Jump to ContentJump to Main Navigation
Show Summary Details
More options …

Cellular and Molecular Biology Letters

Editor-in-Chief: /


IMPACT FACTOR 2016: 1.260
5-year IMPACT FACTOR: 1.506

CiteScore 2016: 1.56

SCImago Journal Rank (SJR) 2016: 0.615
Source Normalized Impact per Paper (SNIP) 2016: 0.470

Online
ISSN
1689-1392
See all formats and pricing
More options …
Volume 16, Issue 2 (Jun 2011)

Rho kinase inhibitors stimulate the migration of human cultured osteoblastic cells by regulating actomyosin activity

Xuejiao Zhang / Cheng Li / Huiling Gao / Hiroaki Nabeka / Tetsuya Shimokawa / Hiroyuki Wakisaka / Seiji Matsuda / Naoto Kobayashi
Published Online: 2011-03-26 | DOI: https://doi.org/10.2478/s11658-011-0006-z

Abstract

We investigated the effects of Rho-associated kinase (ROCK) on migration and cytoskeletal organization in primary human osteoblasts and Saos-2 human osteosarcoma cells. Both cell types were exposed to two different ROCK inhibitors, Y-27632 and HA-1077. In the improved motility assay used in the present study, Y-27632 and HA-1077 significantly increased the migration of both osteoblasts and osteosarcoma cells on plastic in a dose-dependent and reversible manner. Fluorescent images showed that cells of both types cultured with Y-27632 or HA-1077 exhibited a stellate appearance, with poor assembly of stress fibers and focal contacts. Western blotting showed that ROCK inhibitors reduced myosin light chain (MLC) phosphorylation within 5 min without affecting overall myosin light-chain protein levels. Inhibition of ROCK activity is thought to enhance the migration of human osteoblasts through reorganization of the actin cytoskeleton and regulation of myosin activity. ROCK inhibitors may be potentially useful as anabolic agents to enhance the biocompatibility of bone and joint prostheses.

Keywords: Osteoblast; Migration; ROCK inhibitor; Cytoskeleton; Stress fiber; Focal contact; MLC phosphorylation

  • [1] Williams, D.F. On the mechanisms of biocompatibility. Biomaterials 29 (2008) 2941–2953. http://dx.doi.org/10.1016/j.biomaterials.2008.04.023Web of ScienceCrossrefGoogle Scholar

  • [2] Wu, J., Liu, Z.M., Zhao, X.H., Gao, Y., Hu, J. and Gao, B. Improved biological performance of microarc-oxidized low-modulus Ti-24Nb-4Zr-7.9Sn alloy. J. Biomed. Mater. Res. B Appl. Biomater. 92B (2009) 298–306. Google Scholar

  • [3] Park, J.W., Kim, H.K., Kim, Y.J., Jang, J.H., Song, H. and Hanawa, T. Osteoblast response and osseointegration of a Ti-6Al-4V alloy implant incorporating strontium. Acta Biomater. 7 (2010) 2843–2851. http://dx.doi.org/10.1016/j.actbio.2010.01.017Web of ScienceGoogle Scholar

  • [4] Alves, S.F. and Wassall, T. In vitro evaluation of osteoblastic cell adhesion on machined osseointegrated implants. Braz. Oral Res. 23 (2009) 131–136. http://dx.doi.org/10.1590/S1806-83242009000200007CrossrefGoogle Scholar

  • [5] Gurkan, U.A., Cheng, X., Kishore, V., Uquillas, J.A. and Akkus, O. Comparison of morphology, orientation, and migration of tendon derived fibroblasts and bone marrow stromal cells on electrochemically aligned collagen constructs. J. Biomed. Mater. Res. A 94 (2010) 1070–1079. Web of ScienceGoogle Scholar

  • [6] Nakamura, M., Nagai, A., Tanaka, Y., Sekijima, Y. and Yamashita, K. Polarized hydroxyapatite promotes spread and motility of osteoblastic cells. J. Biomed. Mater. Res. A 92 (2009) 783–790. Web of ScienceGoogle Scholar

  • [7] Miranda, L., Carpentier, S., Platek, A., Hussain, N., Gueuning, M.A., Vertommen, D., Ozkan, Y., Sid, B., Hue, L., Courtoy, P.J., Rider, M.H. and Horman, S. AMP-activated protein kinase induces actin cytoskeleton reorganization in epithelial cells. Biochem. Biophys. Res. Commun. 396 (2010) 656–661. http://dx.doi.org/10.1016/j.bbrc.2010.04.151Web of ScienceCrossrefGoogle Scholar

  • [8] Martini, F.J. and Valdeolmillos, M. Actomyosin contraction at the cell rear drives nuclear translocation in migrating cortical interneurons. J. Neurosci. 30 (2010) 8660–8670. http://dx.doi.org/10.1523/JNEUROSCI.1962-10.2010CrossrefWeb of ScienceGoogle Scholar

  • [9] Schwartz, M.A. and Horwitz, A.R. Integrating adhesion, protrusion, and contraction during cell migration. Cell 125 (2006) 1223–1225. http://dx.doi.org/10.1016/j.cell.2006.06.015CrossrefGoogle Scholar

  • [10] Kaibuchi, K., Kuroda, S. and Amano, M. Regulation of the cytoskeleton and cell adhesion by the Rho family GTPases in mammalian cells. Annu. Rev. Biochem. 68 (1999) 459–486. http://dx.doi.org/10.1146/annurev.biochem.68.1.459CrossrefGoogle Scholar

  • [11] Hall, A. Rho GTPases and the actin cytoskeleton. Science 279 (1998) 509–514. http://dx.doi.org/10.1126/science.279.5350.509CrossrefWeb of ScienceGoogle Scholar

  • [12] Gallagher, P.J., Herring, B.P. and Stull, J.T. Myosin light chain kinases. J. Muscle Res. Cell Motil. 18 (1997) 1–16. http://dx.doi.org/10.1023/A:1018616814417CrossrefGoogle Scholar

  • [13] Tokuda, H., Takai, S., Matsushima-Nishiwaki, R., Hanai, Y., Adachi, S., Minamitani, C., Mizutani, J., Otsuka, T. and Kozawa, O. Function of Rhokinase in prostaglandin D2-induced interleukin-6 synthesis in osteoblasts. Prostaglandins Leukot. Essent. Fatty Acids 79 (2008) 41–46. http://dx.doi.org/10.1016/j.plefa.2008.07.004CrossrefGoogle Scholar

  • [14] Harmey, D., Stenbeck, G., Nobes, C.D., Lax, A.J. and Grigoriadis, A.E. Regulation of osteoblast differentiation by Pasteurella multocida toxin (PMT): a role for Rho GTPase in bone formation. J. Bone Miner. Res. 19 (2004) 661–670. http://dx.doi.org/10.1359/JBMR.040105CrossrefGoogle Scholar

  • [15] Kazmers, N.H., Ma, S.A., Yoshida, T. and Stern, P.H. Rho GTPase signaling and PTH 3–34, but not PTH 1–34, maintain the actin cytoskeleton and antagonize bisphosphonate effects in mouse osteoblastic MC3T3-E1 cells. Bone 45 (2009) 52–60. http://dx.doi.org/10.1016/j.bone.2009.03.675CrossrefWeb of ScienceGoogle Scholar

  • [16] Loirand, G., Guerin, P. and Pacaud, P. Rho kinases in cardiovascular physiology and pathophysiology. Circ. Res. 98 (2006) 322–334. http://dx.doi.org/10.1161/01.RES.0000201960.04223.3cCrossrefGoogle Scholar

  • [17] Zohrabian, V.M., Forzani, B., Chau, Z., Murali, R. and Jhanwar-Uniyal, M. Rho/ROCK and MAPK signaling pathways are involved in glioblastoma cell migration and proliferation. Anticancer Res. 29 (2009) 119–123. Google Scholar

  • [18] Hahmann, C. and Schroeter, T. Rho-kinase inhibitors as therapeutics: from pan inhibition to isoform selectivity. Cell. Mol. Life Sci. 67 (2010) 171–177. http://dx.doi.org/10.1007/s00018-009-0189-xWeb of ScienceCrossrefGoogle Scholar

  • [19] Lingor, P., Teusch, N., Schwarz, K., Mueller, R., Mack, H., Bahr, M. and Mueller, B.K. Inhibition of Rho kinase (ROCK) increases neurite outgrowth on chondroitin sulphate proteoglycan in vitro and axonal regeneration in the adult optic nerve in vivo. J. Neurochem. 103 (2007) 181–189. Web of ScienceGoogle Scholar

  • [20] Ikebe, M. and Hartshorne, D.J. Phosphorylation of smooth muscle myosin at two distinct sites by myosin light chain kinase. J. Biol. Chem. 260 (1985) 10027–10031. Google Scholar

  • [21] Totsukawa, G., Yamakita, Y., Yamashiro, S., Hartshorne, D.J., Sasaki, Y. and Matsumura, F. Distinct roles of ROCK (Rho-kinase) and MLCK in spatial regulation of MLC phosphorylation for assembly of stress fibers and focal adhesions in 3T3 fibroblasts. J. Cell Biol. 150 (2000) 797–806. http://dx.doi.org/10.1083/jcb.150.4.797CrossrefGoogle Scholar

  • [22] Gao, S.Y., Li, C.Y., Chen, J., Pan, L., Saito, S., Terashita, T., Saito, K., Miyawaki, K., Shigemoto, K., Mominoki, K., Matsuda, S. and Kobayashi, N. Rho-ROCK signal pathway regulates microtubule-based process formation of cultured podocytes—inhibition of ROCK promoted process elongation. Nephron Exp. Nephrol. 97 (2004) e49–61. http://dx.doi.org/10.1159/000078406CrossrefGoogle Scholar

  • [23] Li, C.Y., Gao, S.Y., Terashita, T., Shimokawa, T., Kawahara, H., Matsuda, S. and Kobayashi, N. In vitro assays for adhesion and migration of osteoblastic cells (Saos-2) on titanium surfaces. Cell Tissue Res. 324 (2006) 369–375. http://dx.doi.org/10.1007/s00441-005-0153-5CrossrefGoogle Scholar

  • [24] Celotti, F., Colciago, A., Negri-Cesi, P., Pravettoni, A., Zaninetti, R. and Sacchi, M.C. Effect of platelet-rich plasma on migration and proliferation of SaOS-2 osteoblasts: role of platelet-derived growth factor and transforming growth factor-beta. Wound Repair Regen. 14 (2006) 195–202. http://dx.doi.org/10.1111/j.1743-6109.2006.00110.xCrossrefGoogle Scholar

  • [25] Tsiridis, E., Upadhyay, N. and Giannoudis, P. Molecular aspects of fracture healing: which are the important molecules? Injury 38Suppl 1 (2007) S11–25. http://dx.doi.org/10.1016/j.injury.2007.02.006Web of ScienceCrossrefGoogle Scholar

  • [26] Klein, M.O., Reichert, C., Koch, D., Horn, S. and Al-Nawas, B. In vitro assessment of motility and proliferation of human osteogenic cells on different isolated extracellular matrix components compared with enamel matrix derivative by continuous single-cell observation. Clin. Oral Implants Res. 18 (2007) 40–45. http://dx.doi.org/10.1111/j.1600-0501.2006.01279.xCrossrefWeb of ScienceGoogle Scholar

  • [27] Jacobs, M., Hayakawa, K., Swenson, L., Bellon, S., Fleming, M., Taslimi, P. and Doran, J. The structure of dimeric ROCK I reveals the mechanism for ligand selectivity. J. Biol. Chem. 281 (2006) 260–268. http://dx.doi.org/10.1074/jbc.M508847200CrossrefGoogle Scholar

  • [28] Tamura, M., Nakao, H., Yoshizaki, H., Shiratsuchi, M., Shigyo, H., Yamada, H., Ozawa, T., Totsuka, J. and Hidaka, H. Development of specific Rhokinase inhibitors and their clinical application. Biochim. Biophys. Acta. 1754 (2005) 245–252. Google Scholar

  • [29] Kim, T.Y., Vigil, D., Der, C.J. and Juliano, R.L. Role of DLC-1, a tumor suppressor protein with RhoGAP activity, in regulation of the cytoskeleton and cell motility. Cancer Metastasis Rev. 28 (2009) 77–83. http://dx.doi.org/10.1007/s10555-008-9167-2Web of ScienceGoogle Scholar

  • [30] Jaganathan, B.G., Ruester, B., Dressel, L., Stein, S., Grez, M., Seifried, E. and Henschler, R. Rho inhibition induces migration of mesenchymal stromal cells. Stem Cells 25 (2007) 1966–1974. http://dx.doi.org/10.1634/stemcells.2007-0167CrossrefWeb of ScienceGoogle Scholar

  • [31] Salhia, B., Rutten, F., Nakada, M., Beaudry, C., Berens, M., Kwan, A. and Rutka, J.T. Inhibition of Rho-kinase affects astrocytoma morphology, motility, and invasion through activation of Rac1. Cancer Res. 65 (2005) 8792–8800. http://dx.doi.org/10.1158/0008-5472.CAN-05-0160CrossrefGoogle Scholar

  • [32] Li, Y., Wu, Y., Wang, Z., Zhang, X.H. and Wu, W.K. Fasudil attenuates lipopolysaccharide-induced acute lung injury in mice through the Rho/Rho kinase pathway. Med. Sci. Monit. 16 (2010) BR112–118. Google Scholar

  • [33] Borensztajn, K., Peppelenbosch, M.P. and Spek, C.A. Coagulation Factor Xa inhibits cancer cell migration via LIMK1-mediated cofilin inactivation. Thromb. Res. 125 (2010) e323–328. http://dx.doi.org/10.1016/j.thromres.2010.02.018Web of ScienceCrossrefGoogle Scholar

  • [34] Koga, T., Awai, M., Tsutsui, J., Yue, B.Y. and Tanihara, H. Rho-associated protein kinase inhibitor, Y-27632, induces alterations in adhesion, contraction and motility in cultured human trabecular meshwork cells. Exp. Eye Res. 82 (2006) 362–370. http://dx.doi.org/10.1016/j.exer.2005.07.006CrossrefGoogle Scholar

  • [35] Honjo, M., Tanihara, H., Kameda, T., Kawaji, T., Yoshimura, N. and Araie, M. Potential role of Rho-associated protein kinase inhibitor Y-27632 in glaucoma filtration surgery. Invest. Ophthalmol. Vis. Sci. 48 (2007) 5549–5557. http://dx.doi.org/10.1167/iovs.07-0878Web of ScienceCrossrefGoogle Scholar

  • [36] Kroening, S., Stix, J., Keller, C., Streiff, C. and Goppelt-Struebe, M. Matrixindependent stimulation of human tubular epithelial cell migration by Rho kinase inhibitors. J. Cell Physiol. 223 (2010) 703–712. Web of ScienceGoogle Scholar

  • [37] Tripathi, B.K. and Zelenka, P.S. Cdk5-dependent regulation of Rho activity, cytoskeletal contraction, and epithelial cell migration via suppression of Src and p190RhoGAP. Mol. Cell. Biol. 29 (2009) 6488–6499. http://dx.doi.org/10.1128/MCB.01098-09Web of ScienceCrossrefGoogle Scholar

  • [38] Kanda, K., Sobue, K. and Kakiuchi, S. Phosphorylation of myosin light chain and the actin-activated ATPase activity of adrenal medullary myosin. J. Biochem. 97 (1985) 961–964. Google Scholar

  • [39] Shutova, M.S., Alexandrova, A.Y. and Vasiliev, J.M. Regulation of polarity in cells devoid of actin bundle system after treatment with inhibitors of myosin II activity. Cell. Motil. Cytoskeleton. 65 (2008) 734–746. http://dx.doi.org/10.1002/cm.20295Web of ScienceCrossrefGoogle Scholar

  • [40] Palazzo, A.F., Cook, T.A., Alberts, A.S. and Gundersen, G.G. mDia mediates Rho-regulated formation and orientation of stable microtubules. Nat. Cell Biol. 3 (2001) 723–729. http://dx.doi.org/10.1038/35087035CrossrefGoogle Scholar

  • [41] Niggli, V., Schmid, M. and Nievergelt, A. Differential roles of Rho-kinase and myosin light chain kinase in regulating shape, adhesion, and migration of HT1080 fibrosarcoma cells. Biochem. Biophys. Res. Commun. 343 (2006) 602–608. http://dx.doi.org/10.1016/j.bbrc.2006.03.022CrossrefGoogle Scholar

  • [42] Smith, A., Bracke, M., Leitinger, B., Porter, J.C. and Hogg, N. LFA-1-induced T cell migration on ICAM-1 involves regulation of MLCKmediated attachment and ROCK-dependent detachment. J. Cell Sci. 116 (2003) 3123–3133. http://dx.doi.org/10.1242/jcs.00606Google Scholar

  • [43] Webb, D.J., Donais, K., Whitmore, L.A., Thomas, S.M., Turner, C.E., Parsons, J.T. and Horwitz, A.F. FAK-Src signalling through paxillin, ERK and MLCK regulates adhesion disassembly. Nat. Cell Biol. 6 (2004) 154–161. http://dx.doi.org/10.1038/ncb1094CrossrefGoogle Scholar

  • [44] Axmann, R., Bohm, C., Kronke, G., Zwerina, J., Smolen, J. and Schett, G. Inhibition of interleukin-6 receptor directly blocks osteoclast formation in vitro and in vivo. Arthritis Rheum. 60 (2009) 2747–2756. http://dx.doi.org/10.1002/art.24781CrossrefGoogle Scholar

About the article

Published Online: 2011-03-26

Published in Print: 2011-06-01


Citation Information: Cellular and Molecular Biology Letters, ISSN (Online) 1689-1392, DOI: https://doi.org/10.2478/s11658-011-0006-z.

Export Citation

© 2011 Versita Warsaw. This work is licensed under the Creative Commons Attribution-NonCommercial-NoDerivatives 3.0 License. BY-NC-ND 3.0

Citing Articles

Here you can find all Crossref-listed publications in which this article is cited. If you would like to receive automatic email messages as soon as this article is cited in other publications, simply activate the “Citation Alert” on the top of this page.

[2]
Brandon D. Riehl, Jeong Soon Lee, Ligyeom Ha, Il Keun Kwon, Jung Yul Lim, and Damian Christopher Genetos
PLOS ONE, 2017, Volume 12, Number 2, Page e0171857
[3]
Agnieszka Strzelecka-Kiliszek, Saida Mebarek, Monika Roszkowska, René Buchet, David Magne, and Slawomir Pikula
Biochimica et Biophysica Acta (BBA) - General Subjects, 2017, Volume 1861, Number 5, Page 1009
[4]
Lijuan Kan, Aubrie Smith, Miao Chen, Benjamin T. Ledford, Huimin Fan, Zhongmin Liu, Jia-Qiang He, and Gangjian Qin
PLOS ONE, 2015, Volume 10, Number 12, Page e0144513
[5]
Shingo Shimada, Kenji Wakayama, Moto Fukai, Tsuyoshi Shimamura, Takahisa Ishikawa, Daisuke Fukumori, Maki Shibata, Kenichiro Yamashita, Taichi Kimura, Satoru Todo, Ikuroh Ohsawa, and Akinobu Taketomi
Artificial Organs, 2016, Volume 40, Number 12, Page 1128
[6]
Xun Xu, Weiwei Wang, Karl Kratz, Liang Fang, Zhengdong Li, Andreas Kurtz, Nan Ma, and Andreas Lendlein
Advanced Healthcare Materials, 2014, Volume 3, Number 12, Page 1991
[7]
C Zucchini, M C Manara, R S Pinca, P De Sanctis, C Guerzoni, M Sciandra, P-L Lollini, G Cenacchi, P Picci, L Valvassori, and K Scotlandi
Oncogene, 2014, Volume 33, Number 15, Page 1912
[8]
A.A. Kramerov, K. Ahmed, and A.V. Ljubimov
Journal of Cellular Biochemistry, 2012, Volume 113, Number 9, Page 2948
[9]
Cunshuan Xu, Yanjie Yang, Junying Yang, Xiaoguang Chen, and Gaiping Wang
Cellular and Molecular Biology Letters, 2012, Volume 17, Number 2
[10]
Eunju O, Seung Woo LEE, Hyun-Sun LEE, Hee-Suk LIM, Hyun-Young AHN, Jong-Chul SHIN, Yonggoo KIM, and Young Ae JOE
Bioscience, Biotechnology, and Biochemistry, 2012, Volume 76, Number 1, Page 172
[11]
Masanori Ichida, Yoshihiro Yui, Kiyoko Yoshioka, Takaaki Tanaka, Toru Wakamatsu, Hideki Yoshikawa, and Kazuyuki Itoh
FEBS Letters, 2011, Volume 585, Number 24, Page 4018

Comments (0)

Please log in or register to comment.
Log in