ROCK inhibitor prevents the dedifferentiation of human articular chondrocytes

Matsumoto, Emi; Furumatsu, Takayuki; Kanazawa, Tomoyuki; Tamura, Masanori; Ozaki, Toshifumi

doi:10.1016/j.bbrc.2012.02.127

Cited by 37 publications

(50 citation statements)

References 35 publications

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“…Attached cells (passage 0) were maintained with Dulbecco's modified Eagle's medium (DMEM, Wako, Osaka, Japan) containing 10% fetal bovine serum (HyClone, South Logan, UT), and penicillin/streptomycin (Sigma). Monolayer-cultured chondrocytes between passage 1 and 7 were used (19). ROCK inhibition was performed by a ROCK-specific inhibitor Y-27632 (Wako).…”

Section: Cells and Cell Culturementioning

confidence: 99%

“…Monolayer culturing of articular chondrocytes leads to dedifferentiation whereby the cells acquire a fibroblastic morphology and lose their chondrocytic properties (15,16). We previously reported that inhibition of the Rho family GTPase ROCK stimulates SOX9-dependent COL2A1 expression and prevents dedifferentiation of human articular chondrocytes (19). Rho GTPases, including RhoA and its downstream effector ROCK, organize the actin cytoskeleton, thereby regulating various biological functions such as cell motility, differentiation and gene expression (20).…”

Section: Introductionmentioning

confidence: 98%

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ROCK inhibition enhances aggrecan deposition and suppresses matrix metalloproteinase-3 production in human articular chondrocytes

Furumatsu

Matsumoto-Ogawa

Tanaka

et al. 2014

Connective Tissue Research

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Homeostasis of articular cartilage is maintained by a balance between catabolism and anabolism. Matrix metalloproteinase-3 (MMP-3) catabolism of cartilaginous extracellular matrix (ECM), including aggrecan (AGN), is an important factor in osteoarthritis progression. We previously reported that inhibition of Rho-associated coiled-coil forming kinase (ROCK), an effector of Rho family GTPases, activates the chondrogenic transcription factor SRY-type high-mobility-group box (SOX) 9 and prevents dedifferentiation of monolayer-cultured chondrocytes. We hypothesized that ROCK inhibition prevents chondrocyte dedifferentiation by altering the transcriptional balance between MMP-3 and AGN. Normal human articular chondrocytes were cultured in the presence or absence of ROCK inhibitor (ROCKi, Y-27632). Expression of MMP-3 and AGN during monolayer cultivation was assessed by quantitative real-time PCR and western blot analysis. Chondrogenic redifferentiation potential of ROCKi-treated chondrocytes was evaluated by immunohistological analysis of pellet cultures. ROCKi treatment suppressed MMP-3 expression in monolayer- and pellet-cultured chondrocytes but increased AGN expression. Chromatin immunoprecipitation revealed that the association between transcription factors E26 transformation specific (ETS)-1 and SOX9 and their target genes MMP-3 and AGN, respectively, was affected by ROCKi treatment. ROCKi decreased the association between ETS-1 and its binding sites on the MMP-3 promoter, whereas ROCKi promoted the interaction between SOX9 and the AGN promoter. Our results suggest that ROCK inhibition may have an important role in modulating the balance between degradation and synthesis of cartilaginous ECM, a finding that may facilitate development of techniques to prepare differentiated chondrocytes for cartilage regeneration therapy.

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Section: Cells and Cell Culturementioning

confidence: 99%

Section: Introductionmentioning

confidence: 98%

ROCK inhibition enhances aggrecan deposition and suppresses matrix metalloproteinase-3 production in human articular chondrocytes

Furumatsu

Matsumoto-Ogawa

Tanaka

et al. 2014

Connective Tissue Research

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“…Normal articular cartilage was obtained at notchplasty in patients (13,18,19, and 21 years of age) undergoing anterior cruciate ligament reconstructions (n 0 4). Unnecessary parts of loose fragments in knee OCD (15,15,15, and 21 years of age) were obtained at arthroscopy (n 0 4).…”

Section: Tissues Cells and Cell Culturementioning

confidence: 99%

“…Proteoglycans in pellets were observed with safranin O staining. Redifferentiation potential was assessed by the Bern pellet histology score (range, 0-9) [18,19]. To assess the effect of tissuederived mesenchymal stem cells (MSCs), we performed in vitro differentiation analysis for the other mesenchymal lineages.…”

Section: In Vitro Differentiation and Histological Analysesmentioning

confidence: 99%

Comparison between normal and loose fragment chondrocytes in proliferation and redifferentiation potential

Sakata

Furumatsu

Miyazawa

et al. 2012

International Orthopaedics (SICOT)

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Purpose Loose fragments in osteochondritis dissecans (OCD) of the knee require internal fixation. On the other hand, loose fragments derived from spontaneous osteonecrosis of the knee (SONK) are usually removed. However, the difference in healing potential between OCD-and SONK-related loose fragments has not been elucidated. In this study, we investigated proliferative activity and redifferentiation potential of normal cartilage-derived and loose fragment-derived chondrocytes. Methods Cells were prepared from normal articular cartilages and loose fragment cartilages derived from knee OCD and SONK. Cellular proliferation was compared. Redifferentiation ability of pellet-cultured chondrocytes was assessed by real-time PCR analyses. Mesenchymal differentiation potential was investigated by histological analyses. Positive ratio of a stem cell marker CD166 was evaluated in each cartilaginous tissue. Results Normal and OCD chondrocytes showed a higher proliferative activity than SONK chondrocytes. Chondrogenic pellets derived from normal and OCD chondrocytes produced a larger amount of safranin O-stained proteoglycans compared with SONK-derived pellets. Expression of chondrogenic marker genes was inferior in SONK pellets. The CD166-positive ratio was higher in normal cartilages and OCD loose fragments than in SONK loose fragments. Conclusions The OCD chondrocytes maintained higher proliferative activity and redifferentiation potential compared with SONK chondrocytes. Our results suggest that chondrogenic properties of loose fragment-derived cells and the amount of CD166-positive cells may affect the repair process of osteochondral defects.

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“…It is well-established that chondrocytes maintain their phenotype when existing in a spherical (rounded) shape, whereas the fibroblast-like appearance with elongated actin stress fiber structure indicated the dedifferentiation of the cells. [32][33][34][35][36] The elongated actin stress fibers exhibited along the chondrocytes were, however, gradually diminished when the cells grew on the scaffolds blended with PHBV, that is, PCL/PHBV:75/25, PCL/PHBV:50/50, PCL/PHBV:25/75. It was also noted that the porcine chondrocytes could maintain their chondrogenic phenotype more efficiently when being cultured on the PCL/PHBV scaffolds blended with higher PHBV contents.…”

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confidence: 99%

PCL/PHBV blended three dimensional scaffolds fabricated by fused deposition modeling and responses of chondrocytes to the scaffolds

Kosorn

Sakulsumbat

Uppanan

et al. 2016

J. Biomed. Mater. Res.

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In this study, poly(ε-caprolactone)/poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PCL/PHBV) blended porous scaffolds were fabricated by fused deposition modeling (FDM). PCL/PHBV filaments, initially prepared at different weight ratios, that is, 100/0, 75/25, 50/50, and 25/75, were fabricated by the lay-down pattern of 0/90/45/135° to obtain scaffolds with dimension of 6.0 × 6.0 × 2.5 mm and average filament diameters and channel sizes in the ranges of 370-390 µm and 190-210 µm, respectively. To enhance the surface hydrophilicity of the materials, the scaffolds were subsequently subjected to a low pressure oxygen plasma treatment. The untreated and plasma-treated scaffolds were comparatively characterized, in terms of surface properties, mechanical strength, and biological properties. From SEM, AFM, water contact angle, and XPS results, the surface roughness, wettability, and hydrophilicity of the blended scaffolds were found to be enhanced after plasma treatment, while the compressive strength of the scaffolds was scarcely changed. It was, however, found to increase with an increasing content of PHBV incorporated. The porcine chondrocytes exhibited higher proliferative capacity and chondrogenic potential when being cultured on the scaffolds with greater PHBV contents, especially when they were plasma-treated. The PCL/PHBV scaffolds were proven to possess good physical, mechanical, and biological properties that could be appropriately used in articular cartilage regeneration. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 105B: 1141-1150, 2017.

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ROCK inhibitor prevents the dedifferentiation of human articular chondrocytes

Cited by 37 publications

References 35 publications

ROCK inhibition enhances aggrecan deposition and suppresses matrix metalloproteinase-3 production in human articular chondrocytes

ROCK inhibition enhances aggrecan deposition and suppresses matrix metalloproteinase-3 production in human articular chondrocytes

Comparison between normal and loose fragment chondrocytes in proliferation and redifferentiation potential

PCL/PHBV blended three dimensional scaffolds fabricated by fused deposition modeling and responses of chondrocytes to the scaffolds

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