Observational Study Medicine ® OPEN Association of myostatin, a cytokine released by muscle, with inflammation in rheumatoid arthritis A cross-sectional study Jessica D. Murillo-Saich, PhDa, Maria Luisa Vazquez-Villegas, MD, PhDb,c, Melissa Ramirez-Villafaña, PhDd, Ana Miriam Saldaña-Cruz, PhDd, Javier A. Aceves-Aceves, MD, MSce, Laura Gonzalez-Lopez, MD, PhDe,f, ∗ Monica Guma, MD, PhDa,g, Jorge I. Gamez-Nava, MD, PhDe,h, Abstract Myostatin is a cytokine produced and released by myocytes that might have an outstanding role not only in muscle wasting during cachexia but also in inflammation. Herein we explore the association between myostatin levels and inflammatory parameters in rheumatoid arthritis (RA). One hundred twenty-seven women without rheumatic diseases and 84 women with a diagnosis of RA were assessed in a cross- sectional study. Outcomes reflecting the activity of the arthritis including Disease Activity Score (DAS28-ESR) and impairment in functioning by the Health Assessment Questionnaire-Disability Index were assessed in RA. We obtained Skeletal muscle mass index (SMI), fat-free mass index (FFMI), and fat mass index using dual-energy x-ray absorptiometry. Serum myostatin was determined by enzyme-linked immunosorbent assay. Myostatin levels were correlated with disease activity and parameters of muscle mass. The SMI was lower and concentration of myostatin was higher in RA patients than in controls (P = .008 and P < .001, respectively). Myostatin significantly positively correlated with C-reactive protein (rho = 0.48, P < .001), erythrocyte sedimentation rate (rho = 0.28, P = .009), and DAS28-ESR (rho = 0.22, P = .04), and negatively correlated with SMI (rho = 0.29, P = .008), (FFMI) (rho = 0.24, P = .027). In the multivariate logistic regression analysis, levels of myostatin remained associated with disease activity in RA (P = .027). In our study, myostatin was associated with disease activity in RA patients, suggesting a mechanistic link between myostatin, muscle wasting and inflammation in RA. Abbreviations: 95% CI = 95% confidence interval, BMI = body mass index, CL = controls, CRP = C-reactive protein, DAS28 = disease activity score, DAS28-ESR = Disease activity score using erythrocyte sedimentation rate, DMARD = disease-modifying anti- rheumatic drug, DXA = Dual-energy X ray absorptiometry, ELISA = enzyme-linked immunosorbent assay, ESR = erythrocyte sedimentation rate, FFMI = fat-free mass index, FMI = fat mass index, GC = glucocorticoids, HAQ-DI = Health assessment questionnaire-disability index, IL-1b = interleukin-1 beta, IL-6 = interleukin-6, Kg = kilograms, mg/day = milligrams per day, mg/dL = milligrams/deciliter, mm = millimeters, mm/h = millimeters/hour, n = number, ng/mL = nanograms/milliliter, NSAID = non-steroidal anti-inflammatory drug, P = P value, RA = rheumatoid arthritis, ROC = receiver-operating characteristic, SMI = skeletal muscle mass index, SMME = skeletal muscle mass of extremities, SPSS = Statistical package for the social science, TGF-b = transforming growth factor-Beta, TNF = tumor necrosis factor, VAS = Visual Analogue Scale. Keywords: diseases activity, inflammation, muscle, myopenia, myostatin, rheumatoid arthritis, rheumatoid cachexia Editor: Bingrong Zhou. The grant was obtained from “Fondo en Investigación en Salud” of Instituto Mexicano del Seguro Social. Grant: FIS/IMSS/PROT/MD16/1565. JDM-S currently is a recipient of a postdoctoral scholarship supported by the National Institutes of Health (T32AR064194 to JDM-S). JDM-S and MLV-V participated equally as first authors. The authors report no conflicts of interest. Supplemental Digital Content is available for this article. The datasets generated during and/or analyzed during the current study are available from the corresponding author on reasonable request. a Division of Rheumatology, Allergy and Immunology, UC San Diego School of Medicine, La Jolla, CA, b Departamento de Epidemiologia, Unidad de Medicina Familiar blica del Departamento de Salud Pu N°, Instituto Mexicano del Seguro Social (IMSS), c Instituto Regional de Investigación en Salud Pu blica del Centro Universitario de Ciencias (CUCS) de la Salud de la Universidad de Guadalajara (UdeG), d Departamento de Fisiologia, CUCS, U de G, e Programa de Doctorado en Farmacología, CUCS, U de G, f Departamento de Medicina Interna-Reumatología, Hospital General Regional 110, IMSS, Guadalajara, Jalisco, Mexico, g Department of Medicine, Autonomous University of Barcelona, Plaça Cívica, Bellaterra, Barcelona, Spain, h Unidad de Investigacion Biomedica 02 IMSS, Guadalajara, Jalisco, Mexico. ∗ Correspondence: Jorge I. Gamez-Nava, Avenida Salto del Agua 2192, Colonia Jardines del Country, Guadalajara Jalisco, CP 44210, Mexico (e-mail: drivangamez@prodigy.net.mx, alternative e-mail: ivangamezacademicoudg@gmail.com). Copyright © 2021 the Author(s). Published by Wolters Kluwer Health, Inc. This is an open access article distributed under the terms of the Creative Commons Attribution-Non Commercial License 4.0 (CCBY-NC), where it is permissible to download, share, remix, transform, and buildup the work provided it is properly cited. The work cannot be used commercially without permission from the journal. How to cite this article: Murillo-Saich JD, Vazquez-Villegas ML, Ramirez-Villafaña M, Saldaña-Cruz AM, Aceves-Aceves JA, Gonzalez-Lopez L, Guma M, Gamez-Nava JI. Association of myostatin, a cytokine released by muscle, with inflammation in rheumatoid arthritis: A cross-sectional study. Medicine 2021;100:3(e24186). Received: 1 February 2020 / Received in final form: 1 December 2020 / Accepted: 12 December 2020 http://dx.doi.org/10.1097/MD.0000000000024186 1 Murillo-Saich et al. Medicine (2021) 100:3 Medicine 1. Introduction (SMME) ([arms + legs]/adjusted by height squared).[10] Rheu- Myostatin which is part of the transforming growth factor-b matoid cachexia was defined using the Engvall et al criteria superfamily, is a cytokine produced and released by myocytes, according to the following values obtained of the comparison that negatively regulates skeletal muscle in humans and animal group: fat-free mass index (FFMI) below the 10th percentile and models.[1] Previous work has linked myostatin with muscle fat mass index (FMI) above the 25th percentile.[11] wasting in several chronic diseases including rheumatoid arthritis (RA).[2,3] RA is an autoimmune disease characterized by synovial 2.3. Quantification of serum myostatin levels inflammation and joint destruction. Fifty percent of patients with Serum concentration of myostatin was measured by enzyme- RA can develop muscle loss that will lead to rheumatoid cachexia linked immunosorbent assay (ELISA) using a commercial kit in around 25% (ranges from 7% to 33%), associated to a poor (MyBioSource, Inc., San Diego, CA). quality of life.[3,4] Yet, it was suggested that myostatin could be also involved in synovitis and joint damage in animal models of RA.[5] Since RA is a multifactorial disease, it is necessary to 2.4. Ethics explore the potential role of other cytokines, such myostatin, to The privacy of the patients was protected, and this study followed understand whether it can participate in the joint inflammation. the recommendations of the 64th Declaration of Helsinki and, it Therefore, the objective of this project was to explore the was conducted under the ethical standards of the Ethics and association of myostatin and parameters of inflammation in RA. Research Board of the Centro Medico Nacional de Occidente of Instituto Mexicano del Seguro Social 13-01with approval code: 2. Methods R-2016-1301-93. 2.1. Study design 2.5. Statistical analysis This was a cross-sectional study. Eighty-four female patients fulfilling the 1987 American We performed all the analyses using SPSS v. 23 statistical College of Rheumatology classification criteria for RA were software (SPSS, Inc., Chicago, IL). We expressed the quantitative recruited. We excluded patients with pregnancy, thyroid disease, variables as medians and ranges, and qualitative variables such as creatinine >1.2 mg/dL, transaminases >2-fold the normal values, frequencies and percentages. We used x2 and Mann–Whitney U cancer, chronic virus infections (such as tuberculosis, human test to compare demographic variables, skeletal muscle mass immunodeficiency virus infections, hepatitis B, hepatitis C). A index, and serum concentrations of myostatin between the RA structured chart review was performed for investigating patients and control group for qualitative and quantitative epidemiological characteristics, comorbid diseases, treatments, variables, respectively. Also, we used Spearman correlation to and other factors potentially related with abnormalities on determine the strength of association between myostatin and musculoskeletal mass such as sedentarism. Sedentarism was clinical variables in RA patients. To evaluate the levels of assessed using a structured questionnaire and it was considered myostatin as a good classifier of disease activity in RA, we when a person practiced <150 minutes of moderate activity per performed a receiver-operating characteristic (ROC) curve. We week or <75 minutes of vigorous-intensity activity per week.[6] performed a multivariate logistic regression analysis including as Clinical assessment included Disease Activity Score (DAS28) a dependent variable moderate or severe disease activity adjusted which is a validated composite index that includes measure joint by age, disease duration, SMI, FMI, and the cutoff value of high swollen count, tender joint count, a patient’s perception of levels of myostatin based on ROC curve analysis. We considered disease severity (assessed with a visual analogue scale that goes P < .05 as a result with significance statistic. from 0 to 100 mm) and erythrocyte sedimentation ratio (ESR).[7] We used DAS28-ESR as disease activity index. This index 3. Results classifies disease activity into 4 categories: remission <2.6; low disease activity ≥2.6 to 3.2; moderate disease activity >3.2 to We included 127 women without rheumatic diseases (controls) 5.1; and high disease activity >5.1. One of the main objectives and 84 women diagnosticated with RA. We show in Table 1 a of treatment in RA is to achieve low disease activity or remission comparison between controls and RA patients. The age was (DAS28-ESR 3.2). Patients with DAS28-ESR >3.2 are similar (60 [24–85] vs 63 [24–89] years’ old, P = .57). RA patients considered with moderate or severe disease activity requiring were more sedentary, and a higher percentage were menopausal. therapeutic modifications.[8] We also included for assessing BMI and other comorbidities including high blood pressure, functional disability the Spanish validated version of the Health diabetes mellitus and smoking were not different between both Assessment Questionnaire-Disability Index (HAQ-DI).[9] groups. Concentrations of myostatin in circulation were For the control group, we included 127 females with similar significantly higher in the RA group than controls (9 [1.2– age and body mass index (BMI) and absence of inflammatory or 140] ng/mL vs 3.5 [1–89.9] ng/mL respectively, P < .001). autoimmune disorders, cancer, chronic kidney diseases, endo- Skeletal muscle mass index was significant diminished in the RA crinopathies, and active infections. group (5.8 [3.4–10.0] vs 6.2 [3.8–9.0] RA vs controls, respectively, P = .008)). As it is described in Table 2, 12 patients (14.3%) met the 2.2. Quantification of skeletal muscle mass criteria for rheumatoid cachexia. To identify whether RA is Skeletal muscle mass index (SMI) was obtained with a dual- associated with high levels of myostatin independently of energy x-ray absorptiometry (DXA) (Lunar Prodigy Advance rheumatoid cachexia, we compared RA patients without 2000 equipment, General Electric). SMI was computed using the rheumatoid cachexia (n = 72) versus controls, and we observed following formula: sum of skeletal muscle mass of extremities that these patients continued to show high levels of myostatin 2 Murillo-Saich et al. Medicine (2021) 100:3 www.md-journal.com Table 1 Table 3 Comparison of selected characteristics, body composition and Correlation of serum myostatin concentrations with epidemiolo- myostatin levels between CL group and RA group. gical and body composition variables in Rheumatoid arthritis. Variables CL (n = 127) RA (n = 84) P Myostatin, n = 84 Age, y 60 (24–85) 63 (24–89) .57 Variables Rho P Alcoholism, n (%) 15 (11.8) 4 (4.9) .09 Age, y 0.09 .37 Smoking, n (%) 11 (8. 7) 8 (9.5) .83 RA disease duration, y 0.24 .02 BMI 27.8 (16–42) 27.3 (18–40) .35 HAQ-DI (score) 0.05 .65 Sedentarism, n (%) 58 (46) 52 (61. 9) .02 DAS28-ESR (score) 0.22 .04 Menopause, n (%) 85 (68) 76 (90.5) <.001 Tender joint counts 0.11 .30 Comorbidities Swollen joint counts 0.19 .08 High blood pressure, n (%) 41 (33. 3) 35 (41. 7) .22 Severity of disease activity 0.03 .79 Diabetes mellitus type 2, n (%) 27 (22.1) 12 (14.3) .16 Pain perceived by the patient (VAS) 0.05 .64 Body composition characteristics Stiffness perceived by the patient (VAS) 0.02 .86 Weight, kg 67 (38–97) 64 (42–98) .15 CRP, mg/dL 0.40 <.001 Skeletal Muscular Mass Index 6.2 (3.8–9) 5.8 (3.4–10) .008 ESR, mm/h 0.28 .009 Laboratory data Weight, kg 0.15 .18 Myostatin, ng/mL 3.5 (1–89.9) 9 (1.2–140) <.001 Skeletal muscle mass index 0.29 .008 Qualitative variables are expressed in frequencies (%) and quantitative variables are expressed in Fat mass index 0.028 .80 median and ranges. BMI = body mass index, CL = control group, RA = rheumatoid arthritis group. Free fat mass index 0.24 .027 Comparison of qualitative variables performed with Chi-square. Comparison of quantitative variables Total glucocorticoid dose, mg/day 0.51 <.001 was performed using Mann–Whitney U test. Statistical significance P < .05. CRP = C-reactive protein, DAS28-ESR = disease activity score using erythrocyte sedimentation rate, ESR = erythrocyte sedimentation rate, ESR = Erythrocyte sedimentation rate, HAQ-DI = Health assessment questionnaire-disability index, Rho = Spearman correlation, VAS = Visual Analogue Scale. Statistical significance P < .05. compared to controls (8.05 [1.2–140] vs 3.46 [1.01–89.9] respectively, P < .001). Table 2 shows the characteristics of the RA patient group. The muscle mass were sedentary and 37 of 60 (61.7%) RA patients median of HAQ-DI was 0.11 (0–2.7), DAS28-ESR was 2.8 (0.9– with normal muscle mass were classified as sedentary (P = 1.0). 7.1), ESR was 20.5 (1–64), CRP was 10.2 (0.3–219). Only 6% However, 11 of 12 (91.7%) RA patients with cachexia were were receiving biological therapy, 96% were receiving a synthetic sedentary, whereas 41 of 72 (56.9%) RA patients without disease-modifying anti-rheumatic drug (DMARD), mostly meth- cachexia were classified as sedentary (P = .025). otrexate, and 71% were on glucocorticoids (GC), with an Table 3 shows the correlation between serum myostatin levels average of 5 mg/day (0–17.5). We observed no significant with quantitative variables in RA patients. Higher myostatin difference in myostatin levels between RA patients with or levels were correlated with low skeletal muscle mass index (P = without cachexia (13.2 [2.5–140.0] vs 8.05 [1.2–140] respec- 0.008). Interestingly, myostatin concentrations correlated with tively; P = .35). Fifteen of 24 (62.5%) RA patients with low- inflammation parameters of DAS28-ESR (rho = 0.22, P = .04), CRP (rho = 0.40, P < .001), and ESR (rho = 0.28, P = .009). Myostatin concentrations also correlated with years since RA Table 2 diagnosis (rho = 0.24, P = .02), SMI (rho = 0.29, P = 0.008), Clinical variables in patients with rheumatoid arthritis. FFMI (rho = 0.24, P = .027), and GC daily doses (rho = 0.51, P < .001). However, we did not observe correlation between FMI Variables n = 84 and myostatin levels (rho = 0.028, P = .80). In data that are not Age, y 63 (24–89) depicted in tables we observed a correlation between SMI with Years since RA diagnosis 13 (0–45) CRP (rho = 0.26, P = .04) and HAQ-DI (rho = 0.21, P = .05). HAQ-DI 0.11 (0–2.7) We show the results of a subanalysis aimed at identifying whether DAS28-ESR 2.8 (0.9–7.1) previously observed correlations between myostatin levels and DAS28-ESR moderate-severe (>3.2), n (%) 31 (36.9) Rheumatoid cachexia, n (%) 12 (14.3) parameters of inflammation were independent of the loss of Treatment muscle mass observed in rheumatoid cachexia (See Supplemental Biological drugs, n (%) 5 (6) Digital Content table S1, http://links.lww.com/MD/F526, show- Glucocorticoids, n (%) 71 (85.5) ing the correlations of serum myostatin concentrations with Dose of glucocorticoids, mg/day 5 (0–17.5) variables associated with inflammatory activity in the subgroup DMARD, n (%) 80 (96.4) of patients without rheumatoid cachexia). For this correlation NSAID, n (%) 78 (95.1) analysis, we excluded the subgroup of patients with rheumatoid Laboratory cachexia (n = 12). In the results of these analyses, the variables of ∗ CRP, mg/dL 10.2 (0.3–219) inflammatory disease activity (DAS28-ESR, CRP, and ESR) ESR, mm/h 20.5 (1–64) remained correlated with myostatin levels. Myostatin, ng/mL 9 (1.2–140) We also performed a ROC curve analysis on myostatin levels Qualitative variables are expressed in frequencies (%) and quantitative variables are expressed in to evaluate levels of myostatin as a classifier for moderate or median and ranges. CRP = C-reactive protein, DAS28-ESR = disease activity score using erythrocyte severe disease activity (DAS28-ESR >3.2) (Fig. 1). The area sedimentation rate, DMARD = synthetic disease-modifying anti-rheumatic drug, ESR = erythrocyte sedimentation rate, HAQ-DI = Health assessment questionnaire-disability index, NSAID = non- under the curve was 0.61. We obtained a sensibility of 61.3% and steroidal anti-inflammatory drug. specificity of 53% to detect moderate or severe disease activity in ∗ Data of 78 of 84 RA patients of CRP were available at the time of the study. RA, with a cutoff point of 13 ng/mL of myostatin. 3 Murillo-Saich et al. Medicine (2021) 100:3 Medicine fluid of patients with RA. These authors also showed that Serum levels of myostatin myostatin is able to induce IL-1b production and to increase TNF expression in synovial fibroblasts, probably through phospha- 100 tidylinositol 3-kinase signaling pathway, suggesting a link between this myokine and synovial inflammation.[13,14] In addition, in an experimental model of chronic kidney disease 80 in mice, Zhang et al administered an anti-myostatin peptibody to produce a pharmacological inhibition of myostatin and identify the effects of myostatin suppression on muscle atrophy and chronic inflammation.[15] A significant increase in muscle mass Sensitivity% 60 and a suppression of circulating inflammatory cytokines including TNF and IL-6 was observed after the administration of this peptibody, supporting our findings of a connection 40 between myostatin levels, muscle wasting and inflammation. Our work shows an association between circulating levels of myostatin and myopenia determined by DXA with inflammatory 20 markers in our RA cohort. This result is similar to the association described in a Chinese RA population, which also showed a significant correlation between muscle loss and joint destruction in RA,[16] although our report would be the first one to show 0 correlation between myostatin levels and components of disease 0 20 40 60 80 100 activity in RA patients. We found our findings also supported by 100% - Specificity% the study performed by Kerschan-Schindl et al, who observed Figure 1. Receiver-operating characteristic curve of serum myostatin levels. that RA patients on remission had lower concentration of The area under the curve was 0.61 with a P = .09. Using cutoff value of ≥13 ng/ circulating myostatin compared with active patients.[17] mL of myostatin, we obtained a sensibility of 61.3% and specificity of 53% to detect moderate or severe disease activity in rheumatoid arthritis patients. We also showed a significant correlation between myostatin and CRP and ESR; these correlations remained significant in the subanalysis after excluding patients with rheumatoid cachexia. These results imply that myostatin can be considered as potential Finally, in the multivariate logistic regression analysis, we biomarker of disease activity independently of severe muscle identified that myostatin serum levels remain associated with mass losses. CRP is an acute-phase response molecule which moderate or severe disease activity (DAS28-ESR >3.2) in RA display a high expression in inflammatory condition such as RA patients after adjusting by age, disease duration, SMI, and FMI and can be induced by some inflammatory cytokines such as IL-6 (P = .027) (Table 4). and IL-1b.[18] Several years ago it was shown that CRP was localized in RA synovial tissue, suggesting a potential pathogenic role of CRP.[19] Interestingly, muscle is a relevant source of IL-6 4. Discussion and this cytokine promotes atrophy and muscle wasting,[20] Myostatin induces muscle atrophy inhibiting myogenesis and which could also be important in this association between developing myopenia, a clinical condition characterize for myostatin, myopenia, and inflammation in our study. Altogether wasting of the muscle fibers due to an illness no matter the myostatin could be a potential player in RA pathogenesis and a age.[12] Yet, previous works have also suggested a link between promising target in RA therapy similar to what has been myostatin and inflammation.[5,13,14] RA synovial tissue expresses suggested in other diseases.[2] higher amounts of myostatin compared to normal synovial tissue, Although this positive correlation between DAS-28 and and was found to accelerate osteoclast formation and joint myostatin could reflect a potential role of myostatin on destruction in animal models of inflammatory arthritis.[5] In inflammation, this could be just a consequence of the sedentarism addition, other reports have shown a positive correlation in patients with high activity and poor response to the treatment. between myostatin and proinflammatory cytokines, such as, Moreover, GC promotes the expression of myostatin leading to IL-1b, and tumor necrosis factor (TNF) expression in synovial muscle atrophy and waste.[21,22] In addition, muscular waste secondary to GC recovers slowly in older individuals. In our study, 85% of the patients were taken glucocorticoids at low Table 4 doses and had a median of 63 years’ old, which could also Multivariate logistic regression analysis assessing the factors contribute to explain our positive correlation between myostatin associated with moderate or severe disease activity. and inflammation. Variables Odds ratio 95% CI P In this study, we observed poor accuracy of myostatin as a Myostatin high levels ≥13 ng/mL 2.938 1.13–7.61 .027 biomarker for moderate-severe disease activity considering a Duration of disease, y 1.052 1.001–1.107 .047 cutoff point of 13 ng/mL However, the positive correlation with Age — No significant in the model — the parameters of inflammation and negative correlation with the SMI — No significant in the model — skeletal muscular mass index, suggest a connection between myostatin, muscle wasting, and inflammation in RA. Patients Multivariate analysis logistic regression analysis. Dependent variable moderate or severe disease activity according DAS28-ESR (>3.2). Method forward stepwise. High myostatin levels were with RA have little physical activity and one possible cause is the considered ≥13 ng/mL (cutoff value obtained of ROC analysis).95% CI = 95% confidence interval, pain and inflammation at the time of inclusion. This poor SMI = skeletal muscle mass index. Statistical significance was considered as P < .05. physical activity leads to continuing sedentary lifestyle that 4 Murillo-Saich et al. Medicine (2021) 100:3 www.md-journal.com contributes to muscle breakdown and increasing circulating Methodology: Laura Gonzalez-Lopez, Jorge I. Gamez-Nava. proinflammatory cytokines, and then, cachexia, among other Resources: Laura Gonzalez-Lopez. comorbidities, which involves patients into a feedback of Revising the manuscript for important intellectual content: Jorge inflammation-muscle waste-sedentarism. Yet, our multivariate I. Gamez-Nava, Laura Gonzalez-Lopez, Monica Guma. analysis suggests that some of the increase in myostatin in RA Statistical analysis: Maria Luisa Vazquez-Villegas, Melissa patients is due to the inflammation and not just sedentarism and/ Ramirez-Villafaña or cachexia. Supervision: Laura Gonzalez-Lopez, Monica Guma, Jorge I. This study has some limitations that should be discussed. First, Gamez-Nava. since this is a cross-sectional design, this work was unable to Validation: Jorge I. Gamez-Nava. identify a temporal relation between serum myostatin levels and Writing – original draft: Jessica D. Murillo-Saich, Jorge I. Gamez- disease activity outcomes in RA patients. Further longitudinal Nava, Laura Gonzalez-Lopez, Monica Guma. studies following myostatin levels in RA are required to solve this Writing – review & editing: Jessica D. Murillo-Saich, Laura limitation. Although these correlations between myostatin levels Gonzalez-Lopez, Monica Guma, Jorge I. Gamez-Nava. and several parameters of inflammation are highly significant we cannot exclude the presence of a possible incidence-prevalence References bias, inherent to any cross-sectional study. Another limitation is the presence of some confounders related with myopenia [1] Rodriguez J, Vernus B, Chelh I, et al. Myostatin and the skeletal muscle atrophy and hypertrophy signaling pathways. Cell Mol Life Sci including disease duration, use of GC and comorbid disease 2014;71:4361–71. that can influence our results. Further follow-up studies assessing [2] Smith RC, Lin BK. Myostatin inhibitors as therapies for muscle wasting myostatin levels in early RA and during different points during associated with cancer and other disorders. Curr Opin Support Palliat the time in the same group of patients with RA are required to Care 2013;7:352–60. [3] Torii M, Hashimoto M, Hanai A, et al. Prevalence and factors associated establish if the changes on myostatin levels precede the joint with sarcopenia in patients with rheumatoid arthritis. Mod Rheumatol inflammation. Finally, this study identified a low frequency of 2019;29:589–95. rheumatoid cachexia in our RA patients. However, there are [4] Santo RCE, Fernandes KZ, Lora PS, et al. Prevalence of rheumatoid several criteria used for cachexia.[11,23,24] We choose to define cachexia in rheumatoid arthritis: a systematic review and meta-analysis. J rheumatoid cachexia based on the criteria used by Engvall Cachexia Sarcopenia Muscle 2018;9:816–25. [5] Dankbar B, Fennen M, Brunert D, et al. Myostatin is a direct regulator of et al.[11] Nevertheless, the low number of patients with presence osteoclast differentiation and its inhibition reduces inflammatory joint of rheumatoid cachexia limited the statistical power to observe destruction in mice. Nat Med 2015;21:1085–90. differences in myostatin levels compared with patients without [6] Organization GWH. Global Recommendations on Physical Activity for this complication. Health. WHO Press. Available at: https://www.who.int/dietphysicalac tivity/physical-activity-recommendations-18-64years.pdf. Published In summary, myostatin was associated with disease activity in 2010. Accessed 06.19.2020, 2020. RA patients, suggesting a mechanistic link between muscle [7] Prevoo ML, van ’t Hof MA, Kuper HH, et al. Modified disease activity wasting and inflammation in RA. Yet, future studies are needed scores that include twenty-eight-joint counts. Development and valida- to further evaluate whether myostatin positively regulates tion in a prospective longitudinal study of patients with rheumatoid synovial inflammation and joint damage. arthritis. Arthritis Rheum 1995;38:44–8. [8] Anderson J, Caplan L, Yazdany J, et al. Rheumatoid arthritis disease activity measures: American College of Rheumatology recommenda- Acknowledgment tions for use in clinical practice. Arthritis Care Res (Hoboken) 2012;64:640–7. The authors thank IMSS Foundation (Fundación IMSS, A.C.) for [9] Cardiel MH, Abello-Banfi M, Ruiz-Mercado R, et al. How to measure the support for the research. health status in rheumatoid arthritis in non-English speaking patients: validation of a Spanish version of the Health Assessment Questionnaire Disability Index (Spanish HAQ-DI). Clin Exp Rheumatol 1993;11: Author contributions 117–21. [10] Baumgartner RN, Koehler KM, Gallagher D, et al. Epidemiology of Agreement to be accountable for all aspects of the work: Jessica sarcopenia among the elderly in New Mexico. Am J Epidemiol D. Murillo-Saich, Maria Luisa Vazquez-Villegas, Melissa 1998;147:755–63. [11] Engvall IL, Elkan AC, Tengstrand B, et al. Cachexia in rheumatoid Ramirez-Villafaña, Ana Miriam Saldaña-Cruz, Javier A. arthritis is associated with inflammatory activity, physical disability, and Aceves-Aceves, Laura Gonzalez-Lopez, Monica Guma, Jorge I. low bioavailable insulin-like growth factor. Scand J Rheumatol Gamez-Nava. 2008;37:321–8. Conceptualization: Jorge I. Gamez-Nava, Monica Guma, Jessica [12] Fearon K, Evans WJ, Anker SD. Myopenia-a new universal term for D. Murillo-Saich, Laura Gonzalez-Lopez. muscle wasting. J Cachexia Sarcopenia Muscle 2011;2:1–3. [13] Su C-M, Hu S-L, Sun Y, et al. Myostatin induces tumor necrosis factor-a Data curation: Maria Luisa Vazquez-Villegas, Melissa Ramirez- expression in rheumatoid arthritis synovial fibroblasts through the PI3K- Villafaña, Ana Miriam Saldaña-Cruz, Javier A. Aceves- Akt signaling pathway. J Cell Physiol 2019;234:9793–801. Aceves. [14] Hu S-L, Chang A-C, Huang C-C, et al. Myostatin Promotes Interleukin- Final approval: Jessica D. Murillo-Saich, Maria Luisa Vazquez- 1b Expression in Rheumatoid Arthritis Synovial Fibroblasts through Inhibition of miR-21-5p. Front Immunol 2017;8:1747–1747. Villegas, Melissa Ramirez-Villafaña, Ana Miriam Saldaña- [15] Zhang L, Rajan V, Lin E, et al. Pharmacological inhibition of myostatin Cruz, Javier A. Aceves-Aceves, Laura Gonzalez-Lopez, suppresses systemic inflammation and muscle atrophy in mice with Monica Guma, Jorge I. Gamez-Nava. chronic kidney disease. FASEB J 2011;25:1653–63. Formal analysis: Maria Luisa Vazquez-Villegas. [16] Lin J-Z, Liang J-J, Ma J-D, et al. Myopenia is associated with joint Founding acquisition and resources: Jorge I. Gamez-Nava, Laura damage in rheumatoid arthritis: a cross-sectional study. J Cachexia Sarcopenia Muscle 2019;10:355–67. Gonzalez-Lopez [17] Kerschan-Schindl K, Ebenbichler G, Föeger-Samwald U, et al. Rheuma- Funding acquisition: Jorge I. 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FEBS J 2013;280: [24] Elkan AC, Engvall IL, Tengstrand B, et al. Malnutrition in women with 4131–48. rheumatoid arthritis is not revealed by clinical anthropometrical [21] Schakman O, Kalista S, Barbé C, et al. Glucocorticoid-induced skeletal measurements or nutritional evaluation tools. Eur J Clin Nutr muscle atrophy. Int J Biochem Cell Biol 2013;45:2163–72. 2008;62:1239–47. 6