets individually in an array format with 99% efficiency. (3)Both loading/trapping islets and fluid flow control were performed using gravity based flow control which eliminated the need for pumps. (4) The oxygen controller can provide fast oxygen microenvironment (less than 30 s) and varying and consistent oxygen concentrations and profiles. (5) As a result of having one layer design and using a very thin glass substrate,the device allowed multiparametric fluorescent and confocal imaging of islet cellular and subcellular metabolic activity and ion signaling such as mitochondrial energetics, ROS levels, calcium influx, and redox activity for high resolution high-content imaging purposes. CONCLUSION: We designed, fabricated, and validated a novel microfluidic platform that allows trapping individual islets with high efficiency. Device also allowed for co-culturing islets and hypoxia studies, showing an improvement over conventional hypoxia chambers. The device allowed high-resolution imaging of islet using fluorescence and confocal microscopy and high-content multiparametric imaging of key insulin stimulator-secretion coupling factors. This work demonstrates the feasibility of array-based cellomics analysis.

Objective: Diazoxide (DZ) has been shown as an effective pharmacological ischemic preconditioning agent. In a previous report, we showed that DZ had a strong protection of rodent islets against ischemia-reperfusion injury. In this follow-up study, we are investigating whether DZ supplementation to University of Wisconsin (UW) solution during pancreas harvesting and preservation has similar cytoprotection in non-human primate. Research Design and Methods: Cynomolgus monkey pancreata were flushed with UW (n = 3) or UW + 150 μM DZ (n = 3) during pancreas harvesting and preserved for 8 hrs prior to islet isolation. Islet yields, in vitro, and in vivo function were evaluated. Results: (i) Significantly higher islet yields were observed in the UW+DZ group than in the UW group (5,7887 vs. 23,574 IEq/pancreas; p = 0.02. and 5,396 vs. 1,646 IEq/gram; p = 0.019). (ii) The DZ-treated islets had higher insulin content (59.22 vs. 40.35 ng/islet; p = 0.006). The islets from the UW + DZ group post culture have significantly higher insulin positive cells per islet (96.15% vs. 82.74%; p = 0.004). (iv)The DZ-treated islets had a significant lower apoptotic cells per islet post culture (1.64% vs. 9.85%, p = 0.0015). (v) The DZ-treated islets had improved in vitro calcium influx in response to glucose challenge and in vivo function reflected in a higher cure rate and shorter days needed to reverse diabetes in a nude mouse transplant model. Conclusion: This study in NHP confirmed our previous observation in rodent model, showing similar DZ cellular protection on islet cells against ischemia-reperfusion injury that provides a strong evidence for future DZ application in human islet and pancreas transplantation.

This chapter discusses various microfluidic technologies that have been developed to study islets and beta-cells. The chapter first introduces key issues in the field of pancreatic islet transplantation as a clinical therapy for Type I diabetes. It then reviews microfluidic technologies that have been developed for the study of pancreatic islet and beta-cell physiology and disease pathophysiology. The chapter then describes the design, fabrication, and application of UIC's microfluidic-based multimodal islet perifusion and live-cell imaging system. Protocols are available at the end of the chapter.