Date of Graduation

Summer 2021

Degree

Master of Science in Cell and Molecular Biology

Department

Biomedical Sciences

Committee Chair

Jianjie Wang

Keywords

P2Y₂R, nucleotide, leukocyte rolling, leukocyte adhesion, microvascular, endothelium

Subject Categories

Cell Biology | Immunology and Infectious Disease | Medical Biochemistry | Medical Cell Biology | Medical Molecular Biology | Molecular Biology

Abstract

Extracellular nucleotides (ATP, UTP) released from cells act on nucleotide receptors to promote vascular inflammation. Increased leukocyte-endothelial interaction is a hallmark of vascular inflammation. The nucleotide P2Y₂ receptor (P2Y₂R), activated by extracellular ATP≈UTP, plays a role in cardiovascular homeostasis and immune regulation. Moreover, accumulating evidence from studies in vitro and in vivo models have implicated the P2Y₂R in the inflammatory response significantly contributing to the progression and pathogenesis of asthma, atherosclerosis, sepsis, and ischemia. I hypothesized that P2Y₂R activation by UTP, an agonist of the receptor, increased leukocyte rolling and adhesion in the microvasculature from baseline. To test the hypothesis, wild type (WT, C57BL/6) and P2Y₂R-deficient (P2Y₂R-/-, C57BL/6 background) mice were utilized. In vivo leukocyte-endothelial interaction (leukocyte total flux, rolling flux, and adhesion) and microvascular hemodynamics in venule of murine skeletal muscle cremaster were assessed using intravital microscopy. A semi-automatic leukocyte tracking methodology was developed by integrating ImageJ and Trackmate for offline image analysis. In WT mice, UTP treatment significantly increased basal leukocyte rolling in a concentration- and time-dependent manner. Adhesion was concentration-dependent and peak adhesion occurred at 5 minutes. Basal rolling leukocytes in P2Y₂R-/- mice was twice as many as those in WT mice and did not change after UTP treatment. Basal adherent leukocytes in P2Y₂R-/- mice were 1.5-fold as many as those in WT mice. Collectively, the findings demonstrate that under resting conditions, P2Y₂R protects excessive interaction between venular endothelium and circulating leukocytes. UTP increases rolling in a P2Y₂R-dependent manner, while the P2Y₂R contribution to UTP-induced increase in adhesion remains elusive.

Copyright

© Spencer E. Thomas

Available for download on Tuesday, August 01, 2023

Open Access

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