推拿重建脑瘫炎症稳态的DNA甲基化调控机制

Cerebral palsy (CP), which its long-term serious harm to the person still has not been successfully solved, is a brain injury syndrome with difficulty of prevention and treatment and high morbidity. The mechanisms of CP have not been fully elucidated and there has not been effective solution measures. Therefore, clarifying the mechanisms and seeking effective prevention strategy of CP have become a top priority. From the human body and animal experiments studies, we previously found that tuina has a better clinical curative effect to treat CP, and can reverse the unbalance of mRNA and protein expression of pro- and anti-inflammatory cytokines to rebuild the steady state of the inflammation of CP. There may be a DNA methylation regulation mechanism of gene expression. Therefore, this project hypothesizes: tuina may rebuild a new inflammatory homeostasis of CP via the control pathway of DNA methylation.This project will establish pup rats models of CP by ischemia and hypoxia treatment and tuina intervenes the models. The inflammatory pathological variation characteristics of the cerebral white matter and cortex and hippocampus tissues in the pup rats will be observed and the effect of tuina intervention will be investigated.We will research the relationship between the mRNA and protein expression and methylation of TNF-α, IL-6 and IL-10 in the issues and the blood mononuclear cells of the pup rats, and study the influence of DNMTs expression on the inflammatory cytokine gene expression in order to clarify the mechanism of effect of tuina intervention to CP via regulating DNA methylation. The project will clarify a novel mechanism of reconstruction of CP inflammatory homeostasis by tuina intervention, which will provide the basis for a new strategy to prevent and treat CP.

脑瘫（CP）对人的长期严重危害依然未得到很好的解决，是防治极为困难和高致残率的脑损伤综合征，其发生机制尚未完全阐明，因此阐明CP发生机制和寻求防治策略已成当务之急。我们前期通过人体和动物实验研究发现：推拿对CP有较好的临床疗效，且能逆转CP中促-抗炎症因子mRNA和蛋白表达平衡失调，重建CP炎症稳态，可能存在DNA甲基化表观遗传调控基因表达的机制。据此本项目提出：推拿可能通过DNA甲基化调控途径重建CP炎症稳态。本项目拟建缺血缺氧性CP幼鼠模型并进行推拿干预，探明CP幼鼠脑白质、皮层和海马组织炎症病理变化特征及推拿干预效果；检测组织与血单个核细胞中TNF-α等炎症因子mRNA与蛋白表达、甲基化和甲基转移酶DNMTs表达的水平，解析甲基化与基因表达的关系，明确推拿通过调节甲基化的变化，进而调控炎症因子基因的表达，阐明推拿重建CP炎症稳态的甲基化调控新机制，为确立防治CP的新策略提供研究基础。

本项目通过建立脑瘫（cerebral palsy，CP）幼鼠模型并进行推拿干预，观察CP幼鼠脑白质、皮层和海马组织炎症病理变化特征及推拿干预效果。研究发现，推拿可以促进脑瘫幼鼠生长发育，提高脑瘫幼鼠学习记忆和运动平衡能力；推拿可以降低脑白质、皮层、海马中促炎细胞因子 IL-6蛋白表达，降低脑瘫幼鼠皮层以及海马促炎细胞因子 TNF-α蛋白表达，提高脑白质、皮层、海马抗炎细胞因子 IL-10 蛋白表达，重建脑瘫炎症相对稳态；推拿可以调控皮层、海马内炎症细胞因子 TNF-α和 IL-10 的基因启动子区域甲基化水平。本研究用动物行为学方法验证了推拿对CP幼鼠良好的康复治疗作用，用分子生物学方法观测到CP幼鼠康复过程中促-抗炎症因子蛋白表达及其DNA甲基化水平的变化，阐明了推拿可以通过调节炎症因子DNA甲基化水平，进而调控炎症因子的表达，重建CP炎症稳态，促进CP肢体功能的康复和脑损伤修复的机制，为临床进一步推广应用中医推拿提供了理论依据。已发表论文21篇，均标注有“国家自然科学基金资助（编号：81460748）”。