Osteoarthritis-Related Inflammation Blocks TGF-β’s Protective Effect on Chondrocyte Hypertrophy via (De)Phosphorylation of the SMAD2/3 Linker Region

Osteoarthritis (OA) is a degenerative joint disease characterized by irreversible cartilage damage, inflammation and altered chondrocyte phenotype. Transforming growth factor-β (TGF-β) signaling via SMAD2/3 is crucial for blocking hypertrophy. The post-translational modifications of these SMAD proteins in the linker domain regulate their function and these can be triggered by inflammation through the activation of kinases or phosphatases. Therefore, we investigated if OA-related inflammation affects TGF-β signaling via SMAD2/3 linker-modifications in chondrocytes. We found that both Interleukin (IL)-1β and OA-synovium conditioned medium negated SMAD2/3 transcriptional activity in chondrocytes. This inhibition of TGF-β signaling was enhanced if SMAD3 could not be phosphorylated on Ser213 in the linker region and the inhibition by IL-1β was less if the SMAD3 linker could not be phosphorylated at Ser204. Our study shows evidence that inflammation inhibits SMAD2/3 signaling in chondrocytes via SMAD linker (de)-phosphorylation. The involvement of linker region modifications may represent a new therapeutic target for OA.

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LncRNA MEG3 Inhibits the Degradation of the Extracellular Matrix of Chondrocytes in Osteoarthritis via Targeting miR-93/TGFBR2 Axis

Cartilage ◽

10.1177/1947603519855759 ◽

2019 ◽

pp. 194760351985575 ◽

Cited By ~ 12

Author(s):

Kang Chen ◽

Hao Zhu ◽

Min-Qian Zheng ◽

Qi-Rong Dong

Keyword(s):

Extracellular Matrix ◽

Transforming Growth Factor ◽

Transforming Growth Factor Β ◽

Cartilage Degradation ◽

Degenerative Joint Disease ◽

Joint Disease ◽

Regulatory Function ◽

Qrt Pcr ◽

Ecm Degradation ◽

Competitive Endogenous Rna

Background As a degenerative joint disease, osteoarthritis (OA) is characterized by articular cartilage degradation. Long noncoding RNAs (lncRNAs) act critical roles in the regulation of OA development, including affecting the proliferation, apoptosis, extracellular matrix (ECM) degradation, and inflammatory response of chondrocytes. The current study’s aim was to investigate the regulatory function and the underlying molecular mechanism of lncRNA MEG3 in ECM degradation of chondrocytes in OA. Methods In the current study, chondrocytes were induced by interleukin-1β (IL-1β) to simulate OA condition, and further assessed cell viability, lncRNA MEG3 and miR-93 expression levels. Overexpression or knockdown of lncRNA MEG3 in chondrocytes treated with IL-1β were performed to investigate the function of MEG3 in regulating cell proliferation, apoptosis and ECM degradation using EdU assay, flow cytometry, quantitative reverse transcription polymerase chain reaction (qRT-PCR), and Western blot. The interaction between MEG3 and miR-93 was assessed using qRT-PCR. Furthermore, overexpression of miR-93 was performed as recovery experiment to explore the functional mechanism of MEG3. Results MEG3 was significantly downregulated in chondrocytes treated with IL-1β, whereas miR-93 was upregulated concomitantly. Overexpression of MEG3 induced the proliferation, suppressed the apoptosis, and relieved the degradation of ECM in IL-1β-induced chondrocytes. By contrast, knockdown of MEG3 suppressed the proliferation, promoted the apoptosis, and aggravated ECM degradation in IL-1β induced chondrocytes. In addition, MEG3 was found to relieve the inhibitive expression of TGFBR2 as a competitive endogenous RNA (ceRNA) of miR-93, and then activated transforming growth factor-β (TGF-β) signaling pathway, regulated chondrocytes ECM degradation in IL-1β induced chondrocytes subsequently. Conclusion LncRNA MEG3 targeted miR-93/TGFBR2 axis, regulated the proliferation, apoptosis and ECM degradation of chondrocytes in OA.

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Post-translational modification of the SMAD2/3 linker region: the missing link between osteoarthritis-related inflammation and disturbed transforming growth factor-β signaling

Osteoarthritis and Cartilage ◽

10.1016/j.joca.2021.02.154 ◽

2021 ◽

Vol 29 ◽

pp. S110-S111

Author(s):

N. Thielen ◽

A. van Caam ◽

M. Neefjes ◽

E. Blaney Davidson ◽

P. van der Kraan

Keyword(s):

Growth Factor ◽

Transforming Growth Factor ◽

Transforming Growth Factor Β ◽

Post Translational Modification ◽

Missing Link ◽

Linker Region

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Selonsertib Alleviates the Progression of Rat Osteoarthritis: An in vitro and in vivo Study

Frontiers in Pharmacology ◽

10.3389/fphar.2021.687033 ◽

2021 ◽

Vol 12 ◽

Author(s):

Jiyuan Yan ◽

Yingchi Zhang ◽

Gaohong Sheng ◽

Bowei Ni ◽

Yifan Xiao ◽

...

Keyword(s):

Therapeutic Potential ◽

Cartilage Damage ◽

Cartilage Degradation ◽

Degenerative Joint Disease ◽

Joint Disease ◽

Therapeutic Effects ◽

Exact Role

Osteoarthritis (OA) is a prevalent degenerative joint disease. Its development is highly associated with inflammatory response and apoptosis in chondrocytes. Selonsertib (Ser), the inhibitor of Apoptosis Signal-regulated kinase-1 (ASK1), has exhibited multiple therapeutic effects in several diseases. However, the exact role of Ser in OA remains unclear. Herein, we investigated the anti-arthritic effects as well as the potential mechanism of Ser on rat OA. Our results showed that Ser could markedly prevent the IL-1β-induced inflammatory reaction, cartilage degradation and cell apoptosis in rat chondrocytes. Meanwhile, the ASK1/P38/JNK and NFκB pathways were involved in the protective roles of Ser. Furthermore, intra-articular injection of Ser could significantly alleviate the surgery induced cartilage damage in rat OA model. In conclusion, our work provided insights into the therapeutic potential of Ser in OA, indicating that Ser might serve as a new avenue in OA treatment.

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Transforming growth factor β-mediated site-specific Smad linker region phosphorylation in vascular endothelial cells

Journal of Pharmacy and Pharmacology ◽

10.1111/jphp.12298 ◽

2014 ◽

Vol 66 (12) ◽

pp. 1722-1733 ◽

Cited By ~ 20

Author(s):

Danielle Kamato ◽

Muhamad Ashraf Rostam ◽

Terence J. Piva ◽

Hossein Babaahmadi Rezaei ◽

Robel Getachew ◽

...

Keyword(s):

Endothelial Cells ◽

Growth Factor ◽

Transforming Growth Factor ◽

Vascular Endothelial Cells ◽

Transforming Growth Factor Β ◽

Vascular Endothelial ◽

Site Specific ◽

Linker Region

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Transforming Growth Factor β-Mediated Transcriptional Repression of c-myc Is Dependent on Direct Binding of Smad3 to a Novel Repressive Smad Binding Element

Molecular and Cellular Biology ◽

10.1128/mcb.24.6.2546-2559.2004 ◽

2004 ◽

Vol 24 (6) ◽

pp. 2546-2559 ◽

Cited By ~ 159

Author(s):

Joshua P. Frederick ◽

Nicole T. Liberati ◽

David S. Waddell ◽

Yigong Shi ◽

Xiao-Fan Wang

Keyword(s):

Growth Factor ◽

Transcriptional Repression ◽

Molecular Mechanisms ◽

Target Genes ◽

Transforming Growth Factor ◽

Transforming Growth Factor Β ◽

Cell Types ◽

Specific Cell ◽

Smad Proteins ◽

Smad Binding Element

ABSTRACT Smad proteins are the most well-characterized intracellular effectors of the transforming growth factor β (TGF-β) signal. The ability of the Smads to act as transcriptional activators via TGF-β-induced recruitment to Smad binding elements (SBE) within the promoters of TGF-β target genes has been firmly established. However, the elucidation of the molecular mechanisms involved in TGF-β-mediated transcriptional repression are only recently being uncovered. The proto-oncogene c-myc is repressed by TGF-β, and this repression is required for the manifestation of the TGF-β cytostatic program in specific cell types. We have shown that Smad3 is required for both TGF-β-induced repression of c-myc and subsequent growth arrest in keratinocytes. The transcriptional repression of c-myc is dependent on direct Smad3 binding to a novel Smad binding site, termed a repressive Smad binding element (RSBE), within the TGF-β inhibitory element (TIE) of the c-myc promoter. The c-myc TIE is a composite element, comprised of an overlapping RSBE and a consensus E2F site, that is capable of binding at least Smad3, Smad4, E2F-4, and p107. The RSBE is distinct from the previously defined SBE and may partially dictate, in conjunction with the promoter context of the overlapping E2F site, whether the Smad3-containing complex actively represses, as opposed to transactivates, the c-myc promoter.

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The human NUPR1/P8 gene is transcriptionally activated by transforming growth factor β via the SMAD signalling pathway

Biochemical Journal ◽

10.1042/bj20120368 ◽

2012 ◽

Vol 445 (2) ◽

pp. 285-293 ◽

Cited By ~ 17

Author(s):

Roxane M. Pommier ◽

Johann Gout ◽

David F. Vincent ◽

Carla E. Cano ◽

Bastien Kaniewski ◽

...

Keyword(s):

Growth Factor ◽

Cancer Progression ◽

Stress Resistance ◽

Nuclear Protein ◽

Transforming Growth Factor ◽

Transforming Growth Factor Β ◽

Transcriptional Level ◽

Smad Proteins ◽

Activation Cascade ◽

Tgfβ Superfamily

NUPR1 (nuclear protein 1), also called P8 (molecular mass 8 kDa) or COM1 (candidate of metastasis 1), is involved in the stress response and in cancer progression. In the present study, we investigated whether human NUPR1 expression was regulated by TGFβ (transforming growth factor β), a secreted polypeptide largely involved in tumorigenesis. We demonstrate that the expression of NUPR1 was activated by TGFβ at the transcriptional level. We show that this activation is mediated by the SMAD proteins, which are transcription factors specifically involved in the signalling of TGFβ superfamily members. NUPR1 promoter analysis reveals the presence of a functional TGFβ-response element binding the SMAD proteins located in the genomic DNA region corresponding to the 5′-UTR (5′-untranslated region). Altogether, the molecular results of the present study, which demonstrate the existence of a TGFβ/SMAD/NUPR1 activation cascade, open the way to consider and investigate further a new mechanism enabling TGFβ to promote tumorigenesis by inducing stress resistance.

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Andrographis paniculata Extract Relieves Pain and Inflammation in Monosodium Iodoacetate-Induced Osteoarthritis and Acetic Acid-Induced Writhing in Animal Models

Processes ◽

10.3390/pr8070873 ◽

2020 ◽

Vol 8 (7) ◽

pp. 873

Author(s):

Donghun Lee ◽

Chae Yun Baek ◽

Ji Hong Hwang ◽

Mi-Yeon Kim

Keyword(s):

Acetic Acid ◽

Weight Bearing ◽

Cartilage Damage ◽

Degenerative Joint Disease ◽

Andrographis Paniculata ◽

Joint Disease ◽

Tnf Α ◽

Monosodium Iodoacetate ◽

History Of

Osteoarthritis (OA), being the most prominent degenerative joint disease is affecting millions of elderly people worldwide. Although Andrographis paniculata is an ethnic medicine with a long history of being used as analgesic agent, no study using a monosodium iodoacetate (MIA) model has investigated its potential activities against OA. In this study, experimental OA was induced in rats with a knee injection of MIA, which represents the pathological characteristics of OA in humans. A. paniculata extract (APE) substantially reversed the loss of hind limb weight-bearing and the cartilage damage resulted from the OA induction in rats. Additionally, the levels of serum pro-inflammatory cytokines, such as IL-1β, IL-6, and TNF-α as well as the concentration of matrix metalloproteinases, including MMP-1, MMP-3, MMP-8, and MMP-13 were decreased by APE administration. Acetic acid-induced writhing responses in mice which quantitatively measure pain were significantly reduced by APE. In vitro, APE inhibited the generation of NO and downregulated the expression of IL-1β, IL-6, COX-2, and iNOS in lipopolysaccharide (LPS)-stimulated RAW264.7 cells. The above results suggest the potential use APE as a therapeutic agent against OA.

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PPARγ preservation via promoter demethylation alleviates osteoarthritis in mice

Annals of the Rheumatic Diseases ◽

10.1136/annrheumdis-2018-214940 ◽

2019 ◽

Vol 78 (10) ◽

pp. 1420-1429 ◽

Cited By ~ 9

Author(s):

Xiaobo Zhu ◽

Fang Chen ◽

Ke Lu ◽

Ai Wei ◽

Qing Jiang ◽

...

Keyword(s):

Dna Methylation ◽

Knockout Mice ◽

Cartilage Damage ◽

Critical Role ◽

Methylation Status ◽

Promoter Hypermethylation ◽

Dna Methyltransferases ◽

Degenerative Joint Disease ◽

Joint Disease ◽

Peroxisome Proliferator

ObjectivesOsteoarthritis (OA) is the most common degenerative joint disease in aged population and its development is significantly influenced by aberrant epigenetic modifications of numerous OA susceptible genes; however, the precise mechanisms that DNA methylation alterations affect OA pathogenesis remain undefined. This study investigates the critical role of epigenetic PPARγ (peroxisome proliferator–activated receptor-gamma) suppression in OA development.MethodsArticular cartilage expressions of PPARγ and bioactive DNA methyltransferases (DNMTs) from OA patients and mice incurred by DMM (destabilisation of medial meniscus) were examined. DNA methylation status of both human and mouse PPARγ promoters were assessed by methylated specific PCR and/or bisulfite-sequencing PCR. OA protections by a pharmacological DNA demethylating agent 5Aza (5-Aza-2'-deoxycytidine) were compared between wild type and PPARγ knockout mice.ResultsArticular cartilages from both OA patients and DMM mice display substantial PPARγ suppressions likely due to aberrant elevations of DNMT1 and DNMT3a and consequential PPARγ promoter hypermethylation. 5Aza known to inhibit both DNMT1 and DNMT3a reversed the PPARγ promoter hypermethylation, recovered the PPARγ loss and effectively attenuated the cartilage damage in OA mice. 5Aza also inhibited the OA-associated excessive inflammatory cytokines and deficit anti-oxidant enzymes, which were blocked by a specific PPARγ inhibitor in cultured chondrocytes. Further, 5Aza-confered protections against the cartilage damage and the associated abnormalities of OA-susceptible factors were significantly abrogated in PPARγ knockout mice.ConclusionEpigenetic PPARγ suppression plays a key role in OA development and PPARγ preservation via promoter demethylation possesses promising therapeutic potentials in clinical treatment of OA and the related joint diseases.

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CREB binding protein is a required coactivator for Smad-dependent, transforming growth factor β transcriptional responses in endothelial cells

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.95.16.9506 ◽

1998 ◽

Vol 95 (16) ◽

pp. 9506-9511 ◽

Cited By ~ 127

Author(s):

James N. Topper ◽

Maria R. DiChiara ◽

Jonathan D. Brown ◽

Amy J. Williams ◽

Dean Falb ◽

...

Keyword(s):

Endothelial Cells ◽

Growth Factor ◽

Transforming Growth Factor ◽

Binding Protein ◽

Transforming Growth Factor Β ◽

Cell Types ◽

Creb Binding Protein ◽

Transcriptional Responses ◽

Smad Proteins ◽

Smad Protein

The transforming growth factor-β (TGF-β) superfamily of growth factors and cytokines has been implicated in a variety of physiological and developmental processes within the cardiovascular system. Smad proteins are a recently described family of intracellular signaling proteins that transduce signals in response to TGF-β superfamily ligands. We demonstrate by both a mammalian two-hybrid and a biochemical approach that human Smad2 and Smad4, two essential Smad proteins involved in mediating TGF-β transcriptional responses in endothelial and other cell types, can functionally interact with the transcriptional coactivator CREB binding protein (CBP). This interaction is specific in that it requires ligand (TGF-β) activation and is mediated by the transcriptional activation domains of the Smad proteins. A closely related, but distinct endothelial-expressed Smad protein, Smad7, which does not activate transcription in endothelial cells, does not interact with CBP. Furthermore, Smad2,4–CBP interactions involve the COOH terminus of CBP, a region that interacts with other regulated transcription factors such as certain signal transduction and transcription proteins and nuclear receptors. Smad–CBP interactions are required for Smad-dependent TGF-β-induced transcriptional responses in endothelial cells, as evidenced by inhibition with overexpressed 12S E1A protein and reversal of this inhibition with exogenous CBP. This report demonstrates a functional interaction between Smad proteins and an essential component of the mammalian transcriptional apparatus (CBP) and extends our insight into how Smad proteins may regulate transcriptional responses in many cell types. Thus, functional Smad–coactivator interactions may be an important locus of signal integration in endothelial cells.

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