人参皂苷CK选择性激动糖皮质激素受体抑制胶原性关节炎T细胞功能及其机制

Ginsenoside metabolite compound K (CK, 20-O-D-glucopyranosyl-20(S)-protopanaxadiol, C36H62O8) belongs to dammarane-type triterpene saponins according to its structure. It is a degradation product of ginsenoside in the intestine by bacteria and is the major form of ginsenoside absorbed into the body and can be transformed from ginsenoside by food microorganisms in vitro. The anti-inflammatory activity of CK has been identified in several studies. Our study demonstrated that CK exerted anti-inflammatory effect on collagen-induced arthritis (CIA) and adjuvant-induced arthritis (AA) animal models, and this effect was due to inhibiting the abnormal activation and differentiation of T cells. However, the mechanism of CK on suppressing T cell activation remains unclear..Classic glucocorticoid receptor (GR) targets gene promoter activation via interaction of homodimeric GR with glucocorticoid response elements (GRE) resulting in physiological effects and adverse effects. Glucocorticoids (GCs) also can block inflammation via interference of the liganded GR with the activity of pro-inflammatory transcription factors NF-κB, a mechanism known as transrepression Our study demonstrated that the effect of CK on inhibiting the abnormal T cell activation can be blocked by GR antagonist, and CK did not affect body weight and blood glucose level in vivo. All these results suggest that the effect of CK on suppressing T cell activation may be related to GR activation, and this activation of GR may not induce GRE transcriptional activation. This study was designed to identify whether CK exerted inhibitory effect on abnormal T cell function via inhibiting NF-κB, and at the same time did not induce GRE transcription activation resulting less adverse effects. To that end, T cells were isolated from collagen induced arthritis rats, the effect of CK on CIA T cell function was assayed, and role of GR was investigated in vitro. Then the potential adverse reaction of CK was investigated in vivo, at the same time, the effect of CK on GR signals was assayed. This study provides an interesting novel insight into the potential mechanism by which CK contributes to the anti-inflammatory effect in autoimmune conditions.

人参皂苷CK（CK）是具有我国自主知识产权的新型单体，课题组研究表明CK有明显抗炎和免疫调节活性，抑制T细胞过度活化是其主要特点，但机制不清。糖皮质激素受体（GR）活化后通过抑制NF-κB发挥抗炎作用，同时诱导GR反应元件（GRE）转录激活而产生不良反应。课题组发现CK对T细胞活化的抑制作用可被GR拮抗剂阻断，且CK不影响实验动物的血糖和体重，提示CK对T细胞活化的抑制作用可能与其激活GR有关，且可能不诱导GRE的转录激活。那么CK可否作为GR的选择性调节剂，通过下调NF-κB抑制T细胞活化，而不诱导GRE的转录激活，较少引起糖皮质激素（GC）样不良反应？未见报道。本项目拟建立胶原性关节炎模型，体外实验探讨GR在CK抑制T细胞功能中的作用及机制，体内实验研究CK是否有GC样不良反应以及与GR信号的关系，为揭示CK的抗炎免疫调节分子机制提供依据，为把CK开发为自主知识产权的原创药物提供基础。

人参皂苷CK（CK）是具有我国自主知识产权的新型单体，课题组前期研究表明CK有明显抗炎和免疫调节作用，抑制T细胞过度活化是其主要特点，但机制不清。本项目建立胶原性关节炎（CIA）模型，发现CK对CIA小鼠体内过度的免疫应答具有下调作用。体外研究发现，CK可以抑制CIA大鼠滑膜细胞对T细胞的活化，且此作用与GR活化有关。CK可以与GR结合，诱导GRα由胞浆到胞核的转移，和GRE的转录；调控GR的下游信号，减少p65、IκBα的磷酸化，抑制NF-κB的活化；CK还可以减少丝裂原活化蛋白激酶磷酸酶(mitogen-activated protein kinase phosphatases，MKP-1）的蛋白表达，负调节MAPK信号。抗炎因子亮氨酸拉链蛋白(gluco-corticoid-induced leucine zipper, GILZ)除了具有抗炎作用之外，还可以增强MSC成骨细胞分化，增加骨形成的作用。CK活化GR后可以诱导GILZ蛋白表达，提示CK对RA的关节骨丢失具有保护作用，不会造成糖皮质激素的骨质疏松的副作用。此外CK不会引起糖代谢紊乱，可能与其减少糖异生的关键酶磷酸烯醇式丙酮酸羧激酶(PEPCK)的蛋白表达有关。这些结果表明CK具有抗炎免疫调节作用，其机制与结合GR，抑制NF-κB和MAPK活化有关；CK不会引起糖皮质激素相关的代谢紊乱，可能与其减少PEPCK，增加GILZ有关。本研究还发现CK通过减少局部PGE2和COX2的蛋白表达发挥镇痛作用。CK联合MTX给药在起效时间、改善关节炎症状、降低血清炎性细胞因子和抗体水平，以及抑制滑膜细胞和巨噬细胞分泌方面优于单用MTX；CK联合CEL在改善脾脏和关节病理以及抑制滑膜细胞上优于单用CEL。抑制免疫细胞（T、B以及DC）活化，恢复细胞免疫和体液免疫平衡是CK发挥抗炎作用的特点。本研究提示CK通过活化GR发挥抗炎免疫调节和镇痛作用，但不会引起类糖皮质激素样副作用，在治疗自身免疫病上具有良好的应用前景。本研究为CK作为治疗自身免疫病药物的开发应用提供了有力的实验依据。