IFITM1基因沉默降低小细胞肺癌细胞顺铂耐药性的作用及机制

doi:10.3760/cma.j.cn371439-20250712-00044

摘要/Abstract

摘要：

目的探讨靶向沉默干扰素诱导的跨膜蛋白1（IFITM1）基因对小细胞肺癌（SCLC）细胞顺铂（cisplatin）耐药性的影响，并探讨作用机制。方法收集2019年1月至2020年12月在云南省玉溪市人民医院肿瘤中心24例接受含顺铂方案化疗SCLC患者的肿瘤组织样本，分为耐药组（n=12）和非耐药组（n=12）。采用实时荧光定量PCR和酶联免疫吸附法分别检测IFITM1的mRNA和蛋白含量。在顺铂耐药SCLC细胞株NCI-H82/cisplatin和SHP-77/cisplatin细胞中靶向沉默IFITM1基因后，检测耐药细胞IC₅₀变化。用低剂量顺铂处理IFITM1基因沉默的上述耐药细胞，分为耐药细胞组、IFITM1沉默对照组、IFITM1沉默组、梯度顺铂处理组、IFITM1沉默+梯度顺铂处理组、顺铂（10 μmol/L）处理组、IFITM1沉默对照+顺铂（10 μmol/L）处理组、IFITM1沉默+顺铂（10 μmol/L）处理组。采用CCK-8法检测细胞增殖；Transwell小室法检测细胞侵袭；TUNEL染色检测细胞凋亡；蛋白质印迹法检测蛋白表达。结果耐药组SCLC患者组织中IFITM1 mRNA、蛋白表达量均高于非耐药组（均P<0.05）。IC₅₀值分析结果显示，NCI-H82/cisplatin细胞耐药细胞组、IFITM1沉默对照组、IFITM1沉默组对于顺铂的IC₅₀值分别为（43.12±8.00）、（41.48±10.57）、（14.82±4.78） μmol/L，差异有统计学意义（F=11.44， P=0.009），IFITM1沉默组的IC₅₀值均小于耐药细胞组、IFITM1沉默对照组（均P<0.05）。CCK-8法结果显示，NCI-H82/cisplatin细胞耐药细胞组、顺铂处理组、IFITM1沉默对照+顺铂处理组和IFITM1沉默+顺铂处理组的增殖活性分别为1.35±0.18、1.15±0.17、1.06±0.18、0.52±0.08，差异有统计学意义（F=15.17， P=0.001），IFITM1沉默+顺铂处理组细胞增殖活性均明显低于耐药细胞组、顺铂处理组和IFITM1沉默对照+顺铂处理组（均P<0.05）。Transwell小室实验结果表明，NCI-H82/cisplatin细胞耐药细胞组、顺铂处理组、IFITM1沉默对照+顺铂处理组和IFITM1沉默+顺铂处理组的侵袭数分别为（1 380.00±166.43）、（1 259.33±198.01）、（1 143.00±169.14）、（563.00±127.39）个，差异有统计学意义（F=15.11， P=0.001），IFITM1沉默+顺铂处理组细胞侵袭数均明显低于耐药细胞组、顺铂处理组和IFITM1沉默对照+顺铂处理组（均P<0.05）。TUNEL染色结果表明，NCI-H82/cisplatin细胞耐药细胞组、顺铂处理组、IFITM1沉默对照+顺铂处理组和IFITM1沉默+顺铂处理组的细胞凋亡率分别为（12.00±2.91）%、（15.83±3.50）%、（16.37±3.59）%、（42.46±4.95）%，差异有统计学意义（F=40.45， P<0.001），IFITM1沉默+顺铂处理组细胞凋亡率均明显高于耐药细胞组、顺铂处理组和IFITM1沉默对照+顺铂处理组（均P<0.05）。SHP-77/cisplatin细胞上述实验结果与NCI-H82/cisplatin细胞一致。蛋白质印迹法结果表明，NCI-H82/cisplatin细胞耐药细胞组、IFITM1沉默对照组和IFITM1沉默组中IFITM1、p-β-catenin、ABCB1、ABCG2蛋白表达量差异均有统计学意义（均P<0.05），与耐药细胞组和IFITM1沉默对照组比较，IFITM1沉默组p-β-catenin蛋白表达量较高，IFITM1、ABCB1和ABCB2蛋白表达量均较低（均P<0.05）。结论 IFITM1在SCLC细胞顺铂耐药机制中起促进作用，靶向沉默IFITM1可通过抑制Wnt信号通路有效降低耐药性。

关键词: 小细胞肺癌, 顺铂, 抗药性，肿瘤, Wnt信号通路, 干扰素诱导的跨膜蛋白1

Abstract:

Objective To investigate the effect of targeted silencing of interferon-induced transmembrane protein 1 （IFITM1） on cisplatin resistance in small cell lung cancer （SCLC） cells and to explore the underlying mechanisms. Methods Tumor tissue samples of 24 patients with SCLC who received chemotherapy with cisplatin regimen at the Tumor Center of People's Hospital of Yuxi City in Yunnan Province from January 2019 to December 2020 were collected and divided into the resistant group （n=12） and the non-resistant group （n=12）. The mRNA and protein expression levels of IFITM1 were detected by real-time quantitative PCR and enzyme-linked immunosorbent assay， respectively. After targeted silencing of IFITM1 in the cisplatin-resistant SCLC cell lines NCI-H82/cisplatin and SHP-77/cisplatin cells， changes in the IC₅₀ of cisplatin were evaluated. IFITM1-silenced cisplatin-resistant cells were treated with low-dose cisplatin and divided into cisplatin-resistant cell group， IFITM1-silencing negative control group， IFITM1-silencing group， gradient cisplatin treatment group， IFITM1-silencing+gradient cisplatin treatment group， cisplatin （10 μmol/L） treatment group， IFITM1-silencing negative control+cisplatin（10 μmol/L） treatment group， IFITM1-silencing+cisplatin （10 μmol/L） treatment group. Cell proliferation was assessed using the CCK-8 assay， cell invasion was evaluated using the Transwell assay， cell apoptosis was detected by TUNEL staining， and protein expression levels were analyzed by Western blotting. Results The expression levels of IFITM1 mRNA and protein in the tissues of SCLC patients in the resistant group were both higher than those in the non-resistant group （both P<0.05）. IC₅₀ analysis results showed that the IC₅₀ values of cisplatin in NCI-H82/cisplatin cells were （43.12±8.00），（41.48±10.57）， and （14.82±4.78）μmol/L in the cisplatin-resistant cell group， IFITM1-silencing negative control group， and IFITM1-silencing group， respectively， with a statistically significant difference （F=11.44， P=0.009）， and the IC₅₀ values in the IFITM1-silencing group were lower than those in the cisplatin-resistant cell group and IFITM1-silencing negative control group （both P<0.05）. CCK-8 assay results showed that， the proliferation activities of NCI-H82/cisplatin cells in the cisplatin-resistant cell group， cisplatin treatment group， IFITM1-silencing negative control+cisplatin treatment group， and IFITM1-silencing+cisplatin treatment group were 1.35±0.18， 1.15±0.17， 1.06±0.18， and 0.52±0.08， respectively， with a statistically significant difference （F=15.17， P=0.001）， and proliferation activity in the IFITM1-silencing+cisplatin treatment group was significantly lower than that in the cisplatin-resistant group， cisplatin treatment group and IFITM1-silencing negative control+cisplatin treatment group （all P<0.05）. Transwell assay results indicated that， the numbers of invading NCI-H82/cisplatin cells in the cisplatin-resistant cell group， cisplatin treatment group， IFITM1-silencing negative control+cisplatin treatment group， and IFITM1-silencing+cisplatin treatment group were 1 380.00±166.43， 1 259.33±198.01， 1 143.00±169.14， and 563.00±127.39， respectively， with a statistically significant difference （F=15.11， P=0.001）， and the numbers of invasion were significantly lower in the IFITM1-silencing+cisplatin treatment group than those in the cisplatin-resistant cell group， cisplatin treatment group and IFITM1-silencing negative control+cisplatin treatment group （all P<0.05）. TUNEL staining results showed that， apoptosis rates of NCI-H82/cisplatin cells in the above four groups were （12.00±2.91）%，（15.83±3.50）%，（16.37±3.59）%， and （42.46±4.95）%， respectively， with a statistically significant difference （F=40.45， P<0.001）， and the apoptosis rate of the IFITM1-silencing+cisplatin treatment group was significantly higher than that of the cisplatin-resistant cell group， cisplatin treatment group and IFITM1-silencing negative control+cisplatin treatment group （all P<0.05）. The above experimental results for SHP-77/cisplatin cells were consistent with those of NCI-H82/cisplatin cells. The results of Western blotting showed that there were statistically significant differences in the protein expression levels of IFITM1， p-β-catenin， ABCB1， and ABCG2 among the NCI-H82/cisplatin resistant cell group， the IFITM1-silencing negative control group， and the IFITM1-silencing group （all P<0.05）. Compared with the cisplatin-resistant cell group and the IFITM1-silencing negative control group， the IFITM1-silencing group exhibited significantly higher p-β-catenin expression and significantly lower IFITM1， ABCB1， and ABCG2 expression levels （all P<0.05）. Conclusions IFITM1 plays a promotive role in cisplatin resistance of SCLC cells. Targeted silencing of IFITM1 effectively reduces cisplatin resistance by inhibiting the Wnt signaling pathway.

Key words: Small cell lung carcinoma, Cisplatin, Drug resistance， neoplasm, Wnt signaling pathway, Interferon-induced transmembrane protein 1

钱海洪, 王雪梅, 刘四艳, 王华, 张恒姣, 杨东海, 孙琼, 赵聃冶. IFITM1基因沉默降低小细胞肺癌细胞顺铂耐药性的作用及机制[J]. 国际肿瘤学杂志, 2026, 53(5): 268-275.

Qian Haihong, Wang Xuemei, Liu Siyan, Wang Hua, Zhang Hengjiao, Yang Donghai, Sun Qiong, Zhao Danye. Effect and mechanism of IFITM1 gene silencing in mitigating cisplatin resistance in small cell lung cancer cells[J]. Journal of International Oncology, 2026, 53(5): 268-275.

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组别	IFITM1蛋白	p-β-catenin蛋白	ABCB1蛋白	ABCB2蛋白
耐药细胞组	2.10±1.02	0.19±0.02	1.00±0.16	0.57±0.11
IFITM1沉默对照组	2.17±0.96	0.19±0.01	1.03±0.15	0.60±0.08
IFITM1沉默组	0.27±0.05^ab	0.89±0.09^ab	0.25±0.05^ab	0.30±0.05^ab
F值	63.91	176.80	34.47	11.22
P值	<0.001	<0.001	<0.001	0.009