Departments of Biochemistry & Molecular Biology and Physiology & Pharmacology, Libin Cardiovascular Institute, Cumming School of Medicine, The University of Calgary, 3330 Hospital Drive N.W., Calgary, AB T2N 4N1, Canada.
Institute of Cardiovascular Disease, Key Laboratory for Arteriosclerosis of Hunan Province, Hengyang Medical College, University of South China, Hengyang 421001, China.
Cells. 2023 May 24;12(11):1461. doi: 10.3390/cells12111461.
(+)-JQ1, a specific chemical inhibitor of bromodomain and extraterminal (BET) family protein 4 (BRD4), has been reported to inhibit smooth muscle cell (SMC) proliferation and mouse neointima formation via BRD4 regulation and modulate endothelial nitric oxide synthase (eNOS) activity. This study aimed to investigate the effects of (+)-JQ1 on smooth muscle contractility and the underlying mechanisms. Using wire myography, we discovered that (+)-JQ1 inhibited contractile responses in mouse aortas with or without functional endothelium, reducing myosin light chain 20 (LC20) phosphorylation and relying on extracellular Ca. In mouse aortas lacking functional endothelium, BRD4 knockout did not alter the inhibition of contractile responses by (+)-JQ1. In primary cultured SMCs, (+)-JQ1 inhibited Ca influx. In aortas with intact endothelium, (+)-JQ1 inhibition of contractile responses was reversed by NOS inhibition (L-NAME) or guanylyl cyclase inhibition (ODQ) and by blocking the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT) pathway. In cultured human umbilical vein endothelial cells (HUVECs), (+)-JQ1 rapidly activated AKT and eNOS, which was reversed by PI3K or ATK inhibition. Intraperitoneal injection of (+)-JQ1 reduced mouse systolic blood pressure, an effect blocked by co-treatment with L-NAME. Interestingly, (+)-JQ1 inhibition of aortic contractility and its activation of eNOS and AKT were mimicked by the (-)-JQ1 enantiomer, which is structurally incapable of inhibiting BET bromodomains. In summary, our data suggest that (+)-JQ1 directly inhibits smooth muscle contractility and indirectly activates the PI3K/AKT/eNOS cascade in endothelial cells; however, these effects appear unrelated to BET inhibition. We conclude that (+)-JQ1 exhibits an off-target effect on vascular contractility.
（+）-JQ1 是一种溴结构域和末端（BET）家族蛋白 4（BRD4）的特异性化学抑制剂，据报道，它通过 BRD4 调节抑制平滑肌细胞（SMC）增殖和小鼠新生内膜形成，并调节内皮型一氧化氮合酶（eNOS）活性。本研究旨在探讨（+）-JQ1 对平滑肌收缩性的影响及其潜在机制。通过线描法，我们发现（+）-JQ1 抑制有或无功能内皮的小鼠主动脉的收缩反应，减少肌球蛋白轻链 20（LC20）磷酸化，且依赖于细胞外 Ca。在缺乏功能内皮的小鼠主动脉中，BRD4 敲除不改变（+）-JQ1 对收缩反应的抑制作用。在原代培养的 SMC 中，（+）-JQ1 抑制 Ca 内流。在完整内皮的主动脉中，NOS 抑制（L-NAME）或鸟苷酸环化酶抑制（ODQ）以及阻断磷脂酰肌醇 3-激酶（PI3K）/蛋白激酶 B（AKT）通路逆转了（+）-JQ1 对收缩反应的抑制作用。在培养的人脐静脉内皮细胞（HUVEC）中，（+）-JQ1 迅速激活 AKT 和 eNOS，PI3K 或 AKT 抑制可逆转这一作用。（+）-JQ1 腹腔注射降低了小鼠的收缩压，这一作用被 L-NAME 共同处理所阻断。有趣的是，（+）-JQ1 对主动脉收缩性的抑制作用及其对 eNOS 和 AKT 的激活作用可被（-）-JQ1 对映体模拟，而（-）-JQ1 对映体在结构上不能抑制 BET 溴结构域。综上所述，我们的数据表明，（+）-JQ1 直接抑制平滑肌收缩性，并间接激活内皮细胞中的 PI3K/AKT/eNOS 级联反应；然而，这些作用似乎与 BET 抑制无关。我们得出结论，（+）-JQ1 对血管收缩性表现出非靶标效应。
