Author

Aparna Tiwari

Date of Graduation

Summer 2014

Degree

Master of Science in Cell and Molecular Biology

Department

Biomedical Sciences

Committee Chair

Jianjie Wang

Keywords

VE-cadherin, adherens junctions, tyrosine phosphorylation at Y658 (pY658), UTP, vascular permeability

Subject Categories

Medical Molecular Biology

Abstract

It was previously demonstrated that UTP (10-5M), an agonist of P2Y2R, increases vascular permeability. However, the molecular mechanism remains to be determined. VE-cadherin, a key component of the adherens junction, is responsible for regulating endothelial permeability by forming stable endothelial cell-cell adhesion. To determine VE-cadherin responseto UTP stimulation, tyrosine phosphorylation of VE-cadherin at Y658 (pY658) was measured using Western blotting in HUVEC. There was a 16% and 19% increase in pY658 level of VE-cadherin in 5 and 15 min UTP-treated samples, respectively,compared to the control sample (n=4, p<0.005 and p<0.05).To determine whether expression level of membrane-associated VE-cadherin decreases in response to 10-5M stimulation, flow cytometry was performed. The membrane-associated and total VE-cadherin expression levels were quantified. There was no change in membrane-associated VE-cadherin expression before and after UTP (10-5M) treatment (n=3, p>>0.05). The findings demonstrate that UTP-stimulation increases pY658 levels of VE-cadherin but does not affect the expression level of membrane-associated VE-cadherin within 15 min of UTP treatment.Further study using alternative approach is needed to validate flow cytometry data.

Copyright

© Aparna Tiwari

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