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Increased Expression and Activity of RhoA Are Associated With Increased DNA Synthesis and Reduced p27^Kip1 Expression in the Vasculature of Hypertensive Rats
Tammy M. Seasholtz, Tong Zhang, Michael R. Morissette, Amy L. Howes, Amy H. Yang and Joan Heller Brown

Circulation Research. 2001;89:488-495, originally published September 14, 2001
https://doi.org/10.1161/hh1801.096337

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Increased Expression and Activity of RhoA Are Associated With Increased DNA Synthesis and Reduced p27^Kip1 Expression in the Vasculature of Hypertensive Rats
Tammy M. Seasholtz, Tong Zhang, Michael R. Morissette, Amy L. Howes, Amy H. Yang, Joan Heller Brown

Circulation Research September 2001, 89 (6) 488-495; DOI: https://doi.org/10.1161/hh1801.096337

By Tammy M. Seasholtz, Tong Zhang, Michael R. Morissette, Amy L. Howes, Amy H. Yang and Joan Heller Brown

Figure 1. Enhanced agonist-stimulated DNA synthesis in vessels and ASMCs from SHR versus WKY. A, Serum- but not thrombin-stimulated [3H]thymidine inco...Show More

Figure 1. Enhanced agonist-stimulated DNA synthesis in vessels and ASMCs from SHR versus WKY. A, Serum- but not thrombin-stimulated [3H]thymidine incorporation was significantly higher (*P<0.01) than control values in vessels from WKY rats. Thrombin (Thr)- and serum-stimulated [3H]thymidine incorporation of SHR vessels was significantly greater than control SHR responses (**P<0.001) and significantly greater than corresponding WKY responses (#P<0.001). Data represent mean±SE from 4 vessels and are expressed as respective control. B, Thrombin produced significant increases in [3H]thymidine incorporation in ASMCs from WKY (*P<0.05) and SHR (**P<0.001) compared with respective control (Ctr) values. The thrombin (Thr)-stimulated response of SHR cells was significantly greater (#P<0.001) than that of WKY cells. C3 treatment fully blocked WKY and SHR Thr-stimulated DNA synthesis. Data represent mean±SE from 2 experiments with 4 to 7 replicates.Show Less
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Increased Expression and Activity of RhoA Are Associated With Increased DNA Synthesis and Reduced p27^Kip1 Expression in the Vasculature of Hypertensive Rats
Tammy M. Seasholtz, Tong Zhang, Michael R. Morissette, Amy L. Howes, Amy H. Yang, Joan Heller Brown

Circulation Research September 2001, 89 (6) 488-495; DOI: https://doi.org/10.1161/hh1801.096337

By Tammy M. Seasholtz, Tong Zhang, Michael R. Morissette, Amy L. Howes, Amy H. Yang and Joan Heller Brown

Figure 2. Selective increase in RhoA expression in vessels from SHR. The expression of RhoA (A), Ras (B), Rho kinase (C), and Rho-GDIα (D) in aortic h...Show More

Figure 2. Selective increase in RhoA expression in vessels from SHR. The expression of RhoA (A), Ras (B), Rho kinase (C), and Rho-GDIα (D) in aortic homogenates from adult SHR and WKY (S and W, respectively) was measured by use of Western blot analysis. RhoA expression was also examined in aortas from 4-week-old SHR and WKY (A). Although RhoA per mg protein was found to be significantly higher (*P<0.01) in SHR aortas, levels of the other proteins were not different. Data represent mean±SE from 6 or 7 rats from each strain.Show Less
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Increased Expression and Activity of RhoA Are Associated With Increased DNA Synthesis and Reduced p27^Kip1 Expression in the Vasculature of Hypertensive Rats
Tammy M. Seasholtz, Tong Zhang, Michael R. Morissette, Amy L. Howes, Amy H. Yang, Joan Heller Brown

Circulation Research September 2001, 89 (6) 488-495; DOI: https://doi.org/10.1161/hh1801.096337

By Tammy M. Seasholtz, Tong Zhang, Michael R. Morissette, Amy L. Howes, Amy H. Yang and Joan Heller Brown

Figure 3. RhoA activity is elevated in aortas from SHR versus WKY rats. A, RhoA activation was assessed by GST-RBD. B, RhoA activation was assessed by...Show More

Figure 3. RhoA activity is elevated in aortas from SHR versus WKY rats. A, RhoA activation was assessed by GST-RBD. B, RhoA activation was assessed by labeling with [35S]GTPγS. The amount of GST-RBD and [35S]GTPγS bound to RhoA was significantly greater in aortic homogenates from SHR than those from WKY (*P<0.05). Data represent mean±SE from 4 aortas from each strain.Show Less
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Increased Expression and Activity of RhoA Are Associated With Increased DNA Synthesis and Reduced p27^Kip1 Expression in the Vasculature of Hypertensive Rats
Tammy M. Seasholtz, Tong Zhang, Michael R. Morissette, Amy L. Howes, Amy H. Yang, Joan Heller Brown

Circulation Research September 2001, 89 (6) 488-495; DOI: https://doi.org/10.1161/hh1801.096337

By Tammy M. Seasholtz, Tong Zhang, Michael R. Morissette, Amy L. Howes, Amy H. Yang and Joan Heller Brown

Figure 4. RhoA expression and Thr-stimulated RhoA activation are increased in ASMCs from SHR. A, Western blot analysis of ASMCs from SHR and WKY revea...Show More

Figure 4. RhoA expression and Thr-stimulated RhoA activation are increased in ASMCs from SHR. A, Western blot analysis of ASMCs from SHR and WKY revealed that total RhoA per mg protein is higher in SHR. B, RhoA-RBD binding was used to measure activation of RhoA. Basal RhoA activity was not different between ASMCs from SHR and WKY rats. Thr (12 nmol/L) produced a significant increase in RhoA-RBD binding in ASMCs from WKY (*P<0.05) and ASMCs from SHR (**P<0.001) compared with respective control (Ctr) values, but the increase was significantly higher in cells from SHR (#P<0.01). Data represent mean±SE from 3 experiments, with each performed in duplicate.Show Less
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Increased Expression and Activity of RhoA Are Associated With Increased DNA Synthesis and Reduced p27^Kip1 Expression in the Vasculature of Hypertensive Rats
Tammy M. Seasholtz, Tong Zhang, Michael R. Morissette, Amy L. Howes, Amy H. Yang, Joan Heller Brown

Circulation Research September 2001, 89 (6) 488-495; DOI: https://doi.org/10.1161/hh1801.096337

By Tammy M. Seasholtz, Tong Zhang, Michael R. Morissette, Amy L. Howes, Amy H. Yang and Joan Heller Brown

Figure 5. RhoA expression and activity are increased in aortas from L-NAME–treated rats. A, Western blot analysis of aortic homogenates from WKY and L...Show More

Figure 5. RhoA expression and activity are increased in aortas from L-NAME–treated rats. A, Western blot analysis of aortic homogenates from WKY and L-NAME–treated rats showed that total RhoA per mg protein is higher in the L-NAME group (*P<0.01). B, RhoA-RBD binding was also significantly greater in aortic homogenates from L-NAME–treated rats (*P<0.01). Data represent mean±SE from 4 to 6 rats per group.Show Less
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Increased Expression and Activity of RhoA Are Associated With Increased DNA Synthesis and Reduced p27^Kip1 Expression in the Vasculature of Hypertensive Rats
Tammy M. Seasholtz, Tong Zhang, Michael R. Morissette, Amy L. Howes, Amy H. Yang, Joan Heller Brown

Circulation Research September 2001, 89 (6) 488-495; DOI: https://doi.org/10.1161/hh1801.096337

By Tammy M. Seasholtz, Tong Zhang, Michael R. Morissette, Amy L. Howes, Amy H. Yang and Joan Heller Brown

Figure 6. Reduced expression of the CKI p27Kip1, but not p21Waf1/Cip1, in aortas from SHR. Western blot analysis of aortic homogenates with use of an...Show More

Figure 6. Reduced expression of the CKI p27Kip1, but not p21Waf1/Cip1, in aortas from SHR. Western blot analysis of aortic homogenates with use of an antibody to p27Kip1 (A) revealed a doublet band at the expected molecular weight (inset: W indicates WKY; S, SHR). The upper band was not significantly different in aortic homogenates between strains (data not shown). The lower band was significantly decreased in homogenates from SHR (*P<0.01). Western blot with an antibody to p21Waf1/Cip1 (B) showed no difference in aortic homogenates from SHR vs WKY rats. Data represent mean±SE from 7 aortas from each strain.Show Less
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Increased Expression and Activity of RhoA Are Associated With Increased DNA Synthesis and Reduced p27^Kip1 Expression in the Vasculature of Hypertensive Rats
Tammy M. Seasholtz, Tong Zhang, Michael R. Morissette, Amy L. Howes, Amy H. Yang, Joan Heller Brown

Circulation Research September 2001, 89 (6) 488-495; DOI: https://doi.org/10.1161/hh1801.096337

By Tammy M. Seasholtz, Tong Zhang, Michael R. Morissette, Amy L. Howes, Amy H. Yang and Joan Heller Brown

Figure 7. Levels of p27Kip1 are reduced by RhoA (L63RhoA) and PI3K (p110CAAX). A, Infection of ASMCs from Sprague-Dawley rats with adenoviral construc...Show More

Figure 7. Levels of p27Kip1 are reduced by RhoA (L63RhoA) and PI3K (p110CAAX). A, Infection of ASMCs from Sprague-Dawley rats with adenoviral constructs encoding for L63RhoA resulted in a significant decrease in p27Kip1, but not p21Waf1/Cip1, expression. B, Adenoviral infection with p110CAAX produced a significant decrease in p27Kip1 expression. Wortmannin (Wort, 10 nmol/L) was added to ASMCs 1 hour before adenoviral infection and again 24 hours later. The L63RhoA-mediated decrease in p27Kip1 was fully inhibited by Wort treatment. There was no difference between AdCMV in the presence or absence of Wort. Data represent mean±SE from 3 or 4 experiments performed in duplicate or triplicate. *P<0.05 compared with AdCMV; #P<0.05 compared with L63RhoA alone.Show Less
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Increased Expression and Activity of RhoA Are Associated With Increased DNA Synthesis and Reduced p27^Kip1 Expression in the Vasculature of Hypertensive Rats
Tammy M. Seasholtz, Tong Zhang, Michael R. Morissette, Amy L. Howes, Amy H. Yang, Joan Heller Brown

Circulation Research September 2001, 89 (6) 488-495; DOI: https://doi.org/10.1161/hh1801.096337

By Tammy M. Seasholtz, Tong Zhang, Michael R. Morissette, Amy L. Howes, Amy H. Yang and Joan Heller Brown

Figure 8. Thr stimulates DNA synthesis and decreases levels of p27Kip1. A, Wort (10 nmol/L) was added to ASMCs from Sprague-Dawley rats 1 hour before...Show More

Figure 8. Thr stimulates DNA synthesis and decreases levels of p27Kip1. A, Wort (10 nmol/L) was added to ASMCs from Sprague-Dawley rats 1 hour before agonist and again 24 hours later. Thr significantly increased [3H]thymidine incorporation in the absence of Wort (*P<0.001 compared with Ctr) but not in the presence of Wort (#P<0.001 compared with Thr alone). Data represent mean±SE from 3 experiments, with each performed in duplicate or triplicate. B, Stimulation with Thr, but not phenylephrine (PE), produced a decrease in p27Kip1 in ASMCs. Data are from 1 experiment performed in triplicate.Show Less