Zhou Ru, Tang Wei, He Pei-Lan, Yang Yi-Fu, Li Yuan-Chao, Zuo Jian-Ping
Laboratory of Immunopharmacology, State Key Laboratory of Drug Research, Shanghai Institute of Materia Medica, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, 555 Zuchongzhi Road, Shanghai 201203, PR China.
Int Immunopharmacol. 2009 Jan;9(1):63-9. doi: 10.1016/j.intimp.2008.09.014. Epub 2008 Oct 23.
(5R)-5-hydroxytriptolide (LLDT-8) displayed immunosuppressive activities both in vitro and in autoimmune disease models. Here, we aim to further clarify the effect of LLDT-8 on the immune responses of human peripheral blood mononuclear cells (PBMC).
Cell proliferation of human PBMC from healthy donors was evaluated by [3H]-thymidine uptake. NK cell cytotoxicity was assayed using K562 cells in a [3H] lysis assay. Cytokine production was determined by enzyme-linked immunosorbent assay. The expression of cell surface molecules was detected with flow cytometry. The mRNA expression and the protein phosphorylation levels were detected by RT-PCR and Western immunoblot assay.
LLDT-8 at 25 and 50 nM significantly inhibited the PHA- and recall antigens-induced T cell proliferation, and suppressed mixed lymphocyte reaction. LLDT-8 reduced cytokines production (IFN-gamma, IL-2, TNF-alpha) in PHA- and Sac-activated PBMC. LLDT-8 did not alter the increased expression of MHC class I/II and B7.1, but reduced B7.2 by approximately 30%. No effect of LLDT-8 was observed for the expression of T cell activation markers (CD69, CD154). However, LLDT-8 significantly reduced IFN-gamma-expressing T cell percentages and IFN-gamma mRNA transcription in PHA-activated T cells. It also inhibited the phosphorylation levels of JNK and p38. LLDT-8 did not affect NK cytotoxic activity against K562 cells.
LLDT-8 was a promising immunosuppressant for human immune-related diseases.
(5R)-5-羟基雷公藤内酯醇（LLDT-8）在体外和自身免疫性疾病模型中均表现出免疫抑制活性。在此，我们旨在进一步阐明LLDT-8对人外周血单个核细胞（PBMC）免疫反应的影响。
通过[3H]-胸腺嘧啶核苷摄取评估健康供体人PBMC的细胞增殖。在[3H]释放试验中使用K562细胞测定NK细胞的细胞毒性。通过酶联免疫吸附测定法测定细胞因子的产生。用流式细胞术检测细胞表面分子的表达。通过RT-PCR和Western免疫印迹法检测mRNA表达和蛋白质磷酸化水平。
25和50 nM的LLDT-8显著抑制PHA和回忆抗原诱导的T细胞增殖，并抑制混合淋巴细胞反应。LLDT-8降低了PHA和Sac激活的PBMC中细胞因子的产生（IFN-γ、IL-2、TNF-α）。LLDT-8未改变MHC I/II类和B7.1表达的增加，但使B7.2降低了约30%。未观察到LLDT-8对T细胞活化标志物（CD69、CD154）表达的影响。然而，LLDT-8显著降低了PHA激活的T细胞中表达IFN-γ的T细胞百分比和IFN-γ mRNA转录。它还抑制了JNK和p38的磷酸化水平。LLDT-8不影响NK对K562细胞的细胞毒性活性。
LLDT-8是一种有前景的用于人类免疫相关疾病的免疫抑制剂。
