Introduction:
Although a causative role for RhoA-Rho kinase signaling has been recognized in the development of human hypertension, the molecular mechanism(s) as well as the RhoA exchange factors (GEFs) responsible for the over activation of RhoA remain unknown. Arhgef1 has been recently identified as a RhoA GEF involved in Ang II-mediated regulation of vascular tone and hypertension in mice.
Hypothesis:
Here we assessed the hypothesis that Arhgef1 is activated and involved in the activation of RhoA-Rho kinase signaling by Ang II in humans.
Methods:
We used in vitro stimulation of human coronary artery smooth muscle cells and human peripheral blood cells (PBMC) by Ang II (0.1 μmol/L), and PBMC isolated from normotensive subjects before and after activation of the renin-angiotensin system by a low-salt diet for 7 days (checked by the increase in plasma aldosterone and active renine). Activation of Arhgef1 was monitored by measuring its tyrosine phosphorylation by western blot and phosphorylation of the Rho kinase target MYPT was used to measure the activity of RhoA/Rho kinase signaling.
Results:
In vitro, Ang II induced activation of Arhgef1 in human vascular smooth muscle cells and PBMC. Silencing of Arhgef1 expression by siRNA in human vascular smooth muscle cells inhibited Ang II-induced activation of RhoA-Rho kinase signaling (0.67±0.12 vs 3.21±0.91relative to unstimulated condition, n=4, P<0.05). In normotensive subjects, activation of the renin-angiotensin system by a low-salt diet stimulated Arhgef1 activity in PBMC (1.26±0.12-fold over basal level, n=39, P<0.05). This activation was associated with an increase activity of RhoA-Rho kinase signaling (1.33±0.14-fold over basal level, n=39, P<0.05).
Conclusions:
Our results show that Ang II stimulates Arhgef1 activity and strongly suggest that Arhgef1 mediates Ang II-induced RhoA activation in humans. Moreover, we show for the first time that measurement of RhoA GEF activity in PBMC might be a useful method to evaluate RhoA GEF activity in humans.
Retraction of: MicroRNA-141 Inhibits the Proliferation of Penile Cavernous Smooth Muscle Cells Associated with Down-Regulation of the Rhoa/Rho Kinase Signaling Pathway
