title: ROCK is involved in vasculogenic mimicry formation in hepatocellular carcinoma cell line.
abstract: Ras homolog family member A (RhoA) and Rho-associated coiled coil-containing protein kinases 1 and 2 (ROCK1 and 2) are key regulators of focal adhesion, actomyosin contraction and cell motility. RhoA/ROCK signaling has emerged as an attractive target for the development of new cancer therapeutics. Whether RhoA/ROCK is involved in regulating the formation of tumor cell vasculogenic mimicry (VM) is largely unknown. To confirm this hypothesis, we performed in vitro experiments using hepatocellular carcinoma (HCC) cell lines. Firstly, we demonstrated that HCC cells with higher active RhoA/ROCK expression were prone to form VM channels, as compared with RhoA/ROCK low-expressing cells. Furthermore, Y27632 (a specific inhibitor of ROCK) rather than exoenzyme C3 (a specific inhibitor of RhoA) effectively inhibited the formation of tubular network structures in a dose-dependent manner. To elucidate the possible mechanism of ROCK on VM formation, real-time qPCR, western blot and immunofluorescence were used to detect changes of the key VM-related factors, including VE-cadherin, erythropoietin-producing hepatocellular carcinoma-A2 (EphA2), phosphoinositide 3-kinase (PI3K), matrix metalloproteinase (MMP)14, MMP2, MMP9 and laminin 5γ2-chain (LAMC2), and epithelial-mesenchymal-transition (EMT) markers: E-cadherin and Vimentin. The results showed that all the expression profiles were attenuated by blockage of ROCK. In addition, in vitro cell migration and invasion assays showed that Y27632 inhibited the migration and invasion capacity of HCC cell lines in a dose-dependent manner markedly. These data indicate that ROCK is an important mediator in the formation of tumor cell VM, and suggest that ROCK inhibition may prove useful in the treatment of VM in HCC.
authors.affiliation: Department of Pharmacy, Shanghai First People's Hospital, School of medicine, Shanghai Jiao Tong University, Shanghai, P.R. China.
authors.forename: Ji-Gang
authors.initials: JG
authors.lastname: Zhang
authors.affiliation: Department of Pharmacy, Shanghai First People's Hospital, School of medicine, Shanghai Jiao Tong University, Shanghai, P.R. China.
authors.forename: Xiao-Yu
authors.initials: XY
authors.lastname: Li
authors.affiliation: Department of Pharmacy, Shanghai First People's Hospital, School of medicine, Shanghai Jiao Tong University, Shanghai, P.R. China.
authors.forename: Yu-Zhu
authors.initials: YZ
authors.lastname: Wang
authors.affiliation: Department of Gastroenterology, Shanghai First People's Hospital, School of medicine, Shanghai Jiao Tong University, Shanghai, P.R. China.
authors.forename: Qi-Di
authors.initials: QD
authors.lastname: Zhang
authors.affiliation: Department of Pharmacy, Shanghai First People's Hospital, School of medicine, Shanghai Jiao Tong University, Shanghai, P.R. China.
authors.forename: Sheng-Ying
authors.initials: SY
authors.lastname: Gu
authors.affiliation: Department of Pharmacy, Shanghai First People's Hospital, School of medicine, Shanghai Jiao Tong University, Shanghai, P.R. China.
authors.forename: Xin
authors.initials: X
authors.lastname: Wu
authors.affiliation: Department of Pharmacy, Shanghai First People's Hospital, School of medicine, Shanghai Jiao Tong University, Shanghai, P.R. China.
authors.forename: Guan-Hua
authors.initials: GH
authors.lastname: Zhu
authors.affiliation: Department of Pharmacy, Shanghai First People's Hospital, School of medicine, Shanghai Jiao Tong University, Shanghai, P.R. China.
authors.forename: Qin
authors.initials: Q
authors.lastname: Li
authors.affiliation: Department of Pharmacy, Shanghai First People's Hospital, School of medicine, Shanghai Jiao Tong University, Shanghai, P.R. China.
authors.forename: Gao-Lin
authors.initials: GL
authors.lastname: Liu
mesh_terms: D000577:Amides / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects; D014408:Biomarkers, Tumor / Q000378:metabolism; D006528:Carcinoma, Hepatocellular / Q000098:blood supply* / Q000235:genetics / Q000378:metabolism; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000187:drug effects; D058750:Epithelial-Mesenchymal Transition; D056945:Hep G2 Cells; D061307:Human Umbilical Vein Endothelial Cells; D006801:Humans; D008113:Liver Neoplasms / Q000098:blood supply* / Q000235:genetics / Q000378:metabolism; D009361:Neoplasm Invasiveness; D009389:Neovascularization, Pathologic / Q000378:metabolism* / Q000473:pathology; D011725:Pyridines / Q000494:pharmacology; D054460:rho-Associated Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism / Q000502:physiology*; D020742:rhoA GTP-Binding Protein / Q000235:genetics / Q000378:metabolism / Q000502:physiology*
chemical_list: D000577:Amides; D014408:Biomarkers, Tumor; D011725:Pyridines; C108830:Y 27632; C515851:ROCK1 protein, human; C515852:ROCK2 protein, human; D054460:rho-Associated Kinases; D020742:rhoA GTP-Binding Protein