FcrIIb基因变异在肾虚类风湿关节炎病证中的作用及从肾论治中药的干预机制研究

Deficiency of kidney is a key of cause and development of Rheumatoid Arthritis (RA) in Traditional Chinese Medicine(TCM) which modern biological mechanism is unclear. B lymphocytes is born and develops in bone marrow. We supposed B lymphocytes has correlation with kidney and RA kidney deficiency would result in defective B lymphocytes according to theory of Traditional Chinese Medicine(TCM) Visceral Manifestation. In the result of our earlier research, the disease activity and risk of autoantibody in RA patients with deficiency of kidney are more than RA with none-deficiency of kidney, which was supposed to link to FcγIIb down-regulation. But its cause and mechanism kept unknown. In this project, to discover the genetic link between defective FcγIIb variants and RA kidney deficiency cause and mechanism of RA kidney deficiency,exon polymorphisms of FcγIIb,disease activity,immune index and joints destruction would be analyzed while CD19、FcγIIb、Lyn and SHP1 of B lymphocytes would be detected by methods like sequencing of exon, flow cytometry and IP-Western. Mechanism of RA kidney deficiency and effect of Chinese medicine would be studied on CD19 and FcγIIb cell signaling pathway by transfecting defective FcγIIb to cell lines. Collagen induced arthritis rats with deficiency of kidney would be made. That would prove the therapy mechanism of Chinese medicine on induction immune tolerance to inhibit joints inflammation and destruction This project would clarify the genetics and immunological biological mechanism of RA deficiency of kidney and new meaning of Visceral Manifestation. The result would help provide scientific information to special Chinese medicine strategy of RA with strengthening kidney treatment.kidney treatment.

肾虚是类风湿关节炎（RA）发生发展的重要因素，其生物学机制尚不明了。B细胞在骨髓发育，按藏象学说“肾主先天，主骨生髓”理论，我们推测肾虚RA可能与B细胞异常发育相关。前期研究显示，肾虚RA病情活动度和产生自身抗体的风险升高，与抑制性受体FcγIIb下调关系密切，但原因和机制仍不清楚。本项目①拟采用巢式PCR、外显子测序和IP-Western等方法，研究FcγIIb基因变异与肾虚RA的关联，观察B细胞CD19、FcγIIb、Lyn和SHP1的变化，探索肾虚RA病证的分子机制；②转染变异FcγIIb基因，观察B细胞CD19激活/FcγIIb抑制通路，研究肾虚RA免疫失耐受机制及从肾论治中药的作用环节；③构建肾虚CIA模型，探索从肾论治中药诱导免疫耐受、抗炎抗骨破坏的机制。本项目对于阐明肾虚RA病证的遗传免疫学物质基础，揭示藏象理论新内涵具有重要意义，将为从肾论治RA的中医特色治疗提供科学依据。

肾虚是类风湿关节炎(RA)发生发展的重要因素，其生物学机制尚不明了。B细胞在骨髓发育，按藏象学说“肾主先天，主骨生髓”理论，我们推测肾虚RA可能与B细胞异常发育相关。前期研究显示，肾虚RA病情活动度和产生自身抗体的风险升高，与抑制性受体FcγRIIB下调关系密切，但原因和机制仍不清楚。本项目分为三个部分来研究肾虚RA与B细胞异常发育的相关性。临床研究证实肾虚RA存在FcγRIIB水平的下调，其可能导致B细胞过度活化和自身抗体水平升高，提示FcγRIIB可能是肾虚类风湿关节炎的重要机制； 细胞实验显示，益肾通痹汤的氯防部位可抑制TNF-α、IL-6 的基因及蛋白表达。通过转染变异FcγRIIB基因、WB等方法证实从肾论治中药益肾通痹汤可通过调节CD19/FcγRIIB、Lyn、SHP1通路来抑制B细胞的异常活化与增殖，促进B细胞的凋亡，从而诱导免疫耐受。动物实验证实益肾通痹汤具有抑制胶原诱导型关节炎症，滑膜中性粒细胞、淋巴细胞、浆细胞浸润，滑膜增生和软骨破坏，以及关节破坏的作用。本项目对于阐明肾虚RA病证的遗传免疫学物质基础，揭示藏象理论新内涵具有重要意义，为从肾论治RA的中医特色治疗提供科学依据。