title: MRC-5 fibroblast-conditioned medium influences multiple pathways regulating invasion, migration, proliferation, and apoptosis in hepatocellular carcinoma.
abstract: BACKGROUND
  Carcinoma associated fibroblasts (CAFs), an important component of tumor microenvironment, are capable of enhancing tumor cells invasion and migration through initiation of epithelial-mesenchymal transition (EMT). MRC-5 fibroblasts are one of the CAFs expressing alpha-smooth muscle actin. It is ascertained that medium conditioned by MRC-5 fibroblasts stimulate motility and invasion of breast cancer cells. However, its role in hepatocellular carcinoma (HCC) is less clear. The aim of our study was to investigate the effect of MRC-5-CM on HCC and explore the underlying mechanisms.
  
  
  RESULTS
  Using a combination of techniques, the role of MRC-5-CM in HCC was evaluated. We determined that MRC-5-CM induced the non-classical EMT in Bel-7402 and MHCC-LM3 cell lines. Initiation of the non-classical EMT was mainly via quintessential redistribution of α-, β- and γ-catenin, P120 catenin, E-cadherin, and N-cadherin, rather than up-regulation of typical EMT-related transcription factors (i.e., Snail, Twist1, ZEB-1 and ZEB2). We also found that MRC-5-CM potentiated both the migration and invasion of Bel-7402 and MHCC-LM3 cells in mesenchymal movement mode through activation of the α6, β3, β4, β7 integrin/FAK pathway and upregulation of MMP2. The flow cytometric analysis showed that MRC-5-CM induced G1 phase arrest in Bel-7402 cells with a concomitant reduction of S phase cells. In contrast, MRC-5-CM induced S phase arrest in MHCC-LM3 cells with a concomitant reduction of cells in the G2/M phase. MRC-5-CM also inhibited apoptosis in Bel-7402 cells while inducing apoptosis in MHCC-LM3 cells.
  
  
  CONCLUSIONS
  Collectively, MRC-5-CM promoted HCC cell motility and invasiveness through initiation of the non-classical EMT, including redistribution of α-, β- and γ-catenin, P120 catenin, E-cadherin, and N-cadherin, activation of the integrin/FAK pathway, and upregulation of MMP2. Hence, MRC-5-CM exerted distinct roles in Bel-7402 and MHCC-LM3 cell viability by regulating cyclins, cyclin dependent kinases (CDKs), CDK inhibitors (CKIs), Bcl-2 family proteins and other unknown mechanosensors.
authors.affiliation: Division of Hepatobiliary and Pancreatic Surgery, Department of Surgery, First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, Zhejiang, People's Republic of China. dsm00012380@126.com.|Key Laboratory of Combined Multi-Organ Transplantation, Ministry of Public Health, Hangzhou, Zhejiang, China. dsm00012380@126.com.|Key Laboratory of Organ Trans-Plantation, Zhejiang Province, Hangzhou, Zhejiang, China. dsm00012380@126.com.
authors.forename: Songming
authors.initials: S
authors.lastname: Ding
authors.affiliation: Division of Hepatobiliary and Pancreatic Surgery, Department of Surgery, First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, Zhejiang, People's Republic of China. guoliangjh@sohu.com.|Key Laboratory of Combined Multi-Organ Transplantation, Ministry of Public Health, Hangzhou, Zhejiang, China. guoliangjh@sohu.com.|Key Laboratory of Organ Trans-Plantation, Zhejiang Province, Hangzhou, Zhejiang, China. guoliangjh@sohu.com.
authors.forename: Guoliang
authors.initials: G
authors.lastname: Chen
authors.affiliation: Division of Hepatobiliary and Pancreatic Surgery, Department of Surgery, First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, Zhejiang, People's Republic of China. zhangwu516@zju.edu.cn.|Key Laboratory of Combined Multi-Organ Transplantation, Ministry of Public Health, Hangzhou, Zhejiang, China. zhangwu516@zju.edu.cn.|Key Laboratory of Organ Trans-Plantation, Zhejiang Province, Hangzhou, Zhejiang, China. zhangwu516@zju.edu.cn.
authors.forename: Wu
authors.initials: W
authors.lastname: Zhang
authors.affiliation: Division of Hepatobiliary and Pancreatic Surgery, Department of Surgery, First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, Zhejiang, People's Republic of China. 1985xcy@gmail.com.|Key Laboratory of Combined Multi-Organ Transplantation, Ministry of Public Health, Hangzhou, Zhejiang, China. 1985xcy@gmail.com.|Key Laboratory of Organ Trans-Plantation, Zhejiang Province, Hangzhou, Zhejiang, China. 1985xcy@gmail.com.
authors.forename: Chunyang
authors.initials: C
authors.lastname: Xing
authors.affiliation: Division of Hepatobiliary and Pancreatic Surgery, Department of Surgery, First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, Zhejiang, People's Republic of China. zjxu@zju.edu.cn.|Key Laboratory of Combined Multi-Organ Transplantation, Ministry of Public Health, Hangzhou, Zhejiang, China. zjxu@zju.edu.cn.|Key Laboratory of Organ Trans-Plantation, Zhejiang Province, Hangzhou, Zhejiang, China. zjxu@zju.edu.cn.
authors.forename: Xiao
authors.initials: X
authors.lastname: Xu
authors.affiliation: Division of Hepatobiliary and Pancreatic Surgery, Department of Surgery, First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, Zhejiang, People's Republic of China. xiehy@zju.edu.cn.|Key Laboratory of Combined Multi-Organ Transplantation, Ministry of Public Health, Hangzhou, Zhejiang, China. xiehy@zju.edu.cn.|Key Laboratory of Organ Trans-Plantation, Zhejiang Province, Hangzhou, Zhejiang, China. xiehy@zju.edu.cn.
authors.forename: Haiyang
authors.initials: H
authors.lastname: Xie
authors.affiliation: Division of Oncology Department, First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, Zhejiang, People's Republic of China. luaililuaili@126.com.
authors.forename: Aili
authors.initials: A
authors.lastname: Lu
authors.affiliation: Division of Hepatobiliary and Pancreatic Surgery, Department of Surgery, First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, Zhejiang, People's Republic of China. ckj263@163.com.|Key Laboratory of Combined Multi-Organ Transplantation, Ministry of Public Health, Hangzhou, Zhejiang, China. ckj263@163.com.|Key Laboratory of Organ Trans-Plantation, Zhejiang Province, Hangzhou, Zhejiang, China. ckj263@163.com.
authors.forename: Kangjie
authors.initials: K
authors.lastname: Chen
authors.affiliation: Division of Hepatobiliary and Pancreatic Surgery, Department of Surgery, First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, Zhejiang, People's Republic of China. guohaijun431@126.com.|Key Laboratory of Combined Multi-Organ Transplantation, Ministry of Public Health, Hangzhou, Zhejiang, China. guohaijun431@126.com.|Key Laboratory of Organ Trans-Plantation, Zhejiang Province, Hangzhou, Zhejiang, China. guohaijun431@126.com.
authors.forename: Haijun
authors.initials: H
authors.lastname: Guo
authors.affiliation: Division of Hepatobiliary and Pancreatic Surgery, Department of Surgery, First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, Zhejiang, People's Republic of China. renzhigang1986@126.com.|Key Laboratory of Combined Multi-Organ Transplantation, Ministry of Public Health, Hangzhou, Zhejiang, China. renzhigang1986@126.com.|Key Laboratory of Organ Trans-Plantation, Zhejiang Province, Hangzhou, Zhejiang, China. renzhigang1986@126.com.
authors.forename: Zhigang
authors.initials: Z
authors.lastname: Ren
authors.affiliation: Division of Hepatobiliary and Pancreatic Surgery, Department of Surgery, First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, Zhejiang, People's Republic of China. shusenzheng@zju.edu.cn.|Key Laboratory of Combined Multi-Organ Transplantation, Ministry of Public Health, Hangzhou, Zhejiang, China. shusenzheng@zju.edu.cn.|Key Laboratory of Organ Trans-Plantation, Zhejiang Province, Hangzhou, Zhejiang, China. shusenzheng@zju.edu.cn.
authors.forename: Shusen
authors.initials: S
authors.lastname: Zheng
authors.affiliation: Division of Hepatobiliary and Pancreatic Surgery, Department of Surgery, First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, Zhejiang, People's Republic of China. linzhou19@163.com.|Key Laboratory of Combined Multi-Organ Transplantation, Ministry of Public Health, Hangzhou, Zhejiang, China. linzhou19@163.com.|Key Laboratory of Organ Trans-Plantation, Zhejiang Province, Hangzhou, Zhejiang, China. linzhou19@163.com.
authors.forename: Lin
authors.initials: L
authors.lastname: Zhou
mesh_terms: D000199:Actins / Q000378:metabolism; D017209:Apoptosis; D015820:Cadherins / Q000378:metabolism; D006528:Carcinoma, Hepatocellular / Q000378:metabolism / Q000473:pathology*; D051177:Catenins / Q000378:metabolism; D045744:Cell Line, Tumor; D002465:Cell Movement; D049109:Cell Proliferation; D002470:Cell Survival; D018920:Coculture Techniques; D017077:Culture Media, Conditioned / Q000737:chemistry*; D058750:Epithelial-Mesenchymal Transition; D005347:Fibroblasts / Q000166:cytology* / Q000378:metabolism; D005434:Flow Cytometry; D020869:Gene Expression Profiling; D006801:Humans; D016023:Integrins / Q000378:metabolism; D008113:Liver Neoplasms / Q000378:metabolism / Q000473:pathology*; D008856:Microscopy, Fluorescence; D009130:Muscle, Smooth / Q000378:metabolism; D009361:Neoplasm Invasiveness; D014157:Transcription Factors / Q000378:metabolism
chemical_list: D000199:Actins; D015820:Cadherins; D051177:Catenins; D017077:Culture Media, Conditioned; D016023:Integrins; D014157:Transcription Factors