Pesquisa de biomarcadores para perda muscular em pacientes com artrite reumatoide : uma análise de metaboloma urinário

Introdução: A artrite reumatoide (AR) é uma doença autoimune que afeta as articulações e evolui com inflamação crônica e destruição local do tecido. Além disso, pacientes com AR podem apresentar manifestações extra-articulares, como alterações na composição corporal. Freqüentemente, perda de músculo esquelético é observada em pacientes com AR. Os métodos de avaliação da perda muscular são caros e pouco disponíveis, limitando seu uso na prática clínica e no desenvolvimento de estudos longitudinais. Estudos recentes de análise do metaboloma têm demonstrado grande potencial na identificação de alterações no perfil metabólico em pacientes com doenças autoimunes e podem fornecer um melhor entendimento dos mecanismos de patogenicidade, diagnóstico precoce e acompanhamento do tratamento. Assim, o perfil metabolômico da urina em pacientes com AR pode ser uma ferramenta útil na identificação de biomarcadores da perda de músculo esquelético. Objetivo: Avaliar o perfil metabolômico urinário de pacientes com artrite reumatoide e associá-lo à perda de músculo esquelético. Métodos: Foram recrutados pacientes com AR de acordo com os critérios de classificação ACR / EULAR 2010, com idade entre 40 e 70 anos. Dados clínicos, atividade da doença e composição corporal foram avaliados e amostras de urina foram coletadas. A atividade da doença foi medida pelo Disease Activity Score-28 com proteína C reativa (DAS28-PCR). A massa muscular foi definida pelo índice de massa magra apendicular (ALMI; kg / altura2), derivado de exames de DXA, que é a soma do tecido magro dos braços e pernas. A análise do metaboloma da urina foi realizada por espectroscopia de Ressonância Magnética Nuclear (RMN) utilizando os softwares Bayesil e Metaboanalyst. Foram utilizados os modelos estatísticos PCA e PLSDA seguidos da correlação de Spearman, e p <0,05 foi considerado estatisticamente significativo. Curva ROC e regressão logística foram utilizadas para estabelecer um modelo diagnóstico. Resultados: Noventa pacientes com AR foram incluídos. A maioria dos pacientes era do sexo feminino (86,7%), com média de idade de 56,0 ± 7,3 anos e mediana do DAS28-CRP de 3,0 (IQR, 1,0–3,0). Nós identificamos 15 metabólitos que mostraram altas pontuações VIP pelo Metaboanalista. Dimetilglicina (r = 0,2; p = 0,053), oxoisovalerato (r = - 0,2; p = 0,055) e ácido isobutírico (r = - 0,249; p = 0,018) apresentaram correlação significativa com ALMI. Área sob a curva ROC (AUC) com base na massa muscular (≤6,0 kg / m2 de ALMI para mulheres e ≤8,1 kg / m2 para 7 homens) regressão logística foi usada para estabelecer um modelo diagnóstico como dimetilglicina (AUC = 0,65), oxoisovalerato ( AUC = 0,49) e ácido isobutírico (AUC = 0,83) com sensibilidade e especificidade significativas. Conclusão: O ácido isobutírico, oxoisovalerato e dimetilglicina observados em amostras de urina foram associados à baixa massa muscular esquelética em pacientes com AR. Esses achados podem sugerir que esse grupo de metabólitos pode ser testado como biomarcador para o diagnóstico de perda muscular.Introduction: Rheumatoid arthritis (RA) is an autoimmune disease that affects joints and progresses with chronic inflammation and local tissue destruction. Additionally, patients with RA may present extra-articular manifestations, such as changes in body composition. Often, skeletal muscle wasting is observed in RA patients. Methods for assessing muscle loss are costly and not widely available, limiting their use in clinical practice and the development of longitudinal studies. Recent studies of metabolome analysis have shown a great potential in identifying changes in the metabolite profile in patients with autoimmune diseases, and can provide a better understanding of pathogenicity mechanisms, early diagnosis, and treatment follow-up. Thus, urine metabolomic profile in RA patients could be a useful tool in identifying skeletal muscle wasting. Objective: To evaluate the urinary metabolomic profile of patients with rheumatoid arthritis and associate it with skeletal muscle loss. Methods: Patients with RA according to the 2010 ACR/EULAR classification criteria, aged between 40 and 70 years, were recruited. Clinical data, disease activity and body composition were evaluated and urine samples were collected. Disease activity was measured by the Disease Activity Score-28 with C-reactive protein (DAS28-PCR). Muscle mass was defined by appendicular lean mass index (ALMI; kg/height2), derived from DXA scans, that is the sum of the lean tissue in the arms and legs. The urine metabolome analysis was performed by Nuclear Magnetic Resonance (NMR) spectroscopy using Bayesil and Metaboanalyst software. The statistical models PCA and PLSDA were used followed by Spearman correlation, and p<0.05 was considered statistically 8 significant. ROC curve and logistic regression was used to establish a diagnostic model. Results: Ninety RA patients was included. Most patients were women (86.7%), with a mean age of 56.0 ± 7.3 years and a median DAS28-CRP of 3.0 (IQR, 1.0–3.0). We identified 15 metabolites that showed high VIP scores by Metaboanalyst. Dimethilglicine ( r=0.2; p=0.053), oxoisovalerate ( r= – 0.2; p=0.055) and isobutiric acid (r= – 0.249; p= 0.018) showed significant correlation with ALMI. Area under the ROC curve (AUC) based on muscle mass (≤6.0 kg/m2 of ALMI for women and ≤8.1 kg/m2 for men) logistic regression were used to establish a diagnostic model as dimethilglycine (AUC=0.65), oxoisovalerate (AUC=0.49) and isobutiric acid (AUC=0.83) with appropriate sensitivity and specificity. Conclusion: Isobutyric acid, oxoisovalerate and dimethilglicine observed in urine samples were associated with low skeletal muscle mass in patients with RA. These findings may suggest that this group of metabolites can be tested as biomarkers for diagnosis of muscle loss

A review of metabolomic profiling in rheumatoid arthritis : bringing new insights in disease pathogenesis, treatment and comorbidities

Metabolomic analysis provides a wealth of information that can be predictive of distinctive phenotypes of pathogenic processes and has been applied to better understand disease development. Rheumatoid arthritis (RA) is an autoimmune disease with the establishment of chronic synovial inflammation that affects joints and peripheral tissues such as skeletal muscle and bone. There is a lack of useful disease biomarkers to track disease activity, drug response and follow-up in RA. In this review, we describe potential metabolic biomarkers that might be helpful in the study of RA pathogenesis, drug response and risk of comorbidities. TMAO (choline and trimethylamine oxide) and TCA (tricarboxylic acid) cycle products have been suggested to modulate metabolic profiles during the early stages of RA and are present systemically, which is a relevant characteristic for biomarkers. Moreover, the analysis of lipids such as cholesterol, FFAs and PUFAs may provide important information before disease onset to predict disease activity and treatment response. Regarding therapeutics, TNF inhibitors may increase the levels of tryptophan, valine, lysine, creatinine and alanine, whereas JAK/STAT inhibitors may modulate exclusively fatty acids. These observations indicate that different disease modifying antirheumatic drugs have specific metabolic profiles and can reveal differences between responders and non-responders. In terms of comorbidities, physical impairment represented by higher fatigue scores and muscle wasting has been associated with an increase in urea cycle, FFAs, tocopherols and BCAAs. In conclusion, synovial fluid, blood and urine samples from RA patients seem to provide critical information about the metabolic profile related to drug response, disease activity and comorbidities

Metabolomic biomarker candidates for skeletal muscle loss in the collagen-induced arthritis (CIA) model

There is no consensus for diagnosis or treatment of RA muscle loss. We aimed to investigate metabolites in arthritic mice urine as biomarkers of muscle loss. DBA1/J mice comprised collagen-induced arthritis (CIA) and control (CO) groups. Urine samples were collected at 0, 18, 35, 45, 55, and 65 days of disease and subjected to nuclear magnetic resonance spectroscopy. Metabolites were identified using Chenomx and Birmingham Metabolite libraries. The statistical model used principal component analysis, partial least-squares discriminant analysis, and partial least-squares regression analysis. Linear regression and Fisher’s exact test via the MetaboAnalyst website were performed (VIP-score). Nearly 100 identified metabolites had CIA vs. CO and disease time-dependent differences (p < 0.05). Twenty-eight metabolites were muscle-associated: carnosine (VIPs 2.8 × 102) and succinyl acetone (VIPs 1.0 × 10) showed high importance in CIA vs. CO models at day 65; CIA pair analysis showed histidine (VIPs 1.2 × 102) days 55 vs. 65, histamine (VIPs 1.1 × 102) days 55 vs. 65, and L-methionine (VIPs 1.1 × 102) days 0 vs. 18. Carnosine was fatigue- (0.039) related, creatine was food intake- (−0.177) and body weight- (−0.039) related, and both metabolites were clinical score- (0.093; 0.050) and paw edema- (0.125; 0.026) related. Therefore, muscle metabolic alterations were detected in arthritic mice urine, enabling further validation in RA patient’s urine, targeting prognosis, diagnosis, and monitoring of RA-mediated muscle loss

Practical screening tools for sarcopenia in patients with systemic sclerosis

Introduction In view of the method of diagnosing sarcopenia being complex and considered to be difficult to introduce into routine practice, the European Working Group on Sarcopenia in Older People (EWGSOP) recommends the use of the SARC-F questionnaire as a way to introduce assessment and treatment of sarcopenia into clinical practice. Only recently, some studies have turned their attention to the presence of sarcopenia in systemic sclerosis (SSc).There is no data about performance of SARC-F and other screening tests for sarcopenia in this population. Objective To compare the accuracy of SARC-F, SARC-CalF, SARC-F+EBM, and Ishii test as screening tools for sarcopenia in patients with SSc. Methods Cross-sectional study of 94 patients with SSc assessed by clinical and physical evaluation. Sarcopenia was defined according to the revised 2019 EWGSOP diagnostic criteria (EWGSOP2) with assessments of dual-energy X-ray absorptiometry, handgrip strength, and short physical performance battery (SPPB). As case finding tools, SARC-F, SARC-CalF, SARC-F+EBM and Ishii test were applied, including data on calf circumference, body mass index, limitations in strength, walking ability, rising from a chair, stair climbing, and self reported number of falls in the last year. The screening tests were evaluated through receiver operating characteristic (ROC) curves. Standard measures of diagnostic accuracy were computed using the EWGSOP2 criteria as the gold standard for diagnosis of sarcopenia. Results Sarcopenia was identified in 15 (15.9%) patients with SSc by the EWGSOP2 criteria. Area under the ROC curve of SARC-F screening for sarcopenia was 0.588 (95% confidence interval (CI) 0.420–0.756, p = 0.283). The results of sensitivity, specificity, positive likelihood ratio (+LR), negative likelihood ratio (-LR) and diagnostic Odds Ratio (DOR) with the EWGSOP2 criteria as the gold standard were 40.0% (95% CI, 19.8–64.2), 81.0% (95% CI, 71.0–88.1), 2.11 (95% CI, 0.98–4.55), 0.74 (95% CI, 0.48–1.13) and 2.84 (95% CI, 0.88–9.22), respectively. SARC-CalF and SARC-F+EBM showed better sensitivity (53.3%, 95% CI 30.1–75.2 and 60.0%, 95% CI 35.7–80.2, respectively) and specificity (84.8%, 95% CI 75.3–91.1 and 86.1%, 95% CI 76.8–92.0, respectively) compared with SARC-F. The best sensitivity was obtained with the Ishii test (86.7%, 95% CI 62.1–96.3), at the expense of a small loss of specificity (73.4%, 95% CI 62.7–81.9). Comparing the ROC curves, SARC-F performed worse than SARC-CalF, SARC-F+EBM and Ishii test as a sarcopenia screening tool in this population (AUCs 0.588 vs. 0.718, 0.832, and 0.862, respectively). Direct comparisons between tests revealed differences only between SARC-F and Ishii test for sensitivity (p = 0.013) and AUC (p = 0.031). Conclusion SARC-CalF, SARC-F+EBM, and Ishii test performed better than SARC-F alone as screening tools for sarcopenia in patients with SSc. Considering diagnostic accuracy and feasibility aspects, SARC-F+EBM seems to be the most suitable screening tool to be adopted in routine care of patients with SSc

Avaliação do efeito antiproliferativo do antagonista do peptídeo liberador de gastrina, RC-3095, e em associação com a temozolamida em modelos experimentais de gliomas

Gliomas apresentam um prognóstico precário apesar das múltiplas modalidades terapêuticas como ressecção neurocirúrgica, radioterapia e quimioterapia, possivelmente pelo seu alto nível de invasividade e resistência aos tratamentos. Estudos recentes demonstraram que o peptídeo liberador de gastrina (GRP), assim como seu receptor, possui efeitos importantes no desenvolvimento de muitos tumores, inclusive de gliomas. O receptor de GRP está super expresso em linhagens celulares de glioblastomas tendo sua função relacionada com a proliferação celular, estimulando o crescimento tumoral. No presente estudo, foi investigado o efeito do RC-3095, um antagonista seletivo do receptor de GRP, como mono terapia e em associação à temozolamida (TMZ), um agente citotóxico alquilante de DNA, em modelos experimentais de células de glioma da linhagem C6 de rato in vitro e in vivo. A proliferação celular foi reduzida tanto por RC-3095 quanto por TMZ, porém quando ambos os agentes foram administrados em associação, a redução da proliferação foi significativamente mais efetiva. Nos experimentos in vivo, o grupo controle apresentou os tumores maiores (52 ± 15.5 mm3), enquanto RC-3095 reduziu o tamanho tumoral, tendo um efeito mais efetivo na dose 0.3 mg/kg (21 ± 9.7 mm3). Porém, a combinação dos dois agents produziu a maior redução no tamanho tumoral (10 ± 7.5 mm3), assim como, apresentou índices histológicos menos agressivos. Nossos resultados demonstram que a combinação de RC-3095 e temozolamida produz uma maior redução tumoral em modelos experimentais de glioma de rato indicando que RC-3095 pode ser um forte candidato para associação à temozolamida potencializando os efeitos de agentes alquilantes de DNA no tratamento de gliomas multiformes.Malignant gliomas have a dismal prognosis despite multi-modality treatments like neurosurgical resection, radiation therapy and chemotherapy, mainly due to their high level of invasion. Recent studies demonstrated that gastrin-releasing peptide (GRP) and its receptors play a role in the development of a variety of cancers including gliomas. The GRP receptor was shown to be over-expressed on glioblastoma cell lines and its function is related with cellular growth. In the present study, we investigated the effects of RC-3095, a selective antagonist of the GRP receptor, alone and in combination with temozolomide (TMZ), a DNA alkylating agent, in vitro and in vivo using experimental rat C6 glioma models. Cellular proliferation was reduced in all treatments (TMZ, RC-3095 and TMZ + RC-3095) with the administration of TMZ together with RC-3095 being the most effective. In in vivo experiments, the control group displayed the largest tumors (52 ± 15.5 mm3), whereas RC-3095 reduced the tumor size, with the most significant effect observed at the dose of 0.3mg/kg (21 ± 9.7 mm3). The combined therapy produced the largest reduction in tumor size (10 ± 7.5 mm3), greater than with either treatment alone. Ours results show that the combination of RC-3095 and TMZ have additive effects on the in vitro and in vivo glioma growth therefore making RC-3095 a candidate drug to potentiate the effects of the DNA alkylanting agent temozolomide in the treatment of malignant gliomas

Estudo da eficácia das drogas temozolamida e RC-3095 em modelos de glioma in vitro e in vivo

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Combinação de temozolamida e RC-3095 em modelos in vivo para tratamento de gliomas

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Transferência gênica utilizando um vetor não viral para Mucopolissacaridose Tipo I

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Transferência gênica utilizando um vetor não viral para Mucopolissacaridose Tipo I

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