Chen Xin, Tharmanathan Tharsika, Mannargudi Baskar, Gou Hong, Uetrecht Jack P
Faculty of Pharmacy, University of Toronto, 144 College Street, Toronto, Canada M5S 3M2.
J Pharmacol Exp Ther. 2009 Dec;331(3):836-41. doi: 10.1124/jpet.109.157362. Epub 2009 Sep 4.
Nevirapine treatment can cause a skin rash. We developed an animal model of this rash and determined that the 12-hydroxylation metabolic pathway is responsible for the rash, and treatment of animals with 12-OH-nevirapine also leads to a rash. In the present study, we investigated the specificity of lymphocytes in nevirapine-induced skin rash. Brown Norway rats were treated with nevirapine or 12-OH-nevirapine to induce a rash. Lymph nodes were removed, and the response of lymphocytes to nevirapine and its metabolites/analogs was determined by cytokine production (enzyme-linked immunosorbent assay, enzyme-linked immunosorbent spot assay, and Luminex) and proliferation (alamar blue assay). Subsets of lymphocytes were depleted to determine which cells were responsible for cytokine production. Lymphocytes from animals rechallenged with nevirapine proliferated to nevirapine, but not to 12-OH-nevirapine or 4-chloro-nevirapine. They also produced interferon-gamma (IFN-gamma) when exposed to nevirapine, significantly less when exposed to 4-chloro-nevirapine, and very little when exposed to 12-OH-nevirapine, even though oxidation to 12-OH-nevirapine is required to induce the rash. Moreover, the specificity of lymphocytes from 12-OH-nevirapine-treated rats was the same, i.e., responding to nevirapine more than to 12-OH-nevirapine, even though these animals had never been exposed to nevirapine. A Luminex immunoassay showed that a variety of other cytokines/chemokines were also produced by nevirapine-stimulated lymphocytes. CD4(+) cells were the major source of IFN-gamma. The specificity of lymphocytes in activation assays cannot be used to determine what initiated an immune response. This has significant implications for understanding the evolution of an immune response and the basis of the pharmacological interaction hypothesis.
奈韦拉平治疗可引起皮疹。我们建立了这种皮疹的动物模型，并确定12-羟基化代谢途径与皮疹有关，用12-羟基奈韦拉平治疗动物也会导致皮疹。在本研究中，我们调查了奈韦拉平诱导的皮疹中淋巴细胞的特异性。用奈韦拉平或12-羟基奈韦拉平处理棕色挪威大鼠以诱导皮疹。切除淋巴结，通过细胞因子产生（酶联免疫吸附测定、酶联免疫斑点测定和Luminex）和增殖（alamar蓝测定）来确定淋巴细胞对奈韦拉平及其代谢物/类似物的反应。去除淋巴细胞亚群以确定哪些细胞负责细胞因子的产生。再次用奈韦拉平攻击的动物的淋巴细胞对奈韦拉平有增殖反应，但对12-羟基奈韦拉平或4-氯奈韦拉平无反应。当暴露于奈韦拉平时，它们还产生γ-干扰素（IFN-γ），暴露于4-氯奈韦拉平时产生的量明显减少，暴露于12-羟基奈韦拉平时产生的量极少，尽管诱导皮疹需要将奈韦拉平氧化为12-羟基奈韦拉平。此外，用12-羟基奈韦拉平处理的大鼠的淋巴细胞特异性相同，即对奈韦拉平的反应比对12-羟基奈韦拉平的反应更强，尽管这些动物从未接触过奈韦拉平。Luminex免疫测定表明，奈韦拉平刺激的淋巴细胞还产生多种其他细胞因子/趋化因子。CD4(+)细胞是IFN-γ的主要来源。激活试验中淋巴细胞的特异性不能用于确定引发免疫反应的因素。这对于理解免疫反应 的演变和药理相互作用假说的基础具有重要意义。
