Smooth Muscle Research Group, Department of Biochemistry & Molecular Biology, Libin Cardiovascular Institute of Alberta, University of Calgary, Calgary, Alberta T2N 4N1, Canada.
J Cell Physiol. 2010 Mar;222(3):556-64. doi: 10.1002/jcp.21970.
The cardiovascular benefits of statins, including atorvastatin (ATV), have been reported to be gender-dependent, but the underlying mechanism is unclear. In this study we examine whether estrogen and its metabolite, 2-methoxyestradiol (2ME), affect the rounding response of human vascular smooth muscle cells (SMCs) induced by ATV. Twenty-four hour treatment with ATV (10-100 microM) induced rounding of cultured human SMCs. Addition of 2ME (1-20 microM), but not 17beta-estradiol, for 2 h induced re-spreading of rounded cells. Our further studies showed that the effects of 2ME were mimicked by microtubule-disrupting drugs and inhibited by taxol. Inhibition of RhoA and ROCK (Rho-kinase) by C3-toxin and H-1152, respectively, blocked 2ME effects. 2ME effects were also blocked by treatment with either actin-interfering drugs, such as cytochalasin D and jasplakinolide, or myosin inhibitor blebbistatin. ML-7 and -9, the inhibitors for myosin light chain kinase, inhibited 2ME effect as well. ATV treatment induced a decrease of F-actin content and Thr18/Ser19 dual phosphorylation of myosin regulatory light chain (MRLC), which was rescued by 2ME or mevalonate. The rescue effects of 2ME on F-actin content and MRLC dual phosphorylation were abolished by taxol or H-1152. In addition, kinesin Eg5 inhibitor monastrol and dynein inhibitor erythro-9-3-(2-hydroxynonyl) adenine (EHNA) significantly blocked 2ME effects. Finally, our results revealed that 2ME inhibited the migration of SMCs induced by ATV (0.1 microM) in wound healing assay and Boyden chamber assay. In summary, our data show that 2ME, but not estrogen, inhibits ATV-induced rounding of human SMCs through induction of microtubule disassembly and activation of the Rho-ROCK-actinomyosin pathway.
他汀类药物（包括阿托伐他汀）的心血管益处已被报道与性别有关，但潜在机制尚不清楚。在这项研究中，我们研究了雌激素及其代谢物 2-甲氧基雌二醇（2ME）是否会影响阿托伐他汀诱导的人血管平滑肌细胞（SMC）的圆化反应。24 小时用阿托伐他汀（10-100μM）处理可诱导培养的人 SMC 圆化。添加 2ME（1-20μM）而非 17β-雌二醇 2 小时可诱导圆化细胞重新铺展。我们的进一步研究表明，2ME 的作用可被微管破坏药物模拟，并被紫杉醇抑制。用 C3 毒素和 H-1152 分别抑制 RhoA 和 ROCK（Rho 激酶）可阻断 2ME 的作用。用肌动蛋白干扰药物（如细胞松弛素 D 和 jasplakinolide）或肌球蛋白抑制剂 blebbistatin 处理也可阻断 2ME 的作用。ML-7 和 -9，肌球蛋白轻链激酶抑制剂，也抑制了 2ME 的作用。阿托伐他汀处理诱导 F-肌动蛋白含量降低和肌球蛋白调节轻链（MRLC）的 Thr18/Ser19 双磷酸化，这可被 2ME 或甲羟戊酸挽救。2ME 对 F-肌动蛋白含量和 MRLC 双磷酸化的挽救作用被紫杉醇或 H-1152 阻断。此外，驱动蛋白 Eg5 抑制剂 monastrol 和动力蛋白抑制剂 erythro-9-（2-羟壬基）腺嘌呤（EHNA）显著阻断了 2ME 的作用。最后，我们的结果表明，2ME 抑制阿托伐他汀（0.1μM）诱导的 SMC 迁移在伤口愈合测定和 Boyden 室测定中。总之，我们的数据表明，2ME，而不是雌激素，通过诱导微管解聚和激活 Rho-ROCK-肌动蛋白通路抑制阿托伐他汀诱导的人 SMC 圆化。
