蜂胶影响细菌内毒素诱导的血管内皮细胞自噬的分子机制研究

Propolis has been reported to possess good anti-inflammatory effect, however, the mechanisms are not fully elaborated. Vascular endothelial cell (VEC) damage or dysfunction is important in many diseases, including inflammation. We previously found that low concentration of propolis could protect VEC induced by lipopolysaccharide (LPS) through decreasing the expression of Toll like receptor 4 (TLR4) and the activity of phosphatidylcholine-specific phospholipase C (PC-PLC), but the protect effect wasn't through inhibiting VEC apoptosis, then we also found that propolis could decrease the level of LC3-Ⅱ, an autophagy marker. We speculated propolis protecting VEC induced by LPS was through autophagy, which could be a new mechanism of propolis on anti-inflammatory effect. To test this hypothesis, we are planning to induce VEC autophagy induced by LPS to copy inflammatory model, and the key factors of VEC autophagy - integrin β4, ANXA7, BIRC2, TLR4 and reactive oxygen species (ROS) are as the breakthrough point to disclose the effect of propolis on autophagy of VEC and clarify the mechanism of propolis on anti-inflammatory effect, which would supply new ideas and new targets for propolis on anti-inflammatory effect.

蜂胶具有较好的抗炎功效，但作用机制尚未彻底阐明。血管内皮细胞（VEC）功能损伤或紊乱会导致包括炎症在内的多种疾病的发生。本课题组前期研究中发现低浓度的蜂胶降低Toll样受体4（TLR4）的表达和磷脂酰胆碱特异性磷脂酶C（PC-PLC）的活性，抑制细菌内毒素（LPS）诱导的VEC损伤，但在此过程中，蜂胶并不是通过抑制VEC凋亡的方式起到保护作用，随后我们发现蜂胶降低LPS诱导的VEC中自噬标志LC3-Ⅱ的水平。我们推测蜂胶可能是通过抑制细胞自噬的方式保护LPS对VEC的损伤，这可能是蜂胶抗炎的一种新的作用机制。为了证实这一假说，本课题拟通过体外LPS诱导VEC自噬性损伤复制炎症模型，以参与VEC自噬的关键因子-膜整连蛋白β4、ANXA7、BIRC2、TLR4、ROS为切入点，从细胞和分子水平揭示蜂胶对VEC自噬的影响，阐明蜂胶的抗炎作用机制，并为蜂胶抗炎作用研究提供新思路和新靶点。

蜂胶具有较好的抗炎功效，但其作用机制尚未彻底阐明。血管内皮细胞（VEC）功能损伤或紊乱会导致包括炎症在内的多种疾病的发生。本项目通过血管内皮细胞（VEC）、巨噬细胞、炎症肿瘤细胞等全方面对蜂胶的抗炎功效进行了深入研究。通过检测在炎症条件下，蜂胶对细胞自噬、炎症信号通路以及对炎症因子分泌全面阐述了蜂胶的抗炎机理。结果，在脂多糖（LPS）诱导的血管内皮细胞中，蜂胶主要通过上调LC3-Ⅱ，抑制P62的表达促进细胞自噬，同时抑制炎症信号通路中的TLR4以及核因子NF-κB p65的活性抑制炎症信号通路；在LPS/IFN-γ共刺激的RAW264.7巨噬细胞炎症模型中，蜂胶显著抑制炎症因子的mRNA转录，抑制炎症因子的产生，阻断NF-κB的活性；在体内，蜂胶通过抑制炎症因子的如IL-6，IL-10，MCP-1，TNF-α，IL-12P70的产生降低肺损伤；在LPS刺激的MDA-MB-231细胞中，蜂胶及其主要组分显著抑制TLR4，MyD88，TRIF, IRAK4 以及核因子NF-κB p654信号通路，促进细胞自噬抑制肿瘤炎症微环境，抑制肿瘤细胞增殖。本项目的研究全面评价了蜂胶的抗炎功效，为蜂胶的抗炎生物学活性的应用奠定了理论基础。