【摘要】 目的 探讨血清叶酸受体1(FOLR1)与卵巢癌患者铂类化疗耐药的关联，明确沉默FOLR1通过调控STAT3信号通路影响上皮性卵巢癌多药耐药细胞顺铂化疗敏感性的分子机制。方法 选取2020-01-01-2022-12-31在重庆大学附属三峡医院接受手术治疗的50例上皮性卵巢癌患者为研究对象，按化疗响应分为铂耐药组(n=20)和铂敏感组(n=30),采用ELISA检测化疗前后血清FOLR1水平；同时选取50例正常卵巢上皮组织、30例良性卵巢上皮肿瘤组织及30例恶性上皮性卵巢癌组织，采用免疫组织化学法检测FOLR1表达。培养SKOV3细胞及顺铂耐药株SKOV3/DDP,qRT-PCR和蛋白质印迹法验证FOLR1表达差异。设置2组细胞实验：(1)SKOV3/DDP细胞随机分为对照组、顺铂组、siRNA-NC组、siRNA-FOLR1组和siRNA-FOLR1+顺铂组；(2)顺铂处理的SKOV3/DDP细胞随机分为顺铂组、siRNA-NC+顺铂组、siRNA-FOLR1+顺铂组和siRNA-FOLR1+Colivelin TFA+顺铂组。CCK8法检测细胞增殖抑制率，流式细胞术检测细胞凋亡率，蛋白质印迹法检测JAK2、p-JAK2、STAT3和p-STAT3蛋白表达。结果 化疗前铂耐药组患者血清FOLR1水平[(0.50±0.14)μg/L]高于铂敏感组[(0.30±0.12)μg/L],t=6.107,P<0.001;化疗后2组均下降，铂耐药组[(0.43±0.12)μg/L]仍高于铂敏感组[(0.14±0.10)μg/L],t=4.215,P<0.001;且铂药组下降幅度更小[(0.07±0.03)μg/L vs(0.16±0.04)μg/L],t=2.109,P=0.042。正常卵巢上皮组织、良性卵巢上皮肿瘤组织FOLR1阳性表达率分别为10.15%和9.88%,差异无统计学意义，χ~2=1.243,P=0.265;恶性卵巢上皮肿瘤组织FOLR1阳性表达率为49.88%,高于正常及良性组织，均P<0.001。SKOV3/DDP细胞的FOLR1 mRNA相对表达量(15.29±0.91 vs 1.00±0.17)和蛋白相对表达量(4.13±0.36 vs 1.01±0.16)均高于SKOV3细胞，t值分别为38.652和16.893,均P<0.001。方差分析结果显示，各组细胞增殖抑制率及凋亡率差异均有统计学意义，均P<0.001。与对照组和siRNA-NC组相比，顺铂组及siRNA-FOLR1组SKOV3/DDP细胞的增殖抑制率与凋亡率均升高(均P<0.001);而siRNA-FOLR1与顺铂联用效果更为明显，其抑制率和凋亡率均明显高于顺铂组及siRNA-FOLR1组(均P<0.001)。此外，在联合用药基础上加入STAT3激动剂Colivelin TFA后，细胞增殖抑制率与凋亡率较siRNA-FOLR1+顺铂组明显下调(均P<0.001),表明干扰FOLR1可能通过调控STAT3信号通路增加SKOV3/DDP细胞对顺铂的敏感性。结论 FOLR1高表达与卵巢癌铂类耐药密切相关，沉默FOLR1可通过抑制STAT3信号通路激活，增强多药耐药细胞对顺铂的敏感性，为逆转卵巢癌化疗耐药提供新靶点。更多还原

【Abstract】 Objective To investigate the association between serum folate receptor 1(FOLR1) and platinum-based chemotherapy resistance in ovarian cancer patients,and to elucidate the molecular mechanism by which silencing FOLR1 influences the cisplatin chemosensitivity of epithelial ovarian cancer multidrug-resistant cells through regulation of the STAT3signaling pathway.Methods Fifty patients with epithelial ovarian cancer who underwent surgical treatment at Chongqing University Three Gorges Hospital from January 1,2020,to December 31,2022,were selected as study subjects.They were divided into a platinum-resistant group(n=20) and a platinum-sensitive group(n=30) based on their response to chemotherapy.Serum FOLR1 levels before and after chemotherapy were detected by using ELISA.Additionally, 50 samples of normal ovarian epithelial tissue,30 samples of b enign ovarian epithelial tumor tissue,and 30 samples of malignant epithelial ovarian cancer tissue were collected.Immunohistochemistry was used to detect FOLR1 expression.SKOV3 cells and the cisplatin-resistant strain SKOV3/DDP were cultured,and FOLR1 expression differences were validated by qRTPCR and western blotting.Two sets of cell experiments were designed:(1) SKOV3/DDP cells were randomly divided into control group,cisplatin group,siRNA-NC group,siRNA-FOLR1 group,and siRNA-FOLR1+cisplatin group;(2) Cisplatin-treated SKOV3/DDP cells were randomly divided into cisplatin group,siRNA-NC+cisplatin group,siRNA-FOLR1+cisplatin group,and siRNA-FOLR1+Colivelin TFA+cisplatin group.The CCK8 assay was used to detect the proliferation inhibition rate,flow cytometry was used to detect the apoptosis rate,and western blotting was used to detect the protein expression levels of JAK 2,p-JAK 2,STAT3,and p-STAT3.Results Before chemotherapy,the serum FOLR1 level in the platinum-resistant group [(0.50±0.14) μg/L] was higher than that in the platinum-sensitive group[(0.30±0.12) μg/L],with a statistically significant difference(t=6.107,P<0.001).After chemotherapy,levels decreased in both groups,but the level in the platinum-resistant group [(0.43±0.12) μg/L] remained higher than that in the platinum-sensitive group [(0.14±0.10) μg/L],t=4.215,P<0.001.The magnitude of decrease in the platinum-resistant group was smaller[(0.07±0.03) μg/L vs(0.16±0.04) μg/L,t=2.109,P=0.042].The positive expression rates of FOLR1 in normal ovarian epithelial tissue and benign ovarian epithelial tumor tissue were 10.15 % and 9.88%,respectively,with no statistically significant difference(χ~2=1.243,P=0.265).The positive expression rate of FOLR1in malignant ovarian epithelial tumor tissue was 49.88%,which was significantly higher than that in normal and benign tissues(both P<0.001).The relative mRNA expression(15.29±0.91 vs 1.00±0.17) and protein expression(4.13±0.36 vs 1.01±0.16) of FOLR1 in SKOV3/DDP cells were significantly higher than those in SKOV3 cells(t values were38.652 and 16.893,respectively,both P<0.001).ANOVA results showed that the differences in cell proliferation inhibition rates and apoptosis rates among all groups were statistically significant(both P<0.001).Compared with the control group and siRNA-NC group,the proliferation inhibition rate and apoptosis rate of SKOV3/DDP cells in the cisplatin group and the siRNA-FOLR1 group were increased(both P<0.001).The combination of siRNA-FOLR1 and cisplatin showed a more pronounced effect,with its inhibition rate and apoptosis rate significantly higher than those in the cisplatin alone group and the siRNA-FOLR1 alone group(all P<0.001).Furthermore,after adding the STAT3 agonist Colivelin TFA to the combination treatment,the cell proliferation inhibition rate and apoptosis rate were significantly downregulated compared to the siRNA-FOLR1+cisplatin group(all P<0.001), indicating that interfering with FOLR1 may increase the sensitivity of SKOV3/DDP cells to cisplatin by regulating the STAT3 signaling pathway.Conclusions High expression of FOLR1 is closely associated with platinum resistance in ovarian cancer.Silencing FOLR1 can enhance the sensitivity of multidrug-resistant cells to cisplatin by inhibiting the activation of the STAT3 signaling pathway,providing a potential new target for reversing chemotherapy resistance in ovarian cancer.更多还原