Investigating the value of urinary biomarkers in relation to lupus nephritis histopathology: present insights and future prospects.

Lupus nephritis (LN), a leading cause of death in Systemic Lupus Erythematosus (SLE) patients, presents significant diagnostic and prognostic challenges. Although renal pathology offers critical insights regarding the diagnosis, classification, and therapy for LN, its clinical utility is constrained by the invasive nature and limited reproducibility of renal biopsies. Moreover, the continuous monitoring of renal pathological changes through repeated biopsies is impractical. Consequently, there is a growing interest in exploring urine as a non-invasive, easily accessible, and dynamic "liquid biopsy" alternative to guide clinical management. This paper examines novel urinary biomarkers from a renal pathology perspective, encompassing cellular components, cytokines, adhesion molecules, auto-antibodies, soluble leukocyte markers, light chain fragments, proteins, small-molecule peptides, metabolomics, urinary exosomes, and ribonucleic acids. We also discuss the application of combined models comprising multiple biomarkers in assessing lupus activity. These innovative biomarkers and models offer insights into LN disease activity, acute and chronic renal indices, fibrosis, thrombotic microangiopathy, podocyte injury, and other pathological changes, potentially improving the diagnosis, management, and prognosis of LN. These urinary biomarkers or combined models may serve as viable alternatives to traditional renal pathology, potentially revolutionizing the method for future LN diagnosis and observation.

Urine Epidermal Growth Factor and Kidney Function Decline in Middle-Aged Adults.

Rationale & Objective: The diagnosis and prognostication of chronic kidney disease (CKD) largely rely on glomerular measures that may not reflect tubular damage. We investigated the associations of urine kidney tubule biomarkers with estimated glomerular filtration rate (eGFR) change among middle-aged adults, when chronic diseases typically emerge. Study Design: An observational cohort study. Setting & Participants: A total of 1,145 participants of the Coronary Artery Risk Development in Young Adults (CARDIA) study without CKD, hypertension, or cardiovascular disease at the year 20 visit. Exposures: Seven different biomarkers of tubular health: urine epidermal growth factor (EGF), alpha-1-microglobulin (α1m), interleukin-18, kidney injury molecule-1, monocyte chemoattractant protein-1, uromodulin, and chitinase-3-like protein 1. Outcomes: Ten-year eGFR change and incident reduced eGFR (new onset of eGFR < 60 mL/min/1.73 m2). Analytical Approach: We examined associations of tubular health biomarkers with 10-year eGFR change and incident reduced eGFR with linear mixed models and interval-censored proportional hazards regression models, respectively. Both minimally and fully adjusted models were controlled for urine creatinine levels. Results: The mean age of participants was 44.8 ± 3.7 years, with 39% African American and 56% female. The average 10-year change in eGFR was -18.6 mL/min/1.73 m2 (95% CI, -19.4 to -17.8). In contrast to the other tubular biomarkers, which showed conflicting results, EGF demonstrated strong, consistent associations with both kidney outcomes. Each 1-standard deviation (SD) higher EGF was associated with a 2.37 mL/min/1.73 m2 (95% CI, 0.64-4.10) smaller 10-year decrease in eGFR and a 42% (95% CI, 4%-64%) lower risk of incident reduced eGFR in the fully adjusted model. Limitations: Observational design, measurements of eGFR were done only at 5-year intervals during follow-up. Conclusions: In middle-aged, community-dwelling adults without hypertension, cardiovascular disease or CKD, higher urine EGF concentrations are associated with slower eGFR decline, whereas other kidney tubule biomarkers lacked a consistent association with kidney function decline.

Current measures of chronic kidney disease (CKD) rely on markers of glomerular health and function. This approach inadequately captures the role of kidney tubule health, a known histopathological predictor of CKD development. We investigated associations of 7 biomarkers of kidney tubule health with 10-year estimated glomerular filtration rate (eGFR) change and incident reduced eGFR. Among 7 biomarkers, only epidermal growth factor showed persistent and inverse associations with both 10-year eGFR change and incident reduced eGFR. These findings suggest that epidermal growth factor has an association with kidney function changes and might play a protective role in kidney disease development.

Application of Proteomics and Machine Learning Methods to Study the Pathogenesis of Diabetic Nephropathy and Screen Urinary Biomarkers.

Diabetic nephropathy (DN) has become the main cause of end-stage renal disease worldwide, causing significant health problems. Early diagnosis of the disease is quite inadequate. To screen urine biomarkers of DN and explore its potential mechanism, this study collected urine from 87 patients with type 2 diabetes mellitus (which will be classified into normal albuminuria, microalbuminuria, and macroalbuminuria groups) and 38 healthy subjects. Twelve individuals from each group were then randomly selected as the screening cohort for proteomics analysis and the rest as the validation cohort. The results showed that humoral immune response, complement activation, complement and coagulation cascades, renin-angiotensin system, and cell adhesion molecules were closely related to the progression of DN. Five overlapping proteins (KLK1, CSPG4, PLAU, SERPINA3, and ALB) were identified as potential biomarkers by machine learning methods. Among them, KLK1 and CSPG4 were positively correlated with the urinary albumin to creatinine ratio (UACR), and SERPINA3 was negatively correlated with the UACR, which were validated by enzyme-linked immunosorbent assay (ELISA). This study provides new insights into disease mechanisms and biomarkers for early diagnosis of DN.

A better understanding of basic science may help our management of LUTS/LUTD in older persons with nocturnal polyuria and nocturia: ICI-RS 2024.

AIMS: To discuss the role of autocrine/paracrine signaling of urothelial arginine vasopressin (AVP) on mammalian bladder capacities and micturition thresholds, impact of distension on water/urea reabsorption from the bladder, review of the literature to better characterize the central/peripheral effects of AVP, desmopressin (dAVP) toxicity, and urine biomarkers of nocturia. METHODS: This review summarizes discussions during an International Consultation on Incontinence-Research Society 2024 think tank with respect to the role of urothelial AVP in aged individuals with nocturnal polyuria, impact of solute and water reabsorption by the bladder on uninterrupted sleep, central effects of AVP, pharmacological basis of dAVP toxicity, and biomarkers in nocturia/lower urinary tract dysfunction (LUTD) with neurological diseases. RESULTS: Consensus recognized AVP function and pathways in the central nervous system (CNS), pre-proAVP localized using immunohistochemistry in bladder sections from adult/aged noncancerous human punch biopsies and rodent bladder sections is likely to accelerate the systemic uptake of water and urea from the bladder of anesthetized mice instilled with 3H-water and 14C-urea. Mechanisms for charged and uncharged solutes and water transport across the bladder, mechanism of dAVP toxicity, and utility of urine biomarkers in those with neurological diseases/nocturia were determined from literature reviews. CONCLUSION: Pre-proAVP is present in human/rodent bladders and may be involved in water reabsorption from bladder that prevents the sensation of fullness for uninterrupted sleep in healthy adults. The mechanism of action of AVP in the CNS was discussed, as was electrolyte/water transport across the bladder, the basis for dAVP toxicity, and feasibility of urine biomarkers to identify nocturia/LUTD with neurological diseases.

Urinary MicroRNA biomarkers of nephrotoxicity in Macaca fascicularis.

Drug-induced kidney injury (DIKI) refers to kidney damage resulting from the administration of medications. The aim of this project was to identify reliable urinary microRNA (miRNAs) biomarkers that can be used as potential predictors of DIKI before disease diagnosis. This study quantified a panel of six miRNAs (miRs-210-3p, 423-5p, 143-3p, 130b-3p, 486-5p, 193a-3p) across multiple time points using urinary samples from a previous investigation evaluating effects of a nephrotoxicant in cynomolgus monkeys. Exosome-associated miRNA exhibited distinctive trends when compared to miRNAs quantified in whole urine, which may reflect a different urinary excretion mechanism of miRNAs than those released passively into the urine. Although further research and mechanistic studies are required to elucidate how these miRNAs regulate signaling in disease pathways, we present, for the first time, data that several miRNAs displayed strong correlations with histopathology scores, thus indicating their potential use as biomarkers to predict the development of DIKI in preclinical studies and clinical trials. Also, these findings can potentially be translated into other non-clinical species or human for the detection of DIKI.

Urine Biomarkers of Kidney Tubule Health and Risk of Incident CKD in Persons Without Diabetes: The ARIC, MESA, and REGARDS Studies.

Rationale & Objective: Tubulointerstitial damage is a feature of early chronic kidney disease (CKD), but current clinical tests capture it poorly. Urine biomarkers of tubulointerstitial health may identify risk of CKD. Study Design: Prospective cohort (Atherosclerosis Risk in Communities [ARIC]) and case-cohort (Multi-Ethnic Study of Atherosclerosis [MESA] and Reasons for Geographic and Racial Differences in Stroke [REGARDS]). Setting & Participants: Adults with estimated glomerular filtration rate (eGFR) ≥60 mL/min/1.73 m2 and without diabetes in the ARIC, REGARDS, and MESA studies. Exposures: Baseline urine monocyte chemoattractant protein-1 (MCP-1), alpha-1-microglobulin (α1m), kidney injury molecule-1, epidermal growth factor, and chitinase-3-like protein 1. Outcome: Incident CKD or end-stage kidney disease. Analytical Approach: Multivariable Cox proportional hazards regression for each cohort; meta-analysis of results from all 3 cohorts. Results: 872 ARIC participants (444 cases of incident CKD), 636 MESA participants (158 cases), and 924 REGARDS participants (488 cases) were sampled. Across cohorts, mean age ranged from 60 ± 10 to 63 ± 8 years, and baseline eGFR ranged from 88 ± 13 to 91 ± 14 mL/min/1.73 m2. In ARIC, higher concentrations of urine MCP-1, α1m, and kidney injury molecule-1 were associated with incident CKD. In MESA, higher concentration of urine MCP-1 and lower concentration of epidermal growth factor were each associated with incident CKD. In REGARDS, none of the biomarkers were associated with incident CKD. In meta-analysis of all 3 cohorts, each 2-fold increase α1m concentration was associated with incident CKD (HR, 1.19; 95% CI, 1.08-1.31). Limitations: Observational design susceptible to confounding; competing risks during long follow-up period; meta-analysis limited to 3 cohorts. Conclusions: In 3 combined cohorts of adults without prevalent CKD or diabetes, higher urine α1m concentration was independently associated with incident CKD. 4 biomarkers were associated with incident CKD in at least 1 of the cohorts when analyzed individually. Kidney tubule health markers might inform CKD risk independent of eGFR and albuminuria.

This study analyzed 3 cohorts (ARIC, MESA, and REGARDS) of adults without diabetes or prevalent chronic kidney disease (CKD) to determine the associations of 5 urinary biomarkers of kidney tubulointerstitial health with incident CKD, independent of traditional measures of kidney health. Meta-analysis of results from all 3 cohorts suggested that higher baseline levels of urine alpha-1-microglobulin were associated with incident CKD at follow-up. Results from individual cohorts suggested that in addition to alpha-1-microglobulin, monocyte chemoattractant protein-1, kidney injury molecule-1, and epidermal growth factor may also be associated with the development of CKD. These findings underscore the importance of kidney tubule interstitial health in defining risk of CKD independent of creatinine and urine albumin.

Comparison of Urinary Biomarkers in Diagnosis of Ureteropelvic Junction Obstruction and Differentiation from Nonobstructive Dilatation.

Background and Aims: Differentiation of nonobstructive dilatation (NOD) from ureteropelvic junction obstruction (UPJO) is a challenge in children with antenatally detected hydronephrosis. The aim of this study is to compare the utility of urinary biomarkers: carbohydrate antigen (CA 19-9), neutrophil gelatinase-associated lipocalin (NGAL), and kidney injury molecule (KIM) in diagnosing UPJO. Methods: A prospective study was conducted after obtaining ethical clearance between 2021 and 2022. Group 1 - control group (n = 30): children with normal antenatal ultrasound with no urinary symptoms. Group 2 - study group (n = 48): children with unilateral hydronephrosis: Group 2a - NOD (n = 24): children stable on ultrasound and diuretic renogram and Group 2b - UPJO (n = 24): children who worsened to Grade 4 hydronephrosis on ultrasound/worsening of differential renal function (10% drop) on renogram who underwent pyeloplasty. Urinary biomarkers NGAL, KIM-1, and CA 19-9 were measured using the enzyme-linked immune absorbent assay method. Results: The urine CA 19-9 level was 128.05 ± 4.08 U/mL in the UPJO group, and this was significantly higher (P = 0.001) than NOD, 70.29 ± 4.41, and controls, 1.91 ± 1.57. The urine NGAL level was 21.41 ± 4.44 pg/mL in UPJO, and this was significantly higher than controls, 2.669 ± 0.513, but not NOD, 24.55 ± 2.67. The urine KIM level was 817 ± 15.84 pg/mL in the UPJO group, and this was significantly higher than controls, 285 ± 8.10, but not NOD, 768.23 ± 15.12. Receiver operating characteristic analysis of CA 19-9 revealed a urine biomarker cutoff of 95 U/mL for diagnosing UPJO (sensitivity 95%; specificity 96%; and area under the curve 0.99). Conclusions: CA 19-9 is a superior marker compared to NGAL and KIM in differentiating UPJO from NOD. Further studies with larger numbers are warranted.

Enhancing breast cancer screening with urinary biomarkers and Random Forest supervised classification: A comprehensive investigation.

OBJECTIVES: Urinary sex hormones are investigated as potential biomarkers for the early detection of breast cancer, aiming to evaluate their relevance and applicability, in combination with supervised machine-learning data analysis, toward the ultimate goal of extensive screening. METHODS: Sex hormones were determined on urine samples collected from 250 post-menopausal women (65 healthy - 185 with breast cancer, recruited among the clinical patients of Candiolo Cancer Institute FPO-IRCCS (Torino, Italy). Two analytical procedures based on UHPLC-MS/HRMS were developed and comprehensively validated to quantify 20 free and conjugated sex hormones from urine samples. The quantitative data were processed by seven machine learning algorithms. The efficiency of the resulting models was compared. RESULTS: Among the tested models aimed to relate urinary estrogen and androgen levels and the occurrence of breast cancer, Random Forest (RF) proved to underscore all the other supervised classification approaches, including Partial Least Squares - Discriminant Analysis (PLS-DA), in terms of effectiveness and robustness. The final optimized model built on only five biomarkers (testosterone-sulphate, alpha-estradiol, 4-methoxyestradiol, DHEA-sulphate, and epitestosterone-sulphate) achieved an approximate 98% diagnostic accuracy on replicated validation sets. To balance the less-represented population of healthy women, a Synthetic Minority Oversampling TEchnique (SMOTE) data oversampling approach was applied. CONCLUSIONS: By means of tunable hyperparameters optimization, the RF algorithm showed great potential for early breast cancer detection, as it provides clear biomarkers ranking and their relative efficiency, allowing to ground the final diagnostic model on a restricted selection five steroid biomarkers only, as desirable for noninvasive tests with wide screening purposes.

Influential factors on urine EV DNA methylation detection and its diagnostic potential in prostate cancer.

The value of Extracellular vesicles (EVs) diagnostic markers is widely recognized. However, current research on EV DNA remains limited. This study investigates the biological properties, preprocessing factors, and diagnostic potential of EV DNA. We found that DNA positive vesicles account for 23.3% ± 6.7% of the urine total EV, with a large amount of DNA attached to the outside. EV DNA fragments are large, there is no significant effect on uEV DNA when store urine less than 6 h at 4°C. In addition, the influence of different EV extraction methods on methylation detection is also minor. More importantly, RASSF1A methylation in urine total EV DNA can distinguish between PCa and BPH, with an AUC of 0.874. Our results suggest the potential of urine EV DNA as a novel marker for PCa diagnosis. This provides a new idea for the study of urinary tumor markers.

Hypermethylation Loci of ZNF671, IRF8, and OTX1 as Potential Urine-Based Predictive Biomarkers for Bladder Cancer.

Bladder cancer (BCa) is a significant health issue and poses a healthcare burden on patients, highlighting the importance of an effective detection method. Here, we developed a urine DNA methylation diagnostic panel for distinguishing between BCa and non-BCa. In the discovery stage, an analysis of the TCGA database was conducted to identify BCa-specific DNA hypermethylation markers. In the validation phase, DNA methylation levels of urine samples were measured with real-time quantitative methylation-specific PCR (qMSP). Comparative analysis of the methylation levels between BCa and non-BCa, along with the receiver operating characteristic (ROC) analyses with machine learning algorithms (logistic regression and decision tree methods) were conducted to develop practical diagnostic panels. The performance evaluation of the panel shows that the individual biomarkers of ZNF671, OTX1, and IRF8 achieved AUCs of 0.86, 0.82, and 0.81, respectively, while the combined yielded an AUC of 0.91. The diagnostic panel using the decision tree algorithm attained an accuracy, sensitivity, and specificity of 82.6%, 75.0%, and 90.9%, respectively. Our results show that the urine-based DNA methylation diagnostic panel provides a sensitive and specific method for detecting and stratifying BCa, showing promise as a standard test that could enhance the diagnosis and prognosis of BCa in clinical settings.

Metabolic profiling in tissues and urine of patients with prostatic lesions and the diagnostic value of urine extracellular vesicles metabolites in prostate cancer.

BACKGROUND: Prostate cancer (PCa) lacks convenient and highly specific diagnostic markers. Although the value of extracellular vesicles (EV) in oncology is widely recognized, the diagnostic value of EV metabolites requires further exploration. This study aimed to explore the diagnostic value of urine EV (u-EV) metabolomics in PCa. METHODS: We first detected metabolites in paired tissues cells (cells), tissue EV (t-EVs), u-EVs, and urine samples in cohort 1 (8 PCa vs. 5 benign prostatic hypertrophy, BPH) to prob the feasibility of EV metabolites as diagnostic markers. We then analyzed the value of u-EVs as markers for PCa diagnosis and typing in the expanded sample cohort (60 PCa vs. 40 BPH). RESULTS: U-EV metabolites were more consistent with those in tissue-derived samples (cells and t-EVs) than those in urine, and more differential metabolites between BPH and PCa were identified in u-EV. Subsequently, we used a random forest model to construct a panel of six metabolites for PCa, which showed an area under the curve (AUC) of 0.833 in training cohort and 0.844 in validation cohort. We also found significantly differentially expressed metabolites between PCa subtypes (Gleason ≤ 7 vs. Gleason > 7 and localized vs. metastasis), demonstrating the value of EV metabolites in PCa typing and prognostic assessment. CONCLUSION: Metabolomic analysis of u-EVs is a promising source of noninvasive markers for PCa diagnosis.

Prenatal antibiotics exposure and preschoolers' internalizing and externalizing problems: A biomonitoring-based prospective birth cohort study.

BACKGROUND: Biomonitoring-based epidemiological studies on prenatal antibiotic exposure and behavioral problems in preschoolers are lacking. The present study aimed to investigate the relationship between prenatal antibiotic exposure and internalizing and externalizing problems in preschoolers. METHODS: Data from 2449 mother-child pairs were analyzed. Urine samples were repeatedly collected across three trimesters, and 43 antibiotics and 2 metabolites were measured, including preferred as veterinary antibiotics (PVAs), VAs, preferred as human antibiotics and human antibiotics. Preschoolers' internalizing and externalizing problems were evaluated by the Achenbach Child Behavior Checklist. Poisson regression models with generalized estimating equations were used to estimate risk ratios (RRs) and 95 % confidence intervals (CIs) for preschoolers' internalizing, externalizing and total problems across tertiles of antibiotic concentrations during three periods of pregnancy, and performed several subgroup analyses. RESULTS: First-trimester urinary oxytetracycline (RR = 1.69, 95%CI: 1.20, 2.39, P-FDR = 0.011), tetracycline (RR = 1.91, 95%CI: 1.36, 2.68, P-FDR < 0.001), doxycycline (RR = 1.66, 95%CI: 1.28, 2.17, P-FDR < 0.001) and PVAs (RR = 1.79, 95%CI: 1.29, 2.48, P-FDR < 0.001) concentrations in the highest tertile were related to an elevated risk of internalizing problems compared with concentrations in the lowest tertile. First-trimester urinary doxycycline concentrations in the third tertile were also associated with an increased risk of externalizing problems compared with the first tertile (RR = 2.00, 95%CI: 1.28, 3.15, P-FDR = 0.042). Compared with concentrations in the lowest tertile, first-trimester urinary doxycycline (RR = 1.63, 95%CI: 1.19, 2.22, P-FDR = 0.028) and PVAs (RR = 1.67, 95%CI: 1.14, 2.43, P-FDR = 0.047) concentrations in the middle tertile were related to an increased risk of total problems. Furthermore, the type of main caregiver and children's outdoor activities time modified the relationships between specific prenatal antibiotic exposure and preschoolers' behavioral problems. CONCLUSIONS: Exposure to specific antibiotics during the first trimester may be related to an increased risk of internalizing and externalizing problems in preschoolers.

Relative validity of habitual sugar and low/no-calorie sweetener consumption assessed by food frequency questionnaire, multiple 24-h dietary recalls and urinary biomarkers: an observational study within the SWEET project.

BACKGROUND: Studies investigating associations between sweeteners and health yield inconsistent results, possibly due to subjective self-report dietary assessment methods. OBJECTIVES: We compared the performance of a food frequency questionnaire (FFQ), multiple 24-h dietary recalls (24hRs), and urinary biomarkers to estimate intake of sugars and low/no-calorie sweeteners (LNCSs). METHODS: Participants (n = 848, age 54 ± 12 y) from a 2-y observational study completed 1 semiquantitative FFQ and ≥ 3 nonconsecutive 24hRs. Both methods assessed intake of sugars (mono- and disaccharides, sucrose, fructose, free and added sugars) and sweetened foods and beverages (sugary foods, fruit juice, and sugar or LNCS-containing beverages [sugar-sweetened beverages and low/no-calorie sweetened beverages (LNCSBs)]); 24hRs also included LNCS-containing foods and tabletop sweeteners (low/no-calorie sweetened foods [LNCSFs]). Urinary excretion of sugars (fructose+sucrose) and LNCSs (acesulfame K+sucralose+steviol glucuronide+cyclamate+saccharin) were simultaneously assessed using ultrapressure liquid chromatography coupled to tandem mass spectrometry in 288 participants with 3 annual 24-h urine samples. Methods were compared using, amongst others, validity coefficients (correlations corrected for measurement error). RESULTS: Median (interquartile range) FFQ intakes ranged from 0 (0-7) g/d for LNCSBs to 94 (73-117) g/d for mono- and disaccharides. LNCSB use was reported by 32% of participants. Median LNCSB+LNCSF intake using 24hRs was 1 (0-50) g/d and reported by 58%. Total sugar excretions were detected in 100% of samples [56 (37-85) mg/d] and LNCSs in 99% of urine samples [3 (1-10) mg/d]. Comparing FFQ against 24hRs showed VCs ranging from 0.38 (fruit juice) to 0.74 (LNCSB). VCs for comparing FFQ with urinary excretions were 0.25 to 0.29 for sugars and 0.39 for LNCSBs; for 24hR they amounted to 0.31-0.38 for sugars, 0.37 for LNCSBs, and 0.45 for LNCSFs. CONCLUSIONS: The validity of the FFQ against 24hRs for the assessment of sugars and LNCSBs ranged from moderate to good. Comparing self-reports and urine excretions showed moderate agreement but highlighted an important underestimation of LNCS exposure using self-reports.

Comparison Shows that Multiplex Polymerase Chain Reaction Identifies Infection-associated Urinary Biomarker-positive Urinary Tract Infections That Are Missed by Standard Urine Culture.

Background: Multiplex polymerase chain reaction (M-PCR) has increased sensitivity for microbial detection compared with standard urine culture (SUC) in cases diagnosed as urinary tract infections (UTIs), leading to questions whether detected microbes are likely causative of UTIs or are incidental findings. Objective: To compare infection-associated biomarker levels against M-PCR and SUC results in symptomatic cases with a presumptive diagnosis of a UTI by a urologist. Design setting and participants: Participants were ≥60 yr old and presented to urology clinics between January and April 2023 with symptoms of UTIs (n = 583). Urine microbial detection was by M-PCR and SUC. Three infection-associated biomarkers (neutrophil gelatinase-associated lipocalin, interleukin-8, and interleukin-1ß) were measured by enzyme-linked immunosorbent assay. Symptomatic cases with elevated biomarkers, detection of uropathogens, and a specialist clinical diagnosis of a UTI were considered definitive UTI cases. Outcome measurements and statistical analysis: Distributions were compared using two-sample Wilcoxon rank sum test, with two-tailed p values of <0.05 considered statistically significant. Results and limitations: In cases with M-PCR-positive/SUC-negative results (n = 80), all median biomarker levels were significantly higher (p < 0.0001) than in cases with M-PCR-negative/SUC-negative results (n = 107). Two or more biomarkers were positive in 76% of M-PCR-positive/SUC-negative specimens. Limitation was an inability to examine associations between each individual organism and inflammation. Conclusions: A significant number of M-PCR-positive/SUC-negative cases had elevated levels of infection-related urinary biomarkers, especially when infection was caused by organisms other than Escherichia coli. This is a strong indication that microbes detected by M-PCR, which would be missed by SUC, are associated with UTIs. Patient summary: We compared infection-associated biomarkers in patients diagnosed with urinary tract infections (UTIs) against the detection of microorganisms by standard urine culture (SUC) and multiplex polymerase chain reaction (M-PCR). We found that most patients with microorganisms detected by M-PCR, which were missed by SUC, had elevated markers of inflammation, indicating that these organisms were likely causative of UTIs.

In Vivo Assessment of Laboratory-Grown Kidney Tissue Grafts.

Directed differentiation of stem cells is an attractive approach to generate kidney tissue for regenerative therapies. Currently, the most informative platform to test the regenerative potential of this tissue is engraftment into kidneys of immunocompromised rodents. Stem cell-derived kidney tissue is vascularized following engraftment, but the connection between epithelial tubules that is critical for urine to pass from the graft to the host collecting system has not yet been demonstrated. We show that one significant obstacle to tubule fusion is the accumulation of fibrillar collagens at the interface between the graft and the host. As a screening strategy to identify factors that can prevent this collagen accumulation, we propose encapsulating laboratory-grown kidney tissue in fibrin hydrogels supplemented with candidate compounds such as recombinant proteins, small molecules, feeder cells, and gene therapy vectors to condition the local graft environment. We demonstrate that the AAV-DJ serotype is an efficient gene therapy vector for the subcapsular region and that it is specific for interstitial cells in this compartment. In addition to the histological evaluation of epithelial tubule fusion, we demonstrate the specificity of two urine biomarker assays that can be used to detect human-specific markers of the proximal nephron (CD59) and the distal nephron (uromodulin), and we demonstrate the deposition of human graft-derived urine into the mouse collecting system. Using the testing platform described in this report, it will be possible to systematically screen factors for their potential to promote epithelial fusion of graft and host tissue with a functional intravital read-out.

Elevated UTI Biomarkers in Symptomatic Patients with Urine Microbial Densities of 10,000 CFU/mL Indicate a Lower Threshold for Diagnosing UTIs.

The literature lacks consensus on the minimum microbial density required for diagnosing urinary tract infections (UTIs). This study categorized the microbial densities of urine specimens from symptomatic UTI patients aged ≥ 60 years and correlated them with detected levels of the immune response biomarkers neutrophil gelatinase-associated lipocalin (NGAL), interleukin-8 (IL-8), and interleukin-1-beta (IL-1ß). The objective was to identify the microbial densities associated with significant elevation of these biomarkers in order to determine an optimal threshold for diagnosing symptomatic UTIs. Biobanked midstream voided urine samples were analyzed for microbial identification and quantification using standard urine culture (SUC) and multiplex-polymerase chain reaction (M-PCR) testing, while NGAL, IL-8, and IL-1ß levels were measured via enzyme-linked immunosorbent assay (ELISA). NGAL, IL-8, and IL-1ß levels were all significantly elevated at microbial densities ≥ 10,000 cells/mL when measured via M-PCR (p < 0.0069) or equivalent colony-forming units (CFUs)/mL via SUC (p < 0.0104) compared to samples with no detectable microbes. With both PCR and SUC, a consensus of two or more elevated biomarkers correlated well with microbial densities > 10,000 cells/mL or CFU/mL, respectively. The association between ≥10,000 cells and CFU per mL with elevated biomarkers in symptomatic patients suggests that this lower threshold may be more suitable than 100,000 CFU/mL for diagnosing UTIs.

Urinary Proteomics for Discovery of Gastric Cancer Biomarkers to Enable Precision Clinical Oncology.

For precision in clinical oncology practice, detection of tumor-derived peptides and proteins in urine offers an attractive and noninvasive alternative for diagnostic or screening purposes. In this study, we report comparative quantitative proteomic profiling of urine samples from patients with gastric cancer and healthy controls using tandem mass tags-based multiplexed mass spectrometry approach. We identified 1504 proteins, of which 246 were differentially expressed in gastric cancer cases. Notably, ephrin A1 (EFNA1), pepsinogen A3 (PGA3), sortilin 1 (SORT1), and vitronectin (VTN) were among the upregulated proteins, which are known to play crucial roles in the progression of gastric cancer. We also found other overexpressed proteins, including shisa family member 5 (SHISA5), mucin like 1 (MUCL1), and leukocyte cell derived chemotaxin 2 (LECT2), which had not previously been linked to gastric cancer. Using a novel approach for targeted proteomics, SureQuant, we validated changes in abundance of a subset of proteins discovered in this study. We confirmed the overexpression of vitronectin and sortilin 1 in an independent set of urine samples. Altogether, this study provides molecular candidates for biomarker development in gastric cancer, and the findings also support the promise of urinary proteomics for noninvasive diagnostics and personalized/precision medicine in the oncology clinic.

Quantitative, Targeted Analysis of Gut Microbiota Derived Metabolites Provides Novel Biomarkers of Early Diabetic Kidney Disease in Type 2 Diabetes Mellitus Patients.

Diabetic kidney disease (DKD) is one of the most debilitating complications of type 2 diabetes mellitus (T2DM), as it progresses silently to end-stage renal disease (ESRD). The discovery of novel biomarkers of early DKD becomes acute, as its incidence is reaching catastrophic proportions. Our study aimed to quantify previously identified metabolites from serum and urine through untargeted ultra-high-performance liquid chromatography coupled with electrospray ionization-quadrupole-time of flight-mass spectrometry (UHPLC-QTOF-ESI+-MS) techniques, such as the following: arginine, dimethylarginine, hippuric acid, indoxyl sulfate, p-cresyl sulfate, L-acetylcarnitine, butenoylcarnitine and sorbitol. The study concept was based on the targeted analysis of selected metabolites, using the serum and urine of 20 healthy subjects and 90 T2DM patients with DKD in different stages (normoalbuminuria-uACR < 30 mg/g; microalbuminuria-uACR 30-300 mg/g; macroalbuminuria-uACR > 300 mg/g). The quantitative evaluation of metabolites was performed with pure standards, followed by the validation methods such as the limit of detection (LOD) and the limit of quantification (LOQ). The following metabolites from this study resulted as possible biomarkers of early DKD: in serum-arginine, dimethylarginine, hippuric acid, indoxyl sulfate, butenoylcarnitine and sorbitol and in urine-p-cresyl sulfate.

Conditional Reprogramming Modeling of Bladder Cancer for Clinical Translation.

The use of advanced preclinical models has become increasingly important in drug development. This is particularly relevant in bladder cancer, where the global burden of disease is quite high based on prevalence and a relatively high rate of lethality. Predictive tools to select patients who will be responsive to invasive or morbid therapies (chemotherapy, radiotherapy, immunotherapy, and/or surgery) are largely absent. Patient-derived and clinically relevant models including patient-derived xenografts (PDX), organoids, and conditional reprogramming (CR) of cell cultures efficiently generate numerous models and are being used in both basic and translational cancer biology. These CR cells (CRCs) can be reprogrammed to maintain a highly proliferative state and reproduce the genomic and histological characteristics of the parental tissue. Therefore, CR technology may be a clinically relevant model to test and predict drug sensitivity, conduct gene profile analysis and xenograft research, and undertake personalized medicine. This review discusses studies that have utilized CR technology to conduct bladder cancer research.

Urinary TNF-α as a potential biomarker for chronic primary low back pain.

Introduction: Over two thirds of individuals with low back pain (LBP) may experience recurrent or persistent symptoms in the long term. Yet, current data do not allow to predict who will develop chronic low back pain and who will recover from an acute episode. Elevated serum levels of the proinflammatory cytokine tumor necrosis factor-α (TNF-α) have been associated with poor recovery and persistent pain following an acute episode of LBP. Inflammatory cytokines may also mediate mechanisms involved in nociplastic pain, and thus, have significant implications in chronic primary low back pain (CPLBP). Methods: This study aimed to investigate the potential of urinary TNF-α levels for predicting outcomes and characterizing clinical features of CPLBP patients. Twenty-four patients with CPLBP and 24 sex- and age-matched asymptomatic controls were recruited. Urinary TNF-α concentrations were measured at baseline and after 4 weeks, during which CPLBP patients underwent spinal manipulative therapy (SMT). Results: Concentrations of TNF-α were found to be elevated in baseline urine samples of CPLBP patients compared to asymptomatic controls. Moreover, these values differed among patients depending on their pain trajectory. Patients with persistent pain showed higher levels of TNF-α, when compared to those with episodic CPLBP. Furthermore, baseline TNF-α concentrations and their changes after 4 weeks predicted alterations in pain intensity and disability following SMT in patients with CPLBP. Discussion: These findings warrant further research on the potential use of urinary TNF-α concentrations as a prognostic biomarker for CPLBP.