Department of Experimental and Clinical Medicine, University of Florence, Florence, Italy.
Department of Liver and Gastrointestinal Diseases, Biodonostia Health Research Institute, CIBERehd, Ikerbasque, San Sebastian, Spain.
Hepatology. 2021 Oct;74(4):2007-2020. doi: 10.1002/hep.31888. Epub 2021 Jul 5.
Cholangiocarcinoma (CCA) is characterized by high resistance to chemotherapy and poor prognosis. Several oncogenic pathways converge on activation of extracellular signal-regulated kinase 5 (ERK5), whose role in CCA has not been explored. The aim of this study was to investigate the role of ERK5 in the biology of CCA.
ERK5 expression was detected in two established (HuCCT-1 and CCLP-1) and two primary human intrahepatic CCA cell lines (iCCA58 and iCCA60). ERK5 phosphorylation was increased in CCA cells exposed to soluble mediators. In both HuCCT-1 and CCLP-1 cells, ERK5 was localized in the nucleus, and exposure to fetal bovine serum (FBS) further increased the amount of nuclear ERK5. In human CCA specimens, ERK5 mRNA expression was increased in tumor cells and positively correlated with portal invasion. ERK5 protein levels were significantly associated with tumor grade. Growth, migration, and invasion of CCA cells were decreased when ERK5 was silenced using specific short hairpin RNA (shRNA). The inhibitory effects on CCA cell proliferation, migration and invasion were recapitulated by treatment with small molecule inhibitors targeting ERK5. In addition, expression of the angiogenic factors VEGF and angiopoietin 1 was reduced after ERK5 silencing. Conditioned medium from ERK5-silenced cells had a lower ability to induce tube formation by human umbilical vein endothelial cells and to induce migration of myofibroblasts and monocytes/macrophages. In mice, subcutaneous injection of CCLP-1 cells silenced for ERK5 resulted in less frequent tumor development and smaller size of xenografts compared with cells transfected with nontargeting shRNA.
ERK5 is a key mediator of growth and migration of CCA cells and supports a protumorigenic crosstalk between the tumor and the microenvironment.
胆管癌（CCA）的特点是对化疗具有高度耐药性和预后不良。几种致癌途径都集中在外源信号调节激酶 5（ERK5）的激活上，但其在 CCA 中的作用尚未得到探索。本研究旨在研究 ERK5 在 CCA 生物学中的作用。
在两种已建立的（HuCCT-1 和 CCLP-1）和两种原发性人肝内 CCA 细胞系（iCCA58 和 iCCA60）中检测到 ERK5 表达。CCA 细胞暴露于可溶性介质后，ERK5 磷酸化增加。在 HuCCT-1 和 CCLP-1 细胞中，ERK5 定位于细胞核中，并且暴露于胎牛血清（FBS）进一步增加了核 ERK5 的含量。在人 CCA 标本中，肿瘤细胞中 ERK5mRNA 表达增加，并与门静脉侵犯呈正相关。ERK5 蛋白水平与肿瘤分级显著相关。使用特异性短发夹 RNA（shRNA）沉默 ERK5 可降低 CCA 细胞的生长、迁移和侵袭。ERK5 的小分子抑制剂处理可重现对 CCA 细胞增殖、迁移和侵袭的抑制作用。此外，ERK5 沉默后血管生成因子 VEGF 和血管生成素 1 的表达减少。ERK5 沉默细胞的条件培养基通过人脐静脉内皮细胞诱导管形成和诱导成纤维细胞和平滑肌细胞/单核细胞/巨噬细胞迁移的能力降低。在小鼠中，与转染非靶向 shRNA 的细胞相比，CCLP-1 细胞沉默 ERK5 后，皮下注射导致肿瘤发生频率降低，异种移植物体积减小。
ERK5 是 CCA 细胞生长和迁移的关键介质，支持肿瘤与微环境之间的促肿瘤相互作用。
