Engineering a Pancreatic Islet Bioinstructive Microenvironment

for Improved Survival, Function, Protection, and Delivery

Clarissa Hernandez Stephens (Sherry Harbin and Craig Goergen, co-advisors)

Abstract: Over 1 million people are living with type 1 diabetes (T1D) in the United States. Islet transplantation is an attractive alternative to standard-of-care exogenous insulin injections or whole pancreas replacement for patients with T1D. Despite successes achieved to date with the widely accepted Edmonton islet transplantation protocol, shortcomings remain including i) inefficiency of islet isolation, in-vitro culture, and preservation and ii) inferior long-term engraftment and function outcomes. As such, a large number of donor pancreases (up to 4) are required per procedure. Current limitations have been largely attributed to the loss of critical microenvironmental cues from surrounding extracellular matrix (ECM) and vasculature as a result of isolation, in-vitro culture, and insufficient oxygen/nutrient supply to engrafted islets. To address these complications and potentially provide new therapies for diabetes, engineering of self-assembling, bioinstructive microenvironments could be performed for improved i) islet survival, function, and protection in vitro and ii) islet delivery and engraftment in vivo. Outcomes of this work are expected to contribute to the development of engineered islet microenvironments that improve yield, viability, and function of cultured islets as well as enhance delivery, vascularization, engraftment and long-term function of transplanted islets.

Purification of protein kinase A (PKA) from cell lysate using magnetic bead conjugation

Mrugesh Parasa (Tami Kinzer-Ursem, advisor)

Abstract: Protein kinase A (PKA) is an important protein, which plays multiple functions in the cell including regulation of transcription, ion channel activation and internalization of membrane proteins. Functional studies of this protein are hampered by the currently used time intensive purification methods. We developed an expedited purification protocol using a myristic acid based bio-orthogonal tag and showed that functionalized magnetic beads can be used to directly purify tagged PKA from lysates. Most significantly, we showed that PKA remains active despite tagging and binding of the tagged PKA to the beads. Using these PKA beads, we performed functional studies to find dissociation constant of ATP from PKA. The methods developed in this study are widely applicable to other proteins as well.

***Bring your lunch to seminar – BMEGSA will provide snacks and drinks***