Urinary Biomarker Levels Research Articles

Trends of Exposure to Acrylamide as Measured by Urinary Biomarkers Levels within the HBM4EU Biomonitoring Aligned Studies (2000-2021).

Acrylamide, a substance potentially carcinogenic in humans, represents a very prevalent contaminant in food and is also contained in tobacco smoke. Occupational exposure to higher concentrations of acrylamide was shown to induce neurotoxicity in humans. To minimize related risks for public health, it is vital to obtain data on the actual level of exposure in differently affected segments of the population. To achieve this aim, acrylamide has been added to the list of substances of concern to be investigated in the HBM4EU project, a European initiative to obtain biomonitoring data for a number of pollutants highly relevant for public health. This report summarizes the results obtained for acrylamide, with a focus on time-trends and recent exposure levels, obtained by HBM4EU as well as by associated studies in a total of seven European countries. Mean biomarker levels were compared by sampling year and time-trends were analyzed using linear regression models and an adequate statistical test. An increasing trend of acrylamide biomarker concentrations was found in children for the years 2014–2017, while in adults an overall increase in exposure was found to be not significant for the time period of observation (2000–2021). For smokers, represented by two studies and sampling for, over a total three years, no clear tendency was observed. In conclusion, samples from European countries indicate that average acrylamide exposure still exceeds suggested benchmark levels and may be of specific concern in children. More research is required to confirm trends of declining values observed in most recent years.

Parabens and Menopause-Related Health Outcomes in Midlife Women: A Pilot Study.

Background: Parabens are antimicrobial agents prevalently found in daily-use products that can interfere with the endocrine and reproductive systems. In this study, we examined the cross-sectional associations of parabens with hot flashes, hormone concentrations, and ovarian volume in a subsample of 101 nonsmoking, non-Hispanic 45- to 54-year-old women from the Midlife Women's Health Study. Materials and Methods: Women self-reported their hot flash history and underwent a transvaginal ultrasound to measure ovarian volume. Participants provided blood for quantification of serum hormones (by enzyme-linked immunosorbent assay or radioimmunoassay) and urine samples for measurements of urinary paraben biomarker levels (by high-performance liquid chromatography negative-ion electrospray ionization-tandem mass spectrometry). Linear or logistic regression models evaluated associations of specific gravity-adjusted paraben biomarker concentrations with hot flashes, hormone concentrations, and ovarian volume. Results: We observed marginal associations of propylparaben, methylparaben, and ∑parabens biomarkers (molar sum of four parabens) with hot flashes and follicle-stimulating hormone (FSH) concentrations, and of these paraben biomarkers and ethylparaben with ovarian volume. For example, women tended to have 32% (95% confidence intervals [CI]: 0.9 to 1.81), 40% (95% CI: 1.0 to 1.95), and 40% (95% CI: 0.98 to 2.01) higher odds of having recent, monthly, and mild hot flashes, respectively, for every two-fold increase in ∑parabens. Similarly, women tended to have 14.54% (95% CI: -0.10 to 31.32) higher FSH concentrations, but 5.67% (95% CI: -12.54 to 1.75) reduced ovarian volume for every two-fold increase in ∑parabens Conclusions: Overall, our preliminary findings suggest that urinary paraben biomarkers may be associated with menopause-related outcomes in midlife women. Additional studies in larger and diverse populations are needed to expand on these findings.

Clinical and subclinical acute kidney injury in children with mild-to-moderate COVID-19.

BackgroundOur aim was to identify acute kidney injury (AKI) and subacute kidney injury using both KDIGO criteria and urinary biomarkers in children with mild/moderate COVID-19.MethodsThis cross-sectional study included 71 children who were hospitalized with a diagnosis of COVID-19 from 3 centers in Istanbul and 75 healthy children. We used a combination of functional (serum creatinine) and damage (NGAL, KIM-1, and IL-18) markers for the definition of AKI and subclinical AKI. Clinical and laboratory features were evaluated as predictors of AKI and subclinical AKI.ResultsPatients had significantly higher levels of urinary biomarkers and urine albumin–creatinine ratio than healthy controls (p < 0.001). Twelve patients (16.9%) developed AKI based on KDIGO criteria, and 22 patients (31%) had subclinical AKI. AKI group had significantly higher values of neutrophil count on admission than both subclinical AKI and non-AKI groups (p < 0.05 for all). Neutrophil count was independently associated with the presence of AKI (p = 0.014).ConclusionsThis study reveals that even children with a mild or moderate disease course are at risk for AKI. Association between neutrophil count and AKI may point out the role of inflammation in the development of AKI.Impact The key message of our article is that not only children with severe disease but also children with mild or moderate disease have an increased risk for kidney injury due to COVID-19.Urinary biomarkers enable the diagnosis of a significant number of patients with subclinical AKI in patients without elevation in serum creatinine.Our findings reveal that patients with high neutrophil count may be more prone to develop AKI and should be followed up carefully.We conclude that even children with mild or moderate COVID-19 disease courses should be evaluated for AKI and subclinical AKI, which may improve patient outcomes.

Prospective study of biomarkers predictive of radiation-induced bladder toxicity in patients treated with radiotherapy for localized prostate cancer: RABBIO (Radiotoxicity Bladder BIOmarkers).

TPS12142 Background: Despite improvements in irradiation techniques, pelvic radiotherapy is responsible for acute and late adverse events in the bladder, defined as radiation cystitis (CysR). The early symptoms of bladder lesions secondary to pelvic irradiation are likely to occur during treatment or after radiotherapy in approximately 50% of irradiated patients. Acute radiation-related injuries are the first step of the fibrosis process. Fibrosis causes loss of bladder function and has a significant impact on the quality of life of the patients.The pathophysiology of CysR is not well understood, in particular because of the risks of complications caused by accessing the bladder tissue after irradiation, thereby limiting our ability to investigate this process and develop treatments. The main objective of our study is to assess the correlation of biologic biomarkers with the intensity of acute CysR and the quality of life of patients, evaluated with the digital telemonitoring platform Cureety. Methods: Patients with intermediate-risk localized prostate cancer and eligible for localized radiotherapy will be included. Inflammatory biomarkers will be analyzed on urine and blood samples before the initiation of radiotherapy, at week 4, 12 and 48 of irradiation, by quantitative methods such as the Multiplex Luminex assay, cytometry in flow and enzyme-linked immunosorbent assay ELISA. We will also characterize the gut and urinary microbiota in stool and urine samples using 16S rRNA sequencing technology. This is in order to assess the impact of the fecal and urinary microbiota in acute CysR. Between sample collection visits, patients will answer various questionnaires relating to the symptoms of radiation cystitis (IPSS), adverse events and quality of life (FACT-P), using the digital telemonitoring platform Cureety. Upon receipt of the questionnaires, an artificial intelligence algorithm will process the information and classify the patients according to the severity of symptoms and adverse reactions reported in accordance with CTCAE / IPSS. This will ultimately allow us to correlate urinary, blood and fecal biomarker levels with the severity of acute CysR symptoms and the quality of life reported by the patients. Conclusion: This prospective study is the first to explore the overexpression of inflammatory proteins in fluid biopsies from patients with symptoms of acute CysR. In addition, the 1-year post-treatment follow-up will allow us to predict which patients are at risk for late CysR and to stratify these patients towards radioprotective treatment. The results of this study will allow us to develop strategies to limit radiation damage to the bladder and improve the quality of life of patients. Clinical trial information: 2021A0319635.

Capillary Electrophoresis Applied to Human Urine Analysis for Clinical Diagnosis: New Trends and Perspectives

Capillary electrophoresis (CE) is an electromigration-based separation technique that emerged in the early 90s and has been evolving since then. Over recent decades, the association of CE with new technologies and detections strategies is enhancing its intrinsic mightiness. Today, CE is a part of the main high-throughput analytical platform used for medical-related applications, together with traditional gas chromatography, high-performance liquid chromatography, and fingerprinting spectroscopic methods. New strategies for assessing human disorders involve a thorough analysis of bodily fluids through said instrumentation. In this context, human urine is considered an excellent alternative, for being rich in information and easily collected with minimal inconvenience for the doners. Overall, since it is an ultrafiltrate of the human bloodstream, urine composition should be changed if any condition jeopardizes human homeostasis. Thus, monitoring the levels of biomarkers in urine by an advantageous technique such as CE can be an interesting choice for diagnostic and other clinical purposes. In this review, we will be commenting on the new tendencies and technologies applied to human urine analysis by CE over the last five years. We will be starting by commenting the applications onto target groups of molecules followed by addressing the CE feasibility for the determination of general chemical profiles through untargeted omics approaches, and finishing with the perspectives on the subject.

Use of Urinary Cytokine and Chemokine Levels for Identifying Bladder Conditions and Predicting Treatment Outcomes in Patients with Interstitial Cystitis/Bladder Pain Syndrome.

Background: Interstitial cystitis/bladder pain syndrome (IC/BPS) is a condition causing bladder inflammation. Urinary biomarkers have been assessed as suitable for the diagnosis and treatment. This study aimed at investigating the role of urinary biomarkers in identifying bladder conditions and predicting the treatment outcome of IC/BPS. Methods: A total of 309 patients with IC/BPS and 30 controls were enrolled in this study. All patients underwent a comprehensive urological workup of symptoms, pain severity, and cystoscopic hydrodistention findings including maximal bladder capacity (MBC) and glomerulation grade. Urine samples were collected to investigate the levels of urinary cytokines and chemokines. According to MBC and glomerulation grade, patients with IC/BPS were further classified into the Hunner’s IC (HIC) and non-HIC groups. The urinary biomarkers between IC/BPS and control groups and HIC and non-HIC groups were compared. Moreover, the treatment response was graded according to global response assessment (GRA) scores, and urinary biomarker levels were analyzed based on different GRAs. Results: Patients with IC/BPS had significantly high urinary monocyte chemoattractant protein-1, eotaxin, tumor necrosis factor -alpha (TNF-α), and prostaglandin E2 levels. Significantly higher levels of urinary interleukin-8, C-X-C motif chemokine ligand 10 (CXCL 10), brain-derived neurotrophic factor, eotaxin, and regulated-on-activation, normal T-cell expressed and secreted (RANTES) were noted in HIC than those with non-HIC and controls. Among all biomarkers, TNF-α had the best sensitivity, specificity, positive predictive value, and negative predictive value. There was a significant correlation between biomarker levels and GRA. Conclusions: Significantly higher urine cytokines and chemokine levels were found in patients with IC/BPS. Most urinary biomarkers were significantly associated with MBC, glomerulation grade, and treatment outcome.

Blood serum levels of selected biomarkers of oxidative stress among printing workers occupationally exposed to low-levels of toluene and xylene.

Printing workers (PWs) are exposed to a mixture of solvents, yet the health risks associated with such exposuer are unknown. This study aimed to compare the serum levels of selected biomarkers of oxidative stress among occupationally exposed PWs to low-level of toluene and xylenes and unexposed controls. Associations between levels of such biomarkers and occupational exposures to toluene and xylene were also investigated. Urinary levels of hippuric acid (HA) and methyl hippuric acids (MHAs) as exposure biomarkers of toluene and xylenes, respectively, and serum levels of oxidative stress biomarkers, including total antioxidant capacity (TAC), malondialdehyde (MDA), and the antioxidant enzymes superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPx), were measured among the 84 subjects, comprising 44 PWs and 40 unexposed subjects. Mean concentrations of urinary HA and MHAs of PWs showed a significant increase compared with the unexposed controls. Although levels of urinary biomarkers of exposure to toluene (HA) and xylenes (MHAs) were well below the biological exposure indices (BEIs; ACGHI), PWs presented significantly increased serum levels of MDA, and significantly decreased serum activities of SOD and GPx compared to the unexposed controls. However, for serum TAC and CAT activity, no significant difference was observed between the two groups. Correlation analyses indicated that urinary levels of HA and MHAs were positively correlated with MDA levels and negatively correlated with GPx and SOD. Our study suggested that the alterations evidenced in serum levels of MDA, SOD, and GPx could be involved in the oxidative stress caused by co-exposure to low levels of toluene and xylene. Further investigation is needed to clarify the effect of low-level occupational exposure to solvents among PWs.

Development and Validation of the Chinese Version Non-Nutritive Sweetener Food Frequency Questionnaire with Urinary Biomarker in Children and Adolescents.

The purpose of this study was to develop a validated food frequency questionnaire (FFQ) to evaluate the intake of non-nutritive sweeteners (NNSs) in child and adolescent Asian populations. Intensive and overall market research was performed to create the applicable NNS-FFQ with 13 food categories and 305 items. Six intense sweeteners, including acesulfame potassium, aspartame, sucralose, glycyrrhizin, steviol glycosides and sorbitol, were investigated. The validity and reproducibility of the NNS-FFQ were evaluated. The validity was further assessed by examining the consistency of reported NNS intake compared with urinary biomarkers using Cohen's κ analysis. This work was considered to be relevant in Asian societies. One hundred and two children and adolescents recruited from several clinics were invited to participate in this study. High content validity indices and high content validity ratio levels were revealed for each sweetener and food category. Reproducibility among subjects was satisfactory. Significant moderate correlations between estimated steviol glycoside/sucralose consumption and sensitive urinary biomarker levels were demonstrated (κ values were 0.59 and 0.45 for steviol glycosides and sucralose, respectively), indicating that the NNS-FFQ can be used to assess an individual's NNS intake. The dietary intense sweetener consumption pattern evaluated in this measurement was similar to those observed in other Asian countries but differed from those observed in Western populations with respect to types and amounts of NNSs. This validated NNS-FFQ can be an applicable and useful tool to evaluate NNS intake in future epidemiological and clinical studies.

Carcinogenic and tobacco smoke-derived particulate matter biomarker uptake and associated healthcare patterns among children.

The objective was to assess the associations of child tobacco smoke exposure (TSE) biomarkers (urinary cotinine, NNAL, and nicotelline N-oxides) and parent-reported smoking and child TSE patterns with total hospital visits, pediatric emergency department (PED) visits, urgent care (UC), revisits, and hospital admissions among 0-9-year-olds. A convenience sample of PED/UC patients (N = 242) who presented to a large, US children's hospital who had baseline urine samples assayed for the TSE biomarkers of interest were included. Biomarker levels were log-transformed, and linear and Poisson regression models were built. The geometric means of child cotinine, creatinine-adjusted NNAL, and N-oxide levels were 11.2 ng/ml, 30.9 pg/mg creatinine, and 24.1 pg/ml, respectively. The mean (SD) number of daily cigarettes smoked by parents was 10.2 (6.1) cigarettes. Each one-unit increase in log-NNAL levels was associated with an increase in total UC visits (aRR = 1.68, 95% CI = 1.18-2.39) among 0-9-year-olds, while controlling for the covariates. Each one-unit increase in child log-NNAL/cotinine ratio (×103) values was associated with an increase in total hospital visits (aRR = 1.39, 95% CI = 1.10-1.75) and UC visits (aRR = 1.56, 95% CI = 1.14-2.13) over 6 months. Systematic screening for child TSE should be conducted during all hospital visits. The comprehensive assessment of TSE biomarkers should be considered to objectively measure young children's exposure. Higher levels of cotinine, a widely used tobacco smoke exposure biomarker, have been associated with higher healthcare utilization patterns among children. Less is known on the associations of carcinogenic and tobacco smoke-derived particulate matter biomarker uptake with child healthcare utilization patterns. This study assessed the associations of several biomarkers with healthcare utilization patterns among pediatric emergency department patients ages 0-9 years who lived with tobacco smokers. Higher urinary NNAL biomarker levels, in individual and ratio form with cotinine, increased children's risk for urgent care visits over 6 months. Higher parent-reported cumulative child tobacco smoke exposure increased children's risk for hospital admissions.

Tubular and glomerular biomarkers of renal tissue function in the urine of fetuses with posterior urethral valves

Posterior urethral valve (PUV) is a congenital malformation characterized by a membranous structure located in the prostatic portion of the male posterior urethra that obstructs the urinary flow. Efforts have been made to determine the degree of impairment of fetal kidney function in this condition. This study aimed to measure the levels of urinary biomarkers of glomerular and tubular functions in fetuses with PUV and to compare with the levels of the same molecules in healthy male premature newborns. Urine samples from 43 fetuses with PUVs were collected and compared with urine samples from 40 healthy male newborns of the same gestational age (controls). Tubular and glomerular biomarkers levels were measured in urine samples by MILLIPLEX® assay kits. Levels of the molecules were related to creatinine (Cr) measurements at same urine samples and expressed as pg/mg Cr. Results were analysed with Graphpad Prism version 7.0 and SPSS version 20.0. Fetuses with PUV showed a significant reduction in urine levels of Epidermal Growth Factor (EGF), Calbindin, Osteoactivin, Molecule Renal Injury 1 (KIM-1 and Factor of trefoil 3 (TFF-3) when compared to controls. On the other hand, urine levels of cystatin C and renin were higher in PUV fetuses. The levels of molecules also differed according to urine osmolality and grade of hydronephrosis. Some urinary excreted molecules may indicate kidney damage in several segments along the nephron, while others may exert important functions. Mechanical and immunological mechanisms related to PUV might significantly modulate the synthesis of cytokines related to glomerular and tubular physiology, leading to alterations in urinary concentrations of those molecules. These biomarkers can be used as future diagnostic and prognostic markers in clinical practice. Early kidney structural and functional impairment influenced the synthesis of glomerular and tubular molecules related to kidney physiological processes in fetuses with PUV.

Variability in Urinary Nicotine Exposure Biomarker Levels Between Waves 1 (2013-2014) and 2 (2014-2015) in the Population Assessment of Tobacco and Health Study.

To date, no studies have evaluated the consistency of biomarker levels in people who smoke over a long-time period in real-world conditions with a large number of subjects and included use behavior and measures of nicotine metabolism. We evaluated the variability of biomarkers of nicotine exposure over approximately a 1-year period in people who exclusively smoke cigarettes, including intensity and recency of use and brand switching to assess impact on understanding associations with product characteristics. Multivariate regression analysis of longitudinal repeated measures of urinary biomarkers of nicotine exposure from 916 adults in the Population Assessment of Tobacco and Health (PATH) Study with demographic characteristics and use behavior variables. Intraclass correlation coefficients (ICCs) were calculated to examine individual variation of nicotine biomarkers and the uncertainty of repeat measures at two time points (Waves 1 and 2). Age, race, and urinary creatinine were significant covariates of urinary cotinine. When including use behavior, recency, and intensity of use were highly significant and variance decreased to a higher extent between than within subjects. The ICC for urinary cotinine decreased from 0.7530 with no use behavior variables in the model to 0.5763 when included. Similar results were found for total nicotine equivalents. Urinary nicotine biomarkers in the PATH Study showed good consistency between Waves 1 and 2. Use behavior measures such as time since last smoked a cigarette and number of cigarettes smoked in the past 30 days are important to include when assessing factors that may influence biomarker concentrations. The results of this study show that the consistency of the nicotine biomarkers cotinine and total nicotine equivalents in spot urine samples from Waves 1 to 2 of the PATH Study is high enough that these data are useful to evaluate the association of cigarette characteristics with biomarkers of exposure under real-world use conditions.

Exposure to polycyclic aromatic hydrocarbons and nicotine, and associations with sperm DNA fragmentation.

BackgroundTobacco smoking has been reported to cause DNA fragmentation and has been suggested to cause mutations in spermatozoa. These effects have been ascribed to the action of polycyclic aromatic hydrocarbons (PAH) present in the smoke. Simultaneously, DNA fragmentation has been associated with mutagenesis.ObjectiveThe aim of this study was to investigate whether levels of urinary biomarkers of PAH and nicotine exposure were associated with sperm DNA fragmentation.MethodsIn the urine of 381 men recruited from two cohorts of young men (17–21 years old) from the general Swedish population, the PAH metabolites 1‐hydroxypyrene and 2‐hydroxyphenanthrene, as well as the nicotine metabolite cotinine, were measured. The sperm DNA fragmentation index (DFI) was analysed using the sperm chromatin structure assay. Associations between the DFI, and PAH metabolite levels as continuous variables as well as in quartiles, were studied by general linear models adjusted for abstinence time. A similar analysis was carried out for cotinine levels, according to which the men were categorised as “non‐smoking” (n = 216) and “smoking” (n = 165).ResultsNo association was found between levels of any of the three biomarkers and DFI, either as a continuous variable (p = 0.87–0.99), or when comparing the lowest and the highest quartiles (p = 0.11–0.61). The same was true for comparison of men categorised as non‐smoking or smoking (DFI 11.1% vs. 11.8%, p = 0.31).DiscussionWe found no evidence of PAH or nicotine exposure to be associated with DFI, which does not exclude that these exposures may have other effects on sperm DNA.ConclusionIn these young men, levels of biomarkers of nicotine and PAH exposure were not associated with DFI.

Urinary miRNA Profiles in Chronic Kidney Injury-Benefits of Extracellular Vesicle Enrichment and miRNAs as Potential Biomarkers for Renal Fibrosis, Glomerular Injury, and Endothelial Dysfunction.

Micro-RNAs (miRNAs) are regulators of gene expression and play an important role in physiological homeostasis and disease. In biofluids, miRNAs can be found in protein complexes or in extracellular vesicles (EVs). Altered urinary miRNAs are reported as potential biomarkers for chronic kidney disease (CKD). In this context, we compared established urinary protein biomarkers for kidney injury with urinary miRNA profiles in obese ZSF1 and hypertensive renin transgenic rats. Additionally, the benefit of urinary EV enrichment was investigated in vivo and the potential association of urinary miRNAs with renal fibrosis in vitro. Kidney damage in both rat models was confirmed by histopathology, proteinuria, and increased levels of urinary protein biomarkers. In total, 290 miRNAs were elevated in obese ZSF1 rats compared with lean controls, whereas 38 miRNAs were altered in obese ZSF1 rats during 14-26 weeks of age. These 38 miRNAs correlated better with disease progression than established urinary protein biomarkers. MiRNAs increased in obese ZSF1 rats were associated with renal inflammation, fibrosis, and glomerular injury. Eight miRNAs were also changed in urinary EVs of renin transgenic rats, including one which might play a role in endothelial dysfunction. EV enrichment increased the number and detection level of several miRNAs implicated in renal fibrosis in vitro and in vivo. Our results show the benefit of EV enrichment for miRNA detection and the potential of total urine and urinary EV-associated miRNAs as biomarkers of altered kidney physiology, renal fibrosis and glomerular injury, and disease progression in hypertension and obesity-induced CKD.

P07 Toxicokinetic profile of VRP-034

BackgroundPolymyxin B (PMB) can be an effective option for treating resistant Gram-negative pathogens, however, its use is severely compromised by its’ dose-limiting toxicity. VRP-034, a novel formulation of PMB, has previously reported a favourable safety profile compared with marketed PMB (Roy et al.1). This study was conducted to understand the toxicokinetic profile of VRP-034 vis-à-vis marketed PMB.Materials and methods30 healthy Sprague-Dawley rats (divided into six groups of n = 5) received one of three doses (3, 9 and 18 mg/kg/day) of either marketed PMB or VRP-034, administered subcutaneously q8 h over 2 days. Blood samples were collected prior to initiation of experiment and at 0.5, 1, 2, 4, 6, 8, 12 and 24 h after the last dose. Plasma concentrations of PMB were quantified by LC-MS. Additionally, renal toxicity was assessed via clinical pathology, kidney injury biomarkers, oxidative stress markers (3-nitrotyrosine and renal tissue lactate), gross and histopathologic observations of kidney. Urine and blood samples were collected at baseline and Day 2 for the assessment of kidney injury (KIM-1, cystatin-C, BUN and serum creatinine) using respective biochemical or ELISA kits. All values are expressed as mean ± SEM.ResultsThe plasma levels of PMB were found to increase in dose-dependent manner in both the groups. At 3 and 9 mg/kg/day, the plasma PMB exposure were similar for both VRP-034 and marketed PMB groups (AUC0–24: 6.47 ± 0.41 μg/mL·h versus 5.66 ± 0.24 μg/mL·h, P>0.05 and 22.90 ± 1.96 μg/mL·h versus 26.84 ± 1.43 μg/mL·h, P>0.05). At the highest dose of 18 mg/kg/day, the AUC0–24 was higher in the VRP-034 group (AUC0–24: 97.83 ± 12.95 μg/mL·h versus 70.48 ± 4.22 μg/mL·h), however this increase was not statistically significant (P>0.05). The levels of early kidney injury urinary biomarkers increased with dose in both the groups (Table 1). On Day 2, Kim-1 levels in the marketed PMB group increased from baseline by 3.96-fold (versus 2.31-fold in VRP-034 group), 8.21-fold (versus 5.41-fold) and 23.41-fold (versus 9.51-fold) at 3, 9, 18 mg/kg/day dose levels respectively. Likewise, cystatin-C levels increased by 6.22-fold (versus 3.26-fold in VRP-034 group), 6.25-fold (versus 3.67-fold) and 13.57-fold (versus 4.33-fold), respectively, at the three dose levels tested. Oxidative stress markers, tissue lactate and 3-nitrotyrosine, were significantly lower (P<0.05) for VRP-034 group compared with marketed PMB group at the highest dose. Histopathology revealed EGTI grade 3 kidney damage in the marketed PMB group compared with grade 1 damage in the VRP-034 group.Table 1.Kidney injury and oxidative stress markers for marketed PMB and VRO-034 groups All values are reported as mean ± SEM.Lactate marker was only measured on Day 2.UCr, urine creatinine.ConclusionsVRP-034 exhibited a more promising toxicokinetic profile as compared with marketed PMB at all dose levels. Our future work will explore the clinical translation of these findings in humans.

Adaptive Mechanisms of Renal Bile Acid Transporters in a Rat Model of Carbon Tetrachloride-Induced Liver Cirrhosis.

Background: Acute kidney injury (AKI) is common in advanced liver cirrhosis, a consequence of reduced kidney perfusion due to splanchnic arterial vasodilation and intrarenal vasoconstriction. It clinically manifests as hepatorenal syndrome type 1, type 2, or as acute tubular necrosis. Beyond hemodynamic factors, an additional mechanism may be hypothesized to explain the renal dysfunction during liver cirrhosis. Recent evidence suggest that such mechanisms may be closely related to obstructive jaundice. Methods: Given the not completely elucidated role of bile acids in kidney tissue damage, this study developed a rat model of AKI with liver cirrhosis induction by carbon tetrachloride (CCl4) inhalation for 12 weeks. Histological analyses of renal and liver biopsies were performed at sacrifice. Organic anion tubular transporter distribution and apoptosis in kidney cells were analyzed by immunohistochemistry. Circulating and urinary markers of inflammation and tubular injury were assayed in 21 treated rats over time (1, 2, 4, 8, and 12 weeks of CCl4 administration) and 5 controls. Results: No renal histopathological alterations were found at sacrifice. Comparing treated rats with controls, organic anion transporters were differentially expressed and localized. High serum bile acid values were detected in cirrhotic animals, while caspase-3 staining was negative in both groups. Increased levels of serum inflammatory and urinary tubular injury biomarkers were observed during cirrhosis progression, with a peak after 4 and 8 weeks of treatment. Conclusions: These findings suggest possible adaptive tubular mechanisms for bile acid transporters in response to cirrhosis-induced AKI.

Preclinical Evaluation of the Renal Toxicity of Oligonucleotide Therapeutics in Mice.

Antisense oligonucleotides (ASO) therapeutics hold great promise for the treatment of numerous diseases, and several ASO drugs have now reached market approval, confirming the potential of this approach. However, some candidates have also failed, due to limited biodistribution/uptake and poor safety profile. In pursuit of better delivery and higher cellular uptake, ASO are being optimized, and new chemistries are developed or conjugated with various ligands. While these developments may lead to candidates with higher potency, it is important to keep the safety aspects in sight and screen for potential toxicity in early phases of preclinical development to avoid subsequent failure in clinical development. Our understanding of ASO-mediated toxicity keeps improving with increased preclinical and clinical data available. In this chapter, we will focus on the assessment of renal toxicity in mice and describe methods to measure the levels of general urinary biomarkers as well as acute kidney injury biomarkers following ASO treatment.

Differential Serum and Urine CRP, IP-10, and TRAIL Levels in Pediatric Urinary Tract Infection.

Background: It is estimated that clinical evaluation and urinalysis are unable to diagnose >10% of urinary tract infections (UTI) in young children. TNF-related apoptosis induced ligand (TRAIL), interferon gamma induced protein-10 (IP-10), and C-reactive protein (CRP) exhibit differential expression in the blood in response to bacterial vs. viral infection. We assessed if the urinary and serum levels of these host biomarkers discriminate UTI, nephronia, and response to antibiotic treatment.Methods: Hospitalized febrile children aged <18 years with suspected UTI based on abnormal urinalysis were recruited prospectively between 2016 and 2018; also, non-febrile controls were recruited. Following urine culture results and hospitalization course, participants were divided into three groups based on AAP criteria and expert adjudication: UTI, viral infection, and indeterminate.Results: Seventy-three children were enrolled, 61 with suspected UTI and 12 non-febrile controls. Of the 61 with suspected UTI, 40 were adjudicated as UTI, 10 viral infection, and 11 as indeterminate. Urinary CRP and IP-10 levels were significantly higher in the UTI group (p ≤ 0.05). Urinary CRP differentiated UTI from non-bacterial etiology in children under and over 3 months of age, with AUCs 0.98 (95% CI: 0.93–1.00) and 0.82 (0.68–0.95), respectively. Similarly, urinary IP-10 discriminated with AUCs of 0.80 (0.59–1.00) and 0.90 (0.80–1.00), respectively. Serum CRP and IP-10 levels were significantly higher in UTI cases with nephronia (p ≤ 0.03). UTI-induced changes in the levels of urinary and serum biomarkers resolved during recovery.Conclusions: CRP, IP-10, and TRAIL represent biomarkers with potential to aid the clinician in diagnosis and management of UTI.

Evaluation of lymphocyte-specific programmed cell death protein 1 receptor expression and cytokines in blood and urine in canine urothelial carcinoma patients.

Urothelial carcinoma (UC) is the most common urinary tumour in dogs. Despite a range of treatment options, prognosis remains poor in dogs. In people, breakthroughs with checkpoint inhibitors have established new standards of care for muscle-invasive bladder cancer patients and elevated levels of programmed cell death protein 1 (PD-1) suggest immune checkpoint blockade may be a novel target for therapy. The goal of this study was to determine if canine UC patients express elevated levels of lymphocyte-specific PD-1 and/or urinary cytokine biomarkers compared to healthy dogs. Paired blood and urine were evaluated in 10 canine UC patients, five cystitis patients and 10 control dogs for lymphocyte-specific PD-1 expression via flow cytometry and relative cytokine expression. In UC patients, PD-1 expression was significantly elevated on CD8+ lymphocytes in urine samples. UC patients had a higher CD4:CD8 ratio in their urine compared to healthy dogs, however, there was no significant variation in the CD8:Treg ratio between any group. Cystitis patients had significantly elevated levels of CD4+ T cells, CD8+ T cells and Tregs in their blood samples compared to UC patients and healthy dogs. Cytokine analysis demonstrated significant elevations in urinary cytokines (granulocyte-macrophage colony-stimulating factor, interferon-gamma [IFN-γ], interleukin (IL)-2, IL-6 IL-7, IL-8 and IL-15, IP-10, KC-like, IL-18, monocyte chemoattractant protein-1 and tumour necrosis factor-alpha). Several of these cytokines have been previously correlated with both lymphocyte-specific PD-1 expression (IFN-γ, IL-2, IL-7 and IL-15) in muscle-invasive urothelial carcinoma in humans. Our results provide evidence of urinary lymphocyte PD-1 expression and future studies could elucidate whether veterinary UC patients will respond favourably to anti-PD-1 immune checkpoint inhibitor therapy.

Study on the daily Ad Libitum smoking habits of active Korean smokers and their effect on urinary smoking exposure and impact biomarkers

Introduction Understanding interactions of smoking topography with biomarkers of exposure to tobacco is essential for accurate smoking risk assessments. Methods In this study, the smoking topography and the levels of tobacco smoke exposure urinary biomarkers of a sample of active Korean smokers were quantified and measured. The results were used to investigate the effect of daily activities and smoking time on the smoking topography. Moreover, correlations between the smoking topography parameters and biomarkers were assessed. Results No significant effect of either the daily activities or time on the smoking topography of the subjects were observed. Synchronic correlations of the cigarette consumption per day (CPD) and the average flow per puff with both urinary cotinine and trans-3’-hydroxycotinine were significant. For the urinary nicotine metabolites, the peak levels appeared when the CPD was over 19 cigarettes per day and the average puff velocity was between 35 and 45 ml/s. Nevertheless, when the average flow was over 60 ml/s, the levels of cotinine and trans-3’-hydroxycotinine significantly dropped. Conclusions The findings of this study may be beneficial for further smoking risk assessments with contributions of both the smoking topography and biomarkers to provide current smokers with applicable cession programs. Clinical significance Smoking habits and levels of urinary biomarkers of Korean smokers are investigated. People with a higher dependency on nicotine smoke cigarettes with slower puffs. Effects of daily activities or time on smoking topography were not significant. Correlations between smoking topography and urinary biomarkers were significant. Peak biomarker levels were observed under certain smoking topography conditions.

Sequential changes in urinary biomarker levels in patients with cirrhosis and severe hepatorenal syndrome.

Whether tubular injury develops in patients with acute kidney injury owing to hepatorenal syndrome (AKI-HRS) is controversial. We performed repeated measurements of biomarkers of tubular injury during a 14-day period in 60 patients with cirrhosis and AKI (34 with AKI-HRS meeting the classical definition of type 1 HRS and 26 with AKI owing to acute tubular necrosis, AKI-ATN). Nineteen of 34 patients had resolution of AKI-HRS, while the remainder had persistent AKI-HRS. The persistence of AKI-HRS was associated with remarkably high short-term mortality. There were no significant differences in urinary NGAL or IL-18 between patients with resolution vs those with persistent AKI-HRS throughout the 14-day period. By contrast, biomarker levels were significantly lower in AKI-HRS, even if persistent, compared to AKI-ATN. These findings are highly suggestive of lack of significant tubular injury in AKI-HRS and could be of value in the clinical decision between combined liver-kidney or liver transplantation alone in patients with cirrhosis and AKI candidates to transplantation.