Lucas Hudgins
Pronouns: He/Him/His
Research Mentor(s): Jason Knight
Research Mentor School/College/Department: Internal Medicine/Rheumatology / Medicine
Program: UROP
Session: Session 3 (11:00am – 11:50am)
Authors: Lucas Hudgins, Chao Liu, Jason Knight
Abstract
Background: Diabetes is a prevalent disease associated with high blood glucose. Diabetes is also associated with a markedly increased risk of clinically relevant cardiovascular disease, as well as preclinical endothelial dysfunction. Patients with type 1 diabetes mellitus experience increased rates of NETosis. NETosis is a process that leads to the release of pro-inflammatory and vasculopathic neutrophil extracellular traps (NETs) by activated neutrophils. NETosis has recently been hypothesized to contribute to the development of cardiovascular dysfunction in diabetic individuals. Akita (AK) mice are a model of type I diabetes that is known to experience endothelial dysfunction. We recently found that AK endothelial dysfunction can be negated by knocking out the genes for either neutrophil elastase (Elane) or peptidylarginine deiminase 4 (Pad4), two factors that are required for NETosis. We have also found that an irreversible inhibitor of neutrophil elastase, GW311616A (GW), prevents neutrophils from undergoing NETosis. Methods and Results: We tested whether GW could improve endothelial dysfunction in aged AK mice. 24-week-old WT or AK mice were fed chow supplemented with either GW or a matching vehicle for 5 weeks. Endothelial function was then determined by exposing precontracted aortic rings to acetylcholine in a wire myograph system. We found that GW-treated AK mice demonstrated superior relaxation in response to acetylcholine as compared with vehicle-treated mice, suggesting a healthier endothelium (remaining force: Akita + vehicle Emax=65.6%; Akita + GW Emax=41.2%; p<0.05). For the second part of this project, we sought to better understand the pathways underlying the protective effects of blocking NETosis. To this end, we submitted aorta samples from standard WT and AK, as well as NE-inhibited (WT-Elane-/- and AK-Elane-/- ) mice for RNA sequencing. Analysis of the RNA-sequencing data suggested multiple differentially regulated genes of interest, including Rnf208, Ccl21a, Ccl21b, and Ccl21d. Validation of the expression of these genes in the aorta sample was then performed by traditional real-time qPCR. The results for Ccl21a, Ccl21b, and Ccl21d were all validated. Conclusion: Pharmacological inhibition of NETosis by GW rescued aortic endothelial dysfunction in AK mice. Some pathways were identified by transcriptomic profiling that may
