小檗碱通过可溶性鸟苷酸环化酶防治肺动脉高压的分子机制研究

Pulmonary hypertension (PH) is a fatal disease with high risk and mortality. The current PH theraputic options are limited and cannot cure the disease. Therefore, there is an urgent need to discover novel medication against PH . Our recent studies have shown that Berberine (BBR), the main functional component of the Traditional Chinese Medicine Huang Lian, can effectively attenuate the PH pathogenesis in either monocrotaline， hypoxia-, or sugen+hypoxia-induced PH in rodents, suggesting that BBR may be a a novel PH medication. Through computer-assisted molecule docking analysis, we further demonstrated that BBR can bind to the functional activation domain H-NOX of the soluble guanylate cyclase (sGC) . These evidence suggest that BBR may protect against PH through its direct binding and activation of sGC. In this study, we aim to further evaluate the beneficial role of BBR on PH, and to elucidate the functional role of sGC in BBR-mediated protection of PH. We propose the following experiments to test our hypothesis: 1) Evaluate whether BBR mediates protection of PH via sGC, and investigate the role of of BBR on sGC downstream signaling pathway；2) Investigate the mechanism underlying the protein interaction between BBR and sGC; 3) Determine the effects of BBR on the PA contraction, the proliferation, migration and intracellular calcium homeostasis of pulmonary arterial smooth muscle cells.This proposal integrates PH animal models, molecular and cell biology strategies to provide a novel PH therapeutic strategy.

肺动脉高压（PH）疾病危害大，治疗药物有限，临床亟需研发新型有效药物。小檗碱是中药黄连的主要有效成分，目前多用于治疗消化道感染。我们前期研究首次发现小檗碱对野百合碱、低氧、Sugen+缺氧诱导的PH大鼠模型具有显著治疗效果。经过计算机分子对接发现其与可溶性鸟苷酸环化酶（sGC）功能启动结构域H-NOX具有结合可能。这些结果提示小檗碱能通过结合并激活sGC从而治疗PH。本项目拟通过下列研究验证我们的假设：1）特异性敲除平滑肌细胞sGCβ1，验证sGC信号通路在小檗碱治疗PH中的作用；2）表达纯化H-NOX蛋白，验证BBR与其直接的分子间相互作用；3）通过离体血管环实验、细胞内钙离子浓度检测实验、及培养原代肺动脉平滑肌细胞，研究小檗碱对肺血管舒缩功能、细胞钙稳态及增殖与迁移的影响。从分子、细胞、组织、动物层面揭示小檗碱治疗PH可能的分子靶点及作用机制，为小檗碱用于临床治疗PH提供理论支持。

本项目探讨小檗碱是否可以通过sGC-cGMP-PKG信号通路减少肺动脉平滑肌细胞 ([Ca2+]i)增加对肺动脉高压起防治作用；验证小檗碱与sGC启动关键结构域H-NOX的结合作用，确定sGC是否为小檗碱的分子作用靶点；明确小檗碱通过sGC调节肺动脉平滑肌细胞钙稳态的具体分子机制。从而揭示小檗碱防治肺动脉高压可能的分子机制和可能的药物靶点，为小檗碱用于临床治疗PH提供理论支持。发表研究论文14篇，其中以通讯作者或第一作者在SCI杂志发表论文7篇，国内核心期刊3篇，国家专利授权3项。培养博士研究生1人，硕士研究生2人。获得广东省科技进步二等奖1项。