基于CD4+CD25+调节性T细胞在单纯疱疹病毒性角膜炎诱导复发中的作用研究益气解毒方的作用机制

Recurrence is the difficult problem in the treatment of herpes simplex keratitis, the mechanism of which is still not clear. And there is no effective and safe medicine for anti-recurrence. According to our research and some other studies, the Chinese medicine of replenishing qi and detoxicating is demonstrated to be effective on the cure and prophylactic of HSK. Meanwhile, we found it was more effective on the treatment of epithelia type than that of stromal and endothelia type, which may indicate the relationship between its effect with virus replication. In view of the important function of regulatory T cell in the aspect of immunosupression, we propose that Yiqi Jiedu formula works in the treatment of recurrent HSK mainly through the way of regulating CD4+CD25+regulatory T cell mediated by STAT5 signal pathway. To verify our hypothesis, we would use the HSK recurrence model of mice to study the relationship between the adjustment of regulatory T cell as well as its downstream effects implemented by Yiqi Jiedu formula with supression of virus reactivation and recurrence of HSK; and we would also adopt cultivated CD4+CD25+Treg cells with medicated serum in vitro, the technological means of siRNA to demonstrate that CD4+CD25+regulatory T cell mediated by STAT5 pathway is the main mechanism in this therapy. Our study is intended to reveal the mechanism of Yiqi Jiedu formula from the aspect of anti-virus replicaiton regulated by immunity of the body , so as to promote clinical application.

单纯疱疹病毒性角膜炎（HSK）的复发是治疗的一大难点，且机制不明，缺乏有效的药物。本课题组及其他学者研究发现，益气解毒中药对治疗和预防HSK复发有效。同时我们在前期研究中发现益气解毒方治疗复发性HSK上皮型优于基质和内皮型，可能和病毒抑制有关。基于调节性T细胞和复发诱因的关系，我们推测：STAT5信号通路介导的CD4+CD25+调节性T细胞对诱导病毒活化起重要作用，而益气解毒方对其的调控是治疗复发性HSK的关键机制。本项目拟通过HSK复发小鼠模型，观察益气解毒方干预CD4+CD25+T细胞增殖与病毒复制、疾病复发的关系；并通过药物血清体外培养，关键转录因子Foxp3基因沉默，证实益气解毒方通过调控STAT5介导的CD4+CD25+T细胞以调整免疫状态，发挥抗病毒作用。旨在以免疫调节对病毒复制的控制角度，明确益气解毒方的作用机制，促进临床应用。

单纯疱疹病毒(HSV-1)感染所致的单纯疱疹病毒性角膜炎（HSK）为常见的眼部致盲性疾病，HSV-1病毒的潜伏和再活化以及免疫系统对病毒的失抑制是复发的关键环节。在前期临床研究中采用玉屏风散加味的益气解毒方治疗复发性 HSK有一定疗效，证实益气解毒方可以缓解患者的症状，改善体征，具有抗复发的作用。本项目从抑制病毒再活化及维持机体免疫平衡角度探讨益气解毒方的作用机制，从中药调控STAT5信号通路及CD4+CD25+调节性T细胞的活化角度展开研究。主要进行如下工作：1、以角膜病毒接种的方式成功建立复发性HSK BALB/C小鼠模型。2、益气解毒方对CD4+CD25+Treg细胞的调控作用。3、STAT5信号通路是否是益气解毒方对Treg的调控作用途径。研究结果发现：1、复发小鼠的CD4+CD25+调节性T细胞含量下降，效应性T细胞CD4/CD8比值失衡，具有抗病毒效力的关键细胞因子IL-2下降，免疫状态异常。2、益气解毒方可上调Treg的活性，上调IL-2，调整CD4/CD8失衡。3、益气解毒方上调STAT5/P-STAT5活性，但这种变化维持时间较短，STAT5可能是中药调控Treg的信号途径。通过上述研究，我们明确了Treg在复发性HSK中的作用及益气解毒方对Treg的调控是中药作用的重要机制，为中药在病毒性角膜炎抗复发上的作用提供科学依据，也为药物研究提供新的靶点。