胞外亲环素A对炎症发生的影响及其抗体在抑制炎症反应中的作用

Effect of extracellular cyclophilin A on inflammatory response and anti-inflammatory activity of antibody against cyclophilin A

摘要:

亲环素A（Cyclophilin A，CypA）是肽基脯氨酰顺/反异构酶家族成员，主要分布于细胞质中，当细胞受到外界刺激时会分泌到细胞间隙，与CD147以配体-受体形式结合，促进炎症细胞的趋化。利用大肠杆菌BL21表达并纯化CypA蛋白，利用该蛋白刺激小鼠的骨髓来源巨噬细胞（BMDM）。实时荧光定量PCR和ELISA试验结果发现，BMDM分泌的IL-1β等炎症因子的表达水平均显著上调，表明胞外CypA具有促进炎症发生的作用。同时以CypA蛋白为免疫原制备多克隆抗体，用于LPS诱导的小鼠急性肺炎的治疗试验。与未治疗组相比，anti-CypA抗体治疗组小鼠的肺组织表面出血点较少，损伤程度减轻，肺组织和血液中IL-1β的表达量显著下降，表明anti-CypA抗体对LPS诱导的炎症具有一定的治疗效果。本研究以胞外CypA-CD147的相互作用为靶点，利用anti-CypA抗体抑制炎症反应，为开发抗炎症药物提供了一种新的思路和策略。

Abstract:

Cyclophilin A (CypA) is a member of peptidyl prolylisomerases (PPIase) family. CypA is best known as a ubiquitously distributed intracellular protein. It has also been shown to be secreted by cells in response to inflammatory stimuli and oxidative stress. Extracellular CypA (eCypA) interacts with CD147 to initiate inflammatory responses via recruiting leucocytes into inflamed tissue. Recombinant CypA was expressed in Escherichia coli and then purified using Superdex 75TM 16/60. The results of Real-time PCR and ELISA showed that the expression levels of proinflammatory cytokines, such as IL-1β, secreted by eCypA stimulated BMDM were significantly up-regulated, indicating that eCypA played an important role in promoting inflammatory responses. In addition, anti-CypA antibody was prepared using purified CypA protein for therapeutic evaluation in a mouse model of LPS-induced acute lung inflammation. Antibody-treated mice showed reduced lung injury and the expression levels of IL-1β in the lung tissue and blood were decreased significantly, indicating that anti-CypA antibody exerted a potent anti-inflammatory activity. Our findings provide a potential therapeutic antibody for inflammation-mediated diseases.