Search

Additional file 1: Supplementary Table 1. Demographic, clinical, laboratory, and cardiovascular disease-related characteristics in patients with axial spondyloarthritis carrying or not the A allele of vaspin rs7159023.

Study partially supported by a grant from Spanish Society of Pulmonology and Thoracic Surgery (SEPAR 474-2017) and from Euronanomed III / Instituto de Salud Carlos III (ISCIII) (AC17/00027) awarded to SC. SR-M is supported by funds of the RETICS Program (RD16/0012/0009) from the ISCIII, co-funded by the European Regional Development Fund. BA-M is recipient of a ‘López Albo’ Post-Residency Programme funded by Servicio Cántabro de Salud. VP-C is supported by a pre-doctoral grant from IDIVAL (PREVAL 18/01). LL-G is supported by funds from IDIVAL (INNVAL 20/06). RP-F is supported by funds of START project (FOREUM18/34). DP-P is a recipient of a Río Hortega programme fellowship from the ISCIII, co-funded by the European Social Fund (ESF, ‘Investing in your future’) (CM20/00006). OG is staff personnel of Xunta de Galicia (Servizo Galego de Saude (SERGAS)) through a research-staff stabilization contract (ISCIII/SERGAS) and his work is funded by ISCIII and the European Union FEDER fund (RD16/0012/0014 (RIER) and PI17/00409). He is beneficiary of project funds from the Research Executive Agency (REA) of the European Union in the framework of MSCA-RISE Action of the H2020 Programme, Project 734899—Olive-Net. RL-M is a recipient of a Miguel Servet type I programme fellowship from the ISCIII, co-funded by the ESF (CP16/00033)., Mucin 1/Krebs von den Lungen-6 (KL-6) is proposed as a serum biomarker of several interstitial lung diseases (ILDs), including connective tissue disorders associated with ILD. However, it has not been studied in a large cohort of Caucasian antisynthetase syndrome (ASSD) patients. Consequently, we assessed the role of MUC1 rs4072037 and serum KL-6 levels as a potential biomarker of ASSD susceptibility and for the differential diagnosis between patients with ILD associated with ASSD (ASSD-ILD +) and idiopathic pulmonary fibrosis (IPF). 168 ASSD patients (149 ASSD-ILD +), 174 IPF patients and 523 healthy controls were genotyped for MUC1 rs4072037 T > C. Serum KL-6 levels were determined in a subgroup of individuals. A significant increase of MUC1 rs4072037 CC genotype and C allele frequencies was observed in ASSD patients compared to healthy controls. Likewise, MUC1 rs4072037 TC and CC genotypes and C allele frequencies were significantly different between ASSD-ILD+ and IPF patients. Additionally, serum KL-6 levels were significantly higher in ASSD patients compared to healthy controls. Nevertheless, no differences in serum KL-6 levels were found between ASSD-ILD+ and IPF patients. Our results suggest that the presence of MUC1 rs4072037 C allele increases the risk of ASSD and it could be a useful genetic biomarker for the differential diagnosis between ASSD-ILD+ and IPF patients., European Union FEDER fund PI17/00409, Servicio Cántabro de Salud, Servizo Galego de Saude, Spanish Society of Pulmonology and Thoracic Surgery SEPAR 474-2017, European Commission EC 734899, CP16/00033, Research Executive Agency, Instituto de Salud Carlos III AC17/00027, CM20/00006, RD16/0012/0009, RD16/0012/0014, European Social Fund, European Regional Development Fund, Foundation for Research in Rheumatology, Instituto de Investigación Marqués de Valdecilla INNVAL 20/06, PREVAL 18/01, Servicio Gallego de Salud, START, Global Change System for Analysis, Research, and Training FOREUM18/34

Osteoarthritis is a high-incidence painful and debilitating disease characterized by progressive degeneration of articular joints, which indicates a breakdown in joint homeostasis favoring catabolic processes. Biomechanical loading, associated with inflammatory and metabolic imbalances of joint, strongly contributes to the initiation and progression of the disease. Obesity is a primary risk factor for disease onset, and mechanical factors increased the risk for disease progression. Moreover, inflammatory mediators, in particular, adipose tissue-derived cytokines (better known as adipokines) play a critical role linking obesity and osteoarthritis. The present article summarizes the knowledge about the role of adipokines in cartilage and bone function, highlighting their contribution to the imbalance of joint homeostasis and, consequently, pathogenesis of osteoarthritis. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 36:594-604, 2018.

Obesity has reached epidemic proportions in the Western society and is increasing in the developing world. It is considered as one of the major contributors to the global burden of disability and chronic diseases, including autoimmune, inflammatory and degenerative diseases. Research conducted on obesity and its complications over the last two decades has transformed the outdated concept of white adipose tissue (WAT) merely serving as an energy depot. WAT is now recognized as an active and inflammatory organ capable of producing a wide variety of factors known as adipokines. These molecules participate through endocrine, paracrine, autocrine, or juxtacrine cross-talk mechanisms in a great variety of physiological or pathophysiological processes, regulating food intake, insulin sensitivity, immunity, and inflammation. Although initially restricted to metabolic activities (regulation of glucose and lipid metabolism), adipokines currently represent a new family of proteins that can be considered key players in the complex network of soluble mediators involved in the pathophysiology of immune/inflammatory diseases. However, the complexity of the adipokine network in the pathogenesis and progression of inflammatory diseases has posed, since the beginning, the important question of whether it may be possible to target the mechanism(s) by which adipokines contribute to disease selectively without suppressing their physiological functions. Here we explore in depth the most recent findings concerning the involvement of adipokines in inflammation and immune responses, in particular in rheumatic, inflammatory and degenerative diseases. We also highlight several possible strategies for therapeutic development and propose that adipokines and their signalling pathways may represent innovative therapeutic strategies for inflammatory disorders. ACKNOWLEDGMENTS: OG and FL are Staff Personnel of Xunta de Galicia (Servizo Galego de Saude, SERGAS) through a research-staff stabilization contract (ISCIII/SERGAS). VF is a “Sara Borrell” Researcher funded by ISCIII and FEDER. RG is a “Miguel Servet” Researcher funded by Instituto de Salud Carlos III (ISCIII) and FEDER. OG, MAGG and RG are members of RETICS Programme, RD16/0012/0014 (RIER: Red de Investigación en Inflamación y Enfermedades Reumáticas) via Instituto de Salud Carlos III (ISCIII) and FEDER. FL is a member of CIBERCV (Centro de Investigación Biomédica en Red de Enfermedades Cardiovasculares). The work of OG (PIE13/00024 and PI14/00016, PI17/00409), FL (PI15/00681 and CB16/11/00226) and RG (PI16/01870 and CP15/00007) was funded by Instituto de Salud Carlos III and FEDER. OG is a beneficiary of a project funded by Research Executive Agency of the European Union in the framework of MSCA-RISE Action of the H2020 Programme (Project number 734899). AM has received funding from the European Commission Framework 7 programme (EU FP7; HEALTH.2012.2.4.5-2, project number 305815; Novel Diagnostics and Biomarkers for Early Identification of Chronic Inflammatory Joint Diseases) plus generous support from the Innovative Medicines Initiative Joint Undertaking under grant agreement No. 115770, resources of which are composed of financial contribution from the European Union’s Seventh Framework programme (FP7/2007-2013) and EFPIA companies’ in kind contribution. The funders had no role in study design, data collection, and analysis, decision to publish, or preparation of the manuscript.

OBJECTIVES: Emerging data suggest that several metabolic factors, released mainly by white adipose tissue (WAT) and joint tissues, and collectively named adipokines, might have a role in the pathophysiology of OA. Recently, novel adipokines such as SERPINE2, WISP2, GPNMB and ITIH5 have been identified in WAT. The main goal of this study was to analyse the expression of these novel adipokines in synovium, infrapatellar fat pad and chondrocytes and to compare the expression of these molecules in healthy and OA tissues. METHODS: Synovial tissues, infrapatellar fat pad and chondrocytes were obtained from 36 OA patients (age 52-85; mean BMI 28.9) who underwent total knee replacement surgery. Healthy synovial tissues and infrapatellar fat pad were obtained from 15 traumatic knee patients (age 23-53; mean BMI 23.5). mRNA and protein expression were determined by qRT-PCR and western blot analysis respectively. RESULTS: All the novel adipokines, matter of our study, are expressed in OA synovium, infrapatellar fat pad and chondrocytes. Moreover, we detected a differential expression of SERPINE2 and ITIH5 in OA synovial tissues as compared to healthy samples. Finally, we also observed an increased expression of WISP2 in OA infrapatellar fat pad in comparison to healthy controls. CONCLUSIONS: In this study we demonstrated for the first time the expression of four novel adipokines in different joint tissues and how these molecules are differentially expressed in healthy and OA joint tissues. Xunta de Galicia/SERGAS Instituto de Salud Carlos III (ISCIII) Fundación Ramón Domínguez Ministerio de Educación de España

Nitric oxide (NO) has been considered a catabolic factor that contributes to OA pathology by inducing chondrocytes apoptosis, matrix metalloproteinases synthesis, and pro-inflammatory cytokines expression. Thus, the research on NO regulation in chondrocytes represents a relevant field which needs to be explored in depth. However, to date, only the murine ATDC-5 cell line and primary chondrocytes are well-established cells to study NO production in cartilage tissues. The goal of this study is to determine whether two commonly used human chondrocytic cell lines: SW-1353 and T/C-28a2 cell lines are good models to examine lipopolysaccharide and/or pro-inflammatory cytokine-driven NO release and iNOS expression. To this aim, we carefully examined NO production and iNOS protein expression in human T/C-28a2 and SW-1353 chondrocytes stimulated with LPS and interleukin (IL)-1 alone or in combination. We also use ATDC-5 cells as a positive control for NO production. NO accumulation has been determined by colorimetric Griess reaction, whereas NOS type II expression was determined by Western Blot analysis. Our results clearly demonstrated that neither human T/C-28a2 nor SW-1353 chondrocytes showed a detectable increase in NO production or iNOS expression after bacterial endotoxin or cytokines challenge with IL-1. Our study demonstrated that T/C-28a2 and SW-1353 human cell lines are not suitable for studying NO release and iNOS expression confirming that ATDC5 and human primary cultured chondrocytes are the best in vitro cell system to study the actions derived from this mediator.

Nesfatin-1 is a recently discovered satiety-inducing adipokine identified in hypothalamic regions that regulates energy balance. So far, no data exist on NUCB2/nesfatin-1 localization in human and murine chondrocytes. Here, we therefore investigated NUCB2/nesfatin-1 gene and protein expression in human and murine chondrocytes and the effect of nesfatin-1 on pro-inflammatory cytokines expression. Peptide localization was performed by laser confocal microscopy, NUCB2 mRNA expression was studied by RT-PCR and protein secretion was measured by XMap technology and Western blot analysis. First, we demonstrated cytoplasmic localization of NUCB2/nesfatin-1 peptide in both human and murine chondrocytes. We present evidence that both mRNA and protein expression of NUCB2 were increased during the differentiation of ATDC5 murine chondrocyte cell line. Furthermore, we demonstrated that nesfatin-1 induces IL-6 and MIP-1alpha mRNA expression and protein secretion in ATDC-5 cells challenged with IL-1, and also increases COX-2 mRNA expression in these cells. Finally, nesfatin-1 provoked a clear induction of pro-inflammatory agents, such as COX-2, IL-8, IL-6, and MIP-1alpha in human primary chondrocytes from OA patients.

Inflammation is a complex mechanism of cell/tissue responses to injuries triggered by multiple causes, including trauma, pathogens or autoimmune abnormal responses. In the last years, a novel line of thought is emerging by giving a more holistic vision of chronic arthropathies through a recently identified group of molecules, called adipokines. Actually, most of these recently identified factors, produced prevalently by white adipose tissue but also by cells of the joints (chondrocytes and synovial fibroblasts) and immune cells, play a significant role in chronic inflammation. Adipokines dysregulation has emerged as a common characteristic of chronic inflammation in rheumatic diseases in particular when obesity or, more precisely, adipose tissue dysfunction is associated with common rheumatic diseases, such as osteoarthritis and rheumatoid arthritis. In this MiniReview, we discuss the role of adipokines in osteoarthritis and rheumatoid arthritis providing an updated overview of their pathophysiological role and potential use as therapeutic targets.

Endocannabinoids has been described to be involved in articular degenerative disease by modulating nociception and immune system. However, the role of the endocannabinoid anandamide on chondrocyte cell viability is still unclear. Therefore, we decided to study anandamide's effects on chondrocytes viability and to evaluate its interactions with the catabolic factor TNF (tumor necrosis factor). Chondrocyte vitality was evaluated by MTT assay. We investigated LDH release, chromatin condensation, cleavage of focal adhesion kinase (FAK), and caspases-3, 8, and 9 activation. c-MYC mRNA levels were determined by RT-PCR. We studied by Western blot the activation patterns of AKT, AMPK, ERK, p38, and JNK kinases. Finally, we evaluate the effect of anandamide in TNF-induced caspase-3 cleavage. Anandamide decreased chondrocyte vitality independently of its receptors. It induced AMPK activation without LDH release. Anandamide induced chromatin condensation, activation of caspase-3, 8, and 9, and FAK cleavage. Surprisingly, despite anandamide inhibited cell proliferation, it increased c-MYC expression. Moreover anandamide inhibited AKT activation, whilst it induced a sustained activation of ERK, JNK, and p38. Finally, anandamide synergized with TNF-alpha in the cleavage of caspase-3. In conclusion, our findings suggest that anandamide, alone or in combination with TNF-alpha, may be a potential destructive agent in cartilage. Instituto de Salud Carlos III Xunta de Galicia (SERGAS) RETICS Programme, RD08/0075 (RIER) via Instituto de Salud Carlos III (ISCIII) “FPU” programme of the Spanish Ministry of Education Instituto de Salud Carlos III (ISCIII)/ “Sara Borrell Programme”

Lipocalin 2 (LCN2) has recently emerged as a novel adipokine involved in different processes including arthritis and chondrocyte inflammatory response. However, little is known about its activity on chondrocyte homeostasis and its regulation by nitric oxide (NO) Hence, we performed a set of experiments aimed to achieve a better understanding of this relationship. Cell vitality was tested in the ATDC5 cell line by the MTT colorimetric assay. Protein expression and gene expression was evaluated by Western blot and real time RT-PCR, respectively. NO production (determined as nitrite accumulation) was assayed by the Griess reaction. First, we demonstrated that LCN2 decreased murine chondrocytes vitality. Next, LCN2 co-stimulation with LPS enhanced NOS2 protein expression by murine chondrocytes. In addition, inhibition of LPS-induced nitric oxide production by aminoguanidine, a selective NOS2 inhibitor, significantly reduced LPS-mediated LCN2 expression. In contrast, treatment of murine chondrocytes with sodium nitroprussiate (SNP), a classic NO donor, scarcely induced LCN2 expression. Intriguingly, SNP addition to LPS-challenged chondrocytes, treated with aminoguanidine, provoked a strong induction of LCN2 expression. Finally, murine ATDC5 cells, co-cultured with LPS pre-challenged macrophages, had higher LCN2 expression in comparison with murine chondrocytes co-cultured with non pre-challenged macrophages. In this work we have described for the first time that NO is able to exert a control on LCN2 expression, suggesting the existence of a feedback loop regulating its expression. Instituto de Salud Carlos III Xunta de Galicia (SERGAS) REDINSCOR Instituto de Salud Carlos III/RETICS Programme

Nesfatin-1, a satiety-inducing peptide identified in hypothalamic regions that regulate energy balance, is an integral regulator of energy homeostasis and a putative glucose-dependent insulin coadjuvant. We investigated its production by human cardiomyocytes and its effects on glucose uptake, in the main cardiac glucose transporter GLUT-4 and in intracellular signaling. Quantitative RT-PCR, Western blots, confocal immunofluorescence microscopy, and ELISA of human and murine cardiomyocytes and/or cardiac tissue showed that cardiomyocytes can synthesize and secrete nesfatin-1. Confocal microscopy of cultured cardiomyocytes after GLUT-4 labeling showed that nesfatin-1 mobilizes this glucose transporter to cell peripherals. The rate of 2-deoxy-D-[(3)H]glucose incorporation demonstrated that nesfatin-1 induces glucose uptake by HL-1 cells and cultured cardiomyocytes. Nesfatin-1 induced dose- and time-dependent increases in the phosphorylation of ERK1/2, AKT, and AS160. In murine and human cardiac tissue, nesfatin-1 levels varied with diet and coronary health. In conclusion, human and murine cardiomyocytes can synthesize and secrete nesfatin-1, which is able to induce glucose uptake and the mobilization of the glucose transporter GLUT-4 in these cells. Nesfatin-1 cardiac levels are regulated by diet and coronary health.

Obesity has been considered a risk factor for osteoarthritis and it is usually accepted that obesity contributes to the development and progression of osteoarthritis by increasing mechanical load of the joints. Nevertheless, recent advances in the physiology of white adipose tissue evidenced that fat cells produce a plethora of factors, called adipokines, which have a critical role in the development of ostearthritis, besides to mechanical effects. In this paper, we review the role of adipokines and highlight the cellular and molecular mechanisms at play in osteoarthritis elicited by adipokines. We also emphasize how defining the role of adipokines has broadned our understanding of the diversity of factors involved in the genesis and progression of osteoarthritis in the hope of modifying it to prevent and treat diseases.

The cloning of leptin in 1994 by Zhang et al. introduced a novel concept about white adipose tissue (WAT) as a very dynamic organ that releases a plethora of immune and inflammatory mediators, such as adipokines and cytokines, which are involved in multiple diseases. Actually, adipokines exert potent modulatory actions on target tissues involved in rheumatic diseases including cartilage, synovial, bone and immune cells. The goal of this paper is to elucidate the recent findings concerning the involvement of adipokines in rheumatic diseases, such as rheumatoid arthritis (RA), osteoarthritis (OA), and systemic lupus erythematosus (SLE).

En este trabajo se ha identificado a los principios activos amitriptilina, naloxona, talidomida y ácido β boswélico como antagonistas del receptor TLR4 capaces de bloquear las vías de señalización de la respuesta inmune innata TLR4, IL1R, NFκβ y NLRP3 en las células articulares de condrocito artrósico humano aislado de pacientes, así como de sinoviocitos y osteoblastos. Estos resultados se obtuvieron por minería de datos, química computacional, ensayos de actividad bioquímica, análisis dirigidos de mRNA y de proteína, así como estudios de proteómica celular y de secretoma. Además, se usó inhibidores farmacológicos (CLI095) y de silenciamiento mRNA (siRNA de TLR4 y NLRP3) para validar el rol de los principios activos como inhibidores del TLR4. Las mismas evidencias en artrosis apuntaron a un potencial uso en gota, que se vio validado por la inhibición de los factores inmunes innatos del líquido sinovial artrósico y gotoso. En este contexto se realizó un paso hacia adelante en la traslación clínica al aunar evidencias in vitro preclínicas con evidencias del minado de datos clínicos. Se observó que el consumo de amitriptilina redujo la necesidad de medicación vinculada a la gota aguda y crónica, específicamente colchicina. Estos resultados muestran el potencial uso del bloqueo de TLR4 por parte de amitriptilina, naloxona, talidomida, ácido β boswélico como herramienta terapéutica para inhibir la respuesta inmune innata mediada por TLR4, IL1R, NFκβ y NLRP3 la cual se encuentra presente en múltiples enfermedades reumáticas como la artrosis y la gota. 2023-03-25

Author(s): Hughes-Austin, Jan M; Deane, Kevin D; Giles, Jon T; Derber, Lezlie A; Zerbe, Gary O; Dabelea, Dana M; Sokolove, Jeremy; Robinson, William H; Holers, V Michael; Norris, Jill M | Abstract: BACKGROUND:Extra-articular manifestations of rheumatoid arthritis (RA), potentially due to systemic inflammation, include cardiovascular disease and sarcopenic obesity. Adiponectin, an adipose-derived cytokine, has been implicated in inflammatory processes in RA, but little is known regarding its association with inflammation in a pre-clinical period. Therefore, we investigated whether adiponectin was associated with inflammatory markers in individuals at risk for RA, and whether RA-related autoimmunity modifies these associations. METHODS:We analyzed samples from 144 first-degree relatives (FDRs) of RA probands, of whom 23 were positive for anti-cyclic citrullinated peptide antibody and/or ≥ 2 rheumatoid factor isotypes (IgM, IgG or IgA). We called this phenotype the 'high risk autoantibody profile (HRP)' as it has been shown in prior work to be g96% specific for future RA. We measured adiponectin, cytokines, and high-sensitivity C-reactive protein (hsCRP). Using linear mixed effects models, we evaluated interaction between HRP positivity and adiponectin on inflammatory markers, adjusting for age, sex, ethnicity, body mass index, pack-years smoking, and use of cholesterol-lowering medications. RESULTS:In everyone, adiponectin concentration was inversely associated with hsCRP and IL-1β in adjusted models, where a 1% higher adiponectin was associated with a 26% lower hsCRP (p = 0.04) and a 26% lower IL-1β (p = 0.04). Significant interactions between HRP and adiponectin for associations with GM-CSF, IL-6, and IL-9 were detected in fully adjusted models (p = 0.0006, p = 0.006, p = 0.01, respectively). In HRP positive FDRs but not HRP negative FDRs, a 1% higher adiponectin was associated with 97% higher GM-CSF, 73% higher IL-6, and 54% higher IL-9 concentrations. CONCLUSIONS:Adiponectin associates with inflammatory markers, and these associations differ in individuals with a high-risk autoantibody profile compared with those without. The interaction between adiponectin and autoimmunity warrants further investigation into the potential systemic effects of RA-related autoantibodies and adiponectin on inflammation in the absence of clinically apparent RA.