Date of Graduation

Summer 2021

Degree

Master of Science in Cell and Molecular Biology

Department

Biomedical Sciences

Committee Chair

Jianjie Wang

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.

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

Copyright

© Spencer E. Thomas

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