Abstract Background Microparticles (MPs) are membrane-derived vesicles released from cells undergoing activation or apoptosis with diverse proinflammatory and prothrombotic activities, that have been implicated in the pathogenesis of systemic sclerosis (SSc). We aimed to evaluate the plasma levels of platelet-derived microparticles (PMPs), endothelial cell-derived microparticles (EMPs), and monocyte-derived microparticles (MMPs) in SSc patients, and the association between MPs and the clinical features of SSc. Methods In this cross-sectional study, 70 patients with SSc and 35 age- and sex-matched healthy controls were evaluated. Clinical and nailfold capillaroscopy (NFC) data were obtained from all patients. Plasma levels of PMPs (CD42+/31+), EMPs (CD105+), and MMPs (CD14+) were quantified by flow cytometry. Results Patients were mainly females (90%), with a mean age of 48.9 years old. PMP, EMP, and MMP levels were significantly increased in SSc patients compared to controls (79.2% ± 17.3% vs. 71.0% ± 19.8%, p = 0.033; 43.5% ± 8.7% vs. 37.8% ± 10.4%, p = 0.004; and 3.5% ± 1.3% vs. 1.1% ± 0.5%, p < 0.0001, respectively). PMP levels were significantly higher in patients with positive anti-topoisomerase-I antibodies (p = 0.030) and in patients with a disease duration > 3 years (p = 0.038). EMP levels were lower in patients with a higher modified Rodnan skin score (p = 0.015), and in those with an avascular score > 1.5 in NFC (p = 0.042). Conclusion The increased levels of PMPs, EMPs and MMPs in scleroderma patients might indicate a possible role for these agents in the pathogenesis of this challenging disease.

Abstract Resolution 658/2022 of the Brazilian Regulatory Agency requires the determination of the permitted daily exposure (PDE) of pharmaceutical agents. Ginkgo biloba L. is used therapeutically to treat memory deficits and other brain diseases. However, published results indicate that more studies are needed to confirm the safety of Ginkgo biloba. This study aimed to evaluate the dry extract of Ginkgo biloba L. leaves PDE as an ingredient in an oral pharmaceutical product in preclinical studies using mice. Acute oral toxicity and repeated dose experiments were performed based on OECD guidelines, as well as genotoxicity tests. The results indicate that Ginkgo biloba L. has low acute toxicity, no liver toxicity, and does not alter blood glucose levels. No changes in weight gain were observed, but food intake decreased in males during the first week of treatment at the highest dose. Hematological parameters were not altered in males, whereas females presented lower leukocyte and lymphocyte counts and higher neutrophil counts at the highest dose. The lipid profile was not altered in males, whereas total cholesterol was increased in females. The estimated PDE was 0.1 mg/day and, when related to the maximum residual concentration, indicates that the cleaning process used is safe and does not require reassessment.

Abstract Background: Annexins are a group of conserved proteins which exert several regulatory functions on various cellular activities. Increased frequency and levels of antibodies against annexin V have already been observed in several autoimmune diseases including systemic sclerosis (SSc), but their role as a vascular biomarker is unknown. The aim of this study was to determine the serum levels and the dynamical behavior of anti-annexin V antibodies over a 24 months follow-up in patients with SSc. Methods: In this bicentric cross-sectional study, 70 patients with SSc were consecutively selected from March 2016 to April 2017. Demographic and clinical features, including the presence of active DUs, were collected. Serum anti-annexin V IgG and IgM antibodies were measured at baseline and after 6, 12 and 24 months of follow-up. Videocapillaroscopy was performed in all patients. Results: Among the 70 SSc patients included anti-annexin V IgG was found in 11 patients (15.7%) (range of 15.88–39.48 U/mL) and anti-annexin V IgM in 10 patients (14.3%) (range of 14.16-22.69 U/mL) at baseline. During follow-up, the number of patients who were positive for anti-annexin V IgG and IgM remained stable over 24 months. Among the patients with positive anti-annexin V IgG at baseline the frequency of patients with necrosis or amputation of extremities, forced vital capacity less than 70% and pulmonary arterial hypertension (PAH) was significantly higher than in patients with negative anti-annexin V IgG antibodies. Patients with anti-annexin V IgG had also a higher Raynaud's Condition Score and a higher Health Assessment Questionnaire Disability Index (HAQ-DI) than patients without these antibodies at baseline. Patients with positive anti-annexin V IgM at baseline presented a higher frequency of PAH, compared to those with negative anti-annexin V IgM at baseline. Conclusions: Anti-annexin V antibodies are stable and do not change their positivity during a 24 month follow-up in SSc patients. Anti-annexin V IgG was associated with more severe interstitial lung involvement and digital microangiopathy, and patients with anti-annexin V IgG or IgM had a higher occurrence of PAH indicating an association of these biomarker with more severe disease.

Abstract Background: Pulmonary arterial hypertension (PAH) is a rare and severe complication of systemic lupus erythematosus (SLE). This study aimed to evaluate clinical and laboratory risk factors associated with PAH in SLE patients. Methods: This was a retrospective case-control study in which patients with SLE with PAH (SLE-PAH) confirmed by right heart catheterization (RHC) were compared with SLE patients without PAH. Clinical and demographic variables related to SLE and PAH and nailfold capillaroscopy were evaluated by reviewing the medical records of the patients. Results: Twenty-one patients with SLE-PAH and 44 patients with SLE without PAH matched for sex and disease duration were included. The scleroderma (SD) pattern on nailfold capillaroscopy was more frequently found in patients with SLE-PAH than in those without PAH (56.3% versus 15.9%, respectively, p = 0.002). By univariate analysis, Raynaud's phenomenon, history of abortion, and SD pattern on capillaroscopy were associated with PAH. Arthritis was a protective factor for PAH development. Multivariate analysis showed that the SD pattern on capillaroscopy was the only variable associated with a significantly higher risk of PAH, with an odds ratio of 6.393 (95% confidence interval, 1.530-26.716; p = 0.011). Conclusion: In this study, SD pattern was associated with a 6.3-fold increased risk for PAH development in SLE patients, suggesting that nailfold capillaroscopy might be useful as a screening method to identify SLE patients with a high risk of developing this severe complication.

Abstract Background/objective: Digital ulcers (DUs) represent a frequent complication of systemic sclerosis (SSc). The aim of this study was to evaluate clinical, serological and capillaroscopy features that are associated with DUs in patients with SSc. Methods: In this bicentric cross-sectional study, 70 patients with SSc were consecutively selected from March 2016 to April 2017. Demographic and clinical features, including the presence of active DUs, were collected. Videocapillaroscopy was performed in all patients. Results: Among the 70 patients included (mean age of 46.8 years, mean disease duration of 9.41 years), 14 (20%) had active DUs. Based on multivariate analysis, the presence of anti-Scl-70 antibodies, the HAQ-DI score, and the capillary loss score were independently associated with DUs with odds ratios of 7.96 (95% CI 1.32-47.99), 55.77 (95% CI 1.76-1764.28), and 16.66 (95% CI 2.07-133.81), respectively. Conclusions: The presence of avascular areas in capillaroscopy, elevation of HAQ-DI score and anti-Scl-70 antibodies were independent factors associated with DUs in patients with SSc.

Abstract Nailfold capillaroscopy (NFC) is a reproducible, simple, low-cost, and safe imaging technique used for morphological analysis of nail bed capillaries. It is considered to be extremely useful for the investigation of Raynaud's phenomenon and for the early diagnosis of systemic sclerosis (SSc). The capillaroscopic pattern typically associated with SSc, scleroderma (“SD”) pattern, is characterized by dilated capillaries, microhemorrhages, avascular areas and/or capillary loss, and distortion of the capillary architecture. The aim of these recommendations is to provide orientation regarding the relevance of NFC, and to establish a consensus on the indications, nomenclature, the interpretation of NFC findings and the technical equipments that should be used. These recommendations were formulated based on a systematic literature review of studies included in the database MEDLINE (PubMed) without any time restriction.

RESUMO Lúpus eritematoso sistêmico (LES) é uma doença autoimune crônica que envolve múltiplos órgãos e sistemas, caracterizada pela produção de auto anticorpos e lesão tecidual. A etiologia do LES é parcialmente conhecida e envolve interação entre fatores genéticos e ambientais. Até 50% dos pacientes com LES apresentam envolvimento neurológico no decorrer da doença. Manifestações neurológicas estão associadas a prejuízo na qualidade de vida e altas taxas de mortalidade e morbidade. Foram identificadas 19 síndromes neuropsiquiátricas em pacientes com LES, divididas entre manifestações do sistema nervoso central e periférico. O objetivo deste artigo é revisar as manifestações neuropsiquiátricas mais importantes. Serão abordadas as características clínicas, os aspectos radiológicos e opções de tratamento dos eventos neuropsiquiátricos.

ABSTRACT Systemic lupus erythematosus (SLE) is a chronic autoimmune disease involving multiple organs, characterized by the production of autoantibodies and the development of tissue injury. The etiology of SLE is partially known, involving multiple genetic and environmental factors. As many as 50% of patients with SLE have neurological involvement during the course of their disease. Neurological manifestations are associated with impaired quality of life, and high morbidity and mortality rates. Nineteen neuropsychiatric syndromes have been identified associated with SLE, and can be divided into central and peripheral manifestations. This article reviews major neuropsychiatric manifestations in patients with SLE and discusses their clinical features, radiological findings and treatment options.

A capilaroscopia periungueal é um método simples, de baixo custo, e de extrema relevância na avaliação de pacientes com fenômeno de Raynaud ou portadores de doenças do espectro da esclerose sistêmica (ES). Além de sua importância para o diagnóstico precoce da ES, constitui instrumento útil na identificação de pacientes esclerodérmicos com risco elevado para o desenvolvimento de complicações vasculares, viscerais e de óbito. A inclusão da capilaroscopia nos novos critérios para classificação da ES do Colégio Americano de Reumatologia (ACR) e da Liga Europeia Contra o Reumatismo (Eular) dá novo impulso para a utilização e disseminação do método. No presente artigo, pretendemos apresentar uma revisão didática, não sistemática, sobre o tema, com ênfase nos avanços recentemente descritos.

Nailfold capillaroscopy is a simple, low-cost method, that is extremely important in the evaluation of patients with Raynaud's phenomenon and of patients with systemic sclerosis (SSc) spectrum diseases. Besides its importance for the early diagnosis of SSc, nailfold capillaroscopy is a useful tool to identify scleroderma patients with high risk for development of vascular and visceral complications and death. The inclusion of capillaroscopy in the new classification criteria for SSc of the American College of Rheumatology (ACR) and European League Against Rheumatism (Eular) gives a new impetus to the use and dissemination of the method. In this paper, we present a didactic, non-systematic review on the subject, with emphasis on advances recently described.

Objetivo Avaliar a segurança e a eficácia da N-acetilcisteína (NAC) por via oral sobre o fluxo sanguíneo da microcirculação digital em pacientes com fenômeno de Raynaud (FRy) secundário à esclerose sistêmica (ES). Métodos Este foi um estudo randomizado, duplo-cego e placebo-controlado, no qual 42 pacientes com ES receberam NAC oral na dose de 600 mg, três vezes ao dia (21 pacientes, idade média 45,6±9,5 anos) ou placebo (21 pacientes, idade média 45,0±12,7 anos) durante quatro semanas. O desfecho primário do estudo foi: melhora no fluxo sanguíneo da microcirculação cutânea antes e após estímulo frio avaliado pelo laser Doppler imaging (LDI) nas semanas 0 e 4. A frequência e a gravidade do FRy e o número de úlceras digitais também foram avaliados nas semanas 0 e 4. Os efeitos adversos foram registrados na quarta semana. Resultados Não houve mudança significativa no fluxo sanguíneo digital avaliado pelo LDI antes ou depois do estímulo frio após quatro semanas de NAC ou placebo. Ambos os grupos apresentaram melhora significativa na frequência e gravidade dos ataques de FRy, sem diferença entre os dois. O grupo placebo apresentou três úlceras digitais enquanto o grupo NAC não apresentou úlceras ao final do estudo. NAC foi bem tolerada e nenhum paciente descontinuou o tratamento. Conclusões NAC por via oral na dose de 1.800mg/dia não demonstrou efeito vasodilatador sobre a microcirculação das mãos após quatro semanas de tratamento em pacientes com FRy secundário à ES.

Objective To evaluate the safety and efficacy of oral N-acetylcysteine (NAC) on digital microcirculation blood flow in patients with Raynaud's phenomenon (RP) secondary to systemic sclerosis (SSc). Methods This was a randomized, double-blind, placebo-controlled trial in which 42 patients with SSc received oral NAC at a dose of 600mg tid (21 patients, mean age 45.6±9.5 years) or placebo (21 patients, mean age 45.0±12.7 years) for four weeks. The primary endpoint was the change in cutaneous microcirculation blood flow before and after cold stimulation measured by laser Doppler imaging (LDI) at weeks 0 and 4. The frequency and severity of RP and the number of digital ulcers were also measured at weeks 0 and 4. The adverse events were recorded in the fourth week. Results There was no significant change in digital blood flow assessed by LDI before or after cold stimulus after four weeks of NAC or placebo. Both groups showed significant improvement in the frequency and severity of RP attacks, with no difference between the two groups. At the end of the study, the placebo group had three digital ulcers, while the NAC group showed no ulcers. NAC was well tolerated and no patient discontinued the treatment. Conclusions NAC orally at a dose of 1800mg/day showed no vasodilator effect on hands’ microcirculation after four weeks of treatment in patients with RP secondary to SSc.

OBJECTIVES: The purpose of this study was to investigate the association between T cell receptor excision circle levels in peripheral blood mononuclear cells and regulatory T cells that co-express CD25 and Foxp3 in healthy children and adolescents of different ages. MATERIALS AND METHODS: The quantification of signal-joint T-cell receptor excision circle levels in the genomic DNA of peripheral blood mononuclear cells was performed using real-time quantitative PCR. The analysis of CD4, CD8, CD25, and Foxp3 expression was performed using flow cytometry. RESULTS: Ninety-five healthy controls (46 females and 49 males) ranging in age from 1 to 18 years were analyzed. The mean T-cell receptor excision circle count in all individuals was 89.095¡36.790 T-cell receptor excision circles per microgram of DNA. There was an inverse correlation between T-cell receptor excision circles counts and age (r = -0.846; p<0.001) as well as between the proportion of CD4+CD25+Foxp3+ T cells and age (r = -0.467; p = 0.04). In addition, we observed a positive correlation between the amount of CD4+CD25+Foxp3+ T cells and the amount of Tcell receptor excision circles per microgram of DNA in individuals of all ages (r = -0.529; p = 0.02). CONCLUSIONS: In this study, we observed a decrease in the thymic function with age based on the fact that the level of T-cell receptor excision circles in the peripheral blood positively correlated with the proportion of regulatory T cells in healthy children and adolescents. These findings indicate that although T-cell receptor excision circles and regulatory T cells levels decrease with age, homeostasis of the immune system and relative regulatory T cells population levels are maintained in the peripheral blood.

OBJECTIVE: To analyze cytokine gene expression in keratinocytes from patients with systemic lupus erythematosus (SLE). INTRODUCTION: Keratinocytes represent 95% of epidermal cells and can secrete several cytokines. METHODS: Keratinocytes were obtained by laser microdissection from 21 patients with SLE (10 discoid and 11 acute lesions) at involved and uninvolved sites. All patients were receiving a low/moderate prednisone dose and 18 were receiving chloroquine diphosphate. IL-2, IL-5, TNF-α and IFN-γ gene expression was evaluated by real-time PCR and expressed as the ratio (R) to a pool of skin samples from 12 healthy volunteers. RESULTS: Heterogeneity in cytokine gene expression was found among patients with SLE. Eighteen of 38 valid SLE samples (47%) presented overexpression (R>1) of at least one cytokine. Lesional skin samples tended to show higher cytokine expression than samples from uninvolved skin (p = 0.06). IL-5 and IFN-γ were the most commonly overexpressed cytokines. Samples with cytokine overexpression corresponded to more extensive and severe lesions. Prednisone dose did not differ between samples without cytokine overexpression (15.71±3.45 mg/day) and those with overexpressed cytokines (12.68±5.41 mg/day) (p = 0.216). Samples from all patients not receiving diphosphate chloroquine had at least one overexpressed cytokine. CONCLUSIONS: The heterogeneous keratinocyte cytokine gene expression reflects the complex immunological and inflammatory background in SLE. Patients with severe/extensive skin lesions showed a higher frequency of cytokine gene overexpression. Increased IFN-γ and IL-5 expression suggests that Th1 and Th2 cells are involved in SLE skin inflammation. The possibility that prednisone and antimalarial drugs may have contributed to low cytokine gene expression in some samples cannot be ruled out.

OBJETIVO: Determinar o comportamento dinâmico do fluxo sanguíneo da microcirculação digital, antes e após dois estímulos frios (EF) de diferentes intensidades, utilizando o método do Laser Doppler Imaging (LDI) em pacientes com esclerose sistêmica (ES) e controles saudáveis. MÉTODOS: Foram incluídos 14 pacientes com ES (51,2 ± 5,5 anos de idade) e 12 controles saudáveis (44,8 ± 9,9 anos). Foram realizados dois protocolos alternativos de EF (submersão das mãos em água a 10 ºC ou 15 ºC, durante 1 minuto). O fluxo médio das quatro polpas digitais da mão esquerda (FPD) foi mensurado com a utilização do LDI (Moor LDI-VR), em condições basais, nos períodos de 1, 4, 10, 25 e 40 minutos após EF. RESULTADOS: O fluxo basal foi significativamente menor em ambos os protocolos em pacientes com ES comparados a controles (312,9 ± 102,7 versus 465,4 ± 135,4 PU, P = 0,006, no protocolo a 15 ºC; 305,2 ± 121,0 versus 437,9 ± 119,8 PU, P = 0,01, no protocolo a 10 ºC). Nos controles houve declínio significativo do FPD após EF, em comparação aos valores basais apenas no tempo de um minuto após EF a 15 ºC (P = 0,001) e nos tempos de 1 e 25 minutos após EF a 10 ºC (P = 0,005; P = 0,001, respectivamente). Nos pacientes com ES, houve declínio significativo do FPD nos tempos de 1, 4 e 10 minutos após ambos EFs (P < 0,000; P = 0,002; P = 0,014, EF a 15 ºC; P < 0,000; P = 0,004; P = 0,001, EF a 10 ºC, respectivamente). CONCLUSÃO: LDI demonstrou baixa perfusão em polpa digital em condições basais e elevado declínio de perfusão com retardo na recuperação após EF na ES. A quantificação do fluxo sanguíneo pelo LDI pode ser útil para o seguimento longitudinal da doença e para a monitoração de intervenções terapêuticas na ES.

OBJECTIVES: The objective of this study was to investigate the dynamic behavior of the blood flow of the microvascular circulation of the fingertips before and after two cold stimuli (CS), using Laser Doppler Imaging with different intensities in patients with systemic sclerosis (SSc) and in healthy individuals. PATIENTS AND METHODS: Fourteen SSc patients (51.2 ± 5.5 years) with Raynaud's phenomenon and 12 healthy controls (44.8 ± 9.0 years) were included in this study. Two CS protocols (submersion of the hands in water at 10 ºC or 15 ºC for 1 minute) were performed on the same day. Mean fingertip blood flow (FBF) of four digits of the left hand was measured using LDI (Moor LDI-VR, Moor Instruments) at baseline and at 1, 4, 10, 25, and 40 minutes after CS. RESULTS: Baseline blood flow was significantly lower in both CS protocols in SSc patients when compared to controls (312.9 ± 102.7 vs 465.4 ± 135.4 PU, P = 0.006 at 15 ºC; 305.2 ± 121.0 vs 437.9 ± 119.8 PU; P = 0.01 at 10 ºC). In the control group, a significant decrease in FBF after CS, when compared to baseline, was observed 1 minute (P = 0.001) after CS at 15 ºC and at 1 (P = 0.005) and 25 minutes (P = 0.001) after CS at 10 ºC. In SSc patients, a significant decrease in FBF was observed in both CS protocols at 1, 4, and 10 minutes (P < 0.000; P = 0.002; P = 0.014, after CS at 15 ºC; P < 0.000; P = 0.004; P = 0.001, after CS at 10 ºC). CONCLUSIONS: Laser Doppler Imaging showed lower baseline fingertip perfusion and further reduction after CS in SSc patients compared to controls. Quantification of fingertip blood flow by LDI may be useful in the longitudinal monitoring of the disease status and therapeutic interventions in SSc.