Department of Biomedical Science and Research Institute of Bioscience and Biotechnology, Hallym University, Chunchon 200-702, Republic of Korea.
Life Sci. 2011 Dec 5;89(23-24):896-904. doi: 10.1016/j.lfs.2011.09.021. Epub 2011 Oct 2.
Cyclophilin A (CypA) is an immunophilin that acts as a receptor for the immunosuppressant drug cyclosporine A (CsA). CypA has emerged as a potential drug target for several inflammatory diseases, although the details of its mechanism are unclear. We examined the protective effects of CypA on inflammation in Raw 264.7 cells and animal models.
A human CypA gene was fused with a protein transduction domain, PEP-1 peptide, to construct a cell permeable PEP-1-CypA protein. The protein expression level of cyclooxygenase-2 (COX-2) and cytokines was detected by Western blot, ELISA and mRNA level of COX-2 and cytokines were measured by RT-PCR. The nuclear factor-kappa B (NF-kB) and mitogen-activated protein kinase (MAPK) activation were analyzed by Western blot and electrophoretic mobility shift assay. Skin inflammation was detected with immunohistochemistry.
Transduced PEP-1-CypA protein markedly inhibited lipopolysaccharide- and 12-O-tetradecanoyl phorbol-13-acetate-induced expression levels of COX-2 as well as pro-inflammatory cytokine levels in vitro and in vivo. Furthermore, transduced PEP-1-CypA protein resulted in a significant reduction in the activation of NF-kB and MAPK.
The results indicate that PEP-1-CypA inhibits inflammatory response cytokines and enzymes by blocking NF-kB and MAPK activation upon stimulation of inflammation in vitro and in vivo. PEP-1-CypA protein may potentially be used as a therapeutic agent against skin diseases-related inflammation.
亲环素 A（CypA）是一种免疫亲和素，作为免疫抑制剂环孢素 A（CsA）的受体发挥作用。CypA 已成为几种炎症性疾病的潜在药物靶点，尽管其机制细节尚不清楚。我们研究了 CypA 对 Raw 264.7 细胞和动物模型中炎症的保护作用。
将人 CypA 基因与蛋白转导结构域 PEP-1 肽融合，构建细胞通透性 PEP-1-CypA 蛋白。通过 Western blot、ELISA 检测环氧化酶-2（COX-2）和细胞因子的蛋白表达水平，通过 RT-PCR 检测 COX-2 和细胞因子的 mRNA 水平。通过 Western blot 和电泳迁移率变动分析检测核因子-κB（NF-κB）和丝裂原活化蛋白激酶（MAPK）的激活。通过免疫组织化学检测皮肤炎症。
转导的 PEP-1-CypA 蛋白显著抑制脂多糖和 12-O-十四烷酰佛波醇-13-乙酸酯诱导的 COX-2 表达水平以及体外和体内的促炎细胞因子水平。此外，转导的 PEP-1-CypA 蛋白导致 NF-κB 和 MAPK 的激活显著减少。
结果表明，PEP-1-CypA 通过阻断 NF-κB 和 MAPK 的激活，抑制炎症刺激时体外和体内的炎症反应细胞因子和酶。PEP-1-CypA 蛋白可能有望成为治疗与皮肤疾病相关的炎症的治疗剂。
