Thrombomodulin and interleukin 6 as potential biomarkers of endothelial dysfunction and inflammation after renal transplant / Trombomodulina e interleucina 6 como potenciais biomarcadores da disfunção endotelial e da inflamação pós-transplante renal

ABSTRACT Introduction: Endothelial dysfunction may contribute to hypercoagulable and inflammation states presents in renal transplant, chronic kidney disease (CKD) and its causes. These disorders can be evaluated by markers, such as thrombomodulin (TM), von Willebrand factor (vWF) and interleukin 6 (IL-6). Objectives: The aim of this study was to assess TM, vWF and IL-6 in renal transplant recipients (RTR) and associate their plasma levels with primary cause of end-stage renal disease (ESRD) and allograft function. Methods: 160 RTR were grouped according to the primary cause of CKD (G1: glomerulopathy; G2: hypertensive nephrosclerosis; G3: diabetic nephropathy; and G4: other causes/unknown etiology); creatinine plasma levels (C1 < 1.4 and C2 ≥ 1.4 mg/dl); and the estimated glomerular filtration rate (eGFR) (R1< 60 and R2 ≥ 60 ml/min/1.73 m2). TM and vWF were determined by the enzyme-linked immunosorbent assay (ELISA) and IL-6 by flow cytometry. The results were presented as median, minimum and maximum; p-value < 0.05 was considered statistically significant. Results: TM levels were significantly higher in the G1 group compared to the others (G1: 8.38; G2: 5.51; G3: 5.88; G4: 6.33 ng/ml, p < 0.0001), and in the R1 group compared to R2 (R1: 6.65; R2: 6.19 ng/ml, p = 0.02). The concentration of IL-6, measured by the mean fluorescence intensity, was higher in C2 group when compared to C1 (C1: 7.9; C2: 13.35, p = 0.03). There was no difference in vWF levels among groups. TM correlated positively with IL-6 and creatinine, and negatively with eGFR. IL-6 also correlated positively with vWF. Conclusion: TM and IL-6 can be identified as potential markers for evaluating renal graft function. TM was more related to the primary cause of CKD compared to vWF and IL-6.

RESUMO Introdução: A disfunção endotelial pode contribuir para estados de hipercoagulabilidade e inflamação presentes no transplante renal e na doença renal crônica (DRC) e suas causas, podendo ser avaliada por marcadores como trombomodulina (TM), fator de von Willebrand (FvW) e interleucina 6 (IL-6). Objetivos: Avaliar TM, FvW e IL-6 em receptores do transplante renal (RTR) e associar seus níveis com a causa primária de DRC pré-transplante e função do enxerto. Métodos: Foram alocados 160 RTR em grupos de acordo com a causa primária da DRC (G1: glomerulopatias; G2: nefroesclerose hipertensiva; G3: nefropatia diabética; e G4: outras causas/etiologia desconhecida), os níveis plasmáticos de creatinina (C1 < 1.4 e C2 ≥ 1.4 mg/dl) e o ritmo de filtração glomerular estimado (eRFG) (R1< 60 e R2 ≥ 60 ml/min/1.73 m2). A TM e o FvW foram determinados pelo ensaio de imunoabsorção enzimática (ELISA) e a IL-6, por citometria de fluxo. Os resultados foram apresentados como mediana, mínimo e máximo; p < 0,05 foi considerado significativo. Resultados: Níveis de TM foram significativamente maiores no grupo G1 em comparação com os demais (G1: 8,38; G2: 5,51; G3: 5,88; G4: 6,33 ng/ml, p < 0,0001), e no grupo R1 comparado com o R2 (R1: 6,65; R2: 6,19 ng/ml, p = 0,02). A concentração de IL-6, avaliada pela intensidade média de fluorescência, foi maior no grupo C2 quando comparada com o C1 (C1: 7,9; C2: 13,35, p = 0,03). Não houve diferença entre os grupos para o FvW. TM correlacionou-se positivamente com IL-6 e creatinina e negativamente com eRFG. A IL-6 foi positivamente correlacionada com o FvW. Conclusão: TM e IL-6 podem ser apontadas como potenciais marcadores para avaliar a função do enxerto renal. A TM relacionou-se mais com a causa primária da DRC, se comparada com FvW e IL-6.

Aspartate aminotransferase to platelet ratio index (APRI) for differentiation of primary and secondary infection by dengue virus / Aplicação do índice de relação aspartato aminotransferase sobre plaquetas (APRI) na diferenciação de infecção primária e secundária pelo vírus da dengue

ABSTRACT INTRODUCTION: Dengue virus (DENV) infection has been considered a major public health problem in tropical countries. The unavailability of serologic testing in public health centers might adversely impact patients' outcome. OBJECTIVE: This study aimed to evaluate the accuracy of mean platelet volume (MPV) and aspartate aminotransferase (AST) to platelet ratio index (APRI) as laboratory markers of DENV infection that could be used to differentiate primary and secondary infections. METHODS: We assessed laboratory results from 503 patients with positive rapid test for DENV infection. RESULTS: Severe thrombocytopenia and increased liver involvement were observed in patients with DENV heterotypic secondary infection. Our data suggest that APRI was able to distinguish patients with primary and secondary infection (p = 0.006) with a relevant sensitivity (75%), specificity (76%) and a cut-off of 1.06. A total of 80 out of 105 (76%) patients with primary DENV infection had APRI ≤ 1.06, and 12 (75%) with secondary DENV infection had APRI > 1.06. On the other hand, MPV did not show significance in the differentiation of types of infection, coming up with poor area under the receiver operating characteristic (ROC) curve (0.61). CONCLUSION: APRI seems to be a powerful tool for early identification of DENV secondary infection cases in health centers.

RESUMO INTRODUÇÃO: A infecção pelo vírus da dengue (DENV) é considerada um grande problema de saúde pública nos países tropicais. A indisponibilidade de testes sorológicos em centros de saúde pública pode afetar negativamente o prognóstico do paciente. OBJETIVO: Este estudo teve como objetivo avaliar a precisão do volume médio de plaquetas (MPV) e o índice da relação de aspartato aminotransferase (AST) sobre plaquetas (APRI) como marcadores laboratoriais de infecção por DENV, que poderiam ser utilizados para diferenciar infecções primárias e secundárias. MÉTODOS: Foram avaliados os resultados laboratoriais de 503 pacientes com teste rápido positivo para infecção por DENV. RESULTADOS: Foram observadas trombocitopenia grave e disfunção hepática em pacientes com infecção secundária heterogênea por DENV. Nossos dados sugerem que o APRI foi capaz de distinguir os pacientes com infecção primária e secundária (p = 0, 006), com relevante sensibilidade (75%) e especificidade (76%) e corte de 1, 06. Um total de 80 de 105 (76%) pacientes com infecção primária por DENV tinha APRI ≤ 1, 06; e 12 (75%) com infecção secundária por DENV, APRI > 1, 06. Por outro lado, o MPV não mostrou significância na diferenciação de tipos de infecção, apresentando baixo valor da área sob a curva de característica de operação do receptor (ROC) (0, 61). CONCLUSÃO: APRI parece ser uma ferramenta poderosa para identificação precoce de casos de infecção secundária de DENV em centros de saúde.

Apolipoprotein E and PAI-1 gene polymorphisms and no association with arterial ischemic stroke and peripheral arterial disease manifestations / Polimorfismos nos genes da apolipoproteína E e do PAI-1 e não associação com as manifestações de acidente vascular cerebral e doença arterial periférica

ABSTRACT Introduction: Arterial thrombosis is considered a multifactorial disease, resulting from the interaction of genetic and acquired risk factors. Objectives: The aim of this study was to investigate the presence of the polymorphism in inhibitor of plasminogen activator type 1 (PAI-1) and apolipoprotein E (ApoE) genes and its interactions with PAI-1 levels and lipids and apolipoprotein profiles, respectively, as well as the frequencies of these polymorphisms and their association with thrombosis. Methods: Ninety-seven patients [48 with arterial ischemic stroke (IS) and 49 with peripheral arterial disease (PAD)], treated at the hematology medical service were included in this study. Polymorphisms were also investigated in 201 control subjects. Polymorphisms were investigated by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). Results: For the PAI-1 polymorphism, there were 54.2% heterozygous (HT) genotypes and 12.5% homozygous (HM) genotypes in the patients' group, and 52.7% HT genotypes and 21.3% HM genotypes in the controls. For the ApoE polymorphism, there were 56.3% (ε3ε3), 6.3% (ε4ε4), 8.3% (ε2ε3), 4.2% (ε2ε4) and 24.9% (ε3ε4) in the patients, and 61.2% (ε3ε3), 4.5% (ε4ε4), 8% (ε2ε3), 4.5% (ε2ε4) and 21.8% (ε3ε4) in the controls. Conclusion: No significant difference was observed by comparing patients and controls. In this study, no association was found between the presence of the evaluated polymorphisms and the occurrence of thrombotic events.

RESUMO Introdução: A trombose arterial é considerada uma doença multifatorial, resultante da interação de fatores de risco genéticos e adquiridos. Objetivos: O objetivo deste estudo foi investigar a presença dos polimorfismos nos genes do inibidor da ativação do plasminogênio tipo 1 (PAI-1) e da apolipoproteína E (ApoE), bem como suas interações com níveis de PAI-1 e lipídios e perfis de apolipoproteína, respectivamente, além das frequências desses polimorfismos e sua associação com trombose. Métodos: Noventa e sete pacientes [48 com acidente vascular cerebral isquêmico arterial (AVC) e 49 com doença arterial periférica (DAP)], tratados no serviço médico de hematologia, foram incluídos neste estudo. Os polimorfismos também foram investigados em 201 indivíduos-controle. Os polimorfismos foram investigados por reação em cadeia da polimerase-fragmento de restrição polimorfismo (PCR-RFLP). Resultados: Para o polimorfismo PAI-1, havia 54,2% genótipos heterozigotos (HT) e 12,5% genótipos de homozigoto (HM) no grupo dos pacientes, e 52,7% genótipos HT e 21,3% genótipos HM nos grupos-controle. Para o polimorfismo da ApoE, havia 56,3% (ε3ε3), 6,3% (ε4ε4), 8,3% (ε2ε3), 4,2% (ε2ε4) e 24,9% (ε3ε4) nos pacientes, e 61,2% (ε3ε3), 4,5% (ε4ε4), 8% (ε2ε3), 4,5% (ε2ε4) e 21,8% (ε3ε4) nos controles. Conclusão: Nenhuma diferença significativa foi observada comparando pacientes e controles. Neste estudo, não foi encontrada associação entre a presença dos polimorfismos avaliados e a ocorrência de eventos trombóticos.

Association of different biomarkers of renal function with D-dimer levels in patients with type 1 diabetes mellitus (renal biomarkers and D-dimer in diabetes)

ABSTRACT Objective This study aimed to evaluate the association between different renal biomarkers with D-Dimer levels in diabetes mellitus (DM1) patients group classified as: low D-Dimer levels (< 318 ng/mL), which included first and second D-Dimer tertiles, and high D-Dimer levels (≥ 318 ng/mL), which included third D-Dimer tertile. Materials and methods D-Dimer and cystatin C were measured by ELISA. Creatinine and urea were determined by enzymatic method. Estimated glomerular filtration rate (eGFR) was calculated using CKD-EPI equation. Albuminuria was assessed by immunoturbidimetry. Presence of renal disease was evaluated using each renal biomarker: creatinine, urea, cystatin C, eGFR and albuminuria. Bivariate logistic regression analysis was performed to assess which renal biomarkers are associated with high D-Dimer levels and odds ratio was calculated. After, multivariate logistic regression analysis was performed to assess which renal biomarkers are associated with high D-Dimer levels (after adjusting for sex and age) and odds ratio was calculated. Results Cystatin C presented a better association [OR of 9.8 (3.8-25.5)] with high D-Dimer levels than albuminuria, creatinine, eGFR and urea [OR of 5.3 (2.2-12.9), 8.4 (2.5-25.4), 9.1 (2.6-31.4) and 3.5 (1.4-8.4), respectively] after adjusting for sex and age. All biomarkers showed a good association with D-Dimer levels, and consequently, with hypercoagulability status, and cystatin C showed the best association among them. Conclusion Therefore, cystatin C might be useful to detect patients with incipient diabetic kidney disease that present an increased risk of cardiovascular disease, contributing to an early adoption of reno and cardioprotective therapies.

Evaluation of the interference of platelets in reticulocyte counting by flow cytometry using thiazole orange / Avaliação da interferência de plaquetas na contagem de reticulócitos por citometria de fluxo, utilizando laranja de tiazol

ABSTRACT Introduction: The reticulocyte count by flow cytometry (FC) - an automated counting method - can present errors due to the presence of interfering factors, contributing to a slight increase in results. However, automated methods have large advantages over the manual method, taken as reference, what justifies efforts to improve their quality. Objective: Evaluate platelet interference with the reticulocyte count by FC, using thiazole orange (TO) (FC/TO). Materials and methods: The method of reticulocyte count by FC/TO and a modified automated equivalent method, which excluded CD61-positive cells (platelets) from analysis (FC/TO/MOD), were compared to the manual method. Conclusion: Results were analyzed according to the recommendations of the Clinical and Laboratory Standards Institute (CLSI) to assess interchangeability between the methods, by linear regression analysis and paired t-test. The exclusion of interfering fragments from result analysis by the modified method produced results in closer proximity to those of the reference method.

RESUMO Introdução: A contagem de reticulócitos por citometria de fluxo (CF) - um método de contagem automatizada - pode apresentar erros devido à presença de interferentes, contribuindo para uma ligeira elevação dos resultados. No entanto, os métodos automatizados possuem grandes vantagens em relação ao manual, tido como referência, o que justifica esforços para a melhoria de sua qualidade. Objetivo: Avaliar a interferência de plaquetas na contagem de reticulócitos por CF, utilizando laranja de tiazol (thiazole orange [TO]) (CF/TO). Materiais e métodos: O método de contagem de reticulócitos por CF/TO e um método equivalente automatizado modificado, no qual se excluíram células CD61-positivas (plaquetas) da análise (CF/TO/MOD), foram comparados com o método manual. Conclusão: Os resultados foram analisados de acordo com as recomendações do Clinical and Laboratory Standards Institute (CLSI) para avaliar a intercambialidade entre os métodos, por meio da análise de regressão linear e do teste t pareado. A exclusão de interferentes da análise dos resultados pelo método modificado demonstrou maior proximidade dos resultados com aqueles do método de referência.

Evaluation of creatinine-based and cystatin C-based equations for estimation of glomerular filtration rate in type 1 diabetic patients

ABSTRACT Objective Several formulas based in different biomarkers may be used to estimate glomerular filtration rate (GRF). However, all of them have some limitations, and it is very important to evaluate their performances in different groups of patients. Therefore, we compared GFR, as estimated by creatinine-based and cystatin C-based equations, according to albuminuria, in type 1 diabetes (T1DM), in an observational case-control study. Subjects and methods T1DM patients were classified according to albuminuria: normoalbuminuric (n = 63), microalbuminuric (n = 30), macroalbuminuric (n = 32). GFR was calculated using creatinine-based and cystatin C-based (aMDRD, CKD-EPIcr, CKD-EPIcys, MacIsaac, Tan and CKD-EPIcrcys) equations. Spearman Correlation was used to evaluate the correlation of GFR estimated by the formulas with albuminuria. ROC curves were constructed to compare AUCs of GFR estimated by equations, in reference to macroalbuminuria. Sensibility, specificity and accuracy were calculated for a cut-off < 60 mL/min/1.73 m2. Results GFR estimated by creatinine-based and cystatin C-based equations significantly differed among normoalbuminuric, microalbuminuric and macroalbuminuric patients. Spearman correlation and AUCs of GFR estimated by creatinine-based and cystatin C-based formulas were very similar to each other, though cystatin C-based equations presented better correlation with albuminuria and higher AUCs than the creatinine-based ones, and the best accuracy to detect macroalbuminuric patients. Conclusion Although GFR estimated by all creatinine-based and cystatin C-based equations permitted the differentiation between T1DM patients, according to albuminuria, cystatin C-based equations presented best accuracy to detect macroalbuminuria in T1DM patients and should be considered in the clinical routine in order to increase the possibility of early diagnostic of chronic renal disease.