Design and use of methods for peptide secretion studies

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A hallmark of Type 2 diabetes is impaired insulin secretion from beta cells of the islets of Langerhans. Type 1 diabetes is characterized by autoimmune attack on beta cells and subsequent loss of insulin secretion. A promising treatment for type 1 diabetes is transplant of islets; however, this treatment is presently limited by inability to adequately assess islets used in transplant. Greater understanding of islets and insulin secretion is of fundamental importance in determining the root causes of diabetes and developing potential treatments. Studies of islets are critically dependent upon instrumentation that can detect their biochemical and physiological dynamics. The overall objective of this work is to develop advanced instrumentation suitable for measuring chemical events related to peptide secretion from islets of Langerhans. Our first two aims are to develop microfluidic tools that will allow single islets to be cultured while simultaneously monitoring hormone and metabolite secretion and metabolism. To allow cell-cell interactions to be studied, the microfluidic chip will incorporate a microcirculatory system that allows two different cell types to be cultured and interact during the measurements. Our third aim is to develop a metabolomic method for simultaneous measurement of polar anionic compounds known to be critically involved in glucose-stimulated insulin secretion. The method will be based on multi-dimensional separations and mass spectrometry detection. Our final aim is to use these methods in collaborative studies including characterization of islets that may be used for transplant with the goal of determining if the measurements may be useful for islet assessment and determining the effect of key gene knock-outs on islet physiology. These methods will offer new opportunities for studying the root causes of diabetes and devising novel treatments. LAY SUMMARY. Diabetes is an enormous medical problem afflicting at least 8 million people and costing over $100 billion in 2002 in the US. Diabetes is characterized by impaired insulin secretion from cell clusters known as islets of Langerhans. Our research will result in novel instrumentation that will allow studies of the chemical changes associated with normal and diseased islet function in unprecedented detail. Assessment of such chemical changes is expected to help identify novel therapeutic targets.