The expression of autophagy-related proteins in peripheral blood mononuclear cells of patients with lupus nephritis is upregulated and related to kidney damage / 细胞与分子免疫学杂志

Objective To identify the relationship between nephritis activity, autophagy and inflammation in patients with SLE. Methods Western blot analysis was used to detect the expression of microtubule-associated protein 1 light chain 3 (LC3) and P62 in peripheral blood mononuclear cells (PBMCs) of SLE patients with lupus nephritis and non-lupus nephritis patients. Tumor necrosis factor α (TNF-α) and interferon γ (IFN-γ) in the serum of SLE patients were determined by ELISA. The correlation between LC3II/LC3I ratio and SLE disease activity score (SLEDAI), urinary protein, TNF-α and IFN-γ levels was analyzed by Pearson method. Results The expression of LC3 was increased and P62 was decreased in SLE patients. TNF-α and IFN-γ were increased in the serum of SLE patients. LC3II/LC3I ratio was positively correlated with SLEDAI (r=0.4560), 24 hour urine protein (r=0.3753), IFN-γ (r=0.5685), but had no correlation with TNF-α (r=0.04 683). Conclusion Autophagy is found in PBMCs of SLE, and the autophagy is correlated with renal damage and inflammation in patients with lupus nephritis.

Correlación clínico patológica y evolución de la nefropatía lúpica en niños y adolescentes / Clinicopathologic correlation and evolution of lupus nephritis in children and adolescents

El lupus eritematoso sistémico (LES) tiene una prevalencia baja en la población general y es menor en la edad pediátrica. La nefropatía lúpica (NL) es más frecuente y de mayor severidad que en adultos, condicionando la morbimortalidad de la enfermedad. Se realizó un estudio descriptivo prospectivo de 20 niños y adolescentes con NL controlados en la Policlínica de Colagenopatías del Centro Hospitalario Pereira Rossell en el período desde octubre de 2003 hasta setiembre de 2013 con el objetivo de describir las características clínico-serológicas y evolutivas de pacientes con NL y correlacionarlas con los hallazgos anátomopatológicos. La NL se observó en el 52,6% de los casos con LES. El 70% fueron de sexo femenino, relación femenino/masculino de 2,3/1, 85% de raza blanca, la mediana del diagnóstico fue de 12 años. Las formas de presentación fueron: alteraciones urinarias menores (AUM) en 14 pacientes (0,7), en cuatro casos síndrome nefrótico (SN), con o sin insuficiencia renal (IR) y/o hipertensión arterial. Un paciente se manifestó con síndrome nefrítico. Un paciente tenía un examen de orina normal. Las formas histopatológicas proliferativas graves se presentaron en 18 (0,9); los casos con AUM presentaban NL grado III-IV en 13 (0,93); todos los casos con SN con o sin IR tenían NL III-IV. No hubo casos de NL aislada como forma de comienzo. En el momento del diagnóstico, los anticuerpos antinucleares fueron positivos en 19 (0,95) y los anti DNA doble cadena en 16 (0,8); C3 y C4 estuvieron descendidos en 19 (0,95) y en 15 (0,75) respectivamente. El seguimiento promedio fue 4,2 años. Al final del seguimiento estaban en remisión 16 pacientes (0,8), cuatro en remisión parcial, todos con función renal normal, excepto un caso que presentó IR extrema, fue trasplantado y tuvo una excelente evolución. Un paciente falleció con hemorragia pulmonar. La sobrevida de la función renal y la de los pacientes fue 0,95 respectivamente. El tratamiento se realizó en base a corticoides, hidroxicloroquina asociados a azatioprina o micofenolato mofetilo. En ocho pacientes con cuadros graves se usó la ciclofosfamida I/V. Esta serie constituye la primera serie nacional de nefropatía lúpica en niños y adolescentes. Conclusión: predominó la presentación clínica con AUM y formas histopatológicas severas, clases III y IV, evidenciando una disociación clínico anatomopatológica. A pesar del elevado porcentaje de NL severas, el manejo adecuado y oportuno y la adherencia al tratamiento y a los controles médicos fueron fundamentales para la evolución favorable de la NL.

Systemic lupus erythematosus (SLE) has a low prevalence in the overall general population and this is lower in children. Child lupus nephropathy (LN) is more frequent and severe than in SLE adult patients, with greater disease morbidity and mortality. A prospective descriptive study of 20 children and adolescents with LN monitored in the Collagen Diseases Office of the Pereira Rossell Hospital between October, 2003 and September, 2013 was performed. The objective of this study was to describe clinical-serological features and the evolution of these patients and to correlate them with its anatomopathological findings. LN was diagnosed in 52,6% of the SLE patients, 70% were female with a female/male correlation of 2,3/1; 85% were Caucasian; median age at diagnosis was 12 years old. The clinical presentations were minor urinary findings (MUF) in 14 patients (0,7) and nephrotic syndrome (NS) in 4 (0,2), and another one nephritic syndrome. One patient presented no symptoms and had normal urinalysis. Severe proliferative classes predominated in18 patients (0.9); 13 (0.93) patients with MUF and all the patients with NS had LN classes III or IV. Isolated LN was not seen in the initial presentation. At the time of diagnosis antinuclear antibodies were positive in 19 patients (0.95); and anti DNA double stranded in 16 (0.8). Low C3 was found in 19 (0.95) and C4 in 15 (0.75), respectively. Average follow-up time was 4.2 years. At the end of follow-up 16 (0.8) were in remission, 4 of them in partial remission; all patients presented normal renal function except for one who evidenced severe renal failure and required hemodialysis and transplantation and had an excellent evolution. One patient died with pulmonary hemorrhage. The renal and patient survivals were 0.95 respectively. Treatment consisted in corticosteroids and hydroxychloroquine associated with azathioprine or mycophenolate mofetil. Cyclophosphamide was administered to 8 patients with severe illness. This is the first national report of LN in children and adolescents. Conclusions: the predominant clinical presentation of LN was MUF with severe anatomopathological findings, classes III and IV, showing a clinical-pathological dissociation. Despite the high percentage of severe LN, early and adequate treatment, as well as a good compliance to it with periodic medical follow-up, were essential to achieve a favorable outcome of LN.

Detecção de podocitúria em pacientes com nefrite lúpica / Detection of podocyturia in patients with lupus nephritis

INTRODUÇÃO: A podocitúria tem sido detectada em doenças glomerulares, tais como em nefrite lúpica (NL), em que a proteinúria é uma manifestação importante, e sua ocorrência parece limitar-se à fase ativa da doença. OBJETIVO: Avaliar a podocitúria por imunofluorescência em pacientes portadores de NL e verificar possível associação com atividade clínica da doença. MÉTODOS: Foram avaliados 56 pacientes com NL. Os pacientes foram divididos em três grupos de acordo com o grau de atividade clínica: Grupo B, sem atividade (n = 17); Grupo C, com atividade discreta (n = 29) e Grupo D, moderada a grave (n = 10). Como grupo controle, foram incluídos 29 indivíduos saudáveis (Grupo A). A podocitúria foi estudada por meio de imunofluorescência indireta, usando-se anticorpos primários antipodocina, nefrina e sinaptopodina, e anticorpo secundário conjugado à FITC. Também foram avaliados os níveis de creatinina sérica e da relação proteína/creatinina (P/C) urinária, assim como a presença de hematúria e leucocitúria. RESULTADOS: A podocitúria com antipodocina e com antissinaptopodina correlacionou-se estatisticamente com a relação P/C (p = 0,001 e p = 0,013, respectivamente). Tanto a podocitúria com antipodocina, quanto a relação P/C, apresentaram correlação significante (p < 0,001) com a graduação de atividade da doença na NL, diferentemente do que se observou com os outros dois anticorpos, antinefrina e antissinaptopodina. CONCLUSÃO: Nossos achados sugerem que a pesquisa de podocitúria com anticorpos antipodocina poderia ser útil no acompanhamento de pacientes com NL, fornecendo dados relevantes quanto à atividade da doença.

INTRODUCTION: The podocyturia has been detected in glomerular diseases, such as lupus nephritis (LN), in which proteinuria is an important manifestation, and its occurrence seems to be limited to the active phase of the disease. OBJECTIVE: To evaluate podocyturia in LN patients, and the possible association with clinical disease activity. METHODS: We evaluated 56 patients with LN, that were classified in three groups according to the degree of clinical activity: Group B, no activity (n = 17), Group C with mild (n = 29) and Group D, moderate to severe activity (n = 10). The control group was composed by 29 healthy subjects (Group A). The podocyturia was studied by indirect immunofluorescence using primary antibodies to podocyte: anti-podocin, nephrin and synaptopodin, and a secondary antibody conjugated with FITC. We also evaluated serum creatinine levels, urinary protein/creatinine (P/C) ratio, hematuria and leucocituria. RESULTS: The podocyturia with anti-podocin and anti-sinaptopodin correlated statistically with the P/C ratio (p = 0.001 and p = 0.013, respectively). The podocyturia with anti-podocin, as well as the P/C ratio showed significant correlation (p < 0.001) with the degree of lupus disease activity, unlike the other two antibodies, anti-nephrin and anti-synaptopodin. CONCLUSION: Our findings show that podocyturia with anti-podocin could be useful in monitoring disease activity in LN patients.

Relação proteína/creatinina na urina versus proteinúria de 24 horas na avaliação de nefrite lúpica / Urinary protein/creatinine ratio versus 24-hour proteinuria in the evaluation of lupus nephritis

INTRODUÇÃO: Tem-se defendido a utilização do índice urinário proteína e creatinina em substituição à determinação de proteinúria de 24 horas para acompanhamento de doenças glomerulares, considerando-se as vantagens de maior facilidade na coleta e o menor custo. Entretanto, há dúvidas quanto à pertinência de usar este índice tanto numa avaliação isolada como no seguimento de pacientes com nefrite lúpica. OBJETIVO: Avaliar as determinações de proteinúria de 24 horas e proteinúria em amostra isolada de urina, fazendo a correção pela creatinina urinária, relação proteinúria/creatininúria, em indivíduos com nefrite lúpica. MÉTODOS: Determinações de proteinúria de 24 horas e relação proteinúria/creatininúria por métodos convencionais (Pirogalol automatizado para proteinúria e picrato alcalino para creatinina). RESULTADOS: Foram comparadas 78 amostras de urina de 41 pacientes com diagnóstico de lúpus eritematoso sistêmico, segundo os critérios da Associação Americana de Reumatologia, com nefrite lúpica, constatando-se uma boa correlação entre proteinúria de 24 horas e relação proteinúria/creatininúria (r = 0,9010 e r² = 0,813). Não se observou, entretanto, uma boa correlação entre proteinúria em amostra isolada (sem correção pela creatinina urinária) versus aquela de 24 horas (r = 0,635 e r² = 0,403) ou versus relação proteinúria/creatininúria (r = 0,754 e r² = 0,569). CONCLUSÃO: Os marcadores de proteinúria de 24 horas e relação proteinúria/creatininúria isoladamente mostraram-se úteis no acompanhamento de cada caso. Porém, observou-se que os seus valores absolutos são diferentes, não possibilitando a substituição de um pelo outro ao longo do seguimento, particularmente quando este resultado é usado para definição de atividade da doença. Se necessário, sugere-se um período de intersecção (duas a três determinações pelos dois métodos) para mudança de um para outro e escolha de um único marcador preferencial para seguimento da proteinúria.

INTRODUCTION: The urinary protein/creatinine ratio has been used instead of 24-hour proteinuria in Nephrology practice for the follow-up of glomerular diseases, considering the advantages of collection and the low cost. However, there are still doubts as to its applicability both for an isolated evaluation and for the follow-up of patients with lupus nephritis. OBJECTIVE: To evaluate 24-hour proteinuria determinations and random urine samples, performing urinary creatinine correction and urinary protein/creatinine ratio in subjects with lupus nephritis. METHODS: 24-hour proteinuria and urinary protein/creatinine ratio were determined by conventional methods (automated Pyrogallol for proteinuria and alkaline picrate for creatinine). RESULTS: Seventy-eight urine samples of 41 patients diagnosed with systemic lupus erythematosus, according to the American Rheumatology Association, with lupus nephritis, were analyzed, and a good correlation between 24-hour proteinuria and urinary protein/creatinine ratio (r = 0.9010 and r² = 0.813) was observed. However, a poor correlation between random proteinuria (without creatinine correction) versus 24-hour proteinuria (r = 0.635 and r² = 0.403) or versus urinary protein/creatinine ratio (r = 0.754 and r² = 0.569) was seen. CONCLUSION: 24-hour proteinuria and urinary protein/creatinine ratio were useful in the follow-up of each case. However, we observed that the absolute values were different, which did not allow the replacement of one for the other during follow-up, especially when this result is used to define the activity of the disease. Based on these results, we suggest a period of intersection from one to the other (two to three determinations by both methods), and the choice of one marker for proteinuria follow-up, if necessary.

Urinary Hypoxia-Inducible Factor-1alpha Levels Are Associated with Histologic Chronicity Changes and Renal Function in Patients with Lupus Nephritis

PURPOSE: Tubulointerstitial hypoxia in the kidney is considered a hallmark of injury and a mediator of the progression of tubulointerstitial fibrosis. Hypoxia-inducible factor-1alpha (HIF-1alpha), a master transcription factor in cellular adaptation to hypoxia, regulates a wide variety of genes, some of which are closely associated with tissue fibrosis. The present study set out to characterize urinary HIF-1alpha expressions in patients with lupus nephritis (LN) and to explore whether urinary HIF-1alpha expressions are associated with histologic chronicity changes and renal function. MATERIALS AND METHODS: Urinary HIF-1alpha levels were measured by enzyme-linked immunosorbent assays in 42 patients with LN and in 30 healthy controls. Activity and chronicity indexes as well as tubular HIF-1alpha expressions were analyzed for each specimen. RESULTS: Urinary HIF-1alpha levels were higher in LN patients than in healthy controls (3.977+/-1.696 vs. 2.153+/-0.554 ng/mL, p0.05). CONCLUSION: Urinary HIF-1alpha levels were elevated in LN patients and were associated with histologic chronicity changes and renal function, indicating that HIF-1alpha might contribute to histologic chronicity in LN.

Caracterização imunofenotípica dos leucócitos presentes na urina de pacientes com nefropatia lúpica em atividade / Immunophenotypic characterization of leukocytes present in urine of patients with active lupus nephropathy

O objetivo deste estudo foi determinar a expressão fenotípica dos leucócitos presentes na urina de pacientes com lúpus eritematoso sistêmico (LES) e nefrite em atividade. Foram selecionados 12 pacientes com glomerulonefrite membranosa. O critério utilizado para classificação de atividade sistêmica foi o SLEDAI (índice de atividade de doença para o LES), cuja média foi de 18,2 pontos. A presença de nefrite ativa foi definida por critérios histológicos, alterações do sedimento urinário, desenvolvimento recente de proteinúria e/ou perda aguda da função renal. A fenotipagem com anticorpos monoclonais (ACMo) e fosfatase alcalina, aplicada em lâminas com citocentrifugado de amostra da urina, foi o método empregado para a classificação das subpopulações de leucócitos urinários. Encontrou-se predomínio de células CD14 positivas (monócitos/macrófagos) nos pacientes com LES 56,8 por cento dos leucócitos urinários). A segunda subpopulação mais frequente foi a de células CD3 CD14 CD20-CD45 + (a maioria provavelmente granulócitos) e os linfócitos T representaram 8,1 por cento do total de leucócitos (teste de Wilcoxon com correção de Bonferroni - p < 0,016). Evidenciou-se uma associação positiva entre exsudação leucocitária glomerular e excreção urinária de linfócitos CD8 + (p = 0,01). Esses resultados reforçam os indícios sugestivos da participação das células de linhagem macrofágica na fisiopatologia da lesão renal proliferativa no LES