地塞米松对顺铂诱导人肺腺癌细胞SPC-A1凋亡的抑制作用及分子机制

The Inhibitory Effects of Dexamethasone on Cisplatin Induced Apoptosis of Human Lung Adenocarcinoma Cell SPC-A1 and Its Molecular Mechanism

研究地塞米松(DEX)对顺铂(CDDP)引起的人肺腺癌细胞SPC-A1凋亡的抑制作用及可能的分子机制。将不同浓度的DEX分别加入人肺腺癌细胞SPC-A1体外诱导培养24h后,再用不同浓度CDDP处理SPC-A1细胞48h。MTT法检测细胞存活率;RT-PCR方法检测1μmol/L DEX诱导培养不同时间后SPC-A1细胞内血清/糖皮质激素-诱导激酶(SGK-1)和有丝分裂原蛋白激酶(MKP-1)的表达;用生物素标记的抗糖皮质激素受体(GR)抗体对SPC-A1细胞行免疫组化染色来检测GR的表达。MTT测定结果显示,SPC-A1细胞经DEX诱导后,对CDDP所致的凋亡有抵抗作用并与DEX浓度呈剂量依赖关系。RT-PCR检测到DEX诱导培养能提高SGK-1在SPC-A1细胞中的表达,表达量随时间延长而增加;但未检测到MKP-1的表达。免疫组化检测显示SPC-A1细胞经DEX诱导后GR上调,细胞内GR表达的阳性细胞数明显高于对照组。结果表明DEX对CDDP引起SPC-A1细胞的凋亡有抑制作用。可能的分子机制是通过上调细胞内GR表达,进而上调其通路下游抗凋亡蛋白SGK-1的表达,导致SPC-A1细胞产生抗凋亡作用。

The aim of this study is to investigate the apoptotic inhibition and its molecular mechanism of dexametha- sone （DEX） acting on cisplatin （CDDP）-induced apoptosis of human lung adenocarcinoma cell SPC-A1. SPC-A1 cells were pre-cultured in vitro for 24 hours with DEX in different concentrations and then CDDP was added in different concentrations for culturing for further 48 hours, The survival rates of the cells were determined by MTT. The ex- pression of serum/glucocorticoid-induced kinase （SGK-1） and mitogen-activated protein kinase phosphatase-1 （MKP- 1） in SPC-A1 cells after being cultured by 1 μmol/L DEX at different time was detected by semi-quantitative RT- PCR technology. The expression of glucocorticoid receptor （GR） in SPC-A1 cells was measured by immunohisto- chemistry （IHC） with biotin-labeled anti-GR. The results of MTT showed that SPC-A1 cells had resistance to CD-DP induced apoptosis with pre-eultured DEX and the resistance intensity presented DEX concentration-dependent. The expressing quantity of SGK-1 in SPC-A1 cells stimulated by DEX could be elevated and increased with intention of time, but the express of MKP-1 was not detected. Up-regulated expression of GR in SPC-A1 cells stimulated by DEX was detected by IHC. The number of cells expressing GR in SPC-A1 cells was significantly higher than that in the control group. The results showed that DEX inhibited apoptosis of SPC-A1 ceils induced by CDDP. The possible molecular mechanism is that elevated expression of GR induced by DEX up-regulates the expression of SGK-1 which locates at the downstream of anti-apoptosis pathway. The apoptosis resistance of SPC-A1 cells may account for all above the factors.