Proteomic identification of early urinary-biomarkers of acute kidney injury in preterm infants.

The immature preterm kidney is likely to be vulnerable to acute kidney injury (AKI). However, the biomarkers currently used for AKI are not sensitive or specific and are also inadequate for the timely detection of AKI in preterm infants. The objectives of this study were to identify novel urinary biomarkers of AKI using proteomic techniques, and to verify and validate that the candidates can serve as early predictive biomarkers for AKI. In total, 1,810 proteins were identified in the discovery phase. Among those proteins, 174 were selected as the 1st targeted proteins. A total of 168 proteins were quantified, and the levels of 6 were significantly increased in the AKI group in the verification phase. Using a clinical assay, the results were confirmed and validated using samples of the first urine after birth from the biorepository. Finally, enzyme-linked immunosorbent assays revealed that the levels of annexin A5, neutrophil gelatinase-associated lipocalin (NGAL), and protein S100-P were significantly higher in the samples of the first urine from patients with AKI than in those from patients without AKI. In conclusion, urinary annexin A5, NGAL and protein S100-P levels are promising biomarkers for early, accurate prediction of AKI in preterm infants.

Urine S100 proteins as potential biomarkers of lupus nephritis activity.

BACKGROUND: Improved, noninvasive biomarkers are needed to accurately detect lupus nephritis (LN) activity. The purpose of this study was to evaluate five S100 proteins (S100A4, S100A6, S100A8/9, and S100A12) in both serum and urine as potential biomarkers of global and renal system-specific disease activity in childhood-onset systemic lupus erythematosus (cSLE). METHODS: In this multicenter study, S100 proteins were measured in the serum and urine of four cSLE cohorts and healthy control subjects using commercial enzyme-linked immunosorbent assays. Patients were divided into cohorts on the basis of biospecimen availability: (1) longitudinal serum, (2) longitudinal urine, (3) cross-sectional serum, and (4) cross-sectional urine. Global and renal disease activity were defined using the Systemic Lupus Erythematosus Disease Activity Index 2000 (SLEDAI-2K) and the SLEDAI-2K renal domain score. Nonparametric testing was used for statistical analysis, including the Wilcoxon signed-rank test, Kruskal-Wallis test, Mann-Whitney U test, and Spearman's rank correlation coefficient. RESULTS: All urine S100 proteins were elevated in patients with active LN compared with patients with active extrarenal disease and healthy control subjects. All urine S100 protein levels decreased with LN improvement, with S100A4 demonstrating the most significant decrease. Urine S100A4 levels were also higher with proliferative LN than with membranous LN. S100A4 staining in the kidney localized to mononuclear cells, podocytes, and distal tubular epithelial cells. Regardless of the S100 protein tested, serum levels did not change with cSLE improvement. CONCLUSIONS: Higher urine S100 levels are associated with increased LN activity in cSLE, whereas serum S100 levels do not correlate with disease activity. Urine S100A4 shows the most promise as an LN activity biomarker, given its pronounced decrease with LN improvement, isolated elevation in urine, and positive staining in resident renal cells.

Cutaneous Burn Injury Modulates Urinary Antimicrobial Peptide Responses and the Urinary Microbiome.

OBJECTIVES: Characterization of urinary bacterial microbiome and antimicrobial peptides after burn injury to identify potential mechanisms leading to urinary tract infections and associated morbidities in burn patients. DESIGN: Retrospective cohort study using human urine from control and burn subjects. SETTING: University research laboratory. PATIENTS: Burn patients. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: Urine samples from catheterized burn patients were collected hourly for up to 40 hours. Control urine was collected from "healthy" volunteers. The urinary bacterial microbiome and antimicrobial peptide levels and activity were compared with patient outcomes. We observed a significant increase in urinary microbial diversity in burn patients versus controls, which positively correlated with a larger percent burn and with the development of urinary tract infection and sepsis postadmission, regardless of age or gender. Urinary psoriasin and ß-defensin antimicrobial peptide levels were significantly reduced in burn patients at 1 and 40 hours postadmission. We observed a shift in antimicrobial peptide hydrophobicity and activity between control and burn patients when urinary fractions were tested against Escherichia coli and Enterococcus faecalis urinary tract infection isolates. Furthermore, the antimicrobial peptide activity in burn patients was more effective against E. coli than E. faecalis. Urinary tract infection-positive burn patients with altered urinary antimicrobial peptide activity developed either an E. faecalis or Pseudomonas aeruginosa urinary tract infection, suggesting a role for urinary antimicrobial peptides in susceptibility to select uropathogens. CONCLUSIONS: Our data reveal potential links for urinary tract infection development and several morbidities in burn patients through alterations in the urinary microbiome and antimicrobial peptides. Overall, this study supports the concept that early assessment of urinary antimicrobial peptide responses and the bacterial microbiome may be used to predict susceptibility to urinary tract infections and sepsis in burn patients.

S100B urine concentrations in late preterm infants are gestational age and gender dependent.

BACKGROUND: Late preterm deliveries (LP, between 34 and 36wks), have considerably increased in the last decades. About 20-25% of LP infants who require intensive care and morbidity on public health are of great magnitude. Therefore, we aimed at offering a reference curve in LP period of a well-established neurotrophic and brain damage marker namely S100B protein. METHODS: We collected, between December 2009 and March 2012, urine samples, at first void (within 6-hours from birth) for S100B assessment, in 277 healthy LP infants consecutively admitted to our units. Standard clinical and laboratory monitoring parameters were also recorded. S100B was measured by using a commercially available immunoluminometric assay. RESULTS: S100B pattern in LP infants was characterized by a slight decrease in protein's concentration from 34 to 35wks. From 35wks onwards S100B started to increase reaching a significant difference (P=0.008) at 36wks. When corrected for gender, significantly higher (P<0.01, for all) S100B concentrations in female were observed from 34 to 36wks. Polynomial type-1 regression analysis showed a significant correlation (R=-0.05; P<0.001) between gestational age and S100B in LP infants considering either the whole study population or when corrected for gender. CONCLUSIONS: S100B in LP infants is gestational age and gender dependent. The present reference curve, for S100B in LP period, offers additional support to protein's neurotrophic role and suggests that gestational age and gender have to be taken into due account, whenever S100B is measured, in order to avoid bias factors.

Role of S100B protein in urine and serum as an early predictor of mortality after severe traumatic brain injury in adults.

S100B is a calcium-binding protein released into the blood from astroglial cells due to brain injury. Some authors have described a correlation between S100B serum concentration and severity of brain damage. There is not much information about the accuracy of urinary S100B for predicting outcome after severe traumatic brain injury (TBI). 55 patients with severe TBI were included in the study. Blood and urine samples were drawn to determine S100B levels on admission and on the subsequent 24, 48, 72 and 96 h. S100B concentrations (serum and urine) were significantly higher in patients who were dead a month after the accident compared to survivors. ROC-analysis showed that S100B at 24h post-severe TBI is a useful tool for predicting mortality (serum: AUC 0.958, urine: AUC 0.778). The best cut-offs for S100B were 0.461 µg/L and 0.025 µg/L (serum and urine respectively), with a sensitivity of 90% for both measurements and a specificity of 88.4% (serum) and 62.8% (urine). We can state that the determination of S100B levels both in urine and serum acts as a sensitive and an effective biomarker for the early prediction of mortality after severe TBI.

Perinatal asphyxia: kidney failure does not affect S100B urine concentrations.

BACKGROUND: S100B protein is a well-established marker of brain damage. Its importance in urine assessment is the convenience of a collection and sampling procedure that can be repeated without risk for the newborn. Since S100B is mainly eliminated by the kidneys and perinatal asphyxia (PA) is often associated with kidney failure we investigated whether S100B release might be kidney-mediated, thereby modifying the protein's reliability as a brain-damage marker. METHODS: We examined a cohort of healthy (n=432) and asphyxiated newborns (n=32) in whom kidney function parameters (blood urea and creatinine concentrations and urine gravity) and urine S100B concentrations were assessed in the first hours after birth. Data were analyzed by multiple logistic regression analysis with S100B as independent variable among a variety of clinical and laboratory monitoring parameters. RESULTS: S100B urine concentrations were significantly higher (P<0.01) in PA newborns than controls. No significant correlations (P>0.05, for all) between total urine S100B levels and kidney function parameters such as creatinine (r=0.03), urea (r=0.04) and urine gravity (r=0.06) were found. Multiple logistic regression analysis of a series of clinical and laboratory monitoring parameters (odds ratio at sampling: 9.47) with S100B as independent variable showed a positive significant correlation only between S100B levels (P<0.001) and the occurrence of PA. CONCLUSION: The present study shows that altered kidney function is not an adverse and/or confounding factor in urine S100B assessment and marks a new step towards the introduction of longitudinal monitoring of brain constituents in clinical practice.

[Markers of brain injury in non-invasive biological fluids]. / Markers of brain injury in non-invasive biological fluids.

Hypoxia-ischemia (H-I) constitutes the main phenomenon responsible for brain-blood barrier permeability modifications leading to cerebral vascular autoregulation loss in newborns. Hypotension, cerebral ischemia, and reperfusion are the main events involved in vascular auto-regulation loss leading to cell death and tissue damage. Reperfusion could be critical since organ damage, particularly of the brain, may be amplified during this period. An exaggerated activation of vasoactive agents, of calcium mediated effects could be responsible for reperfusion injury (R-I), which, in turns, leads to cerebral hemorrhage and damage. These phenomena represent a common repertoire in newborns complicated by perinatal acute or chronic hypoxia treated by risky procedures such as mechanical ventilation, nitric oxide supplementation, brain cooling, and extracorporeal membrane oxygenation (ECMO). Despite accurate monitoring, the post-insult period is crucial, as clinical symptoms and standard monitoring parameters may be silent at a time when brain damage is already occurring and the therapeutic window for pharmacological intervention is limited. Therefore, the measurement of circulating biochemical markers of brain damage, such as vasoactive agents and nervous tissue peptides is eagerly awaited in clinical practice to detect high risk newborns. The present article is aimed at investigating the role of dosage biochemical markers in non-invasive biological fluids such as S100B, a calcium binding protein, activin A, a protein expressed in Central nervous System (CNS).

S-100B in serum and urine after traumatic head injury in children.

BACKGROUND: Children with head trauma are frequently seen in many emergency units. The clinical evaluation of these patients is difficult for a number of reasons and improved diagnostic tools are needed. S-100B, a protein found in glial cells, has previously been shown to be a sensible marker for brain damage after head injury in adults, but few studies have focused on its use in children. METHODS: In this study, 111 children with head trauma were included and venous blood and urine samples were taken at arrival (S1 and U1) and 6 hours later (S2 and U2). S-100B levels were analyzed. Clinical and radiologic evaluations were performed according to hospital routine. Two groups were identified- group 1: no computed tomography (CT) scan performed ora CT scan without any sign of trauma-related intracranial pathology (n = 105). Group 2: A CT scan with signs of trauma-related intracranial pathology (n = 6). RESULTS: In group 1, the median (inter quartile range) serum S-100B value in S1-samples was 0.111 microg/L (0.086-0.153), and in group 2, it was 0.282 microg/L (0.195-1.44) (p < 0.01). Also, S2 values significantly differed between the two groups. Urine values were, however, not significantly differing between the groups. CONCLUSIONS: Serum S-100B values within 6 hours after head trauma in children were significantly higher in patients with intracranial pathology compared with those without intracranial complications. Identification of these high-risk patients already in the emergency department is of major importance, and we suggest that S-100B could be a valuable diagnostic tool in addition to those used in clinical practice today.

Antenatal glucocorticoid treatment affects preterm infants' S100B urine concentration in a dose-dependent manner.

BACKGROUND: Maternal glucocorticoid (GC) treatment is widely used to prevent lung immaturity in preterm infants. There is growing evidence that GCs may be detrimental to the Central Nervous System (CNS). We investigated whether antenatal GC administration affects CNS function in a dose-dependent manner by measuring urine concentrations of a well-established brain damage marker, S100B. METHODS: We conducted a case-control-study in 70 preterm infants (1 GC vs 1 control) whose mothers received a complete GC-course (GC2, n=16), half-course (GC1, n=19), and controls (n=35). At four predetermined time-points, in the first 72 h from birth, we assessed S100B urine concentrations, using a commercially available immunoluminometric assay (Lia-mat Sangtec 100, AB Sangtec Medical, Bromma, Sweden). Data were correlated with primary neonatal outcomes (incidence of respiratory distress syndrome, length of ventilatory support and hospital stay, incidence of intraventricular hemorrhage, adverse 7th day neurological follow-up and neonatal death). RESULTS: S100B in GC2 group at all monitoring time-points was significantly lower (P<0.01) than controls and GC1 group, while no differences (P>0.05) were evident between controls and GC1 group. No significant differences (P>0.05) were shown in primary outcomes between half or complete GC-course treated groups. CONCLUSION: S100B levels of infants antenatally treated with GCs differed in a dose-dependent manner. Data on primary outcomes suggest that lowering antenatal GC-course may be less detrimental for brain without affecting lung maturation. Further clinical trials are needed to elucidate the low GC-course issue.

Evaluation of urinary S100B protein level and lactate/creatinine ratio for early diagnosis and prognostic prediction of neonatal hypoxic-ischemic encephalopathy.

BACKGROUND: Early identification and prevention of hypoxic-ischemic encephalopathy (HIE) in newborns may reduce neonatal mortality and neurological dysfunction. OBJECTIVE: To analyze the diagnostic and prognostic values of urinary S100B level and lactate/creatinine ratio in newborns with HIE. METHODS: Seventy-eight full-term newborns with HIE and 25 normal newborns were enrolled. The Neonatal Behavioral Neurological Assessment (NBNA) and Developmental Screening Test were scored. The concentration of urinary S100B protein was determined using the S100B enzyme-linked immunosorbent assay and the levels of urinary lactate and creatinine were measured with the enzyme colorimetric method. RESULTS: Urinary S100B level on days 1-3 after birth and lactate/creatinine ratio on day 1 were significantly higher in newborns with HIE than those in the control group. Both indexes were positively correlated with the clinical grading of HIE. A cutoff value for the S100B level of 0.47 microg/l on day 3 after birth had a sensitivity of 90% and specificity of 92% for prediction of HIE. A lactate/creatinine ratio of more than 0.55 on day 1 showed the highest sensitivity (92%) and specificity (90%). A combination of both indexes improved the sensitivity and specificity to 99 and 97%, respectively. A negative correlation of both lactate/creatinine ratio on day 1 and S100B level on days 1-3 after birth with the NBNA score was identified on days 3, 7 and 14 after birth. The Developmental Screening Test score of 36 newborns with HIE within 6 months after birth showed that 65% of infants with moderate and high HIE had an abnormal developmental quotient. CONCLUSION: These data suggest that early measurement of both S100B level and lactate/creatinine ratio in the urine of newborns with HIE is a practical convenient and sensitive way to improve diagnosis on the third day of life and prognostic prediction of HIE.

S100B protein and near infrared spectroscopy in preterm and term newborns.

Cerebral monitoring constitutes an emerging issue in perinatal medicine. Near Infrared Spectroscopy (NIRS) monitors brain oxygenation status in sick infants although data in healthy infants are lacking. The present study investigates whether NIRS parameters change according to gestational age and correlate with S100B protein. We recruited 64 healthy newborns (weeks' gestation: 30-42 wks) in which we performed in the first 6-hours after birth routine clinical, radiological and laboratory variables, cerebral oxygen saturation (rSO2), fractional cerebral tissue oxygen extraction (FTOE) values and S100B urine assessment. rSO2 and FTOE correlated (R=-0.73; R=0.51; P less than 0.01, for both) with gestational age. Highest rSO2 and the lowest FTOE peaks (P less than 0.001) were found at 30-33 wks. From 34 wks onwards, rSO2 progressively decreased and FTOE increased reaching their lower dip/peak (P less than 0.001) at 38-39 weeks. A significant correlation between S100B and NIRS parameters (rSO2: r=0.77; FTOE: r=-0.69; P less than 0.01) has been found. The present study shows that NIRS parameters and S100B protein correlation may be of help in brain function monitoring.

Can admission S-100beta predict the extent of brain damage in head trauma patients?

BACKGROUND: As has been shown previously, S-100beta levels in serum can be a useful predictor of brain damage after head trauma. This pilot study was designed to investigate whether urine samples, which are much easier to obtain, could be used for the same purpose instead of serum samples. METHODS: Ninety-six consecutive patients admitted with head trauma were recruited in the study. After exclusion of 54 patients, mostly because of significant additional trauma, S-100beta levels were analyzed in serum and urine of 42 patients using a luminometric assay. A range for normal values was established based on samples from ten healthy volunteers. RESULTS: S-100beta serum levels increased proportional to the severity of the head trauma, as had been previously shown by several other groups. In many patients, initial increases in urine S-100beta levels were seen later than in serum, after which the kinetics of S-100beta levels in urine seemed to follow that established for serum levels. S-100beta values in urine were on average about 54% lower in urine than in serum. CONCLUSIONS: S-100beta levels in urine obtained on admission to the hospital are not a good indicator for the extent of brain damage. However, urine S-100beta levels obtained at later time points might be a useful indicator for the development of secondary brain injury.

Urinary S100A1B and S100BB to predict hypoxic ischemic encephalopathy at term.

Urinary S100A1B and S100BB were measured to detect cases at risk of hypoxic-ischemic encephalopathy (HIE) in asphyxiated newborns. We recruited 42 asphyxiated infants and 63 healthy term neonates. S100A1B and S100BB were measured at first urination (time 0) and at 4 (time 1), 8 (time 2), 12 (time 3), 16 (time 4), 20 (time 5), 24 (time 6), 72 (time 7) hours after birth. 20 infants had no/mild HIE with good prognosis (Group A) and 22 had moderate/severe HIE with a greater risk of neurological handicap (Group B). Urine S100A1B and S100BB levels were significantly (P less than 0.0.01, for all) higher at all monitoring time-points in Group B than Group A and controls, but not between Group A and controls. Both S100A1B and S100BB have great sensitivity and specificity for HIE since their first measurement. In conclusion, S100A1B and S100BB are increased in urine collected from asphyxiated newborns who will develop HIE since first urination, and their measurement may be useful to early predict HIE when monitoring procedures are still of no avail.

Diagnostic accuracy of S100B urinary testing at birth in full-term asphyxiated newborns to predict neonatal death.

BACKGROUND: Neonatal death in full-term infants who suffer from perinatal asphyxia (PA) is a major subject of investigation, since few tools exist to predict patients at risk of ominous outcome. We studied the possibility that urine S100B measurement may identify which PA-affected infants are at risk of early postnatal death. METHODOLOGY/PRINCIPAL FINDINGS: In a cross-sectional study between January 1, 2001 and December 1, 2006 we measured S100B protein in urine collected from term infants (n = 132), 60 of whom suffered PA. According to their outcome at 7 days, infants with PA were subsequently classified either as asphyxiated infants complicated by hypoxic ischemic encephalopathy with no ominous outcome (HIE Group; n = 48), or as newborns who died within the first post-natal week (Ominous Outcome Group; n = 12). Routine laboratory variables, cerebral ultrasound, neurological patterns and urine concentrations of S100B protein were determined at first urination and after 24, 48 and 96 hours. The severity of illness in the first 24 hours after birth was measured using the Score for Neonatal Acute Physiology-Perinatal Extension (SNAP-PE). Urine S100B levels were higher from the first urination in the ominous outcome group than in healthy or HIE Groups (p<0.001 for all), and progressively increased. Multiple logistic regression analysis showed a significant correlation between S100B concentrations and the occurrence of neonatal death. At a cut-off >1.0 microg/L S100B had a sensitivity/specificity of 100% for predicting neonatal death. CONCLUSIONS/SIGNIFICANCE: Increased S100B protein urine levels in term newborns suffering PA seem to suggest a higher risk of neonatal death for these infants.

A comparison of two different assays for determining S-100B in serum and urine.

BACKGROUND: Brain injury after head trauma can be detected by S-100B measurements in serum. Recent preliminary studies indicate that urinary levels of S-100B are also increased after head injury, a finding that is of possible clinical value. The aims of the present study are two-fold: to compare serum measurements of two assays, the Liaison Sangtec 100 system and the Elecsys S100 test, and to investigate to what extent they can detect and measure S-100B in urine. METHODS: A total of 191 serum and 174 urine samples from 107 patients (children aged between 1 and 18 years following head trauma) were measured with both assays. The results were compared using correlation analysis and Bland-Altman difference plots. RESULTS: Serum values of the Sangtec system ranged from 0.02 to 2.28 microg/L, and from 0.005 to 2.13 microg/L for the Elecsys test. Comparisons showed a clear correlation (correlation coefficient 0.80) but not an agreement between the methods. The Sangtec system could only detect S-100B in 20 out of 174 urine samples (range 0.02-0.06 microg/L), whereas the Elecsys test could detect S-100B in 171 samples (range 0.005-0.14 microg/L). No clear relation was observed between the two methods in urine analysis (correlation coefficient 0.60). CONCLUSION: The Sangtec and Elecsys assays are not interchangeable methods when analyzing S-100B in serum or urine samples after head injury.

Emergency department measurement of urinary S100B in children following head injury: can extracranial injury confound findings?

OBJECTIVE: To assess the potential role of urinary S100B as a prognostic biochemical marker following head injury in children in a UK emergency department setting. METHODS: A case-control pilot study was performed in 20 patients with head injury and 15 controls (with extracranial trauma) aged <13 years and within 12 h of their injury recruited over a 4-month period. Urinary S100B levels were measured at presentation to the emergency department. RESULTS: The two groups showed no significant differences in basic characteristics (height, weight, time to sample collection). 50% of the case group had measurable concentrations of S100B following head injury (range 0.02-0.07 microg/l). All patients in the control group had measurable S100B concentrations following extracranial trauma (range 0.02-0.09 microg/l). No significant rise in S100B concentrations occurred in two patients with severe head injuries (Glasgow Coma Score (GCS) <9) and in one patient with a moderate head injury (GCS 10), despite significant injuries on the CT scan. CONCLUSION: Despite detecting measurable S100B levels in urine following head injury, the same levels are measured following extracranial trauma. Urinary S100B is therefore not useful as an early biochemical marker following head injury in children.