维甲酸信号通路对脂多糖作用后A549细胞SP-B表达的影响

Effect of RA Signaling Pathway on the SP-B Expression of A549 Cells after Treatment with LPS

目的探讨在感染相关的肺透明膜病发病中是否有维甲酸受体α(retinoic acid receptor-α,RARα)异常的机制参与以及全反式维甲酸(all trans-retinoic acid,ATRA)对脂多糖(lipopolysaccharide,LPS)引起的A549细胞肺泡活性蛋白-B(surfactant protein B,SP-B)表达下调是否有干预作用。方法将体外培养的人肺腺癌细胞A549分为①对照组:用无血清DMEM培养液培养;②LPS组:含10μg/ml LPS无血清DMEM培养液培养24 h;③LPS+ATRA治疗组:10μg/ml LPS培养24 h,再加入1μmol/ml ATRA培养24 h;④ATRA+LPS预防组:1μmol/ml ATRA培养24 h,再加入10μg/ml LPS培养24 h。应用免疫细胞化学方法(SP法)检测各组A549细胞中SP-B、RARα的表达及分布情况。结果LPS组A549细胞胞质中SP-B表达较对照组明显减弱(P<0.05);RARα胞核内表达减弱而出现明显胞质内表达(P<0.05)。LPS+ATRA治疗组SP-B表达与对照组比较表达强度相近(P>0.05);RARα表达强度及细胞内定位与对照组表达无明显变化(P>0.05)。ATRA+LPS预防组A549细胞质中SP-B表达明显减弱;胞核中RARα的表达明显减弱,而胞质内表达增强,与LPS组结果一致(P>0.05)。结论LPS可引起A549细胞SP-B蛋白水平下调,其机制可能与LPS引起RARα表达和细胞内分布异常有关;ATRA可能通过诱导RARα核内表达而干预LPS引起的A549细胞SP-B表达下调;预先给与ATRA不能减轻LPS引起的A549细胞SP-B蛋白水平下调。

Objective By detecting the expression and distribution changes of the RARα in A549 cells treated with LPS, and observing the expression and distribution changes of SP-B, and RARα in A549 cells, to explore the mechanism by which the retinoic acid receptor abnormalities may be involved in infection-related hyaline membrane disease and the effects of all trans-retinoic acid（ATRA） on the down-regulation of SP-B in A549 cells caused by LPS. Methods The in vitro cultured A549 cells were assigned to control group（serum-free DMEM）, LPS group（serum-free DMEM containing 10 μg/ml LPS for 24 h）, LPS＋ ATRA treatment group（10/μg/ml LPS for 24 h,followed by 1 × 10^-6 μmol/ml ATRA for other 24 h） ,ATRA＋LPS prevention group（1× 10^-6 μmol/ml ATRA for 24 h,followed by 10 μg/ml LPS for other 24 h）. The expression levels of RAR-α and SPB were detected by immunocytochemistry. Results Compared with control group,the expression of SP-B in cytoplasm of A549 cells and that of RARα in nuclei were decreased in LPS group,while the expression of RARα in cytoplasm was increased（P〈 0.05）. Compared with control group,the expression of SP-B and RARα had no changes in LPS＋ATRA treatment group（P〉 0. 05）. Compared with control group,the expression of SP-B in cytoplasm of A549 cells and that of RARa in nuclei were decreased,and the expression of RARα in cytoplasm was increased in ATRA＋ LPS prevention group（P〈0.05）. As compared with LPS group,the expression of SP-B and RARα had no changes in ATRA＋LPS prevention group（P〉0.05）. Conclusion The mechanism by which LPS can lead to the down-regulation of SP-B in A549 cells may be related to the abnormal expression and cellular distribution of RARα. By inducing the expression of RARα in nuclei, ATRA can interfere with the LPS-induced down-regulation of the SP-B expression in A549 cells. Pretreatment of A549 cells with ATRA could not down-regulate the expression of SP-B caused by LPS in A549 cells.