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  • Undergraduate Poster Abstracts
  • SAT-G11 ACTIVATION OF ENOS PHOSPHORYLATION STATE AND NO PRODUCTION VIA ER-α AND ER-β IN UTERINE ARTERY ENDOTHELIAL CELLS

    • Mayra Pastore ;

    SAT-G11

    ACTIVATION OF ENOS PHOSPHORYLATION STATE AND NO PRODUCTION VIA ER-α AND ER-β IN UTERINE ARTERY ENDOTHELIAL CELLS

    Mayra Pastore1, Meghan Conley2, Ronald Magness1.

    1University of Wisconsin-Madison, Madison, WI, 2University of Wisconsin-Parkside, Kenosha, WI.

    Uterine endothelial nitric oxide (NO) production is partly responsible for the maintenance of vasodilatation during physiologic states of high circulating estrogen levels such as pregnancy. Endothelial nitric oxidize synthase (eNOS) has several phosphorylation sites that correlate to its activity state and NO production. However, it is unknown if eNOS regulation depends on ER-α and/or ER-β. We hypothesize that ER-α and ER-β are capable of altering eNOS phosphorylation patterns and increase NO production. A series of experiments were set up. Endothelial cells were treated with the vehicle or increasing concentrations of E2β. The cells were treated with  E2β for 0 to 30 minutes or were pre-treated with inhibitor, ICI-182, 780. We evaluated the eNOS phosphorylation changes at Ser635, Ser1177, and Thr495 via western blotting. Endothelial cells treated with E2β, ATP, PPT (propylpyrazole-triol), and DPN (diarylpropionitrile) were also analyzed for total NO production. E2β treatment increased stimulatory phosphorylations Ser635 and Ser1177 and decreased Thr495 phosphorylation. The increased in eNOS phosphorylation at Ser635 was blocked by ICI-182,780 pre-treatment. Surprisingly, E2β and ICI-182,780 decreased the inhibitory phosphorylation at Thr495. Phosphorylation at Ser635 and Ser1177 were increased starting at 5 minutes of E2β treatment, while the phosphorylation Thr495 was reduced after 30 minutes. Lastly, E2β, ATP, PPT, and DPN treatments increase total NO production starting at 10 minutes and peaking at 30 minutes. These data support the hypothesis that E2β-induced eNOS activation via its phosphorylation state in a dose and time-dependent manner although the inhibitory phosphorylation seemed to occur through an ER-independent mechanism. Our data also support the hypothesis that NO production is shown to increase by the activation of either ER-α or ER-β.