title: ROCK inhibitor prevents the dedifferentiation of human articular chondrocytes.
abstract: Chondrocytes lose their chondrocytic phenotypes in vitro. The Rho family GTPase ROCK, involved in organizing the actin cytoskeleton, modulates the differentiation status of chondrocytic cells. However, the optimum method to prepare a large number of un-dedifferentiated chondrocytes is still unclear. In this study, we investigated the effect of ROCK inhibitor (ROCKi) on the chondrogenic property of monolayer-cultured articular chondrocytes. Human articular chondrocytes were subcultured in the presence or absence of ROCKi (Y-27632). The expression of chondrocytic marker genes such as SOX9 and COL2A1 was assessed by quantitative real-time PCR analysis. Cellular morphology and viability were evaluated. Chondrogenic redifferentiation potential was examined by a pellet culture procedure. The expression level of SOX9 and COL2A1 was higher in ROCKi-treated chondrocytes than in untreated cells. Chondrocyte morphology varied from a spreading form to a round shape in a ROCKi-dependent manner. In addition, ROCKi treatment stimulated the proliferation of chondrocytes. The deposition of safranin O-stained proteoglycans and type II collagen was highly detected in chondrogenic pellets derived from ROCKi-pretreated chondrocytes. Our results suggest that ROCKi prevents the dedifferentiation of monolayer-cultured chondrocytes, and may be a useful reagent to maintain chondrocytic phenotypes in vitro for chondrocyte-based regeneration therapy.
authors.affiliation: Department of Orthopaedic Surgery, Science of Functional Recovery and Reconstruction, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical Sciences, 2-5-1 Shikatacho, Kitaku, Okayama 700-8558, Japan.
authors.forename: Emi
authors.initials: E
authors.lastname: Matsumoto
authors.forename: Takayuki
authors.initials: T
authors.lastname: Furumatsu
authors.forename: Tomoko
authors.initials: T
authors.lastname: Kanazawa
authors.forename: Masanori
authors.initials: M
authors.lastname: Tamura
authors.forename: Toshifumi
authors.initials: T
authors.lastname: Ozaki
mesh_terms: D000577:Amides / Q000494:pharmacology*; D002454:Cell Differentiation / Q000187:drug effects*; D002478:Cells, Cultured; D019902:Chondrocytes / Q000166:cytology / Q000187:drug effects*; D024043:Collagen Type II / Q000378:metabolism; D004742:Enhancer Elements, Genetic; D015870:Gene Expression / Q000187:drug effects; D006801:Humans; D007596:Joints / Q000166:cytology*; D011401:Promoter Regions, Genetic; D011509:Proteoglycans / Q000378:metabolism; D011725:Pyridines / Q000494:pharmacology*; D012038:Regeneration / Q000187:drug effects*; D055758:SOX9 Transcription Factor / Q000378:metabolism; D020741:rho GTP-Binding Proteins / Q000037:antagonists & inhibitors*
chemical_list: D000577:Amides; C499177:COL2A1 protein, human; D024043:Collagen Type II; D011509:Proteoglycans; D011725:Pyridines; D055758:SOX9 Transcription Factor; C527932:SOX9 protein, human; C108830:Y 27632; D020741:rho GTP-Binding Proteins