锚定蛋白AKAP150/PKCα/CaV1.2在有氧运动改善原发性高血压动脉功能中的作用及机制

L-type calcium channels (CaV1.2) on the vascular smooth muscle cells (VSMCs) play an important role in maintaining vascular tone. A-kinase anchoring protein AKAP150 can assemble multifunctional signaling enzymes to precise subcellular compartments, and improve the signal transduction efficiency. Both CaV1.2 channels and AKAP150 are crucially involved in the vascular remodeling of hypertension. Our recent studies have demonstrated that aerobic exercise can significantly inhibit the upregulation of CaV1.2 function in spontaneously hypertensive rats (SHR), and efficiently alleviate the vascular tone increase. However, it is not clear whether such effects are partially mediated by AKAP150 pathway. To clarify this is very important for us to understand the benefits and mechanisms of exercise on the essential hypertension. Therefore, by combining multiple techniques, including patch-clamp, total internal reflection fluorescence (TIRF) microscopy Ca2+ sparklets measurement, in vitro transfection, and AKAP150 knock-in mouse model, we will investigate that (1) whether aerobic exercise inhibits the functional upregulation of CaV1.2 channels in SHR by inhibiting AKAP150, thus suppressing PKCα and calcineurin assembling in the sarcolemma; (2) whether these effects are mediated by angiotensin II and AT1 receptors. Taken together, the present study will explore the mechanism of AKAP150/PKCα/CaV1.2 underlying the aerobic exercise-induced improvement of the arterial function in essential hypertension. The information gained will help us to understand the cellular mechanisms of the exercise therapy on hypertension.

血管平滑肌CaV1.2通道是维持血管张力的重要机制，而A激酶锚定蛋白AKAP150可将多种酶聚集于特定细胞区域以提高信号转导效率，二者在高血压血管重构中均发挥着重要作用。我们近期研究发现有氧运动可有效抑制自发性高血压大鼠SHR动脉CaV1.2功能上调，缓解其张力增高，然而是否与AKAP150调控途径有关尚不清楚，而这对于阐明运动改善原发性高血压的作用机理具有重要意义。本课题拟采用膜片钳和全内反射荧光显微镜钙星检测技术，结合细胞转染和基因敲入等方法，探讨有氧运动是否可以通过抑制AKAP150，从而抑制PKCα和钙调神经磷酸酶于膜上聚集，经兴奋-收缩耦联和兴奋-转录耦联直接和间接抑制SHR的CaV1.2活性上调；并探讨此作用是否由血管紧张素AngII及AT1受体介导，进而揭示AKAP150/PKCα/CaV1.2在运动改善原发性高血压动脉功能中的作用及机制，为高血压运动疗法的细胞机理提供依据。

原发性高血压是脑卒中最重要的危险因素，探求高血压致卒中的发病机制，寻找原发性高血压有效的防治手段，是心血管疾病研究的热点。有氧运动可有效降低血压和卒中罹患率，但其作用机理远未明了。血管平滑肌L型钙通道（CaV1.2）是血管张力和血压调节的重要机制；而A激酶锚定蛋白AKAP150可锚定PKCα转位上膜，耦联激活CaV1.2，还可使CaN聚集膜上，调节NFAT依赖的基因表达，抑制KV2.1功能，经兴奋–收缩和兴奋–转录耦联使血管张力增加；然而AKAP150途径在运动改善高血压动脉功能中的作用及机制目前仍不清楚。本项目选用自发性高血压大鼠（SHR）并构建AKAP150平滑肌条件性过表达敲入鼠（AKAP150 smKI），施以有氧运动，采用膜片钳和TIRF钙星检测，结合生理遥测信号、离体微血管反应性检测，观察有氧运动对高血压动脉功能的改善，着重探讨AKAP150途径在其中的作用及潜在分子机理。研究发现：（1）有氧运动可有效降低SHR血压，改善脑血管功能，其重要机制是有氧运动通过抑制AKAP150，一方面抑制PKCα膜转位，减弱CaV1.2活性，另一方面抑制CaN聚集膜上，阻断KV2.1转录抑制，经兴奋–收缩和兴奋–转录耦联缓解高血压动脉张力。（2）AKAP150体外干扰可减少Ang II刺激下CaV1.2在脑动脉张力作用中的激活，缓解Ang II所致脑血管张力升高。（3）平滑肌AKAP150过表达引起血压升高，AKAP150 smKI小鼠呈现高血压特征。有氧运动可显著降低AKAP150 smKI小鼠血压，抑制血管平滑肌AKAP150表达，下调CaV1.2并上调KV2.1功能，恢复动脉正常功能。该研究为阐明运动调控高血压动脉功能的生理学机制提供了丰富的实验依据，也为高血压的研究和治疗新靶点的发现提供了出色的模型，不仅具有重要的学术价值，也对高血压运动处方和有效治疗靶点的选择具有潜在的指导意义。