linc00958/miR-185-5p/RSF-1 通过 AKT1/GSK3β/VEGFA 通路调节宫颈癌顺铂耐药和血管生成。
linc00958/miR-185-5p/RSF-1 modulates cisplatin resistance and angiogenesis through AKT1/GSK3β/VEGFA pathway in cervical cancer.
机构信息
Department of Gynecological Oncology, Tianjin Medical University Cancer Institute and Hospital, National Clinical Research Center for Cancer, Tianjin, People's Republic of China.
Key Laboratory of Cancer Prevention and Therapy, Tianjin's Clinical Research Center for Cancer, TianjinTianjin, 300060, China.
出版信息
Reprod Biol Endocrinol. 2022 Sep 2;20(1):132. doi: 10.1186/s12958-022-00995-2.
BACKGROUND
Chemoresistance is one of the major obstacles that lead to poor prognosis in cervical cancer. linc00958 was reported to be an oncogene in cervical cancer. However, its role in mediating chemoresistance remains to be revealed.
PURPOSE
To explore the regulatory mechanisms of linc00958 in cisplatin-resistant cervical cancer cells and further validate in xenograft mice.
METHODS
Online bioinformatic tools were used to conduct the pre-investigation of linc00958/miR-185-5p/RSF-1 and predict the associations between RSF-1 and AKT1/GSK3β/VEGFA in cervical cancer. RT-qPCR measured the RNA expression levels of linc00958/miR-185-5p/RSF-1 in SiHa and SiHa/DDP. Cell survival rates were evaluated by CCK8 methods after cells were exposed to differential concentrations of DDP. Dual-luciferase reporter methods were used to measure luciferase activity. Western blot measured RSF-1 protein and phosphorylated changes of AKT1/GSK3β. Immunofluorescence was employed to observe VEGFA secretion in vitro. Tube formation was applied to evaluate the in-vitro changes of angiogenesis. The SiHa/DDP cells stably transfected with pLKO-sh-NC or pLKO-sh-linc00958 plasmids, were injected into mice, establishing xenograft models. The changes in mice weight and tumor volumes were recorded. H&E staining and Immunohistochemistry (IHC) method was further performed.
RESULTS
linc00958 expression was higher in SiHa/DDP cells. High linc00958 expression was associated with low overall survival. In SiHa/DDP cells linc00958/miR-185-5p/RSF-1 axis inhibited the cellular resistance to cisplatin and suppressed VEGFA and the tube formation through AKT1/GSK3β/VEGFA pathway. The knockdown of linc00958 inhibited RSF-1 and Ki67, curbing tumor growth; it also inhibited VEGFA and CD34, decreasing angiogenesis in mice.
CONCLUSION
linc00958/miR-185-5p/RSF-1 modulates cisplatin resistance and angiogenesis through AKT1/GSK3β/VEGFA pathway in cervical cancer.
背景
化疗耐药性是导致宫颈癌预后不良的主要障碍之一。已有研究表明 linc00958 是宫颈癌的癌基因。然而，其在介导化疗耐药性中的作用仍有待揭示。
目的
探讨 linc00958 在顺铂耐药宫颈癌中的调控机制，并进一步在异种移植小鼠中进行验证。
方法
使用在线生物信息学工具对 linc00958/miR-185-5p/RSF-1 进行预调查，并预测宫颈癌中 RSF-1 与 AKT1/GSK3β/VEGFA 之间的关联。RT-qPCR 检测 SiHa 和 SiHa/DDP 细胞中 linc00958/miR-185-5p/RSF-1 的 RNA 表达水平。细胞分别暴露于不同浓度的 DDP 后，通过 CCK8 法评估细胞存活率。双荧光素酶报告基因法检测荧光素酶活性。Western blot 检测 RSF-1 蛋白及 AKT1/GSK3β 的磷酸化变化。免疫荧光法观察体外 VEGFA 的分泌情况。管形成实验用于评估体外血管生成的变化。将稳定转染 pLKO-sh-NC 或 pLKO-sh-linc00958 质粒的 SiHa/DDP 细胞注射到小鼠体内，建立异种移植模型。记录小鼠体重和肿瘤体积的变化。进一步进行 H&E 染色和免疫组化（IHC）方法。
结果
linc00958 在 SiHa/DDP 细胞中的表达较高。高 linc00958 表达与总生存期降低相关。在 SiHa/DDP 细胞中，linc00958/miR-185-5p/RSF-1 轴抑制了细胞对顺铂的耐药性，并通过 AKT1/GSK3β/VEGFA 通路抑制了 VEGFA 和管形成。linc00958 的敲低抑制了 RSF-1 和 Ki67，抑制了肿瘤生长；还抑制了 VEGFA 和 CD34，减少了小鼠的血管生成。
结论
linc00958/miR-185-5p/RSF-1 通过 AKT1/GSK3β/VEGFA 通路调节宫颈癌中的顺铂耐药性和血管生成。
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