Urine microscopy as a biomarker of Acute Kidney Injury following cardiac surgery with cardiopulmonary bypass / Microscopia urinária como biomarcador de lesão renal aguda após cirurgia cardíaca com circulação extracorpórea

Abstract Introduction: Acute kidney injury (AKI) occurs in about 22% of the patients undergoing cardiac surgery and 2.3% requires renal replacement therapy (RRT). The current diagnostic criteria for AKI by increased serum creatinine levels have limitations and new biomarkers are being tested. Urine sediment may be considered a biomarker and it can help to differentiate pre-renal (functional) from renal (intrinsic) AKI. Aims: To investigate the microscopic urinalysis in the AKI diagnosis in patients undergoing cardiac surgery with cardiopulmonary bypass. Methods: One hundred and fourteen patients, mean age 62.3 years, 67.5 % male, with creatinine 0.91 mg/dL (SD 0.22) had a urine sample examined in the first 24 h after the surgery. We looked for renal tubular epithelial cells (RTEC) and granular casts (GC) and associated the results with AKI development as defined by KDIGO criteria. Results: Twenty three patients (20.17 %) developed AKI according to the serum creatinine criterion and 76 (66.67 %) by the urine output criterion. Four patients required RRT. Mortality was 3.51 %. The use of urine creatinine criterion to predict AKI showed a sensitivity of 34.78 % and specificity of 86.81 %, positive likelihood ratio of 2.64 and negative likelihood ratio of 0.75, AUC-ROC of 0.584 (95%CI: 0.445-0.723). For the urine output criterion sensitivity was 23.68 % and specificity 92.11 %, AUC-ROC was 0.573 (95%CI: 0.465-0.680). Conclusion: RTEC and GC in urine sample detected by microscopy is a highly specific biomarker for early AKI diagnosis after cardiac surgery.

Resumo Introdução: Lesão renal aguda (LRA) ocorre em cerca de 22% dos pacientes submetidos a cirurgia cardíaca e 2,3% necessitam de terapia renal substitutiva (TRS). Os atuais critérios diagnósticos para LRA fundamentados no aumento dos níveis de creatinina sérica apresentam limitações e novos biomarcadores estão sendo testados. O sedimento urinário é um biomarcador que pode ajudar a diferenciar a LRA pré-renal (funcional) da LRA renal (intrínseca). Objetivos: Investigar a urinálise microscópica no diagnóstico de LRA em pacientes submetidos a cirurgia cardíaca com circulação extracorpórea. Métodos: Um total de 114 pacientes com idade média de 62,3 anos, 67,5% do sexo masculino e níveis médios de creatinina de 0,91 mg/dL (DP 0,22) tiveram amostras de urina examinadas nas primeiras 24 horas após a cirurgia. A identificação de células epiteliais tubulares renais (CETR) e cilindros granulares (CG) foi associada a desfechos de desenvolvimento de LRA conforme os critérios do KDIGO. Resultados: Vinte e três pacientes (20,17%) desenvolveram LRA pelo critério de creatinina sérica e 76 (66,67%) pelo critério de diurese. Quatro pacientes necessitaram de TRS. A mortalidade foi de 3,51%. O uso da creatinina urinária como critério preditivo para LRA mostrou sensibilidade de 34,78% e especificidade de 86,81%; razão de verossimilhança positiva de 2,64 e razão de verossimilhança negativa de 0,75; e ASC-COR de 0,584 (IC 95%: 0,445-0,723). Para o critério de diurese, a sensibilidade foi de 23,68% e a especificidade 92,11%; a ASC-COR foi 0,573 (IC 95%: 0,465-0,680). Conclusão: A identificação de CETR e CG em amostras de urina por microscopia representa um biomarcador altamente específico para o diagnóstico precoce de LRA após cirurgia cardíaca.

Progress in tear microdesiccate analysis by combining various transmitted-light microscope techniques

BACKGROUND: Tear desiccation on a glass surface followed by transmitted-light microscopy has served as diagnostic test for dry eye. Four distinctive morphological domains (zones I, II, III and transition band) have been recently recognized in tear microdesiccates. Physicochemical dissimilarities among those domains hamper comprehensive microscopic examination of tear microdesiccates. Optimal observation conditions of entire tear microdesiccates are now investigated. One-µl aliquots of tear collected from individual healthy eyes were dried at ambient conditions on microscope slides. Tear microdesiccates were examined by combining low-magnification objective lenses with transmitted-light microscopy (brightfield, phase contrasts Ph1,2,3 and darkfield. RESULTS: Fern-like structures (zones II and III) were visible with all illumination methods excepting brightfield. Zone I was the microdesiccate domain displaying the most noticeable illumination-dependent variations, namely transparent band delimited by an outer rim (Ph1, Ph2), homogeneous compactly built structure (brightfield) or invisible domain (darkfield, Ph3). Intermediate positions of the condenser (BF/Ph1, Ph1/Ph2) showed a structured roughly cylindrical zone I. The transition band also varied from invisibility (brightfield) to a well-defined domain comprising interwoven filamentous elements (phase contrasts, darkfield. CONCLUSIONS: Imaging of entire tear microdesiccates by transmitted-light microscopy depends upon illumination. A more comprehensive description of tear microdesiccates can be achieved by combining illumination methods.

Culture & characterisation of limbal epithelial cells & oral mucosal cells.

Background & objectives: Sources of autologous tissue that can functionally replace the corneal epithelium have been considered as an alternative to allogenous limbal transplants for limbal stem cells deficiency (LSCD). The aim of the present study was to compare the characterization of oral mucosa with limbal epithelial cells by markers using reverse transcriptase polymerase chain reaction (RT-PCR). Methods: Experiments were performed using oral tissue (n=6) obtained from patients who underwent oral mucosal graft for LSCD. Confluent cultures of limbus and oral mucosa epithelial cells were characterized by the pututative stem cell markers using RT-PCR. The morphological characteristics of cultivated epithelial cells were analyzed by haematoxylin and eosin staining and phase contrast microscopy. Results: Confluent sheets of epithelial cells were seen at the end of 14th day resembling the morphological features of limbal epithelia. RT–PCR analysis showed that cultured oral epithelial cells expressed markers such as ABCG2, p63, delta Np63, isoforms of p63, Keratin 3 (K3), membrane protein – Mucin (MUC 1, 4 and 16) and Antimicrobial Peptide - AMP (Human β Defensin – hBD 1, 2 and 3). Interpretation & conclusions: Oral epithelial cultures have morphological features resembling corneal and limbal epithelial cells by expressing similar marker genes. Thus, feasibility of clinical use of oral epithelial cells need be evaluated for allogenous limbal transplants.

Criterios de predicción de urocultivos mediante el campo oscuro / Predictive criteria of urocultures by darkfield

Se hace una valoración de algunos criterios de predicción de urocultivos, tanto negativos, como positivos o contaminados utilizando la microscopia de campo oscuro (CO) para el conteo de leucocitos, gérmenes y cálulas epiteliales. Nuestra conclusión es que la observación de menos de un germen por campo, como promedio, es el único criterio válido para descartar las muestras como negativas y no pueden predecirse ni las positivas ni las contaminadas desde la observación microscópica por campo oscuro