柴胡皂苷A通过调控细胞膜胆固醇转运抑制鸡白痢沙门氏菌侵袭鸡肠道上皮细胞的分子机制

Cellular membrane cholesterol is implicated in the process of actin cytoskeleton rearrangemen caused by Salmonella, and plays an important role in the invasion of Salmonella to the intestinal epithelial cells. In our preliminary study, we found saikosaponin A protected chicken against Salmonella pullorum infection in vivo. In vitro, saikosaponin A decreased the levels of cellular membrane cholesterol in chicken intestinal epithelial cells and inhibited the invasion of Salmonella pullorum to the chicken intestinal epithelial cells. Furthermore, the anti-invasion effect of saikosaponin A was blocked by the addition of water soluble cholesterol. Activation of live X receptor α (LXRα) decreases the levels of cellular membrane cholesterol via promoting cholesterol efflux, but its effect on the invasion of Salmonella pullorum to chicken intestinal epithelial cells remains unclearly.The project will investigate the effects of saikosaponin A, depletion of membrane cholesterol, and activation of LXRα on the invasion of Salmonella pullorum to chicken intestinal epithelial cells, and the effects of saikosaponin A on cellular membrane cholesterol, LXRα signaling pathways, and actin cytoskeleton in chicken intestinal epithelial cells infected with Salmonella pullorum, so as to revelation the key steps of the invasion of Salmonella pullorum modulated by saikosaponin A. These results not only will deepen the understanding of the mechanism of the traditional Chinese medicine on the prevention and treating of pullorum disease, but also will further illuminate the pathogenic mechanism of Salmonella pullorum and provide new targets for the research and development of new antibacterial drugs.

细胞膜胆固醇参与沙门氏菌诱导的肌动蛋白细胞骨架重排，在沙门氏菌侵袭肠道上皮细胞过程中起着重要作用。申请人前期研究发现，柴胡皂苷A具有抗鸡白痢的作用，可降低鸡肠道上皮细胞膜胆固醇含量，添加水溶性胆固醇可阻断柴胡皂苷A对鸡白痢沙门氏菌侵袭鸡肠道上皮细胞的抑制作用。激活LXRα可通过促进胆固醇外流降低细胞膜胆固醇含量，但其对鸡白痢沙门氏菌侵袭鸡肠道上皮细胞的作用尚不清楚。本研究拟建立鸡白痢沙门氏菌侵袭鸡肠道上皮细胞模型，研究消耗细胞膜胆固醇和激活LXRα对鸡白痢沙门氏菌侵袭鸡肠道上皮细胞的作用，明确柴胡皂苷A对鸡白痢沙门氏菌侵袭鸡肠道上皮细胞、细胞膜胆固醇、LXRα信号通路和肌动蛋白细胞骨架的调控作用，揭示柴胡皂苷A抑制鸡白痢沙门氏菌侵袭鸡肠道上皮细胞的关键环节。所获研究结果不仅可以深化对中药防治鸡白痢作用机理的认识，而且可以进一步解析鸡白痢沙门氏菌致病机制，为新型抗菌药的研发提供新靶点。

细胞膜胆固醇参与沙门氏菌诱导的肌动蛋白细胞骨架重排，在沙门氏菌侵袭肠道上皮细胞过程中起着重要作用。激活LXRα可通过促进胆固醇外流降低细胞胆固醇含量，是近年来发现的具有调节胆固醇转运功能的重要靶点，同时也是防治多种疾病的潜力靶点。项目研究发现沙门氏菌可侵袭肠道上皮细胞，诱导细胞骨架重排，进一步研究发现消耗细胞膜胆固醇和激活LXRα抑制鸡白痢沙门氏菌对鸡肠道上皮细胞的侵袭，提示调节胆固醇代谢和激活LXRα可能是防治沙门氏菌病的有效策略。我们对中药多种活性成分进行了深入研究，发现黄芩苷、桑根酮G等中药成分可通过抗毒力、抑菌杀菌、调节宿主等多环节起到抗感染性疾病的作用。我们通过动物实验发现柴胡皂苷A和LXRα激动剂对鸡沙门氏菌病具有保护作用。体外研究发现柴胡皂苷A抑制鸡白痢沙门氏菌对鸡肠道上皮细胞的侵袭，降低细胞胆固醇含量和激活LXRα信号通路，而对细胞骨架调节因子CDC42 转录未见显著作用。最后研究发现添加胆固醇和LXRα抑制剂降低了柴胡皂苷A对沙门氏菌侵袭肠道上皮的抑制作用。这些研究结果表明柴胡皂苷A通过激活LXRα信号通路降低胆固醇含量抑制沙门氏菌对肠道上皮细胞的侵袭，深化了对沙门氏菌利用宿主胆固醇侵袭宿主的致病机制的认识，提出了可采用中药调节宿主胆固醇代谢或激活LXRα信号通路等策略防治感染性疾病。