RESUMEN
PURPOSE: It was reported that gut-kidney axis may play an important role in IgA nephropathy (IgAN). Previous five GWASs of different populations for IgAN have discovered several genes related to intestinal immunity, including DEFA gene. However, the roles of the encoded proteins of DEFA5/6 which were called intestinal antimicrobial peptides HD5 and HD6 were not clear in kidney disease, such as IgAN. The purpose of this study was to clarify the association of HD5 and HD6 with IgAN. METHODS: We measured HD5 and HD6 in serum, urine, and kidney of IgAN patients and normal controls by ELISA, Western blot, and immunofluorescence. The association of HD5 or HD6 levels with clinical and pathologic phenotypes was analyzed. RESULTS: Serum levels of HD5 and HD6 were significantly higher in IgAN patients than those in normal controls. Baseline serum HD5 levels were significantly associated with eGFR (P = 0.002) and tubular atrophy/interstitial fibrosis (P = 0.002) and tubular atrophy/interstitial fibrosis (P = 0.002) and tubular atrophy/interstitial fibrosis (P = 0.002) and tubular atrophy/interstitial fibrosis (. CONCLUSIONS: In IgAN patients, an elevated serum HD5 level at the time of renal biopsy was associated with poor renal outcomes. HD5 rather than HD6 was probably associated with renal function of IgAN patients.
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Glomerulonefritis por IGA/fisiopatología , Regulación hacia Arriba , alfa-Defensinas/sangre , Adulto , Estudios de Casos y Controles , Femenino , Tasa de Filtración Glomerular , Glomerulonefritis por IGA/sangre , Glomerulonefritis por IGA/orina , Humanos , Masculino , Persona de Mediana Edad , Análisis Multivariante , Análisis de Regresión , Factores Sexuales , Análisis de Supervivencia , Adulto Joven , alfa-Defensinas/orinaRESUMEN
BACKGROUND: Pediatricians frequently use urinalysis to diagnose urinary tract infection (UTI) while awaiting urine culture results, but sensitivity and specificity of urinalysis are limited. This study evaluated the diagnostic accuracy of the antimicrobial peptides (AMPs) human α-defensin 5 (HD5) and human neutrophil peptides (HNP) 1-3 as novel UTI biomarkers in children. METHODS: We prospectively enrolled 199 pediatric Emergency Department or Urgent Care patients evaluated for a UTI. Urine concentrations of HD5 and HNP1-3 were measured by enzyme-linked immunosorbent assay. Urine culture was the reference standard. Sensitivities and specificities of leukocyte esterase (LE), HD5, HNP1-3, and test combinations were compared. RESULTS: For predicting positive urine culture, the areas under the receiver-operating characteristic curves for HD5 and HNP1-3 were 0.86 (95% confidence interval (CI): 0.81-0.92) and 0.88 (95% CI: 0.82-0.93), respectively. Compared to LE ≥ trace, the combination test "LE and HD5" increased specificity by 6% (95% CI: 3-10%) without decreasing sensitivity. In the subgroup whose urine was collected by a clean-catch method, combination tests "LE and HD5" and "HD5 and HNP1-3" increased specificity by > 10% compared to LE alone. CONCLUSION: Urine AMP profiles are a promising novel strategy as an adjunct to urinalysis to aid UTI diagnosis in children.
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Infecciones Urinarias/orina , alfa-Defensinas/orina , Adolescente , Biomarcadores/orina , Niño , Preescolar , Registros Electrónicos de Salud , Servicio de Urgencia en Hospital , Femenino , Humanos , Lactante , Masculino , Pediatría , Estudios Prospectivos , Curva ROC , Reproducibilidad de los Resultados , Tamaño de la Muestra , Sensibilidad y Especificidad , UrinálisisRESUMEN
OBJECTIVES: Urinary tract infection (UTI) often represents a diagnostic challenge in the emergency department (ED) where urine culture results are generally not available and other tests demonstrate limited sensitivity and specificity. Antimicrobial peptides (AMPs) are components of the innate immune system that have demonstrated increased urinary levels in response to infection both in children and in adults with chronic UTI. The objective of this study was to determine the relationship between urinary AMP levels and positive urine cultures in adult ED patients with suspected UTI. METHODS: This was a prospective, observational study of adult ED patients with suspected UTI. Enzyme-linked immunosorbent assays were performed to measure urine levels of AMPs: human neutrophil peptides 1-3 (HNP1-3), human α-defensin 5 (HD5), human beta defensin 2 (hBD-2), and cathelicidin (LL-37). Comparisons between positive and negative cultures were performed using Wilcoxon rank sum tests and receiver operating characteristic curves, with calculation of area under the curve (AUC). Data were also analyzed for the older adult subgroup. RESULTS: Of 40 patients enrolled, 23 (58%) were ≥ 65 years, 25 were female (64%), and seven (17%) were nonwhite. Cultures were positive in 13 (32%), including seven in those ≥ 65 years old. HNP1-3, HD5, and hBD-2 levels were significantly higher in those with positive than negative urine cultures. Median HNP1-3 was 5.39 ng/mg (interquartile range [IQR] = 2.74 to 11.09) in positive vs. 0.81 ng/mg (IQR = 0.06 to 3.87) in negative cultures. Median HD5 was 4.75 pg/mg (IQR = 1.6 to 22.7) in positive versus 0.00 pg/mg (IQR = 0 to 2.60) in negative cultures, and median hBD-2 was 0.13 pg/mg (IQR = 0.08 to 0.17) in positive versus 0.02 pg/mg (IQR = 0 to 0.04) in negative cultures (p < 0.05 for all). Findings were similar for adults ≥ 65 years. The AUC was ≥ 0.75 for all three AMPs, both overall and in the older adult subgroup. LL-37 was not significantly higher in patients with positive urine culture. However, LL-37 expression is vitamin D dependent, and inadequate serum levels (< 30 ng/mL) were present in 72% of those tested. CONCLUSIONS: Urinary levels of HNP1-3, HD5, and hBD-2 are significantly greater in the presence of positive urine cultures in ED patients with suspected UTI. These findings are maintained in the high-risk subgroup of older adults.
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Urinálisis/métodos , Infecciones Urinarias/diagnóstico , alfa-Defensinas/orina , Adulto , Anciano , Área Bajo la Curva , Servicio de Urgencia en Hospital , Ensayo de Inmunoadsorción Enzimática , Femenino , Humanos , Masculino , Técnicas Microbiológicas , Persona de Mediana Edad , Proyectos Piloto , Estudios Prospectivos , Curva ROC , Sensibilidad y Especificidad , Estadísticas no ParamétricasRESUMEN
We used protein expression profiles to develop a classification rule for the detection and prognostic assessment of bladder cancer in voided urine samples. Using the Ciphergen PBS II ProteinChip Reader, we analyzed the protein profiles of 18 pairs of samples of bladder tumor and adjacent urothelium tissue, a training set of 85 voided urine samples (32 controls and 53 bladder cancer), and a blinded testing set of 68 voided urine samples (33 controls and 35 bladder cancer). Using t-tests, we identified 473 peaks showing significant differential expression across different categories of paired bladder tumor and adjacent urothelial samples compared to normal urothelium. Then the intensities of those 473 peaks were examined in a training set of voided urine samples. Using this approach, we identified 41 protein peaks that were differentially expressed in both sets of samples. The expression pattern of the 41 protein peaks was used to classify the voided urine samples as malignant or benign. This approach yielded a sensitivity and specificity of 59% and 90%, respectively, on the training set and 80% and 100%, respectively, on the testing set. The proteomic classification rule performed with similar accuracy in low- and high-grade bladder carcinomas. In addition, we used hierarchical clustering with all 473 protein peaks on 65 benign voided urine samples, 88 samples from patients with clinically evident bladder cancer, and 127 samples from patients with a history of bladder cancer to classify the samples into Cluster A or B. The tumors in Cluster B were characterized by clinically aggressive behavior with significantly shorter metastasis-free and disease-specific survival.
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Proteínas de Neoplasias/orina , Proteómica/métodos , Neoplasias de la Vejiga Urinaria/diagnóstico , Neoplasias de la Vejiga Urinaria/orina , Análisis por Conglomerados , Humanos , Peso Molecular , Clasificación del Tumor , Neoplasias de la Vejiga Urinaria/metabolismo , Neoplasias de la Vejiga Urinaria/patología , alfa-Defensinas/orinaRESUMEN
BACKGROUND: The mechanisms that maintain sterility in the urinary tract are incompletely understood. Recent studies have implicated the importance of antimicrobial peptides (AMP) in protecting the urinary tract from infection. Here, we characterize the expression and relevance of the AMP human alpha-defensin 5 (HD5) in the human kidney and urinary tract in normal and infected subjects. METHODOLOGY/PRINCIPAL FINDINGS: Using RNA isolated from human kidney, ureter, and bladder tissue, we performed quantitative real-time PCR to show that DEFA5, the gene encoding HD5, is constitutively expressed throughout the urinary tract. With pyelonephritis, DEFA5 expression significantly increased in the kidney. Using immunoblot analysis, HD5 production also increased with pyelonephritis. Immunostaining localized HD5 to the urothelium of the bladder and ureter. In the kidney, HD5 was primarily produced in the distal nephron and collecting tubules. Using immunoblot and ELISA assays, HD5 was not routinely detected in non-infected human urine samples while mean urinary HD5 production increased with E.coli urinary tract infection. CONCLUSIONS/SIGNIFICANCE: DEFA5 is expressed throughout the urinary tract in non-infected subjects. Specifically, HD5 is expressed throughout the urothelium of the lower urinary tract and in the collecting tubules of the kidney. With infection, HD5 expression increases in the kidney and levels become detectable in the urine. To our knowledge, our findings represent the first to quantitate HD5 expression and production in the human kidney. Moreover, this is the first report to detect the presence of HD5 in infected urine samples. Our results suggest that HD5 may have an important role in maintaining urinary tract sterility.
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Riñón/metabolismo , Infecciones Urinarias/metabolismo , alfa-Defensinas/análisis , Infecciones por Escherichia coli , Humanos , Riñón/química , Túbulos Renales Colectores/química , Túbulos Renales Colectores/metabolismo , Reacción en Cadena de la Polimerasa , Pielonefritis/metabolismo , Sistema Urinario/química , Sistema Urinario/metabolismo , Urotelio/química , Urotelio/metabolismo , alfa-Defensinas/genética , alfa-Defensinas/orinaRESUMEN
Urinary biomarkers or profiles that allow noninvasive detection of recurrent transitional cell carcinoma (TCC) of the bladder are urgently needed. We obtained duplicate proteomic (SELDI) profiles from 227 subjects (118 TCC, 77 healthy controls and 32 controls with benign urological conditions) and used linear mixed effects models to identify peaks that are differentially expressed between TCC and controls and within TCC subgroups. A Random Forest classifier was trained on 130 profiles to develop an algorithm to predict the presence of TCC in a randomly selected initial test set (n = 54) and an independent validation set (n = 43) several months later. Twenty two peaks were differentially expressed between all TCC and controls (p < 10(-7)). However potential confounding effects of age, sex and analytical run were identified. In an age-matched sub-set, 23 peaks were differentially expressed between TCC and combined benign and healthy controls at the 0.005 significance level. Using the Random Forest classifier, TCC was predicted with 71.7% sensitivity and 62.5% specificity in the initial set and with 78.3% sensitivity and 65.0% specificity in the validation set after 6 months, compared with controls. Several peaks of importance were also identified in the linear mixed effects model. We conclude that SELDI profiling of urine samples can identify patients with TCC with comparable sensitivities and specificities to current tumor marker tests. This is the first time that reproducibility has been demonstrated on an independent test set analyzed several months later. Identification of the relevant peaks may facilitate multiplex marker assay development for detection of recurrent disease.