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1.
J Oral Biosci ; 66(1): 68-75, 2024 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-38266705

RESUMO

OBJECTIVES: Cellular differentiation is based on the effects of various growth factors. Transforming growth factor (TGF)-ß1 plays a pivotal role in inducing osteogenic differentiation of mesenchymal stem cells (MSCs). In this study, we investigated the influence of connective tissue growth factor (CTGF), known to function synergistically with TGF-ß1, on osteogenic differentiation in MSCs. METHODS: UE7T-13 cells were treated with TGF-ß1 and/or CTGF. Subsequently, protein levels of intracellular signaling pathway molecules were determined through western blot analysis. The mRNA expression levels of osteogenic differentiation markers were investigated using reverse transcription-quantitative polymerase chain reaction. Bone matrix mineralization was evaluated through alizarin red staining. RESULTS: Co-treatment with TGF-ß1 and CTGF resulted in the suppression of TGF-ß1-induced phosphorylation of extracellular signal-regulated kinase 1/2, an intracellular signaling pathway molecule in MSCs, while significantly enhancing the phosphorylation of p38 mitogen-activated protein kinase (MAPK). In MSCs, co-treatment with CTGF and TGF-ß1 led to increased expression levels of alkaline phosphatase and type I collagen, markers of osteogenic differentiation induced by TGF-ß1. Osteopontin expression was observed only after TGF-ß1 and CTGF co-treatment. Notably, bone sialoprotein and osteocalcin were significantly upregulated by treatment with CTGF alone. Furthermore, CTGF enhanced the TGF-ß1-induced mineralization in MSCs, with complete suppression observed after treatment with a p38 MAPK inhibitor. CONCLUSIONS: CTGF enhances TGF-ß1-induced osteogenic differentiation and subsequent mineralization in MSCs by predominantly activating the p38 MAPK-dependent pathway.


Assuntos
Células-Tronco Mesenquimais , Proteína Quinase 14 Ativada por Mitógeno , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/farmacologia , Fator de Crescimento Transformador beta1/farmacologia , Fator de Crescimento do Tecido Conjuntivo/genética , Fator de Crescimento do Tecido Conjuntivo/metabolismo , Fator de Crescimento do Tecido Conjuntivo/farmacologia , Osteogênese , Diferenciação Celular , Células-Tronco Mesenquimais/metabolismo
2.
Mol Biol Rep ; 50(2): 1595-1602, 2023 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-36526849

RESUMO

BACKGROUND: Temporomandibular joint osteoarthritis (TMJ-OA) causes cartilage degeneration, bone cavitation, and fibrosis of the TMJ. However, the mechanisms underlying the fibroblast-like synoviocyte (FLS)-mediated inflammatory activity in TMJ-OA remain unclear. METHODS AND RESULTS: Reverse transcription-quantitative polymerase chain reaction analysis revealed that the P2Y1, P2Y12, and P2Y13 purinergic receptor agonist adenosine 5'-diphosphate (ADP) significantly induces monocyte chemotactic protein 1 (MCP-1)/ C-C motif chemokine ligand 2 (CCL2) expression in the FLS1 synovial cell line. In contrast, the uracil nucleotide UTP, which is a P2Y2 and P2Y4 agonist, has no significant effect on MCP-1/CCL2 production in FLS1 cells. In addition, the P2Y13 antagonist MRS 2211 considerably decreases the expression of ADP-induced MCP-1/CCL2, whereas ADP stimulation enhances extracellular signal-regulated kinase (ERK) phosphorylation. Moreover, it was found that the mitogen-activated protein kinase/ERK kinase (MEK) inhibitor U0126 reduces ADP-induced MCP-1/CCL2 expression. CONCLUSION: ADP enhances MCP-1/CCL2 expression in TMJ FLSs via P2Y13 receptors in an MEK/ERK-dependent manner, thus resulting in inflammatory cell infiltration in the TMJ. Collectively, the findings of this study contribute to a partial clarification of the signaling pathway underlying the development of inflammation in TMJ-OA and can help identify potential therapeutic targets for suppressing ADP-mediated purinergic signaling in this disease.


Assuntos
Receptores Purinérgicos P2 , Sinoviócitos , Camundongos , Animais , Quimiocina CCL2/genética , Quimiocina CCL2/metabolismo , MAP Quinases Reguladas por Sinal Extracelular , Difosfatos , Sinoviócitos/metabolismo , Ligantes , Receptores Purinérgicos P2/metabolismo , Quinases de Proteína Quinase Ativadas por Mitógeno , Articulação Temporomandibular , Fibroblastos/metabolismo , Adenosina , Difosfato de Adenosina/farmacologia , Difosfato de Adenosina/metabolismo , Células Cultivadas
3.
J Oral Biosci ; 65(1): 97-103, 2023 03.
Artigo em Inglês | MEDLINE | ID: mdl-36584898

RESUMO

OBJECTIVES: Temporomandibular joint osteoarthritis (TMJ-OA) is a multifactorial disease caused by inflammation and oxidative stress. It has been hypothesized that mechanical stress-induced injury of TMJ tissues induces the generation of reactive oxygen species (ROS), such as hydroxyl radical (OH∙), in the synovial fluid (SF). In general, the overproduction of ROS contributes to synovial inflammation and dysfunction of the subchondral bone in OA. However, the mechanism by which ROS-injured synoviocytes recruit inflammatory cells to TMJ-OA lesions remains unclear. METHODS: Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) was performed to evaluate the mRNA expression of chemoattractant molecules. The phosphorylation levels of intracellular signaling molecules were evaluated using western blot analysis. RESULTS: Hydrogen peroxide (H2O2) treatment significantly promoted mRNA expression of neutrophil chemoattractant CXCL15/Lungkine in a dose-dependent manner (100-500 µM) in fibroblast-like synoviocytes (FLSs) derived from mouse TMJ. H2O2 (500 µM) significantly upregulated the phosphorylation of extracellular signal-regulated kinase (ERK)1 and ERK2 in FLSs. Intriguingly, the mitogen-activated protein (MAP)/ERK kinase (MEK) inhibitor U0126 (10 µM) nullified H2O2-induced increase in CXCL15/Lungkine mRNA expression. Additionally, H2O2 (500 µM) administration significantly upregulated OH∙ production in FLSs, as assessed by live-cell permeant fluorescent probe targeted against OH∙ under fluorescence microscopy. Furthermore, the ROS inhibitor N-acetyl-l-cysteine (5 mM) partially but significantly reversed H2O2-mediated phosphorylation of ERK1/2. CONCLUSIONS: H2O2-induced oxidative stress promoted the expression of CXCL15/Lungkine mRNA in a MEK/ERK-dependent manner in mouse TMJ-derived FLSs, suggesting that FLSs recruit neutrophils to TMJ-OA lesions through the production of CXCL15/Lungkine and exacerbate the local inflammatory response.


Assuntos
Osteoartrite , Sinoviócitos , Animais , Camundongos , Fatores Quimiotáticos/metabolismo , Fibroblastos/metabolismo , Fibroblastos/patologia , Peróxido de Hidrogênio/efeitos adversos , Peróxido de Hidrogênio/metabolismo , Inflamação/induzido quimicamente , Inflamação/metabolismo , Inflamação/patologia , Quinases de Proteína Quinase Ativadas por Mitógeno/metabolismo , Osteoartrite/metabolismo , Osteoartrite/patologia , Estresse Oxidativo , Espécies Reativas de Oxigênio/efeitos adversos , Espécies Reativas de Oxigênio/metabolismo , RNA Mensageiro/metabolismo , Sinoviócitos/metabolismo , Sinoviócitos/patologia , Articulação Temporomandibular/metabolismo , Articulação Temporomandibular/patologia
4.
Exp Ther Med ; 20(3): 1967-1974, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32782506

RESUMO

Osteoarthritis (OA)-related fibrosis is a possible cause of temporomandibular joint (TMJ) stiffness. However, the molecular mechanisms underlying the fibrogenic activity in fibroblast-like synoviocytes (FLSs) remain to be clarified. The present study examined the effects of receptor tyrosine kinase (RTK) ligands, such as fibroblast growth factor (FGF)-1 and epidermal growth factor (EGF), on myofibroblastic differentiation of the FLS cell line FLS1, which is derived from the mouse TMJ. The present study revealed that both FGF-1 and EGF dose-dependently suppressed the expression of the myofibroblast (MF) markers, including α-smooth muscle actin (α-SMA) and type I collagen, in FLS1 cells. Additionally, both FGF-1 and EGF activated extracellular signal-regulated kinase (ERK) in FLS1 cells. In addition, the mitogen-activated protein kinase (MAPK)/ERK kinase (MEK) inhibitor U0126 abrogated the FGF-1- and EGF-mediated suppression of MF marker expression. On the other hand, inflammatory cytokines, such as interleukin-1ß and tumor necrosis factor-α, also suppressed the expression of MF markers in FLS1 cells. Importantly, U0126 abrogated the inflammatory cytokine-mediated suppression of MF marker expression. Interestingly, RTK ligands and inflammatory cytokines additively suppressed the expression of type I collagen. These results suggested that RTK ligands and inflammatory cytokines cooperatively inhibited the fibrogenic activity in FLSs derived from the TMJ in a MEK/ERK-dependent manner. The present findings partially clarify the molecular mechanisms underlying the development of OA-related fibrosis in the TMJ and may aid in identifying therapeutic targets for this condition. Additionally, FGF-1 and EGF could be therapeutically utilized to prevent OA-related fibrosis around the inflammatory TMJ.

5.
Int J Mol Med ; 42(3): 1484-1494, 2018 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-29901090

RESUMO

Mechanosensitive (MS) neurons in the periodontal ligament (PDL) pass information to the trigeminal ganglion when excited by mechanical stimulation of the tooth. During occlusal tooth trauma of PDL tissues, MS neurons are injured, resulting in atrophic neurites and eventual degeneration of MS neurons. Nerve growth factor (NGF), a neurotrophic factor, serves important roles in the regeneration of injured sensory neurons. In the present study, the effect of pro­inflammatory cytokines, including interleukin 1ß (IL­1ß) and tumor necrosis factor α (TNF­α), on transforming growth factor ß1 (TGF­ß1)­induced NGF expression was evaluated in rat PDL­derived SCDC2 cells. It was observed that TGF­ß1 promoted NGF expression via Smad2/3 and p38 mitogen­activated protein kinase (MAPK) activation. IL­1ß and TNF­α suppressed the TGF­ß1­induced activation of Smad2/3 and p38 MAPK, resulting in the abrogation of NGF expression. NGF secreted by TGF­ß1­treated SCDC2 cells promoted neurite extension and the expression of tyrosine hydroxylase, a rate­limiting enzyme in dopamine synthesis in rat pheochromocytoma PC12 cells. These results suggested that pro­inflammatory cytokines suppressed the TGF­ß­mediated expression of NGF in PDL­derived fibroblasts through the inactivation of TGF­ß­induced Smad2/3 and p38 MAPK signaling, possibly resulting in the disturbance of the regeneration of injured PDL neurons.


Assuntos
Fibroblastos/metabolismo , Interleucina-1beta/farmacologia , Fator de Crescimento Neural/metabolismo , Ligamento Periodontal/citologia , Proteínas Smad/metabolismo , Fator de Crescimento Transformador beta/farmacologia , Fator de Necrose Tumoral alfa/farmacologia , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Trifosfato de Adenosina/análogos & derivados , Trifosfato de Adenosina/farmacologia , Animais , Fibroblastos/efeitos dos fármacos , Humanos , Fator de Crescimento Neural/genética , Neuritos/efeitos dos fármacos , Neuritos/metabolismo , Células PC12 , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos , Receptores de Fatores de Crescimento Transformadores beta/metabolismo , Transdução de Sinais/efeitos dos fármacos , Tirosina 3-Mono-Oxigenase/metabolismo
6.
Mol Med Rep ; 17(3): 3448-3454, 2018 03.
Artigo em Inglês | MEDLINE | ID: mdl-29257332

RESUMO

Surface pre-reacted glass­ionomer (S­PRG)-containing dental materials, including composite and coating resins have been used for the restoration and/or prevention of dental cavities. S­PRG is known to have the ability to release aluminum, boron, fluorine, silicon, and strontium ions. Aluminum ions are known to be inhibitors whereas boron, fluorine, silicon, and strontium ions are known to be promoters of mineralization, via osteoblasts. However, it remains to be clarified how an aqueous eluate obtained from S­PRG containing these ions affects the ability of mesenchymal stem cells (MSCs), which are known to be present in dental pulp and bone marrow, to differentiate into osteogenic cell types. The present study demonstrated that 200­ to 1,000­fold­diluted aqueous eluates obtained from S­PRG significantly upregulated the mRNA expression level of the osteogenic differentiation marker alkaline phosphatase in human MSCs (hMSCs) without exhibiting the cytotoxic effect. In addition, the 500­ to 1,000­fold­diluted aqueous eluates obtained from S­PRG significantly and clearly promoted mineralization of the extracellular matrix of hMSCs. It was additionally demonstrated that hMSCs cultured on the cured resin composites containing S­PRG fillers exhibited osteogenic differentiation in direct correlation with the weight percent of S­PRG fillers. These results strongly suggested that aqueous eluates of S­PRG fillers promoted hard tissue formation by hMSCs, implicating that resins containing S­PRG may act as a useful biomaterial to cover accidental exposure of dental pulp.


Assuntos
Resinas Acrílicas/farmacologia , Diferenciação Celular/efeitos dos fármacos , Dióxido de Silício/farmacologia , Resinas Acrílicas/química , Fosfatase Alcalina/genética , Fosfatase Alcalina/metabolismo , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Materiais Dentários/química , Matriz Extracelular/metabolismo , Humanos , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/metabolismo , Osteoblastos/citologia , Osteoblastos/metabolismo , Osteogênese/efeitos dos fármacos , RNA Mensageiro/metabolismo , Dióxido de Silício/química , Regulação para Cima/efeitos dos fármacos , Água/química
7.
Exp Cell Res ; 358(2): 411-420, 2017 09 15.
Artigo em Inglês | MEDLINE | ID: mdl-28712928

RESUMO

Immunosuppressive/anti-inflammatory macrophage (Mφ), M2-Mφ that expressed the typical M2-Mφs marker, CD206, and anti-inflammatory cytokine, interleukin (IL)-10, is beneficial and expected tool for the cytotherapy against inflammatory diseases. Here, we demonstrated that bone marrow-derived lineage-positive (Lin+) blood cells proliferated and differentiated into M2-Mφs by cooperation with the bone marrow-derived mesenchymal stem cells (MSCs) under hypoxic condition: MSCs not only promoted proliferation of undifferentiated M2-Mφs, pre-M2-Mφs, in the Lin+ fraction via a proliferative effect of the MSCs-secreted macrophage colony-stimulating factor, but also promoted M2-Mφ polarization of the pre-M2-Mφs through cell-to-cell contact with the pre-M2-Mφs. Intriguingly, an inhibitor for intercellular adhesion molecule (ICAM)-1 receptor/lymphocyte function-associated antigen (LFA)-1, Rwj50271, partially suppressed expression of CD206 in the Lin+ blood cells but an inhibitor for VCAM-1 receptor/VLA-4, BIO5192, did not, suggesting that the cell-to-cell adhesion through LFA-1 on pre-M2-Mφs and ICAM-1 on MSCs was supposed to promoted the M2-Mφ polarization. Thus, the co-culture system consisting of bone marrow-derived Lin+ blood cells and MSCs under hypoxic condition was a beneficial supplier of a number of M2-Mφs, which could be clinically applicable to inflammatory diseases.


Assuntos
Medula Óssea/metabolismo , Comunicação Celular , Ativação de Macrófagos/fisiologia , Macrófagos/metabolismo , Células-Tronco Mesenquimais/citologia , Animais , Anti-Inflamatórios/farmacologia , Diferenciação Celular/imunologia , Hipóxia Celular , Células Cultivadas , Técnicas de Cocultura , Macrófagos/imunologia , Camundongos , Molécula 1 de Adesão de Célula Vascular/metabolismo
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