Cao Fei, Zhang Zhi-Hui, Wu Ping, Li Li, Huang Jiang-Ming
Department of Medical Oncology, Sichuan Provincial Cancer Hospital and Institute, Chengdu 610041, China.
Sichuan Da Xue Xue Bao Yi Xue Ban. 2013 Nov;44(6):902-6.
To study the apoptotic inhibition and its molecular mechanism of dexamethasone (Dex) on cisplatin (CDDP)-induced human lung adenocarcinoma cells.
The human lung adenocarcinoma cell The human lung adenocarcinoma cell line, SPCA/I, was pre-cultured in vitro for 24 hours with Dex in different concentration and then different concentration of CDDP was added. The cells were cultured for another 48 hours. The survival rate of the cells was determined by MTT colorimetry. The appototic rate of SPCA/I cells measured by flow cytometer. Using 1 micromol/ L Dex to stimulate the SPCA/I cells RNAs of the cells at different time points (1 h, 2 h, 4 h, 6 h, 12 h) were extracted respectively. Semi-quantitative RT-PCR technology was used to detect the expression of the serum and glucocorticoid-induced kinase (SGK-1) and mitogen-activated protein kinase phosphatase-1(MKP-1) in SPCA/I cells. Simultaneously the glucocorticoid receptor (GR) of the SPCA/I cell line cells were measured by using biotin-labeled anti-glucocorticoid receptor antibody with immunohistochemistry assay.
SPCA/I cells showed resistance to CDDP-induced apoptosis while pre-cultured with Dex and the resistance intensity was Dex concentration-dependent. After Dex stimulating the SPCA/I cells, SGK-1 expressed increased and reached the peak at 12 h. But the expression of MKP-1 was not detected. Immunohistochemistry results showed that upregulated GR in SPCA/I cells after stimulation with Dex. The number of intracellular GR was significantly higher than that of control group.
The experimental results in vitro demonstrated that Dex inhibits apoptosis on CDDP-induced human lung adenocarcinoma cell line, SPCA/I. This anti-apoptosis effect might due to Dex increasing the expression of SGK-1, an anti-apoptotic protein, in its downstream signal pathway through the increasement of intracellular GR of SPCA/I cells.
研究地塞米松（Dex）对顺铂（CDDP）诱导的人肺腺癌细胞凋亡的抑制作用及其分子机制。
将人肺腺癌细胞系SPCA/I在体外分别用不同浓度的Dex预培养24小时，然后加入不同浓度的CDDP，再培养48小时。采用MTT比色法测定细胞存活率。用流式细胞仪检测SPCA/I细胞的凋亡率。用1μmol/L Dex刺激SPCA/I细胞，分别在不同时间点（1小时、2小时、4小时、6小时、12小时）提取细胞RNA。采用半定量RT-PCR技术检测SPCA/I细胞中血清和糖皮质激素诱导激酶（SGK-1）和丝裂原活化蛋白激酶磷酸酶-1（MKP-1）的表达。同时用生物素标记的抗糖皮质激素受体抗体通过免疫组织化学法检测SPCA/I细胞系细胞的糖皮质激素受体（GR）。
SPCA/I细胞在用Dex预培养后对CDDP诱导的凋亡表现出抗性，且抗性强度呈Dex浓度依赖性。Dex刺激SPCA/I细胞后，SGK-1表达增加，并在12小时达到峰值。但未检测到MKP-1的表达。免疫组织化学结果显示，Dex刺激后SPCA/I细胞中GR上调。细胞内GR的数量明显高于对照组。
体外实验结果表明，Dex抑制CDDP诱导的人肺腺癌细胞系SPCA/I的凋亡。这种抗凋亡作用可能是由于Dex通过增加SPCA/I细胞内GR，在其下游信号通路中增加抗凋亡蛋白SGK-1的表达所致。
