基于脏器微血管通透性的调控，探讨生脉散及其主要补气和收摄成份固摄作用的机理

lipopolysaccharide (LPS) provokes multiorgan injury and shock, named as the prostration syndrome in Chinese medicine, in pathology of which microvascular damage and albumin extravasation play a critical role. In Chinese medicine, ShengmaiSan, a medicine capable of invigorating qi for relieving desertion, has been used to treat prostration syndrome with preferable outcomes in clinic. Our previous studies demonstrated that Yiqifumai injection (YQFM, a kind of ShengMaiSan preparation) attenuates albumin leakage from rat mesenteric venules after LPS challenge, ginsenoside Rb1 (Rb1), as a major tonifying qi component of YQFM, is able to reduce the number of caveolae in vascular endothelial cell, inhibit the expression and phosphorylation of tight junction proteins ZO-1 and VE-Cadherin. However, it is not clear whether YQFM and its major tonifying qi component Rb1 and perturbation component schisandrin B (Sch B) may confer consolidation effect attenuating vascular hyperpermeability and multiorgan injury after initiation of albumin extravasation induced by LPS in brain, lungs and mesentery. The present study is designed to investigate the effect of YQFM, Rb1, Sch B and Rb1+Sch B on LPS-induced albumin extravasation from venules and organ injury in rat brain, lungs and mesentery. By observing the modulation in aquaporin, caveolae, intercellular junction and basement membrane of the endothelial cells, the present study intends to disclose the effect and underling mechanism of YQFM, Rb1 and Sch B on albumin extravasation from venules and multiorgan injury, to explore if Rb1+Sch B has a synergy and its mechanism, with hoping to reveal the scientific basis for the potential of ShengMaiSan to invigorate qi for relieving desertion.

微血管通透性异常是内毒素引起的休克脱证的关键环节。基于固摄治法组成的生脉散救治休克脱证有良好的疗效。生脉散制剂 (YQFM) 可改善脂多糖 (LPS) 引起的大鼠肠系膜细静脉血浆白蛋白的漏出,其补气成分人参皂苷Rb1 (Rb1) 可以抑制LPS诱导的血管内皮细胞质膜微囊、连接蛋白ZO-1、VE-Cadherin的表达和磷酸化。但是，YQFM、Rb1和收涩成分五味子乙素 (Sch B) 能否固摄LPS诱导的脑、肺细静脉血浆白蛋白漏出，改善多脏器损伤；Rb1和Sch B联合使用能否增效，及其机理尚不清晰。为此，本课题观察YQFM、Rb1、SchB、Rb1+Sch B对LPS引起的大鼠脑、肺、肠系膜细静脉血浆白漏出和器官损伤的改善作用，通过水通道、质膜微囊、细胞间连接、血管基底膜及其相关调控蛋白，探讨YQFM、Rb1、Sch B固摄作用及其机理，揭示生脉散固摄作用科学内涵。

本研究获得了以下结果：.1. YQFM前、后给药均可以抑制LPS连续滴注90 min引发的大鼠脑、肺、肠系膜细静脉血浆白蛋白的漏出，抑制了脑、肺、肠组织炎症反应、微血管周围水肿和多脏器损伤。YQFM抑制了血管内皮细胞粘附连接蛋白和紧密连接蛋白的低表达，抑制了内皮细胞F-actin的解聚、ATP含量的降低、ATP的分解以及ATP合成酶亚基的低表达。YQFM还可以抑制组织和细胞中Src激酶和NF-κB的活化，进而抑制脂膜微囊蛋白Cav-1的表达和磷酸化。YQFM还可以抑制LPS诱导的微血管基底膜胶原蛋白Collagen IV的降解。.2. YQFM中的有效成分Rb1对LPS诱导的细静脉血浆白蛋白的漏出和多脏器损伤也有部分的抑制作用。Rb1抑制了LPS引发内皮细胞连接蛋白的低表达和F-actin的解聚、ATP含量的降低、ATP的分解以及ATP合成酶亚基的低表达。部分地阐明了Rb1抑制LPS引发的微血管白蛋白渗出的作用与其调节能量代谢，增加内皮细胞骨架蛋白稳定性，从而增强细胞间连接的作用相关。Rb1也可以通过抑制Src激酶的活化抑制Cav-1的磷酸化水平。.3. YQFM中的有效成分Sch对LPS引发的细静脉血浆白蛋白的漏出抑制作用不明显，但是，Sch可以部分地抑制LPS诱导的组织水肿和多脏器损伤。Sch可以显著地抑制NF-κB的活化，抑制炎性因子的表达，抑制组织中MMP的活化，从而抑制血管基底膜胶原蛋白Collagen IV的降解。Sch还可以抑制组织和血管内皮MLC的磷酸化。Sch抑制LPS引发的微血管渗透性的增加与其抑制炎症反应，进而抑制MMP的活化和胶原降解以及抑制MLC磷酸化从而抑制应力纤维丝的形成相关。.4. Rb1+Sch联合给药组在抑制LPS引发的细静脉血浆白蛋白的漏出以及组织水肿和多脏器损伤中的作用明显优于单纯Rb1和Sch给药组。表明以改善能量为主的补气固摄和以抗炎为主的收涩固摄联合使用在抑制和改善LPS引发的微血管渗透性增加和组织损伤中的协同作用。.以上结果为生脉散制剂注射用益气复脉冻干在救治以微血管渗透性增加为主要病理基础的休克脱证方面的应用提供了科学依据，阐明了其补气固摄和收涩固摄的物质基础和作用机理，揭示了补气固摄和收涩固摄理论的科学内涵。