Date of Graduation

Fall 2024

Degree

Master of Science in Cell and Molecular Biology

Department

Biomedical Sciences

Committee Chair

Jianjie Wang

Abstract

Inflammatory mediators profoundly affect endothelial permeability by disrupting its barrier function, resulting in increased permeability (hyperpermeability). Interestingly, hyperpermeability is involved in a wide range of disorders and diseases, and can pose significant risks to life in diseases, such as acute respiratory distress syndrome and systemic capillary leak syndrome. Though vascular leakage, a hallmark of inflammation, profoundly exacerbates the pathogenesis of diseases, the direct mechanism is not fully understood. In vivo study has shown that activation of nucleotide P2Y2 receptor (P2Y2R) increases permeability in murine venule. The goal of this study was to determine the molecular mechanism of P2Y2R-induced increase in permeability in primary cultured murine skeletal muscle microvascular endothelial cells (MSMEC). Wild type (WT) and P2Y2R knock out (KO) MSMEC were used to measure permeability response to P2Y2R agonist, uridine triphosphate (UTP, 10-5M), stimulation. Transwell permeability assay was used to measure endothelial permeability to fluorescence labeled-dextran (70 kDa). It was shown that the basal permeability in WT MSMEC was comparable to that in KO MSMEC, and unstimulated WT MSMEC responded similarly to UTP stimulated WT MSMEC. Normalized data from UTP-stimulated WT and KO MSMEC showed no significant difference in permeability over a time course. However, during the period between 30 and 90 minutes of UTP stimulation, there was a noticeable trend of increased permeability in WT MSMEC compared to KO MSMEC. Based on the results from current literature and our previous in vivo work, the undetected permeability response to UTP observed in this study may be attributed to the limited sensitivity of the macromolecule tracking approach using Transwell inserts, which may fail to capture transient changes in permeability upon initial stimulation with UTP in WT MSMEC. Identifying the mechanistic involvement of MSMEC P2Y2R in modulation of permeability will advance the understanding of vascular permeability during inflammatory response and contribute to the treatment of a wide range of diseases.

Keywords

P2Y2 receptor, purinergic receptors, G protein-coupled receptor, microvascular endothelial permeability, murine primary endothelial cells

Subject Categories

Medicine and Health Sciences

Copyright

© Dana S. James

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Open Access

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