Tian Guodong, Zuo Lu, Li Jie, Zheng Xin, Gao Feng
Department of Hepatopancreatobilary Surgery, The First College of Clinical Medical Sciences, China Three Gorges University, No.183 Yiling Avenue, Wujiagang District, Yichang, 443000, Hubei, China.
Department of Geriatric, The First College of Clinical Medical Sciences, China Three Gorges University, Yichang, 443000, China.
Biochem Genet. 2025 Jan 8. doi: 10.1007/s10528-025-11020-7.
Accumulating evidence has demonstrated that Keratin18 (KRT18) functions as a pivotal gene in the progression of various cancers. However, its role in cholangiocarcinoma (CCA) remains unexplored. Our study elucidated the biological functions and underlying mechanisms of KRT18 in CCA. Bioinformatic databases were used to identify potential miRNAs and lncRNAs. The cellular localization of KRT18 and lncRNA HCG18 was examined through subcellular fractionation. Expression levels of genes were assessed by qRT-PCR, while protein levels were measured via western blot. Cell viability was analyzed using CCK-8 assays. Colony formation and EdU assays assessed cell proliferation, and sphere formation assays evaluated stem cell properties. The interactions between HCG18, miR-194-5p, and KRT18 were explored through RNA immunoprecipitation, RNA pulldown, and luciferase reporter assays. A xenograft tumor model was conducted to evaluate the in vivo function. In CCA tissues and cell lines, KRT18 expression was elevated. Functionally, silencing KRT18 reduced cell proliferation and stemness and inhibited cell cycle. Mechanistically, miR-194-5p directly targeted KRT18. HCG18, which was upregulated in CCA, interacted with miR-194-5p. Overexpression of KRT18 negated the effects of HCG18 suppression on CCA cell proliferation and stemness. Activation of MAPK signaling reversed the antitumor effects of KRT18 downregulation on CCA in vitro. Moreover, HCG18 was found to activate MAPK signaling through the miR-194-5p/KRT18 pathway. The in vivo assay demonstrated that HCG18 knockdown inhibited tumor growth by the miR-194-5p/KRT18/MAPK axis. HCG18 can promote cell proliferation and stem cell characteristics in CCA through the miR-194-5p/KRT18/MAPK signaling.
越来越多的证据表明，角蛋白18（KRT18）在多种癌症的进展中起着关键基因的作用。然而，其在胆管癌（CCA）中的作用仍未得到探索。我们的研究阐明了KRT18在CCA中的生物学功能及潜在机制。利用生物信息学数据库鉴定潜在的微小RNA（miRNA）和长链非编码RNA（lncRNA）。通过亚细胞分级分离检测KRT18和lncRNA HCG18的细胞定位。通过qRT-PCR评估基因表达水平，通过蛋白质印迹法测量蛋白质水平。使用CCK-8法分析细胞活力。集落形成和EdU法评估细胞增殖，成球试验评估干细胞特性。通过RNA免疫沉淀、RNA下拉和荧光素酶报告基因试验探索HCG18、miR-194-5p和KRT18之间的相互作用。建立异种移植肿瘤模型以评估体内功能。在CCA组织和细胞系中，KRT18表达升高。在功能上，沉默KRT18可降低细胞增殖和干性并抑制细胞周期。机制上，miR-194-5p直接靶向KRT18。在CCA中上调的HCG18与miR-194-5p相互作用。KRT18的过表达抵消了HCG18抑制对CCA细胞增殖和干性的影响。丝裂原活化蛋白激酶（MAPK）信号的激活逆转了KRT18下调对体外CCA的抗肿瘤作用。此外，发现HCG18通过miR-194-5p/KRT18途径激活MAPK信号。体内试验表明，敲低HCG18通过miR-194-5p/KRT18/MAPK轴抑制肿瘤生长。HCG18可通过miR-194-5p/KRT18/MAPK信号促进CCA中的细胞增殖和干细胞特性。
