“治乱安危”--以Cyr61为核心靶标探讨三草软膏在银屑病“瘥后防复”中的作用机制

Psoriasis is a chronic inflammatory disease that requires long-term standard treatment. The weak glucocorticoids are commonly used topical formulations to treat mild to moderate psoriasis, however the mechanism of relapse after discontinuation is still unknown. The latest study shows that upstream proinflammatory cytokine Cyr61 can be used as a new intervention target, which might be through the regulation ofAkt/NF-κB signaling pathway . The expression of Cyr61 was high in patients with psoriasis. In addition, the animal model of psoriasis treated with external application of Three Grass ointment showed that the inflammation was relieved and the expression of Cyr61 mRNA was decreased, but the glucocorticoid had no effect on Cyr61. Based on the above observations, we suggested a hypothesis that “glucocorticoids inhibit the downstream inflammatory responsebut has no effect on expression of Cyr61 , which may be one of the important factors for the recurrence of psoriasis after glucocorticoid withdrawal. Prophylactic use of Three Grass ointment can reduce proinflammatory cytokine Cyr61, improving the Akt/NF-κB nuclear transcription activation, preventing the recurrence of inflammation". The project intends to use CRISPR/Cas9 gene knockout plasmid, plasmid transfection, neutralizing monoclonal antibody, immunofluorescence technique to explain why prophylactic use of Three Grass ointment inhibits psoriasis inflammation. This study will enrich the scientific connotation of Chinese medicine external treatment, playing a positive role on holistic concept "manage chaos and appease danger" of traditional Chinese medicine.

银屑病是一种慢性炎症性疾病，需要长期规范治疗。弱效糖皮质激素是轻中度银屑病的常用外用制剂，但停药后反弹复发的机制仍未明了。最新研究提示细胞核内转录调控信号通路Akt/NF-κB的上游前炎症因子Cyr61可作为干预新靶点。课题组外用三草软膏预处理的银屑病动物模型显示炎症减轻，并伴有Cyr61mRNA表达降低，而糖皮质激素对Cyr61无影响。基于以上差异，课题组提出“糖皮质激素抑制下游炎症反应，但皮损中Cyr61仍持续高表达，这可能是糖皮质激素停药后银屑病复发的重要因素之一。预防性使用三草软膏，可降低前炎症因子Cyr61，改善Akt/NF-κB核转录激活，预防炎症复发”的科学假说。本项目拟运用CRISPR/Cas9基因敲除、质粒转染、单克隆抗体中和、活性荧光标记技术，阐释预防性外用三草软膏对银屑病炎症的抑制。本研究对丰富中医外治法的科学内涵，体现祖国医学“治乱安危”的整体思想具有积极意义。

银屑病是一种慢性炎症性疾病，需要长期规范治疗。轻中度银屑病现有治疗方案首选弱效糖皮质激素外用制剂，但停药后疾病复发的机制仍未明了。本项目研究的三草软膏，由紫草、茜草、生甘草与大豆油熬制而成，是沪上夏氏外科传人孙世道教授数十年的临床经验处方，精选的三味药有清热凉血、化瘀消斑之功，临床常用于银屑病患者的皮损治疗、预防及日常护理。本研究进行了临床试验，结果提示三组在治疗期银屑病皮损均有一定好转，三草软膏与玉泽身体乳疗效相当，使用外用药维持治疗比不使用药物复发可能性更小，使用三草软膏比玉泽身体乳在维持治疗中复发的可能性更小。免疫组化数据表明银屑病皮损组织样本的Cyr61阳性表达率高于正常皮肤样本，同时，银屑病皮损中Cyr61阳性细胞数目较正常皮肤样本显著增加。动物实验数据表明，三草软膏提取物干预处理可显著改善造模小鼠背部皮肤的炎症表型、PASI评分和病理表现。与正常对照组相比，咪喹莫特造模组Cyr61 mRNA表达呈现显著上调，而三草软膏提取物干预组能够下调银屑病小鼠模型皮肤中Cyr61 mRNA表达水平。体外细胞实验结果表明，与对照组相比，三草软膏提取物可抑制角质形成细胞的增殖；在IFN-γ诱导的炎症细胞模型中，Cyr61、ICAM-1 mRNA和蛋白表达上调，而三草软膏提取物可下调Cyr61与ICAM-1的表达。中草药制剂与目前市面常用的皮肤屏障修复剂作用具有一定的等效性，在缓解期维持治疗阶段皮损复发情况可能更少，初步证实三草软膏在银屑病患者“瘥后防复”中具有一定的运用价值。促炎因子Cyr61在银屑病皮损中过表达，阳性率显著升高，是银屑病炎症进程的重要环节。三草软膏提取物干预能显著降低咪喹莫特诱导的银屑病样模型小鼠皮损炎症反应，其作用机制与下调Cyr61、ICAM-1的mRNA和蛋白表达，抑制角质形成细胞增殖有关，这是三草软膏在临床上能有效干预银屑病皮损病理进程的重要原因。本研究对丰富中医外治法的科学内涵，体现祖国医学“治乱安危”的整体思想具有积极意义。