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RhoA/ROCK signaling mediates plasticity of scirrhous gastric carcinoma motility

Abstract

The small guanosine triphosphatase (GTPase) Rho and its downstream effector Rho-associated kinase (ROCK) is one of a key mediator involved in controlling focal adhesions and the dynamics of actin stress fibers. The molecular mechanisms for the function of Rho/ROCK pathway leading to the progression in scirrhous gastric carcinoma cells have not been defined. The activation of RhoA in several gastric carcinoma cells was examined. The role of RhoA/ROCK pathway in the metastatic processes of gastric carcinoma cells, using a human scirrhous gastric cancer cell line, OCUM-2MD3 was investigated by in vitro adhesion and invasion assay. The effect of ROCK inhibitor, Y-27632 on the mRNA expression of the integrin family and MMP in gastric carcinoma cells was subsequently examined by Reverse transcriptional (RT)-PCR analysis. Finally, Random OCUM-2MD3 cell motility was evaluated using Time-lapse microscopy. ROCK inhibitor significantly increased the adhesion of OCUM-2MD3 cells to the extracellular matrix (ECM) protein matrigel. Further examination using ECM components showed enhanced binding ability was obtained only in laminin and Integrin subunits α3-integrin was clearly up-regulated by treatment with Y-27632 in OCUM-2MD3 cells. ROCK inhibitor also enhanced the invasion of OCUM-2MD3 cells through matrigel and the expression of membrane-type 1 matrix metalloproteinase (MT1-MMP). Time-lapse microscopy showed conversion of OCUM-2MD3 cells from round to more elongated morphology in the presence of Y-27632, suggesting that inhibition of RhoA/ROCK pathway undergo a so-called ‘amoeboid to mesenchymal’ transition. The fact that Rac1 inhibitor decreased the facilitated invasion by ROCK inhibitor suggested the possibility that increased invasion ability of OCUM-2MD3 cells was related to Rac activity. These data may suggest that RhoA/ROCK regulate plasticity of metastatic gastric carcinoma via mesenchymal-amoeboid transition, leading to provide new insights for designing a new and effective treatment for this type of refractory carcinoma.

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Acknowledgments

This study is partially founded by Grants-in Aid for Scientific Research (Nos. 20591573 and 22390262) from the Ministry of Education, Science, Sports, Culture and Technology of Japan. We thank Masako Shinkawa for her help in doing cell preparations and several assays.

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Correspondence to Tasuku Matsuoka.

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Matsuoka, T., Yashiro, M., Kato, Y. et al. RhoA/ROCK signaling mediates plasticity of scirrhous gastric carcinoma motility. Clin Exp Metastasis 28, 627–636 (2011). https://doi.org/10.1007/s10585-011-9396-6

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Keywords

  • Rho/ROCK
  • Gastric carcinoma
  • Integrin
  • AMT