A740003 Can Reverse Chondrocyte Apoptosis and Prevent Extracellular Matrix Degradation by NF-KB Pathway in Facet Joint Osteoarthritis

Abstract Facet joint osteoarthritis (FJOA) is one of the common causes of low back pain, but the molecular mechanism is still unclear. Previous studies have found that P2X7 receptor has been proved to play an important role in skeletal and joint diseases. The purpose of this study was to explore the role of A740003, selective P2X7R antagonist, in the development of FJOA. Our study found that A740003 can inhibit the expression of P2X7R in OA chondrocytes. It can lead to anti-inflammatory and anti-apoptotic effects in primary chondrocytes by IL-1β/BzATP. Our results imply that decreased P2X7R can reverse chondrocyte apoptosis and prevent extracellular matrix degradation by NF-KB pathway. Moreover, in our present work, we found that A740003 inhibit the abrrently activation of NF-KB pathway by preventing the activated P65 translocation to nucleus. Our results indicate that P2X7R is a therapeutic target for the treatment of FJOA, and that A740003 could be a therapeutic candidate for this clinical application.

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Paeonol Ameliorates Inflammation and Cartilage of Chondrocytes in Osteoarthritis by Upregulating SIRT1

10.21203/rs.3.rs-104024/v1 ◽

2020 ◽

Author(s):

Peng Shang ◽

Ying Liu ◽

Junqing Jia

Keyword(s):

Extracellular Matrix ◽

Caspase 3 ◽

Type Ii Collagen ◽

Blue Staining ◽

Chondrocyte Apoptosis ◽

Matrix Degradation ◽

Extracellular Matrix Degradation ◽

Tnf Α ◽

And Cartilage

Abstract Objective: To explore the possible role of paeonol on chondrocyte inflammation and cartilage protection in osteoarthritis (OA).Methods: Primary chondrocytes were isolated from rat stifle joints, and were identified through toluidine blue staining and immunofluorescence staining of type II collagen. The chondrocytes were transfected with sh-SIRT1 or/and paeonol (0, 20, 50, 100, 200, 1000 mg/L) before OA modeling induced by IL-1β. ELISA determined the expressions of TNF-α, IL-17 and IL-6, and apoptotic rate was examined by flow cytometry. qRT-PCR and Western blot quantified the expressions of MMP-1, MMP-3, MMP-13, TIMP-1, cleaved-caspase-3, Bax, Bcl-2, and the proteins related to NF-κB pathway.Results: Increases in TNF-α, IL-17, IL-6, MMP-1, MMP-3 and MMP-13 and decrease in TIMP-1 were found in IL-1β stimulated chondrocytes. The apoptotic rate as well as the expressions of cleaved-caspase-3 and Bax was up-regulated, and Bcl-2 expression was suppressed in response to IL-1β treatment. NF-κB pathway was activated in IL-1β-stimulated chondrocytes. Paeonol enhanced SIRT1 expression to inactivate NF-κB pathway, thus ameliorating the secretion of inflammatory cytokines, extracellular matrix degradation and chondrocyte apoptosis.Conclusion: Paeonol inhibits IL-1β induced inflammation and extracellular matrix degradation in chondrocytes through up-regulating SIRT1 and suppressing NF-κB pathway.

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The phosphatase Shp1 interacts with and dephosphorylates cortactin to inhibit invadopodia function

Cell Communication and Signaling ◽

10.1186/s12964-021-00747-6 ◽

2021 ◽

Vol 19 (1) ◽

Author(s):

Alessia Varone ◽

Chiara Amoruso ◽

Marcello Monti ◽

Manpreet Patheja ◽

Adelaide Greco ◽

...

Keyword(s):

Extracellular Matrix ◽

Tyrosine Phosphatase ◽

Melanoma Cells ◽

Matrix Degradation ◽

Extracellular Matrix Degradation ◽

Invadopodia Formation ◽

Interference Rna

Abstract Background Invadopodia are actin-based cell-membrane protrusions associated with the extracellular matrix degradation accompanying cancer invasion. The elucidation of the molecular mechanisms leading to invadopodia formation and activity is central for the prevention of tumor spreading and growth. Protein tyrosine kinases such as Src are known to regulate invadopodia assembly, little is however known on the role of protein tyrosine phosphatases in this process. Among these enzymes, we have selected the tyrosine phosphatase Shp1 to investigate its potential role in invadopodia assembly, due to its involvement in cancer development. Methods Co-immunoprecipitation and immunofluorescence studies were employed to identify novel substrate/s of Shp1AQ controlling invadopodia activity. The phosphorylation level of cortactin, the Shp1 substrate identified in this study, was assessed by immunoprecipitation, in vitro phosphatase and western blot assays. Short interference RNA and a catalytically-dead mutant of Shp1 expressed in A375MM melanoma cells were used to evaluate the role of the specific Shp1-mediated dephosphorylation of cortactin. The anti-invasive proprieties of glycerophosphoinositol, that directly binds and regulates Shp1, were investigated by extracellular matrix degradation assays and in vivo mouse model of metastasis. Results The data show that Shp1 was recruited to invadopodia and promoted the dephosphorylation of cortactin at tyrosine 421, leading to an attenuated capacity of melanoma cancer cells to degrade the extracellular matrix. Controls included the use of short interference RNA and catalytically-dead mutant that prevented the dephosphorylation of cortactin and hence the decrease the extracellular matrix degradation by melanoma cells. In addition, the phosphoinositide metabolite glycerophosphoinositol facilitated the localization of Shp1 at invadopodia hence promoting cortactin dephosphorylation. This impaired invadopodia function and tumor dissemination both in vitro and in an in vivo model of melanomas. Conclusion The main finding here reported is that cortactin is a specific substrate of the tyrosine phosphatase Shp1 and that its phosphorylation/dephosphorylation affects invadopodia formation and, as a consequence, the ability of melanoma cells to invade the extracellular matrix. Shp1 can thus be considered as a regulator of melanoma cell invasiveness and a potential target for antimetastatic drugs.

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