Shotgun Proteomics of Isolated Urinary Extracellular Vesicles for Investigating Respiratory Impedance in Healthy Preschoolers.

Urine proteomic applications in children suggested their potential in discriminating between healthy subjects from those with respiratory diseases. The aim of the current study was to combine protein fractionation, by urinary extracellular vesicle isolation, and proteomics analysis in order to establish whether different patterns of respiratory impedance in healthy preschoolers can be characterized from a protein fingerprint. Twenty-one 3-5-yr-old healthy children, representative of 66 recruited subjects, were selected: 12 late preterm (LP) and 9 full-term (T) born. Children underwent measurement of respiratory impedance through Forced Oscillation Technique (FOT) and no significant differences between LP and T were found. Unbiased clustering, based on proteomic signatures, stratified three groups of children (A, B, C) with significantly different patterns of respiratory impedance, which was slightly worse in group A than in groups B and C. Six proteins (Tripeptidyl peptidase I (TPP1), Cubilin (CUBN), SerpinA4, SerpinF1, Thy-1 membrane glycoprotein (THY1) and Angiopoietin-related protein 2 (ANGPTL2)) were identified in order to type the membership of subjects to the three groups. The differential levels of the six proteins in groups A, B and C suggest that proteomic-based profiles of urinary fractionated exosomes could represent a link between respiratory impedance and underlying biological profiles in healthy preschool children.

Urinary ERdj3 and mesencephalic astrocyte-derived neutrophic factor identify endoplasmic reticulum stress in glomerular disease.

Podocyte injury and endoplasmic reticulum (ER) stress have been implicated in the pathogenesis of various glomerular diseases. ERdj3 (DNAJB11) and mesencephalic astrocyte-derived neurotrophic factor (MANF) are ER chaperones lacking the KDEL motif, and may be secreted extracellularly. Since podocytes reside in the urinary space, we examined if podocyte injury is associated with secretion of KDEL-free ER chaperones from these cells into the urine, and if chaperones in the urine reflect ER stress in glomerulonephritis. In cultured podocytes, ER stress increased ERdj3 and MANF intracellularly and in culture medium, whereas GRP94 (KDEL chaperone) increased only intracellularly. ERdj3 and MANF secretion was blocked by the secretory trafficking inhibitor, brefeldin A. Urinary ERdj3 and MANF increased in rats injected with tunicamycin (in the absence of proteinuria). After induction of passive Heymann nephritis (PHN) and puromycin aminonucleoside nephrosis (PAN), there was an increase in glomerular ER stress, and appearance of ERdj3 and MANF in the urine, coinciding with the onset of proteinuria. Rats with PHN were treated with the chemical chaperone, 4-phenyl butyrate (PBA), starting at the time of disease induction, or after disease was established. In both protocols, 4-PBA reduced proteinuria and urinary ER chaperone secretion, compared with PHN rats treated with saline (control). In conclusion, urinary ERdj3 and MANF reflect glomerular ER stress. 4-PBA protected against complement-mediated podocyte injury and the therapeutic response could be monitored by urinary ERdj3 and MANF.

Urinary Biomarkers in Overactive Bladder: Revisiting the Evidence in 2019.

CONTEXT: In overactive bladder (OAB), after an initial outbreak of research, it is more consensual that biomarkers may be better used to phenotype patients. Herein, we revisit this topic, including some of the most promising biomarkers. OBJECTIVE: To provide a comprehensive analysis of the actual role of biomarkers in OAB. EVIDENCE ACQUISITION: A PubMed-based literature search was conducted, including the most relevant articles published in the last 15 yr, on nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), adenosine triphosphate (ATP), genomics, and microbiota as OAB biomarkers. Articles with no full text available or not written in English were excluded. Additional reviews were included. EVIDENCE SYNTHESIS: Urinary NGF, BDNF, and ATP are increased in many OAB patients. These biomarkers can help identify OAB phenotypes and select the ideal candidates for new therapies directed to neurotrophic and purinergic pathways. Circulating urinary miRNA may be useful for establishing the ideal moment for bladder outlet obstruction relief and will eventually lead to the development of therapeutic agents that inhibit or reverse fibrotic pathways in the bladder. Urinary microbiota seems to be related to OAB symptoms, in particular urgency urinary incontinence, and may have strong implications in the prevention, diagnosis, and treatment of OAB. CONCLUSIONS: In the future, physicians may consider the use of biomarkers to identify distinct OAB phenotypes, with distinct causal mechanisms, selecting patients for specific target therapies with expected better outcomes. PATIENT SUMMARY: Overactive bladder biomarkers can be useful for phenotype patients and for selecting more effective target therapies.

Urinary nerve growth factor: a biomarker for detrusor overactivity in children? A meta-analysis and trail sequential analysis.

PURPOSE: Based on, previously, a systematic review, urinary nerve growth factor (NGF) has emerged as one potentially noninvasive biomarker for detrusor overactivity (DO) in adults. We performed this systematic review to explore if NGF is a biomarker for DO in children. METHODS: A literature search was conducted in PubMed, Embase, Web of science, and Cochrane Library. Copies of all relevant articles were retrieved for quality assessment and data abstraction by two reviewers. Primary outcome was pooled standardized mean difference (SMD) for NGF/Cr (NGF normalized to urine creatinine) level between DO group and controls. RESULTS: Three case-control studies published from 2012 to 2016 were included with 74 patients and 70 controls. Children with DO had a significant higher baseline urinary NGF/Cr level compared to controls (SMD = 2.48, 95%CI = 0.85-4.10, P < 0.01). After treatment, the level of NGF/Cr decreased significantly compared to baseline level at 6th month time points (SMD = 0.94, 95%CI = 0.03-1.86, P = 0.04). We calculated the required information size to 99 patients for comparison of urinary NGF/Cr level between DO and controls by trail sequential analysis (TSA). CONCLUSION: Based on this systematic review, NGF/Cr may be a noninvasive biomarker for DO in children in the future. However, based on TSA, more original studies are needed to clarify the role of NGF/Cr in the biomarker effect.

[Sensitivity and specificity of urinary and serum neurotrophins in the diagnosis of neurogenic detrusor overactivity in multiple sclerosis].

Lower urinary tract dysfunction is common among neurological patients. Traditionally, the basic method of diagnosis is a complex urodynamic study. In recent years, many studies have focused on the search for new non-invasive diagnostic modalities. In particular, neurotrophins are considered as potential biological markers of a neurogenic bladder. AIM: To estimate the sensitivity and specificity of the serum and urinary nerve growth factor (NGF) and brain neurotrophic factor (BDNF) in MS patients as markers of detrusor overactivity. MATERIALS AND METHODS: The study comprised 20 patients with multiple sclerosis, who complained of voiding problems. The control group consisted of 20 people without neurological diseases, lower urinary tract symptoms and detrusor overactivity estimated by filling cystometry. Apart from standard laboratory tests, diagnostic evaluation included a complex urodynamic study, ultrasound of the urinary tract, cystoscopy, testing serum and urinary NGF and BDNF using the enzyme immunoassay. The diagnostic significance of neurotrophins was evaluated using ROC analysis. RESULTS: According to the ROC analysis, the diagnostic sensitivity and specificity of serum NGF as a marker of detrusor hyperactivity was 57% and 93%, respectively (for serum NGF more or equal 26 pg/ml). The quality of the test according to the expert scale of AUC values was "very good" (AUC=0.806). Detecting NGF in patients urine was less effective. The sensitivity and specificity were 52% and 40%, respectively (for NGF more or equal 6 pg/ml). The quality of the test according to the expert scale of AUC values was "average" (AUC=0.64). The serum BDNF demonstrated high sensitivity (90%) and low specificity (23%), AUC=0.56. The urinary BDNF was more informative, (AUC=0.65). The combination of all four markers provides a sensitivity of 85.7% and a specificity of 66.7% (AUC=0.824). CONCLUSIONS: Testing serum and urinary neurotrophins in patients with multiple sclerosis can be used to diagnose detrusor overactivity. The NGF is a highly specific biomarker, while the BDNF is highly sensitive. Combined testing for serum NGF and BDNF is most informative.

Urinary Levels of IL-1ß and GDNF in Preterm Neonates as Potential Biomarkers of Motor Development: A Prospective Study.

Objectives. To evaluate the association between inflammatory biomarkers, neurotrophic factors, birth conditions, and the presence of motor development abnormalities in preterm neonates. Methods. Plasma and urinary levels of cytokines (IL-1ß, IL-6, IL-10, TNF, and IL-12p70), chemokines (CXCL8/IL-8, CCL2/MCP-1, CCL5/RANTES, CXCL10/IP-10, and CXCL9/MIG), and neurotrophic factors (BDNF and GDNF) were evaluated in 40 preterm neonates born between 28 and 32 incomplete weeks of gestation, at four distinct time points: at birth (umbilical cord blood) (T0), at 48 (T1), at 72 hours (T2), and at 3 weeks after birth (T3). Biomarkers levels were compared between different time points and then associated with Test of Infant Motor Performance (TIMP) percentiles. Results. Maternal age, plasma, and urinary concentrations of inflammatory molecules and neurotrophic factors were significantly different between groups with normal versus lower than expected motor development. Higher levels of GDNF were found in the group with lower than expected motor development, while IL-1ß and CXCL8/IL-8 values were higher in the group with typical motor development. Conclusion. Measurements of cytokines and neurotrophic factors in spot urine may be useful in the follow-up of motor development in preterm neonates.

Association of urinary nerve growth factor levels with erectile function in young men with type 2 diabetes mellitus.

We investigated urine nerve growth factor (NGF) levels and erectile dysfunction in diabetic men <45 years of age. Urinary NGF levels were measured in 72 diabetic men and 20 control subjects without lower urinary tract symptoms or erectile dysfunction. Participants were evaluated using the International Prostate Symptom Score, quality of life index, Overactive Bladder Symptom Score (OABSS), the five-item version of the International Index of Erectile Function questionnaire (IIEF-5), the patient perception of bladder condition questionnaire, measurement of flow rate and post-void residual urine volume. The results showed that the diabetic men had significantly higher urinary normalized NGF/creatinine (Cr) levels compared to the healthy controls (0.48±1.2 vs 0.01±0.01, P=0.04). The increased urinary NGF/Cr levels correlated negatively with the IIEF-5 total score (P=0.03, coefficient=-0.26, -0.02 to -0.47). The 42 patients with urinary NGF/Cr levels <0.05 had higher IIEF-5 scores than the 30 patients with urinary NGF/Cr level ⩾0.05 (20.2±4.6 vs 16.9±6.7, P=0.03). We conclude that urinary NGF levels were associated with erectile dysfunction in the men with type 2 <45 years of age.

Efficacy of urinary midkine as a biomarker in patients with acute kidney injury.

BACKGROUND: The mortality and morbidity associated with acute kidney injury (AKI) remains high, despite advances in interventions. A multifunctional heparin-binding growth factor, midkine (MK), is involved in the pathogenesis of ischemic kidney injury. However, the clinical relevance of MK has not yet been elucidated. The present study investigated whether urinary MK can serve as a novel biomarker of AKI. METHODS: We initially compared the predictive value of MK with other urinary biomarkers, including N-acetyl-ß-D-glucosaminidase (NAG), interleukin (IL)-18, and neutrophil gelatinase-associated lipocalin (NGAL), for the detection and differential diagnosis of established AKI (549 patients). Subsequently, the reliability of MK for the early detection of AKI was prospectively evaluated in 40 patients undergoing elective abdominal aortic aneurysm surgery. Urine samples were obtained at baseline, the period of aortic cross-clamping and declamping, the end of the surgery, and on post-operative day 1. RESULTS: The areas under the receiver operating characteristic curves for the diagnosis of AKI in various kidney diseases were 0.88, 0.70, 0.72, and 0.84 for MK, NAG, IL-18, and NGAL, respectively. When the optimal cutoff value of urinary MK was set at 11.5 pg/mL, the sensitivity and specificity were 0.87 and 0.85, respectively. In the second study, urinary MK peaked at the period of aortic declamping, about 1 h after cross-clamping in patients with AKI. Interestingly, the rise of MK in AKI patients was very precipitous compared with other biomarker candidates. CONCLUSION: Urinary MK was prominent in its ability to detect AKI and may allow the start of preemptive medication.

Urinary markers of acute kidney injury in newborns with perinatal asphyxia (.).

BACKGROUND: Acute kidney injury (AKI) affects up to 60% of severely asphyxiated neonates. The diagnosis of AKI can be and is further challenged by a lack of good biomarkers. We studied the role of novel markers for AKI, neutrophil gelatinase-associated lipocalin (NGAL), interleukin-8 (IL-18), Netrin-1 (NTN-1), and sodium hydrogen exchanger isoform 3 (NHE3) on development and early diagnosis of AKI in newborns with perinatal asphyxia (PA). METHODS: Forty-one newborns with a diagnosis of PA (15 with AKI and 26 without AKI) and 20 healthy matched controls were involved to the study. Urinary samples were obtained on postnatal days 1 and 4 for patients with PA and on postnatal day 1 for the control subjects. AKI was defined using a serum creatinine-based modification of the acute kidney injury network criteria. RESULTS: The levels of NGAL, NTN-1, NHE3, and IL-18 on the first postnatal day urine samples were higher in patients compared to controls (p < 0.001, p <0.001, p  <0.02, p  <0.001, respectively). In patients with AKI, the levels of NGAL and IL-18 were higher when compared to patients without AKI (p = 0.002, p  <0.001, respectively). The levels of NTN-1 and NHE3 were similar in both groups. For the samples obtained on postnatal day 4, only NGAL levels were significantly higher in patients with AKI (p = 0.004) compared to those without AKI. CONCLUSION: To our knowledge, this is the largest study, which evaluated the utility of urinary biomarkers in the diagnosis of AKI in newborns with PA. First day, urine NGAL and IL-18 levels have an important diagnostic power in such patients.

Urinary Netrin-1: A New Biomarker for the Early Diagnosis of Renal Damage in Obese Children.

OBJECTIVE: Urinary netrin-1 is a new marker to demonstrate early tubular damage. The aim of this study was to determine whether urinary netrin-1 is increased in obese children. METHODS: A total of 68 normoalbuminuric and normotensive obese patients and 65 controls were included in the study. Urine samples were collected for assessment of urinary phosphorus, sodium, potassium, creatinine, albumin, and netrin-1. Blood samples were collected for measurements of fasting glucose, insulin, lipid, phosphorus, sodium, potassium, and creatinine levels. Homeostatic model assessment insulin resistance index was calculated. RESULTS: Gender and age were similar between obese and control groups (12.01±3.03 vs. 11.7±3.2 years, p=0.568 and 33 vs. 35 girls, p=0.543, respectively). Obese patients had significantly higher netrin-1 excretion than the controls (841.68±673.17 vs. 228.94±137.25 pg/mg creatinine, p=0.000). Urinary netrin-1 level was significantly higher in obese subjects with insulin resistance compared to those without insulin resistance (1142±1181 vs. 604.9±589.91 pg/mg creatinine, p=0.001). CONCLUSION: In normotensive and normoalbuminuric obese children, urinary netrin-1 level can increase before onset of albuminuria. Urinary netrin-1 excretion appears to be affected predominantly by insulin resistance and hyperinsulinemia. Urinary netrin-1 may be a new biomarker for determining early tubular injury in obese children.

Mesencephalic Astrocyte-Derived Neurotrophic Factor as a Urine Biomarker for Endoplasmic Reticulum Stress-Related Kidney Diseases.

Endoplasmic reticulum (ER) stress and disrupted proteostasis contribute to the pathogenesis of a variety of glomerular and tubular diseases. Thus, it is imperative to develop noninvasive biomarkers for detecting ER stress in podocytes or tubular cells in the incipient stage of disease, when a kidney biopsy is not yet clinically indicated. Mesencephalic astrocyte-derived neurotrophic factor (MANF) localizes to the ER lumen and is secreted in response to ER stress in several cell types. Here, using mouse models of human nephrotic syndrome caused by mutant laminin ß2 protein-induced podocyte ER stress and AKI triggered by tunicamycin- or ischemia-reperfusion-induced tubular ER stress, we examined MANF as a potential urine biomarker for detecting ER stress in podocytes or renal tubular cells. ER stress upregulated MANF expression in podocytes and tubular cells. Notably, urinary MANF excretion concurrent with podocyte or tubular cell ER stress preceded clinical or histologic manifestations of the corresponding disease. Thus, MANF can potentially serve as a urine diagnostic or prognostic biomarker in ER stress-related kidney diseases to help stratify disease risk, predict disease progression, monitor treatment response, and identify subgroups of patients who can be treated with ER stress modulators in a highly targeted manner.

[Diagnostic values of urinary netrin-1 and kidney injury molecule-1 for acute kidney injury induced by neonatal asphyxia].

OBJECTIVE: To investigate the values of urinary netrin-1 and kidney injury molecule-1 (KIM-1) in the early diagnosis of acute kidney injury (AKI) induced by neonatal asphyxia. METHODS: A total of 80 full-term neonates with asphyxia were enrolled (mild asphyxia: 34 neonates; severe asphyxia: 46 neonates). Forty normal full-term neonates were selected as the control group. Urinary samples were collected from the neonates in the three groups within 12 hours and 13-48 hours after birth. ELISA was applied to measure urinary levels of netrin-1 and KIM-1. Peripheral venous blood samples were also collected to measure serum creatinine (Scr) level. RESULTS: Compared with the control group, the asphyxia group had significantly higher urinary levels of netrin-1 and KIM-1 within 48 hours after birth and a significantly higher Scr level within 13-48 hours after birth (P<0.05). The neonates in the AKI group had significantly higher urinary levels of netrin-1 and KIM-1 and Scr level within 48 hours after birth than those in the non-AKI group (P<0.05). The areas under the receiver operating characteristic curve for urinary netrin-1 and KIM-1 levels within 12 hours after birth to predict AKI after asphyxia were 0.878 (95% CI: 0.775-0.981; P<0.01) and 0.899 (95% CI: 0.829-0.969; P<0.01), respectively. Any two indicators of urinary netrin-1 level, urinary KIM-1 level, and Scr level within 12 hours after neonatal asphyxia had a positive correlation (P<0.05). CONCLUSIONS: Urinary netrin-1 and KIM-1 levels increase significantly when neonates with asphyxia develop AKI. Urinary netrin-1 and KIM-1 can be used as indicators for the early diagnosis of AKI after asphyxia.

Netrin-1 and semaphorin 3A predict the development of acute kidney injury in liver transplant patients.

Acute kidney injury (AKI) is a serious complication after liver transplantation. Currently there are no validated biomarkers available for early diagnosis of AKI. The current study was carried out to determine the usefulness of the recently identified biomarkers netrin-1 and semaphorin 3A in predicting AKI in liver transplant patients. A total of 63 patients' samples were collected and analyzed. AKI was detected at 48 hours after liver transplantation using serum creatinine as a marker. In contrast, urine netrin-1 (897.8 ± 112.4 pg/mg creatinine), semaphorin 3A (847.9 ± 93.3 pg/mg creatinine) and NGAL (2172.2 ± 378.1 ng/mg creatinine) levels were increased significantly and peaked at 2 hours after liver transplantation but were no longer significantly elevated at 6 hours after transplantation. The predictive power of netrin-1, as demonstrated by the area under the receiver-operating characteristic curve for diagnosis of AKI at 2, 6, and 24 hours after liver transplantation was 0.66, 0.57 and 0.59, respectively. The area under the curve for diagnosis of AKI was 0.63 and 0.65 for semaphorin 3A and NGAL at 2 hr respectively. Combined analysis of two or more biomarkers for simultaneous occurrence in urine did not improve the AUC for the prediction of AKI whereas the AUC was improved significantly (0.732) only when at least 1 of the 3 biomarkers in urine was positive for predicting AKI. Adjusting for BMI, all three biomarkers at 2 hours remained independent predictors of AKI with an odds ratio of 1.003 (95% confidence interval: 1.000 to 1.006; P = 0.0364). These studies demonstrate that semaphorin 3A and netrin-1 can be useful early diagnostic biomarkers of AKI after liver transplantation.

Urinary netrin-1 and KIM-1 as early biomarkers for septic acute kidney injury.

BACKGROUND: Acute kidney injury (AKI) during sepsis is associated with poor outcome. However, diagnosis of AKI with serum creatinine (SCr) level change is neither highly sensitive nor specific. Therefore, identification of novel biomarkers for early diagnosis of AKI is desirable. AIMS: To evaluate the capacity of combining urinary netrin-1 and human kidney injury molecule type 1 (KIM-1) in the early diagnosis of septic AKI. METHODS: We prospectively recruited 150 septic patients from Jun 2011 to Jun 2013 at Zhejiang Provincial People's Hospital, China. SCr, urinary netrin-1, and KIM-1 levels were recorded at 0, 1, 3, 6, 24, and 48 h of ICU admission and compared between AKI and non-AKI patients. In addition, we investigated the prognostic value of netrin-1 and KIM-1 between non-survivors and survivors in septic AKI patients. RESULTS: SCr levels started to show elevation after 24 h of ICU admission. However, netrin-1 levels increased significantly as early as 1 h, peaked at 3-6 h and remained elevated up to 48 h of ICU admission in septic AKI patients. KIM-1 increased significantly by 6 h, peaked at 24 h and remained significantly elevated until 48 h of ICU admission. Furthermore, we observed significant higher urinary KIM-1 levels at 24 h and 48 h in non-survivors compared to survivors in AKI patients. CONCLUSIONS: Our results suggest that both netrin-1 and KIM-1 are clinically useful as early biomarkers in the diagnosis of septic AKI. In addition, persistent elevation of urinary KIM-1 level may be associated with poor prognosis.

Epigenetic inactivation of VGF associated with Urothelial Cell Carcinoma and its potential as a non-invasive biomarker using urine.

BACKGROUND: To identify new epigenetic markers and further characterize Urothelial Cell Carcinoma (UCC), we tested the promoter methylation (PM) status of 19 genes previously identified as cancer specific methylated genes in other solid tumors. METHODS: We used bisulfite sequencing, methylation specific PCR and quantitative methylation specific PCR (QMSP) to test the PM status of 19 genes in urothelial cancer cell lines. RESULTS: Among the 19 genes tested, VGF was found to be completely methylated in several UCC cell lines. VGF QMSP analysis showed that methylation values of almost all the primary 19 UCC tissues were higher than the paired normal tissues (P=0.009). In another cohort, 12/35 (34.3%) of low grade UCC cases displayed VGF methylation. As a biomarker for non-invasive detection of UCC, VGF showed a significantly higher frequency of methylation in urine from UCC cases (8/20) compared to controls (1/20) (P=0.020). After treatment of cell lines with 5-Aza-2'-deoxycytidine, VGF was robustly re-expressed. Forced expression of VGF in bladder cancer cell lines inhibited cell growth. CONCLUSION: Selection of candidates from genome-wide screening approach in other solid tumors successfully identified UCC specific methylated genes.

Netrin-1 promotes medulloblastoma cell invasiveness and angiogenesis, and demonstrates elevated expression in tumor tissue and urine of patients with pediatric medulloblastoma.

Invasion and dissemination of medulloblastoma within the central nervous system is the principal factor predicting medulloblastoma treatment failure and death. Netrin-1 is an axon guidance factor implicated in tumor and vascular biology, including in invasive behaviors. We found that exogenous netrin-1 stimulated invasion of human medulloblastoma cells and endothelial cells in contrast to VEGF-A, which promoted invasion of endothelial cells but not medulloblastoma cells. Furthermore, medulloblastoma cells expressed endogenous netrin-1 along with its receptors, neogenin and UNC5B. Blockades in endogenous netrin-1, neogenin, or UNC5B reduced medulloblastoma invasiveness. Neogenin blockade inhibited netrin-1-induced endothelial cells tube formation and recruitment of endothelial cells into Matrigel plugs, two hallmarks of angiogenesis. In patients with pediatric medulloblastoma, netrin-1 mRNA levels were increased 1.7-fold in medulloblastoma tumor specimens compared with control specimens from the same patient. Immunohistochemical analyses showed that netrin-1 was elevated in medulloblastoma tumors versus cerebellum controls. Notably, urinary levels of netrin-1 were 9-fold higher in patients with medulloblastoma compared with control individuals. Moreover, urinary netrin-1 levels were higher in patients with invasive medulloblastoma compared with patients with noninvasive medulloblastoma. Finally, we noted that urinary netrin-1 levels diminished after medulloblastoma resection in patients. Our results suggest netrin-1 is a candidate biomarker capable of detecting an invasive, disseminated phenotype in patients with medulloblastoma and predicting their disease status.

Age-related differences in kidney injury biomarkers induced by cisplatin.

Acute kidney injury (AKI) occurs in a half of cisplatin (CDDP)-treated patients. Traditional biomarkers including blood urea nitrogen (BUN) and serum creatinine (SCr) are still used for detection of CDDP-induced AKI, but these biomarkers are not specific or sensitive. The aim of this study was to identify the specific and sensitive biomarkers against CDDP-induced renal injury between young (3-week-old) and old (20-week-old) rats. All animals were intraperitoneally injected once with CDDP (6 mg/kg). After 3 days, all animals were sacrificed and serum, urine, and kidney tissues were collected. Urinary and serum biomarkers as well as histological changes were measured. CDDP-induced proximal tubular damage was apparent from histopathological examination, being more severe in 3-week-old rats accompanied by increased number of TUNEL-positive apoptotic cells. This was associated with elevated urinary kidney injury molecule-1 (KIM-1), glutathione-S-transferase alpha (GST-α), vascular endothelial growth factor (VEGF), and tissue inhibitor of metalloproteinases-1 (TIMP-1). In contrast, the levels of neutrophil gelatinase-associated lipocalin (NGAL) and osteopontin were significantly increased in 20-week-old rats after CDDP treatment. These results indicate that the use of age-specific urinary biomarkers is necessary to diagnosis of CDDP-induced AKI. Especially, urinary KIM-1, GST-α, TIMP-1, and VEGF levels may help in the early diagnosis of young patients with CDDP-induced AKI.

Netrin-1, a urinary proximal tubular injury marker, is elevated early in the time course of human diabetes.

BACKGROUND: Netrin-1 was recently identified as an early diagnostic biomarker of chronic kidney disease (CKD) in an experimental animal model. However, its usefulness for early diagnosis of CKD in humans is unknown. The current study evaluated whether netrin-1 is increased in urine from human diabetic patients. METHODS: Spot urine samples from healthy volunteers, diabetes without microalbuminuria, diabetes with microalbuminuria and diabetes with macroalbuminuria were collected after receiving consent. Netrin-1 in urine was quantified by enzyme-linked immunosorbent assay and the data analyzed to determine whether urinary netrin-1 significantly correlates with disease progression. RESULTS: Urinary netrin-1 levels were significantly increased in normoalbuminuric diabetic patients compared to healthy controls, and still further elevated in patients with microalbuminuria and overt nephropathy. Urinary netrin-1 was significantly associated with albuminuria and estimated glomerular filtration rate, independently of age and sex. CONCLUSION: Netrin-1 is detectable in urine from diabetic patients and may serve as a useful early diagnostic biomarker predicting the development of CKD in diabetes.

Urine protein biomarkers for the diagnosis and prognosis of necrotizing enterocolitis in infants.

OBJECTIVES: To test the hypothesis that an exploratory proteomics analysis of urine proteins with subsequent development of validated urine biomarker panels would produce molecular classifiers for both the diagnosis and prognosis of infants with necrotizing enterocolitis (NEC). STUDY DESIGN: Urine samples were collected from 119 premature infants (85 NEC, 17 sepsis, 17 control) at the time of initial clinical concern for disease. The urine from 59 infants was used for candidate biomarker discovery by liquid chromatography/mass spectrometry. The remaining 60 samples were subject to enzyme-linked immunosorbent assay for quantitative biomarker validation. RESULTS: A panel of 7 biomarkers (alpha-2-macroglobulin-like protein 1, cluster of differentiation protein 14, cystatin 3, fibrinogen alpha chain, pigment epithelium-derived factor, retinol binding protein 4, and vasolin) was identified by liquid chromatography/mass spectrometry and subsequently validated by enzyme-linked immunosorbent assay. These proteins were consistently found to be either up- or down-regulated depending on the presence, absence, or severity of disease. Biomarker panel validation resulted in a receiver-operator characteristic area under the curve of 98.2% for NEC vs sepsis and an area under the curve of 98.4% for medical NEC vs surgical NEC. CONCLUSIONS: We identified 7 urine proteins capable of providing highly accurate diagnostic and prognostic information for infants with suspected NEC. This work represents a novel approach to improving the efficiency with which we diagnose early NEC and identify those at risk for developing severe, or surgical, disease.

Current developments in early diagnosis of acute kidney injury.

Acute kidney injury (AKI) is a very frequent and serious clinical problem, accounting for overall high morbidity and mortality. Up to date, mortality due to AKI is virtually unchanged over the past 50 years. This may partly be explained due to a delay in initiating renal protective and appropriate therapeutic measures since until now there are no reliable early-detecting biomarkers. The gold standard, serum creatinine, displays poor specificity and sensitivity with regard to identification of the incipient phase of AKI, and this is also true for cystatin C. We aimed to review novel biomarkers of AKI in urine and serum which have now progressed to the clinical phase. The main focus refers to their diagnostic and prognostic value. For this purpose, a web-based literature search using PubMed was performed comprising the following terms: renal failure, acute kidney injury and biomarkers. New molecules such as neutrophil gelatinase-associated lipocalin (NGAL), kidney injury molecule-1 (KIM-1), N-acetyl-ß-D-glucosaminidase (NAG), monocyte chemotactic peptide (MCP-1), Il-18, liver-type fatty acid-binding protein (L-FABP) and Netrin-1 are available and represent promising new markers that, however, need to be further evaluated in the clinical setting for suitability. In clinical settings with incipient AKI, not only the development and the implementation of more sensitive, practicable and accurate biomarkers are required for well-timed treatment initiation. Just as important is a substantial improvement of refined and applicable prophylactic therapeutic options in these situations. Before full adoption in clinical practice can be accomplished, adequately powered clinical trials testing a row of biomarkers are strongly warranted.