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1.
Nat Commun ; 15(1): 8179, 2024 Sep 18.
Artigo em Inglês | MEDLINE | ID: mdl-39294178

RESUMO

Deposition of monosodium urate and calcium pyrophosphate (MSU and CPP) micro-crystals is responsible for painful and recurrent inflammation flares in gout and chondrocalcinosis. In these pathologies, the inflammatory reactions are due to the activation of macrophages responsible for releasing various cytokines including IL-1ß. The maturation of IL-1ß is mediated by the multiprotein NLRP3 inflammasome. Here, we find that activation of the NLRP3 inflammasome by crystals and concomitant production of IL-1ß depend on cell volume regulation via activation of the osmo-sensitive LRRC8 anion channels. Both pharmacological inhibition and genetic silencing of LRRC8 abolish NLRP3 inflammasome activation by crystals in vitro and in mouse models of crystal-induced inflammation. Activation of LRRC8 upon MSU/CPP crystal exposure induces ATP release, P2Y receptor activation and intracellular calcium increase necessary for NLRP3 inflammasome activation and IL-1ß maturation. We identify a function of the LRRC8 osmo-sensitive anion channels with pathophysiological relevance in the context of joint crystal-induced inflammation.


Assuntos
Inflamassomos , Inflamação , Interleucina-1beta , Macrófagos , Proteínas de Membrana , Proteína 3 que Contém Domínio de Pirina da Família NLR , Animais , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/genética , Macrófagos/metabolismo , Camundongos , Inflamassomos/metabolismo , Interleucina-1beta/metabolismo , Humanos , Proteínas de Membrana/metabolismo , Proteínas de Membrana/genética , Inflamação/metabolismo , Inflamação/patologia , Ácido Úrico/metabolismo , Ácido Úrico/farmacologia , Camundongos Endogâmicos C57BL , Cálcio/metabolismo , Masculino
2.
Ann Rheum Dis ; 2024 Aug 12.
Artigo em Inglês | MEDLINE | ID: mdl-39134394

RESUMO

OBJECTIVES: Inflammatory mediators such as interleukin 6 (IL-6) are known to activate catabolic responses in chondrocytes during osteoarthritis (OA). This study aimed to investigate the role of a downstream target gene of IL-6, the serine protease inhibitor SerpinA3N, in the development of cartilage damage in OA. METHODS: RNA sequencing was performed in murine primary chondrocytes treated with IL-6, and identified target genes were confirmed in human and murine OA cartilage samples. Male cartilage-specific Serpina3n-deficient mice and control mice underwent meniscectomy (MNX) or sham surgery at 10 weeks of age. Intra-articular injections of SerpinA3N or sivelestat (an inhibitor of leucocyte elastase (LE), a substrate for SerpinA3N) were performed in wild-type mice after MNX. Joint damage was assessed 3-9 weeks after surgery by histology and micro-CT. The effect of sivelestat was assessed in cartilage explants exposed to macrophage-derived conditioned media. RESULTS: RNA sequencing revealed that SerpinA3N is a major target gene of IL-6 in chondrocytes. The expression of SerpinA3N is increased in OA cartilage. Conditional loss of SerpinA3N in chondrocytes aggravated OA in mice, while intra-articular injection of SerpinA3N limited joint damage. Chondrocytes did not produce serine proteases targeted by SerpinA3N. By contrast, macrophages produced LE on IL-6 stimulation. Sivelestat limited the cartilage catabolism induced by conditioned media derived from IL-6-stimulated macrophages. Additionally, an intra-articular injection of sivelestat is protected against OA in the MNX model. CONCLUSIONS: SerpinA3N protects cartilage against catabolic factors produced by macrophages, including LE. SerpinA3N and LE represent new therapeutic targets to dampen cartilage damage in OA.

3.
Ann Rheum Dis ; 83(7): 945-956, 2024 Jun 12.
Artigo em Inglês | MEDLINE | ID: mdl-38373842

RESUMO

INTRODUCTION: Untreated gout is characterised by monosodium urate (MSU) crystal accumulation responsible for recurrent flares that are commonly separated by asymptomatic phases. Both phases are inflammatory conditions of variable intensity. Gout flares are self-limited inflammatory reactions involving multiple mediators. This study aimed to characterise the inflammatory profiles of gout at different phases. METHODS: Using the Olink targeted proteomics, levels of 92 inflammation-related proteins were measured in plasma samples of a prospective gout population (GOUTROS), collected at gout flare (T1), the intercritical phase (T2) and after reaching the target serum urate level under urate-lowering therapy (T3). Results were validated in an independent cohort (OLT1177-05) with plasmas collected at T1 and T2. Ex vivo and in vitro experiments were performed to assess the inflammatory properties of new biomarkers. RESULTS: In total, 21 inflammatory new biomarkers were differentially expressed during the three time-points of gout disease. The levels of four of these proteins (interleukin 6 (IL-6), colony-stimulating factor 1, vascular endothelial growth factor A and tumour necrosis factor superfamily 14 (TNFSF14)) were increased during gout flare in an independent cohort. IL-6 and TNFSF14 had the highest fold change in expression during T1 versus T2 or T3. TNFSF14 was produced at the inflamed joint and enhanced the inflammatory response induced by lipopolysaccharide and MSU crystal stimulation. Conversely, TNFSF14 blockade reduced the inflammatory response. Additionally, single nucleotide polymorphisms of TNFSF14 affected the ability of myeloid cells to produce inflammatory cytokines. CONCLUSION: Gout flare involves multiple inflammatory mediators that may be used as potential therapeutic targets.


Assuntos
Biomarcadores , Gota , Membro 14 da Superfamília de Ligantes de Fatores de Necrose Tumoral , Humanos , Gota/tratamento farmacológico , Gota/sangue , Biomarcadores/sangue , Masculino , Pessoa de Meia-Idade , Feminino , Membro 14 da Superfamília de Ligantes de Fatores de Necrose Tumoral/sangue , Exacerbação dos Sintomas , Citocinas/sangue , Supressores da Gota/uso terapêutico , Idoso , Ácido Úrico/sangue , Estudos Prospectivos , Interleucina-6/sangue , Adulto , Proteômica/métodos , Fator A de Crescimento do Endotélio Vascular/sangue
4.
Nat Commun ; 15(1): 910, 2024 Jan 30.
Artigo em Inglês | MEDLINE | ID: mdl-38291039

RESUMO

Acquired mutations in the UBA1 gene were recently identified in patients with severe adult-onset auto-inflammatory syndrome called VEXAS (vacuoles, E1 enzyme, X-linked, autoinflammatory, somatic). However, the precise physiological and clinical impact of these mutations remains poorly defined. Here we study a unique prospective cohort of VEXAS patients. We show that monocytes from VEXAS are quantitatively and qualitatively impaired and display features of exhaustion with aberrant expression of chemokine receptors. In peripheral blood from VEXAS patients, we identify an increase in circulating levels of many proinflammatory cytokines, including IL-1ß and IL-18 which reflect inflammasome activation and markers of myeloid cells dysregulation. Gene expression analysis of whole blood confirms these findings and also reveals a significant enrichment of TNF-α and NFκB signaling pathways that can mediate cell death and inflammation. This study suggests that the control of the nflammasome activation and inflammatory cell death could be therapeutic targets in VEXAS syndrome.


Assuntos
Inflamassomos , Monócitos , Síndromes Mielodisplásicas , Dermatopatias Genéticas , Adulto , Humanos , Inflamassomos/genética , Estudos Prospectivos , Células Mieloides , Mutação
5.
Semin Arthritis Rheum ; 63: 152306, 2023 12.
Artigo em Inglês | MEDLINE | ID: mdl-37976811

RESUMO

INTRODUCTION: Ectopic calcifications (ECs) and heterotopic ossifications (HOs) form in non-mineralized tissues, most often in subcutaneous and muscular areas. Local and systemic complications can cause severe disability. Systemic administration of sodium thiosulfate (STS) gives promising results but is difficult to use in clinical practice. OBJECTIVE: Evaluation of the efficacy and safety of topical STS in ECs and HOs. METHODS: Retrospective analysis of the CATSS-O registry that included patients receiving topical STS 25 % prepared by the pharmacy of Limoges hospital during 2014-2020. The efficacy of STS was assessed by imaging (radiography or CT) after at least 6 months' treatment. RESULTS: Among 126 patients who received STS 25 %, 35 had complete clinical and radiographic data for analysis (28 with ECs and 7 with HOs; 18 children [mean age 8.9 years, range 1.5-16], 17 adults [mean age 52.4 years, range 24-90]). Calcifications or ossifications were due to dermatomyositis (8 children, 6 adults), systemic scleroderma (6 adults) or pseudo-hypoparathyroidism 1A (7 children). They were single (37.1 %) or multiple (62.9 %). Treated regions were in the lower limbs (31.4 %), upper limbs (37.1 %) or both (28.6 %) and the axial region (2.9 %). Topical STS was clinically effective in 9/28 (32.1 %) patients with ECs and 2/7 (28.6 %) children with HOs. Three patients experienced complete disappearance of their calcifications. Response for ECs was better in children than adults (54.5% vs 17.6 %, p = 0.035). Topical STS was well tolerated. CONCLUSION: Local STS seems effective for ossifications, particularly pediatric calcifications or ossifications. Randomized and experimental studies are needed to confirm this observation and to identify the underlying mechanisms.


Assuntos
Calcinose , Ossificação Heterotópica , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Criança , Pré-Escolar , Humanos , Lactente , Pessoa de Meia-Idade , Adulto Jovem , Calcinose/diagnóstico por imagem , Calcinose/tratamento farmacológico , Calcinose/etiologia , Ossificação Heterotópica/diagnóstico por imagem , Ossificação Heterotópica/tratamento farmacológico , Ossificação Heterotópica/complicações , Osteogênese , Estudos Retrospectivos
6.
Sci Rep ; 13(1): 11080, 2023 07 08.
Artigo em Inglês | MEDLINE | ID: mdl-37422514

RESUMO

Spectral photon-counting computed tomography (SPCCT) is a new technique with the capability to provide mono-energetic (monoE) images with high signal to noise ratio. We demonstrate the feasibility of SPCCT to characterize at the same time cartilage and subchondral bone cysts (SBCs) without contrast agent in osteoarthritis (OA). To achieve this goal, 10 human knee specimens (6 normal and 4 with OA) were imaged with a clinical prototype SPCCT. The monoE images at 60 keV with isotropic voxels of 250 × 250 × 250 µm3 were compared with monoE synchrotron radiation CT (SR micro-CT) images at 55 keV with isotropic voxels of 45 × 45 × 45 µm3 used as benchmark for cartilage segmentation. In the two OA knees with SBCs, the volume and density of SBCs were evaluated in SPCCT images. In 25 compartments (lateral tibial (LT), medial tibial, (MT), lateral femoral (LF), medial femoral and patella), the mean bias between SPCCT and SR micro-CT analyses were 101 ± 272 mm3 for cartilage volume and 0.33 mm ± 0.18 for mean cartilage thickness. Between normal and OA knees, mean cartilage thicknesses were found statistically different (0.005 < p < 0.04) for LT, MT and LF compartments. The 2 OA knees displayed different SBCs profiles in terms of volume, density, and distribution according to size and location. SPCCT with fast acquisitions is able to characterize cartilage morphology and SBCs. SPCCT can be used potentially as a new tool in clinical studies in OA.


Assuntos
Cistos Ósseos , Cartilagem Articular , Osteoartrite do Joelho , Osteoartrite , Humanos , Articulação do Joelho/diagnóstico por imagem , Cartilagem/diagnóstico por imagem , Microtomografia por Raio-X/métodos , Cistos Ósseos/diagnóstico por imagem , Osteoartrite do Joelho/diagnóstico por imagem , Cartilagem Articular/diagnóstico por imagem
7.
Arthritis Rheumatol ; 75(10): 1703-1713, 2023 10.
Artigo em Inglês | MEDLINE | ID: mdl-37494275

RESUMO

OBJECTIVE: Calcium pyrophosphate deposition (CPPD) disease is prevalent and has diverse presentations, but there are no validated classification criteria for this symptomatic arthritis. The American College of Rheumatology (ACR) and EULAR have developed the first-ever validated classification criteria for symptomatic CPPD disease. METHODS: Supported by the ACR and EULAR, a multinational group of investigators followed established methodology to develop these disease classification criteria. The group generated lists of candidate items and refined their definitions, collected de-identified patient profiles, evaluated strengths of associations between candidate items and CPPD disease, developed a classification criteria framework, and used multi-criterion decision analysis to define criteria weights and a classification threshold score. The criteria were validated in an independent cohort. RESULTS: Among patients with joint pain, swelling, or tenderness (entry criterion) whose symptoms are not fully explained by an alternative disease (exclusion criterion), the presence of crowned dens syndrome or calcium pyrophosphate crystals in synovial fluid are sufficient to classify a patient as having CPPD disease. In the absence of these findings, a score >56 points using weighted criteria, comprising clinical features, associated metabolic disorders, and results of laboratory and imaging investigations, can be used to classify as CPPD disease. These criteria had a sensitivity of 92.2% and specificity of 87.9% in the derivation cohort (190 CPPD cases, 148 mimickers), whereas sensitivity was 99.2% and specificity was 92.5% in the validation cohort (251 CPPD cases, 162 mimickers). CONCLUSION: The 2023 ACR/EULAR CPPD disease classification criteria have excellent performance characteristics and will facilitate research in this field.


Assuntos
Calcinose , Pirofosfato de Cálcio , Condrocalcinose , Reumatologia , Humanos , Condrocalcinose/diagnóstico por imagem , Síndrome , Estados Unidos
8.
Ann Rheum Dis ; 82(10): 1248-1257, 2023 10.
Artigo em Inglês | MEDLINE | ID: mdl-37495237

RESUMO

OBJECTIVE: Calcium pyrophosphate deposition (CPPD) disease is prevalent and has diverse presentations, but there are no validated classification criteria for this symptomatic arthritis. The American College of Rheumatology (ACR) and EULAR have developed the first-ever validated classification criteria for symptomatic CPPD disease. METHODS: Supported by the ACR and EULAR, a multinational group of investigators followed established methodology to develop these disease classification criteria. The group generated lists of candidate items and refined their definitions, collected de-identified patient profiles, evaluated strengths of associations between candidate items and CPPD disease, developed a classification criteria framework, and used multi-criterion decision analysis to define criteria weights and a classification threshold score. The criteria were validated in an independent cohort. RESULTS: Among patients with joint pain, swelling, or tenderness (entry criterion) whose symptoms are not fully explained by an alternative disease (exclusion criterion), the presence of crowned dens syndrome or calcium pyrophosphate crystals in synovial fluid are sufficient to classify a patient as having CPPD disease. In the absence of these findings, a score>56 points using weighted criteria, comprising clinical features, associated metabolic disorders, and results of laboratory and imaging investigations, can be used to classify as CPPD disease. These criteria had a sensitivity of 92.2% and specificity of 87.9% in the derivation cohort (190 CPPD cases, 148 mimickers), whereas sensitivity was 99.2% and specificity was 92.5% in the validation cohort (251 CPPD cases, 162 mimickers). CONCLUSION: The 2023 ACR/EULAR CPPD disease classification criteria have excellent performance characteristics and will facilitate research in this field.


Assuntos
Calcinose , Condrocalcinose , Reumatologia , Humanos , Estados Unidos , Condrocalcinose/diagnóstico por imagem , Pirofosfato de Cálcio , Síndrome
9.
Sci Rep ; 10(1): 16740, 2020 10 07.
Artigo em Inglês | MEDLINE | ID: mdl-33028903

RESUMO

Chondrocyte apoptosis may have a pivotal role in the development of osteoarthritis. Interest has increased in the use of anti-apoptotic compounds to protect against osteoarthritis development. In this work, we investigated the effect of adrenomedullin (AM), a 52 amino-acid hormone peptide, and a 31 amino-acid truncated form, AM(22-52), on chondrocyte apoptosis. Bovine articular chondrocytes (BACs) were cultured under hypoxic conditions to mimic cartilage environment and then treated with Fas ligand (Fas-L) to induce apoptosis. The expression of AM and its calcitonin receptor-like receptor (CLR)/receptor activity-modifying protein (RAMP) (receptor/co-receptor) was assessed by immunostaining. We evaluated the effect of AM and AM(22-52) on Fas-L-induced chondrocyte apoptosis. FAS expression was appreciated by RT-qPCR and immunostainings. The expression of hypoxia-inducible factor 1α (HIF-1α), CLR and one co-receptor (RAMP2) was evidenced. With BACs under hypoxia, cyclic adenosine monophosphate production increased dose-dependently with AM stimulation. AM significantly decreased caspase-3 activity (mean 35% decrease; p = 0.03) as a marker of Fas-L-induced apoptosis. Articular chondrocytes treated with AM showed significantly reduced cell death, along with downregulated Fas expression and production, as compared with AM(22-52). AM decreased articular chondrocyte apoptosis by downregulating a Fas receptor. These findings may pave the way for novel therapeutic approaches in osteoarthritis.


Assuntos
Adrenomedulina/farmacologia , Apoptose/efeitos dos fármacos , Condrócitos/efeitos dos fármacos , Proteína Ligante Fas/farmacologia , Fragmentos de Peptídeos/farmacologia , Adrenomedulina/metabolismo , Animais , Proteína Semelhante a Receptor de Calcitonina/metabolismo , Cartilagem Articular/metabolismo , Bovinos , Condrócitos/metabolismo , Regulação para Baixo/efeitos dos fármacos , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Proteína 2 Modificadora da Atividade de Receptores/metabolismo , Transdução de Sinais/efeitos dos fármacos , Receptor fas/metabolismo
10.
Ann Rheum Dis ; 79(11): 1506-1514, 2020 11.
Artigo em Inglês | MEDLINE | ID: mdl-32699039

RESUMO

OBJECTIVE: Macrophage activation by monosodium urate (MSU) and calcium pyrophosphate (CPP) crystals mediates an interleukin (IL)-1ß-dependent inflammation during gout and pseudo-gout flare, respectively. Since metabolic reprogramming of macrophages goes along with inflammatory responses dependently on stimuli and tissue environment, we aimed to decipher the role of glycolysis and oxidative phosphorylation in the IL-1ß-induced microcrystal response. METHODS: Briefly, an in vitro study (metabolomics and real-time extracellular flux analysis) on MSU and CPP crystal-stimulated macrophages was performed to demonstrate the metabolic phenotype of macrophages. Then, the role of aerobic glycolysis in IL-1ß production was evaluated, as well in vitro as in vivo using 18F-fluorodeoxyglucose positron emission tomography imaging and glucose uptake assay, and molecular approach of glucose transporter 1 (GLUT1) inhibition. RESULTS: We observed that MSU and CPP crystals led to a metabolic rewiring toward the aerobic glycolysis pathway explained by an increase in GLUT1 plasma membrane expression and glucose uptake on macrophages. Also, neutrophils isolated from human synovial fluid during gout flare expressed GLUT1 at their plasma membrane more frequently than neutrophils isolated from bloodstream. Both glucose deprivation and treatment with either 2-deoxyglucose or GLUT1 inhibitor suppressed crystal-induced NLRP3 activation and IL-1ß production, and microcrystal inflammation in vivo. CONCLUSION: In conclusion, we demonstrated that GLUT1-mediated glucose uptake is instrumental during the inflammatory IL-1ß response induced by MSU and CPP crystals. These findings open new therapeutic paths to modulate crystal-related inflammation.


Assuntos
Pirofosfato de Cálcio , Gota/metabolismo , Ativação de Macrófagos/fisiologia , Macrófagos/metabolismo , Ácido Úrico , Animais , Pirofosfato de Cálcio/imunologia , Pirofosfato de Cálcio/metabolismo , Pirofosfato de Cálcio/farmacologia , Transportador de Glucose Tipo 1/imunologia , Transportador de Glucose Tipo 1/metabolismo , Glicólise/efeitos dos fármacos , Glicólise/fisiologia , Gota/imunologia , Humanos , Interleucina-1beta/imunologia , Interleucina-1beta/metabolismo , Ativação de Macrófagos/efeitos dos fármacos , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Camundongos , Proteína 3 que Contém Domínio de Pirina da Família NLR/imunologia , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Ácido Úrico/imunologia , Ácido Úrico/metabolismo , Ácido Úrico/farmacologia
11.
Joint Bone Spine ; 87(5): 395-404, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32422338

RESUMO

OBJECTIVE: To develop French Society of Rheumatology-endorsed recommendations for the management of urate-lowering therapy (ULT). METHODS: Evidence-based recommendations were developed by 9 rheumatologists (academic or community-based), 3 general practitioners, 1 cardiologist, 1 nephrologist and 1 patient, using a systematic literature search, one physical meeting to draft recommendations and two Delphi rounds to finalize them. RESULTS: A set of 3 overarching principles and 5 recommendations was elaborated. The overarching principles emphasize the importance of patient education, especially the need for explaining the objective of lowering serum urate (SU) level to obtain crystal dissolution, clinical symptoms disappearance and avoidance of complications. ULT is indicated as soon as the diagnosis of gout is established. SU level must be decreased below 300µmol/l (50mg/l) in all gout patients or at least below 360µmol/l (60ml/l) when the 300µmol/l target cannot be reached, and must be maintained at these targets and monitored life-long. The choice of the ULT primarily relies on renal function: in patients whose estimated glomerular filtration rate (eGFR) is above 60ml/min/1.73m2, first-line ULT is allopurinol; in those with eGFR between 30 and 60ml/min/1.73m2, allopurinol use must be cautious and febuxostat can be considered as an alternative; and in those whose eGFR is below 30ml/min/1.73m2, allopurinol must be avoided and febuxostat should be preferred. Prophylaxis of ULT-induced gout flares involves progressive increase of ULT dosage and low-dose colchicine for at least 6 months. Cardiovascular risk factors and diseases, the metabolic syndrome and chronic kidney disease must be screened and managed. CONCLUSION: These recommendations aim to provide simple and clear guidance for the management of ULT in France.


Assuntos
Gota , Reumatologia , Alopurinol/uso terapêutico , França/epidemiologia , Gota/diagnóstico , Gota/tratamento farmacológico , Supressores da Gota/uso terapêutico , Humanos , Ácido Úrico
12.
Int J Mol Sci ; 21(4)2020 Feb 22.
Artigo em Inglês | MEDLINE | ID: mdl-32098291

RESUMO

Mechanical overload and aging are the main risk factors of osteoarthritis (OA). Galectin 3 (GAL3) is important in the formation of primary cilia, organelles that are able to sense mechanical stress. The objectives were to evaluate the role of GAL3 in chondrocyte primary cilium formation and in OA in mice. Chondrocyte primary cilium was detected in vitro by confocal microscopy. OA was induced by aging and partial meniscectomy of wild-type (WT) and Gal3-null 129SvEV mice (Gal3-/-). Primary chondrocytes were isolated from joints of new-born mice. Chondrocyte apoptosis was assessed by Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL), caspase 3 activity and cytochrome c release. Gene expression was assessed by qRT-PCR. GAL3 was localized at the basal body of the chondrocyte primary cilium. Primary cilia of Gal3-/- chondrocytes were frequently abnormal and misshapen. Deletion of Gal3 triggered premature OA during aging and exacerbated joint instability-induced OA. In both aging and surgery-induced OA cartilage, levels of chondrocyte catabolism and hypertrophy markers and apoptosis were more severe in Gal3-/- than WT samples. In vitro, Gal3 knockout favored chondrocyte apoptosis via the mitochondrial pathway. GAL3 is a key regulator of cartilage homeostasis and chondrocyte primary cilium formation in mice. Gal3 deletion promotes OA development.


Assuntos
Apoptose/genética , Cartilagem Articular/metabolismo , Condrócitos/metabolismo , Cílios/metabolismo , Galectina 3/genética , Mitocôndrias/metabolismo , Animais , Animais Recém-Nascidos , Cartilagem Articular/patologia , Caspase 3/metabolismo , Células Cultivadas , Condrócitos/citologia , Galectina 3/deficiência , Marcação In Situ das Extremidades Cortadas , Camundongos da Linhagem 129 , Camundongos Knockout , Osteoartrite/genética , Osteoartrite/metabolismo
13.
Biofactors ; 45(4): 583-597, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31132316

RESUMO

HFE-hemochromatosis is a disease characterized by a systemic iron overload phenotype mainly associated with mutations in the HFE protein (HFE) gene. Osteoarthritis (OA) has been reported as one of the most prevalent complications in HFE-hemochromatosis patients, but the mechanisms associated with its onset and progression remain incompletely understood. In this study, we have characterized the response to high iron concentrations of a primary culture of articular chondrocytes isolated from newborn Hfe-KO mice and compared the results with that of a similar experiment developed in cells from C57BL/6 wild-type (wt) mice. Our data provide evidence that both wt- and Hfe-KO-derived chondrocytes, when exposed to 50 µM iron, develop characteristics of an OA-related phenotype, such as an increased expression of metalloproteases, a decreased extracellular matrix production, and a lower expression level of aggrecan. In addition, Hfe-KO cells also showed an increased expression of iron metabolism markers and MMP3, indicating an increased susceptibility to intracellular iron accumulation and higher levels of chondrocyte catabolism. Accordingly, upon treatment with 50 µM iron, these chondrocytes were found to preferentially differentiate toward hypertrophy with increased expression of collagen I and transferrin and downregulation of SRY (sex-determining region Y)-box containing gene 9 (Sox9). In conclusion, high iron exposure can compromise chondrocyte metabolism, which, when simultaneously affected by an Hfe loss of function, appears to be more susceptible to the establishment of an OA-related phenotype.


Assuntos
Condrócitos/metabolismo , Compostos Férricos/farmacologia , Proteína da Hemocromatose/genética , Hemocromatose/metabolismo , Sobrecarga de Ferro/metabolismo , Ferro/metabolismo , Osteoartrite/metabolismo , Agrecanas/genética , Agrecanas/metabolismo , Animais , Cartilagem Articular/metabolismo , Cartilagem Articular/patologia , Proteínas de Transporte de Cátions/genética , Proteínas de Transporte de Cátions/metabolismo , Diferenciação Celular , Condrócitos/efeitos dos fármacos , Condrócitos/patologia , Colágeno Tipo I/genética , Colágeno Tipo I/metabolismo , Modelos Animais de Doenças , Matriz Extracelular/efeitos dos fármacos , Matriz Extracelular/metabolismo , Matriz Extracelular/patologia , Feminino , Ferritinas/genética , Ferritinas/metabolismo , Regulação da Expressão Gênica , Hemocromatose/complicações , Hemocromatose/genética , Hemocromatose/patologia , Proteína da Hemocromatose/deficiência , Humanos , Sobrecarga de Ferro/complicações , Sobrecarga de Ferro/genética , Sobrecarga de Ferro/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Osteoartrite/etiologia , Osteoartrite/genética , Osteoartrite/patologia , Oxirredutases/genética , Oxirredutases/metabolismo , Cultura Primária de Células , Receptores da Transferrina/genética , Receptores da Transferrina/metabolismo , Fatores de Transcrição SOX9/genética , Fatores de Transcrição SOX9/metabolismo , Proteína da Região Y Determinante do Sexo , Transdução de Sinais
15.
Cell Death Dis ; 10(3): 190, 2019 02 25.
Artigo em Inglês | MEDLINE | ID: mdl-30804327

RESUMO

Fas-associated death domain (FADD) is a key adaptor molecule involved in numerous physiological processes including cell death, proliferation, innate immunity and inflammation. Therefore, changes in FADD expression have dramatic cellular consequences. In mice and humans, FADD regulation can occur through protein secretion. However, the molecular mechanisms accounting for human FADD secretion were still unknown. Here we report that canonical, non-canonical, but not alternative, NLRP3 inflammasome activation in human monocytes/macrophages induced FADD secretion. NLRP3 inflammasome activation by the bacterial toxin nigericin led to the proinflammatory interleukin-1ß (IL-1ß) release and to the induction of cell death by pyroptosis. However, we showed that FADD secretion could occur in absence of increased IL-1ß release and pyroptosis and, reciprocally, that IL-1ß release and pyroptosis could occur in absence of FADD secretion. Especially, FADD, but not IL-1ß, secretion following NLRP3 inflammasome activation required extracellular glucose. Thus, FADD secretion was an active process distinct from unspecific release of proteins during pyroptosis. This FADD secretion process required K+ efflux, NLRP3 sensor, ASC adaptor and CASPASE-1 molecule. Moreover, we identified FADD as a leaderless protein unconventionally secreted through microvesicle shedding, but not exosome release. Finally, we established human soluble FADD as a new marker of joint inflammation in gout and rheumatoid arthritis, two rheumatic diseases involving the NLRP3 inflammasome. Whether soluble FADD could be an actor in these diseases remains to be determined. Nevertheless, our results advance our understanding of the mechanisms contributing to the regulation of the FADD protein expression in human cells.


Assuntos
Artrite Reumatoide/metabolismo , Proteína de Domínio de Morte Associada a Fas/metabolismo , Gota/metabolismo , Inflamassomos/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Piroptose/efeitos dos fármacos , Animais , Caspase 1/metabolismo , Linhagem Celular Tumoral , Micropartículas Derivadas de Células/metabolismo , Micropartículas Derivadas de Células/ultraestrutura , Proteína de Domínio de Morte Associada a Fas/genética , Humanos , Imunidade Inata/efeitos dos fármacos , Inflamação/metabolismo , Interleucina-1beta/metabolismo , Macrófagos/metabolismo , Monócitos/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/genética , Nigericina/farmacologia , Canais de Potássio/metabolismo
16.
Joint Bone Spine ; 86(2): 233-238, 2019 03.
Artigo em Inglês | MEDLINE | ID: mdl-30243785

RESUMO

OBJECTIVES: Beta-2-microglobulin (ß2M) dialysis-related amyloidosis (DRA), a disabiliting joint disease, has been initially reported in patients under long-term dialysis. The incidence and prevalence has significantly decreased with the improvement in dialysis techniques. Here, we attempted to clarify the clinical and MRI features to improve the diagnosis. METHODS: We retrospectively reviewed the files of 19 patients under dialysis treatment referred for suspicion of ß2M DRA. The diagnosis was based on MRI criteria (low signal intensity on both T1- and T2-weighted MR sequences). MRI analysis included a scoring of the several joint lesions. Scores were quantified according to a severity scale (0 to 3). RESULTS: Patients had a mean age of 66.0 ± 10.5 years and mean dialysis duration of 23.7 ± 10.5 years. DRA affected mainly large joints (shoulder in 73.7%, hip in 47.3%) and spine (36.8%). MRI images for 8 shoulders, 8 hips, and 3 spines were analysed. Amyloid synovitis was present in all cases, with high mean scores in the three sites. In all joints, the most common lesions were tendon thickening (68.4%) and bone erosions (68.4%). The mean tendon thickening score was high, particularly at the shoulders and also at the spine. Bone erosions were most frequent in the shoulder and pelvis. CONCLUSION: In patients under long-term dialysis, ß2M DRA involves large joints but also the spine. Special awareness should be drawn by the thickening of the tendon. MRI is required to characterize the pattern of the lesions and to achieve the diagnosis.


Assuntos
Amiloidose/etiologia , Artrite/etiologia , Diálise Renal/efeitos adversos , Tendinopatia/epidemiologia , Tendinopatia/etiologia , Microglobulina beta-2/efeitos adversos , Adulto , Fatores Etários , Idoso , Amiloidose/diagnóstico por imagem , Amiloidose/epidemiologia , Amiloidose/patologia , Artrite/diagnóstico por imagem , Artrite/fisiopatologia , Estudos de Coortes , Feminino , França , Humanos , Incidência , Imageamento por Ressonância Magnética/métodos , Masculino , Pessoa de Meia-Idade , Placa Amiloide/epidemiologia , Placa Amiloide/etiologia , Placa Amiloide/patologia , Prognóstico , Diálise Renal/métodos , Insuficiência Renal Crônica/diagnóstico , Insuficiência Renal Crônica/terapia , Estudos Retrospectivos , Medição de Risco , Índice de Gravidade de Doença , Fatores Sexuais , Sinovite/epidemiologia , Sinovite/etiologia , Sinovite/patologia , Tendinopatia/diagnóstico por imagem , Tendinopatia/patologia , Microglobulina beta-2/metabolismo
17.
Front Immunol ; 9: 2248, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30356764

RESUMO

Background: Calcium pyrophosphate (CPP) microcrystal deposition is associated with wide clinical phenotypes, including acute and chronic arthritis, that are interleukin 1ß (IL-1ß)-driven. Two CPP microcrystals, namely monoclinic and triclinic CPP dihydrates (m- and t-CPPD), have been identified in human tissues in different proportions according to clinical features. m-CPP tetrahydrate beta (m-CPPTß) and amorphous CPP (a-CPP) phases are considered as m- and t-CPPD crystal precursors in vitro. Objectives: We aimed to decipher the inflammatory properties of the three crystalline phases and one amorphous CPP phase and the intracellular pathways involved. Methods: The four synthesized CPP phases and monosodium urate crystals (MSU, as a control) were used in vitro to stimulate the human monocytic leukemia THP-1 cell line or bone marrow-derived macrophages (BMDM) isolated from WT or NLRP3 KO mice. The gene expression of pro- and anti-inflammatory cytokines was evaluated by quantitative PCR; IL-1ß, IL-6 and IL-8 production by ELISA; and mitogen-activated protein kinase (MAPK) activation by immunoblot analysis. NF-κB activation was determined in THP-1 cells containing a reporter plasmid. In vivo, the inflammatory potential of CPP phases was assessed with the murine air pouch model via cell analysis and production of IL-1ß and CXCL1 in the exudate. The role of NF-κB was determined by a pharmacological approach, both in vivo and in vitro. Results:In vitro, IL-1ß production induced by m- and t-CPPD and m-CPPTß crystals was NLRP3 inflammasome dependent. m-CPPD crystals were the most inflammatory by inducing a faster and higher production and gene expression of IL-1ß, IL-6, and IL-8 than t-CPPD, m-CPPTß and MSU crystals. The a-CPP phase did not show an inflammatory property. Accordingly, m-CPPD crystals led to stronger activation of NF-κB, p38, extracellular signal-regulated kinase 1/2 (ERK1/2) and c-Jun N-terminal kinase (JNK) MAPKs. Inhibition of NF-κB completely abrogated IL-1ß and IL-8 synthesis and secretion induced by all CPP crystals. Also, inhibition of JNK and ERK1/2 MAPKs decreased both IL-1ß secretion and NF-κB activation induced by CPP crystals. In vivo, IL-1ß and CXCL1 production and neutrophil infiltration induced by m-CPPD crystals were greatly decreased by NF-κB inhibitor treatment. Conclusion: Our results suggest that the inflammatory potential of different CPP crystals relies on their ability to activate the MAPK-dependent NF-κB pathway. Studies are ongoing to investigate the underlying mechanisms.


Assuntos
Pirofosfato de Cálcio/imunologia , Inflamação/imunologia , Sistema de Sinalização das MAP Quinases/imunologia , NF-kappa B/imunologia , Animais , Pirofosfato de Cálcio/química , Pirofosfato de Cálcio/metabolismo , Linhagem Celular , Células Cultivadas , Cristalização , Citocinas/genética , Citocinas/imunologia , Citocinas/metabolismo , Humanos , Inflamação/genética , Inflamação/metabolismo , Mediadores da Inflamação/química , Mediadores da Inflamação/imunologia , Mediadores da Inflamação/metabolismo , Macrófagos/imunologia , Macrófagos/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Knockout , NF-kappa B/metabolismo , Células THP-1
18.
JCI Insight ; 3(17)2018 09 06.
Artigo em Inglês | MEDLINE | ID: mdl-30185659

RESUMO

Sarcomas are still unsolved therapeutic challenges. Cancer stem cells are believed to contribute to sarcoma development, but lack of specific markers prevents their characterization and targeting. Here, we show that calpain-6 expression is associated with cancer stem cell features. In mouse models of bone sarcoma, calpain-6-expressing cells have unique tumor-initiating and metastatic capacities. Calpain-6 levels are especially high in tumors that have been successfully propagated in mouse to establish patient-derived xenografts. We found that calpain-6 levels are increased by hypoxia in vitro and calpain-6 is detected within hypoxic areas in tumors. Furthermore, calpain-6 expression depends on the stem cell transcription network that involves Oct4, Nanog, and Sox2 and is activated by hypoxia. Calpain-6 knockdown blocks tumor development in mouse and induces depletion of the cancer stem cell population. Data from transcriptomic analyses reveal that calpain-6 expression in sarcomas inversely correlates with senescence markers. Calpain-6 knockdown suppresses hypoxia-dependent prevention of senescence entry and also promotion of autophagic flux. Together, our results demonstrate that calpain-6 identifies sarcoma cells with stem-like properties and is a mediator of hypoxia to prevent senescence, promote autophagy, and maintain the tumor-initiating cell population. These findings open what we believe is a novel therapeutic avenue for targeting sarcoma stem cells.


Assuntos
Autofagia , Calpaína/metabolismo , Senescência Celular/fisiologia , Proteínas Associadas aos Microtúbulos/metabolismo , Células-Tronco Neoplásicas/metabolismo , Sarcoma/metabolismo , Animais , Biomarcadores , Calpaína/genética , Carcinogênese/metabolismo , Linhagem Celular Tumoral , Modelos Animais de Doenças , Perfilação da Expressão Gênica , Técnicas de Silenciamento de Genes , Humanos , Hipóxia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Proteínas Associadas aos Microtúbulos/genética , Proteína Homeobox Nanog/metabolismo , Neoplasias , Fator 3 de Transcrição de Octâmero/metabolismo , Fatores de Transcrição SOXB1/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto
19.
Eur Radiol ; 28(2): 478-486, 2018 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-28884296

RESUMO

OBJECTIVES: To assess the efficacy of bisphosphonate therapy on bone pain in patients with osteoid osteoma (OO) (main objective), and to describe bisphosphonate-induced changes in nidus mineralisation and regional bone-marrow oedema (BMO). METHODS: A prospective, observational study was conducted from 2011 to 2014. Patients with risk factors for complications of percutaneous or surgical ablation or recurrence after ablation, were offered once monthly intravenous bisphosphonate treatment until significant pain alleviation was achieved. RESULTS: We included 23 patients. The first two patients received pamidronate and the next 21 zoledronic acid (mean, 2.95 infusions per patient). Bisphosphonate therapy was successful in 19 patients (83%), whose mean pain visual analogue scale score decreased by 76.7%; this pain-relieving effect persisted in 17 patients (74%) with a mean follow-up time of 36 months. Computed tomography (CT) demonstrated a mean nidus density increase of 177.7% (p = 0.001). By magnetic resonance imaging (MRI), mean decreases were 38.4% for BMO surface area and 30.3% for signal intensity (p = 0.001 and p = 0.000, respectively). CONCLUSIONS: In 17/23 patients with painful OO managed conservatively with bisphosphonates, long-term final success was achieved. Bisphosphonates may accelerate the spontaneous healing of OO. KEY POINTS: • 19/23 patients with OO managed with bisphosphonates experienced significant pain relief • Pain relief was sustained in 17/23 patients, mean follow-up of 36 months • CT demonstrated a significant increase in nidus mineralisation • MRI demonstrated a significant decrease in bone marrow oedema • Bisphosphonate therapy may accelerate the spontaneous healing of OO.


Assuntos
Neoplasias Ósseas/tratamento farmacológico , Difosfonatos/uso terapêutico , Imageamento por Ressonância Magnética/métodos , Dor Musculoesquelética/diagnóstico , Osteoma Osteoide/tratamento farmacológico , Tomografia Computadorizada por Raios X/métodos , Adolescente , Adulto , Conservadores da Densidade Óssea/uso terapêutico , Neoplasias Ósseas/complicações , Neoplasias Ósseas/patologia , Neoplasias Ósseas/cirurgia , Criança , Feminino , Humanos , Masculino , Dor Musculoesquelética/tratamento farmacológico , Dor Musculoesquelética/etiologia , Osteoma Osteoide/complicações , Osteoma Osteoide/patologia , Estudos Prospectivos , Resultado do Tratamento , Adulto Jovem
20.
Semin Arthritis Rheum ; 47(3): 451-455, 2017 12.
Artigo em Inglês | MEDLINE | ID: mdl-28779847

RESUMO

INTRODUCTION: Tumoral calcinosis (TC) is a difficult-to-treat complication that can occur during several diseases such as dermatomyositis or genetic hyperphosphatemia. It is a painful and disabling condition that can lead to local complications including joint mobility reduction, cutaneous ulceration and superinfection. For the largest lesions, the treatment relies essentially on surgery. Intravenous sodium thiosulfate (STS) is efficient to treat calciphylaxis in patients undergoing hemodialysis. Local injections of STS seem efficient in superficial calcifications. OBJECTIVE: To report the efficacy and safety of intra-lesional injections of STS in tumoral calcinosis. RESULTS: We report two cases of successful intra-lesional injections of STS. A 44-year-old woman, with a history of dermatomyositis, presenting large subcutaneous calcifications in the right elbow, and a 42-year-old man, with a history of familial tumoral calcinosis, presenting large intramuscular calcifications in the right buttock, received weekly intra-lesional of 1-3g STS injections for 12 and 21 months, respectively. In both cases, the treatment relieved pain and greatly reduced the tumoral calcinosis with a very significant functional improvement without specific adverse effects. In case 1, TC size decreased from 28.7*56.0mm at baseline to 21.5*30.6mm at M12 treatment (59% reduction). In case 2, TC reduced from 167.5*204.3mm at baseline to 86.2*85.2mm at M21 treatment (79% reduction). CONCLUSION: Local injection of STS could be a promising therapeutic strategy for large and deep TC lesions and could therefore be an alternative to surgery.


Assuntos
Calcinose/tratamento farmacológico , Quelantes/administração & dosagem , Dermatomiosite/tratamento farmacológico , Hiperfosfatemia/tratamento farmacológico , Tiossulfatos/administração & dosagem , Adulto , Calcinose/etiologia , Dermatomiosite/complicações , Dermatomiosite/diagnóstico por imagem , Feminino , Fator de Crescimento de Fibroblastos 23 , Humanos , Hiperfosfatemia/complicações , Hiperfosfatemia/genética , Injeções Intralesionais , Imageamento por Ressonância Magnética , Masculino , Síndrome de Sjogren/complicações
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