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Small-interfering RNAs (siRNAs) offer promising prospects for treating pyroptosis-related autoimmune diseases. However, poor stability and off-target effects during in vivo transportation hinder their practical clinical applications. Precision delivery and adaptive release of siRNAs into inflamed tissues and immune cells could unleash their full therapeutic potential. This study establishes a pyroptotic-spatiotemporally selective siRNA delivery system (PMRC@siGSDME) that selectively targets inflammatory tissues, responds to pyroptosis, and exhibits remarkable therapeutic efficacy against various autoimmune diseases. Novel hybrid nanovesicles (NVs) are designed as a combination of pyroptotic macrophage membranes (PMs) and R8-cardiolipin-containing nanovesicles (RC-NVs). Evidence provides that PM-derived proteins involved in cell-cell interactions and membrane trafficking may contribute to the specificity of NVs to inflammatory tissue. In addition, cardiolipin anchored in the hybrid NVs increases its affinity for activated gasdermin E (GSDME) and achieves pyroptosis-adaptive release of siGSDME for the spatiotemporally selective suppression of immune responses. More importantly, PMRC@siGSDME displays significant anti-inflammatory and therapeutic effects in multiple mouse autoimmune disease models, including arthritis and inflammatory bowel disease (IBD). Collectively, an innovative siRNA delivery strategy precisely tailored for pyroptotic cells has been developed, paving the way for new treatments for autoimmune inflammatory diseases with minimal side effects and wide clinical applicability.
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The knee joint has long been considered a closed system. The pathological effects of joint diseases on distant organs have not been investigated. Herein, our clinical data showed that post-traumatic joint damage, combined with joint bleeding (hemarthrosis), exhibits a worse liver function compared with healthy control. With mouse model, hemarthrosis induces both cartilage degeneration and remote liver damage. Next, we found that hemarthrosis induces the upregulation in ratio and differentiation towards Th17 cells of CD4+ T cells in peripheral blood and spleen. Deletion of CD4+ T cells reverses hemarthrosis-induced liver damage. Degeneration of cartilage matrix induced by hemarthrosis upregulates serological type II collagen (COL II), which activates CD4+ T cells. Systemic application of a COL II antibody blocks the activation. Furthermore, bulk RNAseq and single-cell qPCR analysis revealed that the cartilage Akt pathway is inhibited by blood treatment. Intra-articular application of Akt activator blocks the cartilage degeneration and thus protects against the liver impairment in mouse and pig models. Taken together, our study revealed a pathological joint-liver axis mediated by matrikine-activated CD4+ T cells, which refreshes the organ-crosstalk axis and provides a new treatment target for hemarthrosis-related disease. Intra-articular bleeding induces cartilage degradation through down-reulation of cartilage Akt pathway. During this process, the soluble COL II released from the damaged cartilage can activate peripheral CD4+ T cells, differention into Th17 cells and secretion of IL-17, which consequently induces liver impairment. Intra-articular application of sc79 (inhibitor of Akt pathway) can prevent the cartilage damage as well as its peripheral influences.
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Linfócitos T CD4-Positivos , Fígado , Animais , Camundongos , Humanos , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/patologia , Fígado/patologia , Fígado/metabolismo , Hemartrose/genética , Hemartrose/patologia , Masculino , Modelos Animais de Doenças , Células Th17/imunologia , Células Th17/patologia , Colágeno Tipo II/genética , Venenos Elapídicos/farmacologia , Feminino , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismoRESUMO
Nanoliposomes have a broad range of applications in the treatment of autoimmune inflammatory diseases because of their ability to considerably enhance drug transport. For their clinical application, nanoliposomes must be able to realize on-demand release of drugs at disease sites to maximize drug-delivery efficacy and minimize side effects. Therefore, responsive drug-release strategies for inflammation treatment have been explored; however, no specific design has been realized for a responsive drug-delivery system based on pyroptosis-related inflammation. Herein, we report a pioneering strategy for self-adaptive pyroptosis-responsive liposomes (R8-cardiolipin-containing nanoliposomes encapsulating dimethyl fumarate, RC-NL@DMF) that precisely release encapsulated anti-pyroptotic drugs into pyroptotic cells. The activated key pyroptotic protein, the N-terminal domain of gasdermin E, selectively integrates with the cardiolipin of liposomes, thus forming pores for controlled drug release, pyroptosis, and inflammation inhibition. Therefore, RC-NL@DMF exhibited effective therapeutic efficacies to alleviate autoimmune inflammatory damages in zymosan-induced arthritis mice and dextran sulfate sodium-induced inflammatory bowel disease mice. Our novel approach holds great promise for self-adaptive pyroptosis-responsive on-demand drug delivery, suppressing pyroptosis and treating autoimmune inflammatory diseases.
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BACKGROUND: Extracellular vesicles (EVs) derived from mesenchymal stromal cells (MSCs) offer promising prospects for stimulating cartilage regeneration. The different formation mechanisms suggest that exosomes and ectosomes possess different biological functions. However, little attention has been paid to the differential effects of EV subsets on cartilage regeneration. METHODS: Our study compared the effects of the two EVs isolated from adipose-derived MSCs (ASCs) on chondrocytes and bone marrow-derived MSCs (BMSCs) in vitro. Additionally, we loaded the two EVs into type I collagen hydrogels to optimize their application for the treatment of osteochondral defects in vivo. RESULTS: In vitro experiments demonstrate that ASC-derived exosomes (ASC-Exos) significantly promoted the proliferation and migration of both cells more effectively than ASC-derived ectosomes (ASC-Ectos). Furthermore, ASC-Exos facilitated a stronger differentiation of BMSCs into chondrogenic cells than ASC-Ectos, but both inhibited chondrocyte apoptosis to a similar extent. In the osteochondral defect model of rats, ASC-Exos promoted cartilage regeneration in situ better than ASC-Ectos. At 8 weeks, the hydrogel containing exosomes group (Gel + Exo group) had higher macroscopic and histological scores, a higher value of trabecular bone volume fraction (BV/TV), a lower value of trabecular thickness (Tb.Sp), and a better remodeling of extracellular matrix than the hydrogel containing ectosomes group (Gel + Ecto group). At 4 and 8 weeks, the expression of CD206 and Arginase-1 in the Gel + Exo group was significantly higher than that in the Gel + Ecto group. CONCLUSION: Our findings indicate that administering ASC-Exos may be a more effective EV strategy for cartilage regeneration than the administration of ASC-Ectos.
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Micropartículas Derivadas de Células , Exossomos , Células-Tronco Mesenquimais , Ratos , Animais , Exossomos/metabolismo , Cartilagem/metabolismo , Células-Tronco Mesenquimais/metabolismo , HidrogéisRESUMO
Lumbar disc herniation (LDH) is rare in juveniles. LDH occurring at age 20 years or younger is referred to as juvenile disc herniation (JDH). While adult LDH is regarded as an advanced stage of disc degeneration, it remains unclear why intervertebral discs rupture in youth. This study aimed to characterize magnetic resonance imaging (MRI) findings of JDH and investigate possible etiological factors. From 2013 to 2020, JDH patients and controls were identified and interviewed to assess demographics, general lifestyles, and family histories. MRIs were evaluated for disc degeneration, epiphyseal ring separation, Modic changes and endplate lesions. The relationships between JDH and suspected risk factors were examined. A total of 297 JDH patients (199 boys and 98 girls, age 17.3 ± 2.1 years) and 185 controls (age 17.1 ± 2.4 years) were studied. Age, body mass index, exposures to daily physical labor, regular exercise, and daily sitting time were similar between JDH cases and controls. A family medical history of serious back pain was more common in JDH patients than in controls (59.4% vs. 26.5%, p < 0.001), as well as family history of clinically established LDH (45.0% vs. 12.4%, p < 0.001). Epiphyseal ring separation was identified in 102 (29.2%) herniated discs in 91 (36.4%) JDH patients, while occurring in only 5 (1.4%) control participants (p < 0.001). Overall, severe disc degeneration was not a prominent finding in JDH patients. In conclusion, epiphyseal ring separation was a common magnetic resonance feature in JDH. Findings suggest a genetically mediated developmental model of JDH, rather than a model of premature disc degeneration.
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Degeneração do Disco Intervertebral , Deslocamento do Disco Intervertebral , Disco Intervertebral , Adulto , Masculino , Feminino , Adolescente , Humanos , Adulto Jovem , Deslocamento do Disco Intervertebral/diagnóstico por imagem , Deslocamento do Disco Intervertebral/etiologia , Deslocamento do Disco Intervertebral/patologia , Degeneração do Disco Intervertebral/diagnóstico por imagem , Degeneração do Disco Intervertebral/etiologia , Degeneração do Disco Intervertebral/patologia , Estudos de Casos e Controles , Vértebras Lombares/diagnóstico por imagem , Vértebras Lombares/patologia , Disco Intervertebral/patologia , Imageamento por Ressonância Magnética/métodosRESUMO
OBJECTIVE: Osteoarthritis (OA) is one of the most common degenerative joint diseases and is associated with autophagy suppression. However, the molecular mechanism of autophagy regulation in the context of OA is not fully understood. In this study, we sought to determine the role that HECTD1 plays in the pathogenesis of OA. METHODS: We used RNA sequencing analysis to explore the differential expression of E3 ubiquitin ligase genes in healthy human cartilage and human cartilage affected by OA. Using surgery- and aging-induced OA mouse models, we comprehensively analyzed the function of the screened gene Hectd1 in the development of OA; furthermore, we dissected the mechanism by which HECTD1 regulates autophagy and OA progression using a combination of molecular biologic, cell biologic, and biochemical approaches. RESULTS: HECTD1 was significantly down-regulated in human OA cartilage samples compared to healthy cartilage samples. Overexpression of HECTD1 in mouse joints alleviated OA pathogenesis, whereas conditional depletion of Hectd1 in cartilage samples aggravated surgery- and aging-induced OA pathogenesis. Mechanistically, HECTD1 bound to Rubicon and ubiquitinated Rubicon at lysine residue 534, which targets Rubicon for proteasomal degradation. More importantly, HECTD1-mediated Rubicon degradation regulated chondrocyte autophagy, leading to mitigation of stress-induced chondrocyte death and the subsequent progression of OA. CONCLUSION: HECTD1 plays a crucial role in the pathogenesis of OA, in that HECTD1 regulates chondrocyte autophagy by ubiquitinating and targeting Rubicon for proteasomal degradation.
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Produtos Biológicos , Osteoartrite , Humanos , Animais , Camundongos , Ubiquitinação , Condrócitos , Autofagia/genética , Osteoartrite/genética , Ubiquitina-Proteína Ligases/genéticaRESUMO
Rheumatoid arthritis (RA) is a common autoimmune disease leading to pain, disability, and even death. Although studies have revealed that aberrant activation of STING was implicated in various autoimmune diseases, the role of STING in RA remains unclear. In the current study, we demonstrated that STING activation was pivotal in RA pathogenesis. As the accumulation of dsDNA, a specific stimulus for STING, is a feature of RA, we developed a spherical polyethyleneimine-coated mesoporous polydopamine nanoparticles loaded with STING antagonist C-176 (PEI-PDA@C-176 NPs) for treating RA. The fabricated NPs with biocompatibility had high DNA adsorption ability and could effectively inhibit the STING pathway and inflammation in macrophages. Intra-articular administration of PEI-PDA@C-176 NPs could effectively reduce joint damage in mice models of dsDNA-induced arthritis and collagen-induced arthritis by inhibiting STING pathway. We concluded that materials with synergistic effects of STING inhibition might be an efficacious strategy to treat RA.
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Osteoarthritis (OA) is a progressive joint disease characterized by the degradation and destruction of articular cartilage, which is involved with pathological microenvironmental alterations induced by damaged chondrocytes and inflammatory macrophages. However, the current therapies cannot effectively alleviate the progression of OA. Our previous studies have shown that the pathological process of OA progression is accompanied by DNA damage, and inhibition of STING, a key molecule in DNA damage, may become a potential method for the treatment of OA. Itaconate, a metabolite highly expressed in macrophages under inflammatory conditions, has shown a wide range of anti-inflammatory effects, but its effect on OA and its underlying mechanism has not yet been studied. In this study, we found that exogenous supplementation of itaconate can activate Nrf2, and accordingly inhibit the STING-dependent NF-κB pathway, thereby alleviating the inflammation, ECM degeneration and senescence of chondrocytes stimulated by IL-1ß. In addition, itaconate can regulate the polarization of RAW264.7 macrophages, further reducing the apoptosis of chondrocytes. In vivo, intra-articular injection of itaconate reduces the degradation of cartilage and inflammation of synovial membrane in the mouse OA model. In conclusion, the present work suggests that exogenous supplementation of itaconate inhibits the inflammation, senescence and ECM degeneration of chondrocytes through the Nrf2/STING/NF-κB axis and regulates the polarization of synovial macrophages, thereby ameliorating the progression of OA, which supports that itaconate as a potential drug for the treatment of OA.
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Cartilagem Articular , Osteoartrite , Animais , Cartilagem Articular/metabolismo , Condrócitos , Modelos Animais de Doenças , Inflamação/metabolismo , Interleucina-1beta/metabolismo , Macrófagos/metabolismo , Camundongos , Fator 2 Relacionado a NF-E2/metabolismo , NF-kappa B/metabolismo , Osteoartrite/patologia , SuccinatosRESUMO
Osteoarthritis (OA), a degenerative disorder, is considered to be one of the most common forms of arthritis. Limonin (Lim) is extracted from lemons and other citrus fruits. Limonin has been reported to have anti-inflammatory effects, while inflammation is a major cause of OA; thus, we propose that limonin may have a therapeutic effect on OA. In this study, the therapeutic effect of limonin on OA was assessed in chondrocytes in vitro in IL-1ß induced OA and in the destabilization of the medial meniscus (DMM) mice in vivo. The Nrf2/HO-1/NF-κB signaling pathway was evaluated to illustrate the working mechanism of limonin on OA in chondrocytes. In this study, it was found that limonin can reduce the level of IL-1ß induced proinflammatory cytokines such as INOS, COX-2, PGE2, NO, TNF-α, and IL-6. Limonin can also diminish the biosynthesis of IL-1ß-stimulated chondrogenic catabolic enzymes such as MMP13 and ADAMTS5 in chondrocytes. The research on the mechanism study demonstrated that limonin exerts its protective effect on OA through the Nrf2/HO-1/NF-κB signaling pathway. Taken together, the present study shows that limonin may activate the Nrf2/HO-1/NF-κB pathway to alleviate OA, making it a candidate therapeutic agent for OA.
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Artrite Experimental/tratamento farmacológico , Condrócitos/efeitos dos fármacos , Inflamação/tratamento farmacológico , Interleucina-1beta/toxicidade , Limoninas/farmacologia , Fator 2 Relacionado a NF-E2/metabolismo , Osteoartrite/tratamento farmacológico , Animais , Artrite Experimental/etiologia , Artrite Experimental/metabolismo , Artrite Experimental/patologia , Condrócitos/metabolismo , Condrócitos/patologia , Regulação da Expressão Gênica , Heme Oxigenase (Desciclizante)/genética , Heme Oxigenase (Desciclizante)/metabolismo , Inflamação/etiologia , Inflamação/metabolismo , Inflamação/patologia , Masculino , Meniscos Tibiais/cirurgia , Camundongos , Camundongos Endogâmicos C57BL , Fator 2 Relacionado a NF-E2/genética , NF-kappa B/genética , NF-kappa B/metabolismo , Osteoartrite/etiologia , Osteoartrite/metabolismo , Osteoartrite/patologiaRESUMO
Lumbar disc degeneration is a common cause of chronic low back pain and an important contributor to various degenerative lumbar spinal disorders. However, currently there is currently no effective therapeutic strategy for treating disc degeneration. The pro-inflammatory cytokine interleukin-1ß (IL-1ß) mediates disc degeneration by inducing apoptotic death of nucleus pulposus (NP) cells and degradation of the NP extracellular matrix. Here, we confirmed that extracellular secretion of IL-1ß via secretory autophagy contributes to disc degeneration, and demonstrate that a thermosensitive reactive oxygen species (ROS)-responsive hydrogel loaded with a synthetic growth hormone-releasing hormone analog (MR409) can protect against needle puncture-induced disc degeneration in rats. Methods: The expression levels of proteins related to secretory autophagy such as tripartite motif-containing 16 (TRIM16) and microtubule-associated protein light chain 3B (LC3B) were examined in human and rat disc tissues by histology and immunofluorescence. The effects of TRIM16 expression level on IL-1ß secretion were examined in THP-1 cells transfected with TRIM16 plasmid or siRNA using ELISA, immunofluorescence, and immunoblotting. The in vitro effects of MR409 on IL-1ß were examined in THP-1 cells and primary rat NP cells using ELISA, immunofluorescence, immunoblotting, and qRT-PCR. Further, MR409 was subcutaneously administered to aged mice to test its efficacy against disc degeneration using immunofluorescence, X-ray, micro-CT, and histology. To achieve controllable MR409 release for intradiscal use, MR409 was encapsulated in an injectable ROS-responsive thermosensitive hydrogel. Viscosity, rheological properties, release profile, and biocompatibility were evaluated. Thereafter, therapeutic efficacy was assessed in a needle puncture-induced rat model of disc degeneration at 8 and 12 weeks post-operation using X-ray, magnetic resonance (MR) imaging, histological analysis, and immunofluorescence. Results: Secretory autophagy-related proteins TRIM16 and LC3B were robustly upregulated in degenerated discs of both human and rat. Moreover, while upregulation of TRIM16 facilitated, and knockdown of TRIM16 suppressed, secretory autophagy-mediated IL-1ß secretion from THP-1 cells under oxidative stress, MR409 inhibited ROS-induced secretory autophagy and IL-1ß secretion by THP-1 cells as well as IL-1ß-induced pro-inflammatory and pro-catabolic effects in rat NP cells. Daily subcutaneous injection of MR409 inhibited secretory autophagy and ameliorated age-related disc degeneration in mice. The newly developed ROS-responsive MR409-encapsulated hydrogel provided a reliable delivery system for controlled MR409 release, and intradiscal application effectively suppressed secretory autophagy and needle puncture-induced disc degeneration in rats. Conclusion: Secretory autophagy and associated IL-1ß secretion contribute to the pathogenesis of disc degeneration, and MR409 can effectively inhibit this pathway. The ROS-responsive thermosensitive hydrogel encapsulated with MR409 is a potentially efficacious treatment for disc degeneration.
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Autofagia/genética , Interleucina-1beta/metabolismo , Degeneração do Disco Intervertebral/genética , Proteínas Associadas aos Microtúbulos/metabolismo , Núcleo Pulposo/efeitos dos fármacos , Fragmentos de Peptídeos/farmacologia , Proteínas com Motivo Tripartido/genética , Ubiquitina-Proteína Ligases/genética , Adulto , Idoso , Animais , Autofagia/efeitos dos fármacos , Feminino , Imunofluorescência , Humanos , Hidrogéis , Degeneração do Disco Intervertebral/diagnóstico por imagem , Degeneração do Disco Intervertebral/patologia , Imageamento por Ressonância Magnética , Masculino , Camundongos , Pessoa de Meia-Idade , Núcleo Pulposo/diagnóstico por imagem , Núcleo Pulposo/metabolismo , Núcleo Pulposo/patologia , Ratos , Espécies Reativas de Oxigênio/metabolismo , Células THP-1 , Proteínas com Motivo Tripartido/metabolismo , Microtomografia por Raio-XRESUMO
Damaged deoxyribonucleic acid (DNA) is a primary pathologic factor for osteoarthritis (OA); however, the mechanism by which DNA damage drives OA is unclear. Previous research demonstrated that the cyclic GMP-AMP synthase (cGAS)-stimulator of interferon genes (STING) participates in DNA damage response. As a result, the current study aimed at exploring the role STING, which is the major effector in the cGAS-STING signaling casacde, in OA progress in vitro, as well as in vivo. In this study, the expression of STING was evaluated in the human and mouse OA tissues, and in chondrocytes exposed to interleukin-1 beta (IL-1ß). The influences of STING on the metabolism of the extracellular matrix (ECM), apoptosis, and senescence, were assessed in STING overexpressing and knocking-down chondrocytes. Moreover, the NF-κB-signaling casacde and its role in the regulatory effects of STING on ECM metabolism, apoptosis, and senescence were explored. The STING knockdown lentivirus was intra-articularly injected to evaluate its therapeutic impact on OA in mice in vivo. The results showed that the expression of STING was remarkably elevated in the human and mouse OA tissues and in chondrocytes exposed to IL-1ß. Overexpression of STING promoted the expression of MMP13, as well as ADAMTS5, but suppressed the expression of Aggrecan, as well as Collagen II; it also enhanced apoptosis and senescence in chondrocytes exposed to and those untreated with IL-1ß. The mechanistic study showed that STING activated NF-κB signaling cascade, whereas the blockage of NF-κB signaling attenuated STING-induced apoptosis and senescence, and ameliorated STING-induced ECM metabolism imbalance. In in vivo study, it was demonstrated that STING knockdown alleviated destabilization of the medial meniscus-induced OA development in mice. In conclusion, STING promotes OA by activating the NF-κB signaling cascade, whereas suppression of STING may provide a novel approach for OA therapy.
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Condrócitos , Proteínas de Membrana/fisiologia , Osteoartrite/metabolismo , Idoso , Animais , Apoptose , Células Cultivadas , Senescência Celular , Condrócitos/metabolismo , Condrócitos/patologia , Matriz Extracelular/metabolismo , Feminino , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Pessoa de Meia-Idade , Cultura Primária de CélulasRESUMO
Osteoarthritis (OA) is an age-related degenerative disease and currently cannot be cured. Transcription factor EB (TFEB) is one of the major transcriptional factors that regulates autophagy and lysosomal biogenesis. TFEB has been shown to be an effective therapeutic target for many diseases including OA. The current study explores the therapeutic effects of 20-Deoxyingenol (20-DOI) on OA as well as its working mechanism on TFEB regulation. The in vitro study showed that 20-DOI may suppress apoptosis and senescence induced by oxidative stress in chondrocytes; it may also promote the nuclear localization of TFEB in chondrocytes. Knock-down of TFEB compromised the effects of 20-DOI on apoptosis and senescence. The in vivo study demonstrated that 20-DOI may postpone the progression of OA in mouse destabilization of the medial meniscus (DMM) model; it may also suppress apoptosis and senescence and promote the nuclear localization of TFEB in chondrocytes in vivo. This work suggests that 20-Deoxyingenol may alleviate osteoarthritis by activating TFEB in chondrocytes, while 20-DOI may become a potential drug for OA therapy.
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Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/agonistas , Condrócitos/efeitos dos fármacos , Diterpenos/farmacologia , Osteoartrite/tratamento farmacológico , Envelhecimento/efeitos dos fármacos , Animais , Apoptose/efeitos dos fármacos , Autofagia/efeitos dos fármacos , Células Cultivadas , Modelos Animais de Doenças , Diterpenos/uso terapêutico , Imunofluorescência , Masculino , Camundongos , Camundongos Endogâmicos C57BLRESUMO
OBJECTIVES: Diabetes is a risk factor for intervertebral disc degeneration (IVDD). Studies have demonstrated that diabetes may affect IVDD through transcriptional regulation; however, whether post-transcriptional regulation is involved in diabetic IVDD (DB-IVDD) is still unknown. This study was performed to illustrate the role of HuR, an RNA-binding protein, in DB-IVDD development and its mechanism. MATERIALS AND METHODS: The expression of HuR was evaluated in nucleus pulposus (NP) tissues from diabetic IVDD patients and in high glucose-treated NP cells. Senescence and autophagy were assessed in HuR over-expressing and downregulation NP cells. The mRNAs that were regulated by HuR were screened, and immunoprecipitation was applied to confirm the regulation of HuR on targeted mRNAs. RESULTS: The results showed that the expression of HuR was decreased in diabetic NP tissues and high glucose-treated NP cells. Downregulation of HuR may lead to increased senescence in high glucose-treated NP cells, while autophagy activation attenuates senescence in HuR deficient NP cells. Mechanistic study showed that HuR prompted Atg7 mRNA stability via binding to the AU-rich elements. Furthermore, overexpression of Atg7, but not HuR, may ameliorate DB-IVDD in rats in vivo. CONCLUSIONS: In conclusion, HuR may suppress senescence through autophagy activation via stabilizing Atg7 in diabetic NP cells; while Atg7, but not HuR, may serve as a potential therapeutic target for DB-IVDD.
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Proteína 7 Relacionada à Autofagia/metabolismo , Autofagia , Senescência Celular , Proteína Semelhante a ELAV 1/metabolismo , Degeneração do Disco Intervertebral/patologia , Regiões 3' não Traduzidas , Animais , Autofagia/efeitos dos fármacos , Proteína 7 Relacionada à Autofagia/genética , Células Cultivadas , Senescência Celular/efeitos dos fármacos , Diabetes Mellitus Experimental/induzido quimicamente , Diabetes Mellitus Experimental/complicações , Proteína Semelhante a ELAV 1/antagonistas & inibidores , Proteína Semelhante a ELAV 1/genética , Glucose/farmacologia , Humanos , Degeneração do Disco Intervertebral/etiologia , Degeneração do Disco Intervertebral/metabolismo , Masculino , Proteínas Associadas aos Microtúbulos/metabolismo , Núcleo Pulposo/citologia , Núcleo Pulposo/metabolismo , Interferência de RNA , RNA Mensageiro/metabolismo , RNA Interferente Pequeno/metabolismo , Ratos , Ratos Sprague-Dawley , Proteína Sequestossoma-1/metabolismoRESUMO
Intervertebral disc degeneration (IVDD) has been reported to be a major cause of low back pain. Studies have demonstrated that IVDD may be dysregulated at the transcriptional level; however, whether post-transcriptional regulation is involved is still unknown. The current study aimed to illustrate the role of Human antigen R (HuR), an RNA binding protein involved in post-transcriptional regulation, in IVDD. The results showed that the expression of HuR was decreased in degenerative nucleus pulposus (NP) tissues as well as in TNF-α-treated NP cells. Downregulation of HuR may lead to increased inflammation and extracellular matrix (ECM) degradation in TNF-α-treated NP cells; however, these effects were not reversed in HuR overexpressed NP cells. Inhibition of the NF-κB signaling pathway attenuates inflammation and ECM degradation in HuR-deficient NP cells. A mechanism study showed that HuR prompted NKRF mRNA stability via binding to its AU-rich elements, and upregulation of NKRF suppressed inflammation and ECM degradation in HuR-deficient NP cells. Furthermore, we found that NKRF, but not HuR, overexpression ameliorated the process of IVDD in rats in vivo. In conclusion, HuR suppressed inflammation and ECM degradation in NP cells via stabilizing NKRF and inhibiting the NF-κB signaling pathway; NKRF, but not HuR, may serve as a potential therapeutic target for IVDD.
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The limited molecular classifications and disease signatures of osteoarthritis (OA) impede the development of prediagnosis and targeted therapeutics for OA patients. To classify and understand the subtypes of OA, we collected three types of tissue including cartilage, subchondral bone, and synovium from multiple clinical centers and constructed an extensive transcriptome atlas of OA patients. By applying unsupervised clustering analysis to the cartilage transcriptome, OA patients were classified into four subtypes with distinct molecular signatures: a glycosaminoglycan metabolic disorder subtype (C1), a collagen metabolic disorder subtype (C2), an activated sensory neuron subtype (C3), and an inflammation subtype (C4). Through ligand-receptor crosstalk analysis of the three knee tissue types, we linked molecular functions with the clinical symptoms of different OA subtypes. For example, the Gene Ontology functional term of vasculature development was enriched in the subchondral bone-cartilage crosstalk of C2 and the cartilage-subchondral bone crosstalk of C4, which might lead to severe osteophytes in C2 patients and apparent joint space narrowing in C4 patients. Based on the marker genes of the four OA subtypes identified in this study, we modeled OA subtypes with two independent published RNA-seq datasets through random forest classification. The findings of this work contradicted traditional OA diagnosis by medical imaging and revealed distinct molecular subtypes in knee OA patients, which may allow for precise diagnosis and treatment of OA.
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Osteoarthritis is a chronic degenerative disease characterized by cartilage destruction. It is the fourth most disabling disease worldwide and is currently incurable. Inflammation and extracellular matrix (ECM) degradation are considered to be substantial reasons for accelerating the progression of OA. ß-Hydroxyisoamylshikonin (ß-HIVS) is a natural naphthoquinone compound with anti-inflammatory and antioxidant activity. However, the effect of ß-HIVS on OA is still unclear. In this study, we found that ß-HIVS can down-regulate the expression of NO, PEG2, IL-6, TNF-α, COX-2, and iNOS, suggesting its anti-inflammatory effects in chondrocytes; we also found that ß-HIVS may down-regulate the expression of ADAMTS5 and MMP13 and up-regulate the expression of aggrecan and collagen II to inhibit the degradation of ECM. Mechanistically, ß-HIVS inhibited the NFκB pathway by activating the Nrf2/HO-1 axis, thereby exerting its anti-inflammatory and inhibitory effects on ECM degradation. In vivo experiments also proved the therapeutic effects of ß-HIVS on OA in mice, and Nrf2 is the target of ß-HIVS. These findings indicate that ß-HIVS may become a new drug for the treatment of OA.
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Anti-Inflamatórios/administração & dosagem , Condrócitos/efeitos dos fármacos , Interleucina-1beta/imunologia , Fator 2 Relacionado a NF-E2/imunologia , Naftoquinonas/administração & dosagem , Osteoartrite/tratamento farmacológico , Animais , Condrócitos/imunologia , Heme Oxigenase-1/genética , Heme Oxigenase-1/imunologia , Humanos , Interleucina-1beta/genética , Masculino , Metaloproteinase 13 da Matriz/genética , Metaloproteinase 13 da Matriz/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Fator 2 Relacionado a NF-E2/genética , NF-kappa B/genética , NF-kappa B/imunologia , Osteoartrite/genética , Osteoartrite/imunologiaRESUMO
As an ordinary joint vestigial disease, osteoarthritis (OA) contributes to a considerable proportion of disability cases worldwide. Inflammation, as the main pathological factor, mediates the occurrence and development of OA. Akebia Saponin D (ASD), also known as Asperosaponin VI, is one of the active components extracted from Dipsaci Radix and is rich in Dipsacus loose tea. It has shown sound therapeutic effects on various diseases; nevertheless, its role in OA therapy is not completely understood. This study demonstrated the anti-inflammatory activity of ASD in OA through a series of in vivo and in vitro experiments. In vitro experiments revealed that ASD might prohibit the production of inflammatory mediators in IL-1ß treated chondrocytes such as COX-2, iNOS, NO, PGE2, IL-6, and TNF-α. Meanwhile, it may also inhibit the production of ADAMTS-5 and MMP13 and promote the production of Aggrecan and Collagen II. The mechanism study demonstrated that ASD exerted an anti-inflammatory effect by activating the NRF2 target, upregulating the expression of HO-1, and preventing P65 from binding to DNA. In vivo experiments demonstrated that ASD might improve the progression of OA in a DMM mouse model. These research results provide evidence for the potential application of ASD in OA therapy.
Assuntos
Condrócitos/efeitos dos fármacos , Condrócitos/metabolismo , Heme Oxigenase (Desciclizante)/metabolismo , Inflamação/tratamento farmacológico , Fator 2 Relacionado a NF-E2/metabolismo , NF-kappa B/metabolismo , Osteoartrite/tratamento farmacológico , Saponinas/farmacologia , Animais , Anti-Inflamatórios/farmacologia , Ciclo-Oxigenase 2/metabolismo , Feminino , Mediadores da Inflamação/metabolismo , Interleucina-1beta/metabolismo , Masculino , Metaloproteinase 13 da Matriz/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , NF-kappa B/genética , Óxido Nítrico Sintase Tipo II/metabolismoRESUMO
Multilayer scaffolds fabricated by 3D printing or other techniques have been used to repair osteochondral defects. However, it remains a challenge to regenerate the articular cartilage and subchondral bone simultaneously with higher performance. In the present study, we enhanced the repair efficiency of osteochondral defects by developing a bi-layer scaffold: an interleukin-4 (IL-4)-loaded radially oriented gelatin methacrylate (GelMA) scaffold printed with digital light processing (DLP) in the upper layer and a porous polycaprolactone and hydroxyapatite (PCL-HA) scaffold printed with fused deposition modeling (FDM) in the lower layer. An in vitro test showed that both layers supported cell adhesion and proliferation, as the lower layer promoted osteogenic differentiation and the upper layer with IL-4 relieved the negative effects of inflammation on murine chondrocytes, which were induced by interleukin-1ß (IL-1ß) and M1 macrophages. In a rabbit osteochondral defect repair model, the IL-4-loaded bi-layer scaffold group obtained the highest histological score (24 ± 2) compared to the nontreated (11 ± 1) and pure bi-layer scaffold (16 ± 1) groups after 16 weeks of implantation, which showed that the IL-4-loaded bi-layer scaffold promoted regeneration of both cartilage and subchondral bone with increased formation of neocartilage and neobone tissues. Thus, the IL-4-loaded bi-layer scaffold is an attractive candidate for repair and regeneration of osteochondral defects.
Assuntos
Cartilagem Articular , Alicerces Teciduais , Animais , Condrócitos , Interleucina-4 , Camundongos , Osteogênese , Impressão Tridimensional , Coelhos , Engenharia TecidualRESUMO
In this study, we used murine chondrocytes as an in vitro model and mice exhibiting destabilization of the medial meniscus (DMM) as an in vivo model to investigate the mechanisms through which S-allyl cysteine (SAC) alleviates osteoarthritis (OA). SAC significantly reduced apoptosis and senescence and maintained homeostasis of extracellular matrix (ECM) metabolism in tert-butyl hydroperoxide (TBHP)-treated chondrocytes. Molecular docking analysis showed a -CDOCKER interaction energy value of 203.76 kcal/mol for interactions between SAC and nuclear factor erythroid 2-related factor 2 (Nrf2). SAC increased the nuclear translocation of Nrf2 and activated the Nrf2/HO1 signaling pathway in TBHP-treated chondrocytes. Furthermore, Nrf2 knockdown abrogated the antiapoptotic, antisenescence, and ECM regulatory effects of SAC in TBHP-treated chondrocytes. SAC treatment also significantly reduced cartilage ossification and erosion, joint-space narrowing, synovial thickening and hypercellularity in DMM model mice. Collectively, these findings show that SAC ameliorates OA pathology in TBHP-treated chondrocytes and DMM model mice by activating the Nrf2/HO1 signaling pathway.
RESUMO
Mesenchymal stem cells (MSCs) are an important cell source for tissue homeostasis and repair due to their stemness characteristic. Lots of intrinsic signaling pathways have been reported to regulate MSC stemness, but the extrinsic signals such as sodium lactate, particularly in physiological conditions, are poorly understood. Herein, we evaluated the effect of sodium lactate on human MSC stemness regulation by examining colony-forming ability, energy metabolism, multi-lineage differentiation ability, and pluripotent gene and protein expression. The underlying mechanism was further investigated with gene knockdown as well as small molecule interference and rescue experiments. We found that: (1) low concentration (1 mM) of sodium lactate promoted the stemness of human MSCs; (2) the upregulation of glycolysis was responsible for the MSC stemness promotion; (3) lysine demethylase 6B (KDM6B) was the key regulator which mediated sodium lactate-induced glycolysis and human MSC stemness enhancement. This study indicated that sodium lactate played an important role in human MSC stemness maintenance in physiological conditions, which could be related to KDM6B mediated metabolic regulation. It would provide new insight into stem cell biology, and contribute to cell transplantation and tissue regeneration strategies.