The Regulation of Release of Dipeptidyl Peptidase IV from Healthy and Diabetic Skeletal Muscle
Type of DegreePhD Dissertation
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Dipeptidyl peptidase IV (DPP-IV) is a multifaceted enzyme that can be shed from skeletal muscle membrane via matrixmetalloproteinases (MMPs). Tissue inhibitors of MMPs (TIMP) within the extracellular matrix (ECM) modulate the activation of MMPs. Hyperglycemia was shown to alter ECM proteins, likely causing a disruption in the shedding of DPP-IV from the membrane. However, little is known regarding the shedding of DPP-IV from skeletal muscle with stimulation and what role ECM proteins play in the mechanism. Additionally, it is not well known how hyperglycemia affects this system. In Part 1 of the study, C2C12 skeletal muscle cells were stimulated following differentiation. To determine the role of ECM proteins, MMP2 was inhibited in one set and MMP9 was inhibited in another for 6 hours prior to stimulation. For Part 2, C2C12 cells were exposed to normal (5mM), pre-diabetic (15mM), or diabetic (25mM) glucose conditions for 1, 7, or 10 days. At 10 days after differentiation, cells were stimulated and fractionated. DPP-IV released into the media was determined via fluorometric assay of pre and post media samples. MMP2, pro-MMP9, TIMP2, and DPP-IV of the membrane fraction of the cells were measured via ELISAs. In Part 1, no statistically significant changes with stimulation were seen except for the significant decrease in pro-MMP9 when cells were treated with an MMP9 inhibitor. In the second part, there were no significant differences in the DPP-IV shed following stimulation. MMP2 was increased in 25mM glucose after 7 and 10 days of exposure, pro-MMP9 was not significantly changed, TIMP2 was decreased after 1 and 7 days of exposure to 15mM glucose, and membrane bound DPP-IV was not significantly altered with glucose exposure. From this study, it remains unclear if DPP-IV is shed from skeletal muscle; however, it was shown that ECM proteins play a role in the process and can be altered with hyperglycemia.
- Neidert Final Dissertation 2017.pdf