基于AMPK信号通路调控TRPV1的电针治疗干眼症的机制研究

Dry eye disease is the most common ocular surface disease at present. Immune-related inflammation are considered as the main pathogenesis of dry eye. TRPV1 is involved in the inflammatory process of dry eye by mediating the communication between sensory nerve endings and immune cells.The AMPK signaling pathway regulates TRPV1 and inflammatory responses.The previous work of our research group confirmed the exact efficacy of electroacupuncture in the treatment of dry eye.On this basis, an experimental dry eye model was prepared to observe the effects of electroacupuncture intervention on the amount of tear secretion, tear film rupture time, cornea fluorescein staining and the regulation of corneal nerve dysfunction in rats with dry eye. Using Western blot, fluorescence quantitative PCR, and the laser confocal and other technologies, to discuss the relationship of onset of dry eyes and electricity for its intervention effect through observing the expression of TRPV1, NGF, SP, CGRP, and Drp1, CaMKKβ, AMPK and p-AMPK.This study is going to prove that electroacupuncture can reduce TNF-α, excessive expression of MMP9 and reduce dry eyes ocular surface inflammation by regulating the AMPK pathway and TRPV1 expression in the cornea and conjunctiva and promote restoration of the ocular surface and it appears.To provide scientific basis for electroacupuncture treatment of dry eye.

干眼症是目前最常见的眼表疾病，发病率逐年升高。免疫相关的炎症被认为是干眼症最主要的发病机制。TRPV1通过介导感觉神经末梢、免疫细胞之间的沟通而参与了干眼症的炎症过程。AMPK信号通路对TRPV1及炎症因子有调控作用。本课题组前期工作证实电针治疗干眼症疗效确切，在此基础上拟制备实验性干眼症模型，观察电针干预对干眼症大鼠泪液分泌量、泪膜破裂时间、角膜荧光素染色及对角膜神经功能异常的调节作用。采用Western blot、荧光定量PCR、激光共聚焦等技术，通过探讨角、结膜中TRPV1、NGF、SP、CGRP、Drp1、CaMKKβ、AMPK、p-AMPK等的表达与干眼症发病的关系及电针对其的干预作用。拟证实电针通过调节干眼症角结膜中AMPK信号通路及TRPV1表达，减少TNF-α、MMP9过度表达而减轻干眼症眼表炎症、促进眼表和泪膜修复的目的。为电针治疗干眼症提供科学依据。

干眼（Dry eye）是目前最常见的眼表疾病，发病率逐年升高且趋于低龄化。干眼的治疗策略已经从单纯的湿润眼表转向改变潜在发病机制的过程。近年来越来越多的研究证实针刺治疗干眼临床疗效肯定并无副作用。电针疗法可以通过延长泪膜破裂时间，促进泪液分泌改善干眼症状。干眼症状中的干燥、痒、疼痛的分子机制主要与眼表炎症、神经肽及相关的离子通道密切相关，TRPV1通过介导感觉神经末梢、免疫细胞之间的沟通而参与了干眼症的炎症过程。因此，本项目以TRPV1及其下游信号通路为切入点探讨电针治疗干眼的作用机制。结果发现：①电针干预可以提高干眼大鼠泪液分泌量、泪膜破裂时间，降低角膜荧光素染色评分和拭眼次数。干眼大鼠角膜中央神经受到损伤，神经密度降低；经电针干预后均可促进干眼大鼠角膜中央神经损伤的修复。提示电针可能通过抑制TRPV1离子通道的异常激活修复干眼大鼠眼表功能和角膜中央神经损伤。②干眼大鼠角结膜和三叉神经节组织中TRPV1、SP、CGRP、IL-1β、TNF-α蛋白表达升高，经电针干预后可下调TRPV1、SP、CGRP、IL-1β、TNF-α蛋白的表达；提示电针可能通过抑制TRPV1离子通道的异常激活，下调神经肽SP、CGRP的表达，缓解DE大鼠神经源性炎症。③电针可以抑制干眼大鼠角膜组织中TRPV1、CaMKKβ、AMPK、IL-1β、TNF-α蛋白的表达，提示电针干预可能通过调控TRPV1/AMPK信号通路发挥缓解眼表神经源性炎症的作用。结果从TRPV1及其下游信号通路调控角度揭示了电针治疗干眼症的信号转导机制和关键信息分子，为电针治疗干眼症的疗效机制研究提供实验依据与理论基础，有助于推动电针疗法在干眼症等眼科疾病中的进一步应用。