Abstract for presentation at Australia and New Zealand Society of Nephrology Annual Scientific Conference

Protease Activated Receptor-1 Regulates Thrombospondin in Unilateral Ureteral Obstruction

  • Melissa Sam, Nephrology, Prince of Wales Hospital and University of New South Wales, Australia
  • Dr Sean Kennedy, Nephrology, Prince of Wales Hospital and University of New South Wales, Australia
  • Dr Philip Peake, Nephrology, Prince of Wales Hospital, Australia
  • Prof John Charlesworth, Prince of Wales Hospital and University of New South Wales, Australia
  • Dr Jonathan Erlich, Nephrology, Prince of Wales Hospital and University of New South Wales, Australia

    • Thrombospondin-1 (TSP-1) contributes to fibrosis by a variety of mechanisms including caspase-3 induced apoptosis. We investigated whether thrombin signalling via protease activated receptor-1 (PAR-1) causes TSP-1 upregulation in unilateral ureteral obstruction (UUO).
    Methods. The left ureter of wild type (C57BL/6) mice and PAR-1 deficient mice (PAR-1) was ligated and the right underwent a sham procedure. Real-time RTPCR quantitated the expression of mRNA in kidneys collected at 3, 7 and 14 days for TSP-1, CD36 and caspase-3 relative to 18S rRNA. For in-vitro experiments, human proximal tubular cells (HK2) were grown to confluence in serum free medium and incubated with thrombin for varying periods of time. TSP-1, TGF-ß and 18S mRNA was quantitated and compared to controls. Results are expressed as mean ± SEM.
    Results. TSP-1 was upregulated in the ligated kidney of C57BL/6 but not PAR-1 mice at 3 days. The expression of TSP-1 in left relative to right kidneys was greater in C57BL/6 than PAR-1 at 7 (6.58±1.31 vs. 2.38±1.40, C57BL/6 vs. PAR-1; p=0.03) and 14 days (7.64+/-1.25 vs. 3.46+/-0.5; p=0.03). Caspase-3 was increased in ligated kidneys of C57BL/6 mice at 3 days (2.4±0.45 vs. 0.77±0.59, C57BL/6 vs. PAR-1; p=0.04). CD36 expression was not altered by UUO in either group at 3 days. Incubation of HK2 cells with 5U/mL of thrombin for 4 hours caused greater than 3-fold increase in TSP-1 mRNA without altering TGF-ß.
    Conclusion. PAR-1 signalling in UUO increases TSP-1 expression and apoptotic signalling, possibly by a direct effect of thrombin on tubular epithelium.
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    Conference Organiser - ICMS Pty Ltd