Data_Sheet_1_Large-conductance Ca2 +-activated K+ channel β1-subunit maintains the contractile phenotype of vascular smooth muscle cells.docx

BackgroundVascular smooth muscle cells (VSMCs) phenotype switching is very important during the pathogenesis and progression of vascular diseases. However, it is not well understood how normal VSMCs maintain the differentiated state. The large-conductance Ca2+-activated K+ (BKCa) channels are widely expressed in VSMCs and regulate vascular tone. Nevertheless, there is limited understanding of the role of the BKCa channel in modulation of the VSMC phenotype.
Methods and resultsWe assessed BKCa channel expression levels in normal and injured carotid arteries from rats of the balloon-injury model. A strong decrease of BKCa-β1 was seen in the injured carotid arteries, accompanied by a parallel decrease of the VSMC contractile markers. BKCa-β1 in primary rat aortic VSMCs was decreased with the increase of passage numbers and the stimulation of platelet-derived growth factor (PDGF)-BB. Conversely, transforming growth factor β upregulated BKCa-β1. Meanwhile, the BKCa-β1 level was positively associated with the levels of VSMC contractile proteins. Intravenous injection of PDGF-BB induced downregulation of BKCa-β1 expression in the carotid arteries. Knockdown of BKCa-β1 favored VSMC dedifferentiation, characterized by altered morphology, abnormal actin fiber organization, decreased contractile proteins expression and reduced contractile ability. Furthermore, the resultant VSMC dedifferentiated phenotype rendered increased proliferation, migration, enhanced inflammatory factors levels, and matrix metalloproteinases activity. Studies using primary cultured aortic VSMCs from human recapitulated key findings. Finally, protein level of BKCa-β1 was reduced in human atherosclerotic arteries.
ConclusionBKCa-β1 is important in the maintenance of the contractile phenotype of VSMCs. As a novel endogenous defender that prevents pathological VSMC phenotype switching, BKCa-β1 may serve as a potential therapeutic target for treating vascular diseases including post-injury restenosis and atherosclerosis.

血管平滑肌细胞（Vascular smooth muscle cells, VSMCs）表型转换在血管疾病的发病机制与疾病进展过程中发挥关键作用。然而，目前对于正常VSMCs如何维持其分化状态的分子机制仍未完全明晰。大电导钙激活钾离子（large-conductance Ca2+-activated K+, BKCa）通道广泛表达于VSMCs中，可通过调控血管张力参与心血管生理功能的维持。但目前学界对于BKCa通道在调控VSMC表型中所扮演的角色仍了解有限。
方法与结果 本研究以球囊损伤模型大鼠为研究对象，检测了正常颈动脉与损伤颈动脉组织中BKCa通道的表达水平。结果显示，损伤颈动脉组织中BKCa-β1的表达水平显著下调，同时伴随VSMC收缩标志物表达的平行降低。在原代大鼠主动脉VSMCs中，BKCa-β1的表达水平随细胞传代次数的增加以及血小板衍生生长因子（platelet-derived growth factor, PDGF）-BB的刺激而呈下降趋势。反之，转化生长因子β（transforming growth factor β）可上调BKCa-β1的表达。与此同时，BKCa-β1的表达水平与VSMC收缩蛋白的表达呈正相关。静脉注射PDGF-BB可诱导大鼠颈动脉组织中BKCa-β1的表达下调。敲低BKCa-β1可促进VSMC去分化，具体表现为细胞形态发生改变、肌动蛋白纤维排列异常、收缩蛋白表达水平降低以及收缩能力减弱。进一步研究发现，发生去分化的VSMC表型可导致细胞增殖、迁移能力增强，炎症因子分泌水平升高以及基质金属蛋白酶活性增加。利用原代培养的人主动脉VSMCs开展的实验验证了上述核心研究结果。最后，人类动脉粥样硬化病变动脉组织中BKCa-β1的蛋白水平显著降低。
结论 BKCa-β1在维持VSMC收缩表型的过程中具有重要作用。作为一种可阻断病理性VSMC表型转换的新型内源性保护因子，BKCa-β1有望成为治疗损伤后再狭窄及动脉粥样硬化等血管疾病的潜在治疗靶点。