摘要目的 探讨半胱氨酰白三烯受体在人角质形成细胞巾的功能.方法 两步酶消化法分离、培养人角质形成细胞.分别采用激光共聚焦和RT-PCR技术观察培养细胞CysLTR的定位和表达;Fura2/AM法检测胞内钙流,MTT法检测LTD4、MK571和BAYu9773分别作用后细胞增殖的变化.结果 人角质形成细胞可见CysLTR1和CysLTR2表达,主要定位于细胞膜和部分胞质及胞核.LTD4刺激后角质形成细胞表达CysLTR较对照组明显增强,尤其是胞核中CysLTR表达增强(P＜0.05).胞内钙流峰值浓度达42.27±3.0 mmol/L,与基础值比较有差异统计学意义(P＜0.01);细胞增殖速率也加快,与对照组比较差异有统计学意义(P＜0.01).MK571、BAYu9773可完全阻断以上反应,且两组间差异无统计学意义.结论 人角质形成细胞存在功能性CysLTR表达,并可促进胞内钙流增加及细胞增殖.

abstractsObjective To detect the expression and function of cysteinyl leukotriene receptors (CysLTRs)in keratinocytes.Methods Human keratinocytes were isolated from the tissue of foreskin by digestion with dispase Ⅱ and trypsin,and subjected to primary culture.By using confocal laser scanning microscopy and reverse transcriptase PCR,the localization and expression of CysLTRs were studied in kemtinocytes.respectively.Some primarily cultured keratinocytes were pretreated with leukotriene D4 (30 nmoi/L),MK571(300 nmol/L),and BAYu9773 for 5 minutes followed by the detection of intmcellular calcium level using the Ca2+ indicator dye Fura-2/AM as well as cell proliferation bv MTT assay.Results The expressions of CysLTR1 and CysLTR2 were observed in cultured keratinocytes,and they were mainly located on cell membrane,partly in cytoplasm and nuclei.Compared with non.stimulated cells,a significant increase Was noted in the expression of CysLTRs,especially in the nuclei of keratinocytes stimulated by LTD4(P＜0.05),together with an elevation in intracellular calcium level(42.27±3.00 mmol/L,P＜0.01)and acceleration in cell proliferation (P＜0.01).However,both MK571 and BAYu9773 could completely block the effect of LTD4 on intmcellular calcium level and cell proliferation.and there was no significant difference in the blocking effect between MK571 and BAYu9773.Conclusions Functional CysLTRs are expressed in human keratinocytes.and they carl increase the intracellular calcium level in,and cell proliferation of,keratinocytes.