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Rnd3 accelerates termination of thrombin‐induced Rho activation in endothelial cells.
Author(s) -
Kurtz Kristine M,
Breslin Jerome W
Publication year - 2009
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.23.1_supplement.762.14
Subject(s) - umbilical vein , transfection , microbiology and biotechnology , thrombin , chemistry , endothelial stem cell , rhoa , intravital microscopy , in vivo , signal transduction , in vitro , biology , platelet , biochemistry , immunology , gene
Rnd3, recently implicated as an endogenous inhibitor of Rho signaling, is present in cultured endothelial cells but its functional significance is unknown. We tested the hypothesis that Rnd3 inhibits Rho signaling in endothelial cells during an inflammatory challenge. We assessed in vivo expression of Rnd3 in rat mesenteric microvessels by immunofluorescence confocal microscopy. RhoA activation was assessed in human umbilical vein endothelial cells (HUVEC) treated with thrombin (1U/ml) for 5, 30, or 60 min. using a commercially available G‐LISA kit. HUVEC were mock‐transfected or transfected with pMAT‐FLAG‐Rnd3, Rnd3 siRNA, or siCONTROL RNA. The results show that Rnd3 is present in vivo within endothelial cells. Thrombin significantly increased Rho activity in all groups treated, however the time course varied, particularly at 60 min. Rho‐GTP levels were significantly lower in pMAT‐FLAG‐Rnd3 cells compared to mock at 60 min. In contrast, in Rnd3 siRNA cells, Rho‐GTP levels at 60 min. were significantly higher than siCONTROL cells. The data indicate that Rnd3 accelerates termination of thrombin‐induced Rho activation, which may serve as an important negative feedback signal for concluding inflammatory signaling in endothelial cells. Supported by NIH RR018766 and a grant from the American Heart Association.