原薯蓣皂素 A 通过 NF-κB 通路诱导移植物中 T 细胞靶向大鼠心脏移植的免疫抑制。
Protosappanin A induces immunosuppression of rats heart transplantation targeting T cells in grafts via NF-kappaB pathway.
机构信息
Department of Cardiology, The Second Affiliated Hospital of Harbin Medical University, Harbin, Heilongjiang Province, China.
出版信息
Naunyn Schmiedebergs Arch Pharmacol. 2010 Jan;381(1):83-92. doi: 10.1007/s00210-009-0461-5. Epub 2009 Nov 19.
Protosappanin A as one major and effective ingredient from Caesalpinia sappan L. exhibited antirejection activity obviously in heart-transplanted rat. The present study was designed to screen out the potential target genes of protosappanin A with microarray technology and reveal some molecular mechanism of immunosuppressive effect. Rats performed with ectopic peritoneal heart transplantation were randomized into three groups receiving different treatments for 7 days: protosappanin A group (25 mg kg(-1)), cyclosporine A group (10 mg kg(-1)), and control group. The differentially expressed genes responding to protosappanin A were analyzed with microarrays. Among common differentially expressed genes, the ones of interest were selected for further evaluation by real-time quantitative reverse transcriptase polymerase chain reaction (qRT-PCR), Western blot, immunochemistry, immunofluorescence, and ELISA. Among the 146 common differentially expressed genes, NF-kappaB and related genes like IkappaBa, IFN-r, and IP10 were selected for verification. The results of qRT-PCR, Western blot, immunochemistry, and ELISA showed that protosappanin A significantly reduced the expression of NF-kappaB, IFN-r, and IP10 (p < 0.05) and increased IkappaBa expression (p < 0.05) in graft. Moreover, the immunochemistry staining of NF-kappaB and IkappaBa was mainly observed in infiltrating mononuclear cells. Strikingly, immunofluorescent staining localized NF-kappaB to the TCR-positive T cells in graft. Furthermore, protosappanin A exhibited inhibitory effect on T cell proliferation in recipients after 7-day treatment. In conclusion, protosappanin A might act on T cells through inhibiting NF-kappaB activation and downstream gene expressions of IFN-r and IP10, meanwhile reducing T cell proliferation responding to alloantigen, so as to induce immunosuppressive effect. The results encourage a potential therapeutic evaluation of protosappanin A for clinical organ transplantation or other T cell-mediated immune disorders. Additionally, our study also verified the feasibility of microarray utilization in Chinese herb research to explore molecular mechanism and promote development of scientific theories.
从苏木中提取的主要有效成分之一原巴西苏木素 A 对大鼠异位心脏移植有明显的抗排斥作用。本研究旨在采用基因芯片技术筛选出原巴西苏木素 A 的潜在靶基因，揭示其免疫抑制作用的部分分子机制。将异位腹腔心脏移植大鼠随机分为 3 组，分别给予不同的药物处理 7 天：原巴西苏木素 A 组（25mg/kg）、环孢素 A 组（10mg/kg）和对照组。采用基因芯片分析原巴西苏木素 A 作用后的差异表达基因。对共同差异表达基因进行实时定量逆转录聚合酶链反应（qRT-PCR）、Western blot、免疫组化、免疫荧光和 ELISA 检测，从中筛选出感兴趣的基因进一步验证。在 146 个共同差异表达基因中，选择 NF-κB 及其相关基因如 IkappaBa、IFN-r 和 IP10 进行验证。qRT-PCR、Western blot、免疫组化和 ELISA 结果表明，原巴西苏木素 A 可显著降低移植物中 NF-κB、IFN-r 和 IP10 的表达（p<0.05），增加 IkappaBa 的表达（p<0.05）。免疫组化染色显示 NF-κB 和 IkappaBa 主要存在于浸润的单核细胞中。免疫荧光染色显示 NF-κB 定位于移植物中 TCR 阳性 T 细胞上。此外，原巴西苏木素 A 还可抑制接受者 T 细胞的增殖。综上所述，原巴西苏木素 A 可能通过抑制 NF-κB 激活及其下游 IFN-r 和 IP10 基因的表达，同时减少 T 细胞对同种抗原的增殖反应，从而诱导免疫抑制作用。研究结果为原巴西苏木素 A 用于临床器官移植或其他 T 细胞介导的免疫性疾病的治疗提供了依据。此外，本研究还验证了基因芯片技术在中药研究中探索分子机制和促进科学理论发展的可行性。
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