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Antimicrobial peptides (AMPs) are critical to the protection of the urinary tract of humans and other animals from pathogenic microbial invasion. AMPs rapidly destroy pathogens by disrupting microbial membranes and/or augmenting or inhibiting the host immune system through a variety of signaling pathways. We have previously demonstrated that alpha-defensins 1-3 (

Background Mortality in severe malaria remains high in children treated with intravenous artesunate. Acute kidney injury (AKI) is a common complication of severe malaria, but the interactions between AKI and other complications on the risk of mortality in severe malaria are not well characterized. Methods Between 2014 and 2017, 600 children aged 6–48 months to 4 years hospitalized with severe malaria were enrolled in a prospective clinical cohort study evaluating clinical predictors of mortality in children with severe malaria. Results The mean age of children in this cohort was 2.1 years (standard deviation, 0.9 years) and 338 children (56.3%) were male. Mortality was 7.3%, and 52.3% of deaths occurred within 12 hours of admission. Coma, acidosis, impaired perfusion, AKI, elevated blood urea nitrogen (BUN), and hyperkalemia were associated with increased mortality (all P Conclusions AKI interacts with coma, acidosis, or impaired perfusion to significantly increase the risk of death in severe malaria. Among children with AKI, those who have hyperkalemia or elevated BUN have a higher risk of death. A better understanding of the causes of these complications of severe malaria, and development and implementation of measures to prevent and treat them, such as dialysis, are needed to reduce mortality in severe malaria.

Emergency departments (EDs) often rely on urinalysis (UA) to rapidly identify urinary tract infections (UTIs) in children. However, the suboptimal test characteristics of UA can lead to false-positive results. Novel urinary biomarkers may increase the diagnostic precision of UA. In this study, we compared the concentrations of 6 pre-selected proteins: BH3 interacting domain death agonist (BID), B-cell lymphoma 6 protein, ras GTPase-activating protein 1, cathepsin S (CTSS), 3-hydroxyanthranilate 3,4-dioxygenase, and transgelin-2. In a pediatric ED, we prospectively enrolled 167 children with UA and urine culture collected. Pyuria was defined as either ≥ 5 white blood cells per high-power field on microscopy or positive leukocyte esterase (LE). The urine culture was considered positive if it yielded ≥ 50,000 colony-forming units per milliliter of any single urinary pathogen. Urine protein levels were measured by enzyme-linked immunosorbent assay and normalized to urine creatinine. BID was significantly higher in the UTI group compared to the culture-negative pyuria group with a mean ratio of 1.42 (95% confidence interval (CI), 1.15, 1.76) when uncorrected for creatinine concentration. When corrected for creatinine concentration, CTSS was significantly elevated in the UTI group compared to the culture-negative pyuria group with a mean ratio of 2.11 (95% CI, 1.39, 3.21). BID and CTSS concentrations were elevated in the urine of children with UTI compared to those with culture-negative pyuria. These proteins deserve further research into their utility to serve as novel biomarkers for UTI.

Antimicrobial peptides (AMPs) are critical to the protection of the urinary tract of humans and other animals from pathogenic microbial invasion. AMPs rapidly destroy pathogens by disrupting microbial membranes and/or augmenting or inhibiting the host immune system through a variety of signaling pathways. We have previously demonstrated that alpha-defensins 1-3 ( DEFA1A3 ) are AMPs expressed in the epithelial cells of the human kidney collecting duct in response to uropathogens. We also demonstrated that DNA copy number variations in the DEFA1A3 locus are associated with UTI and pyelonephritis risk. Because DEFA1A3 is not expressed in mice, we utilized human DEFA1A3 gene transgenic mice ( DEFA 4/4 ) to further elucidate the biological relevance of this locus in the murine urinary tract. We demonstrate that the kidney transcriptional and translational expression pattern is similar in humans and the human gene transgenic mouse upon uropathogenic Escherichia coli (UPEC) stimulus in vitro and in vivo. We also demonstrate transgenic human DEFA 4/4 gene mice are protected from UTI and pyelonephritis under various UPEC challenges. This study serves as the foundation to start the exploration of manipulating the DEFA1A3 locus and alpha-defensins 1-3 expression as a potential therapeutic target for UTIs and other infectious diseases.

Hospital readmissions experienced by kidney transplant recipients may be secondary to a range of conditions, including infections and rejection episodes. The objective was to identify trends in patients with kidney transplant complications, in regard to hospital discharges, ED visits, and charges over the years available from 1993 to 2015. Using the Healthcare Cost and Utilization Project database, trends were identified in hospitalizations, emergency department visits, and charges from 1993 to 2015 for complications following kidney transplantation. Hospital discharges have significantly increased over time and at a faster rate than the increase in number of kidney transplants performed, while emergency department visits numbers and rates remain unchanged. The type of kidney transplant complications experienced were analyzed by incidence and proportion of total charges. Rejection made up the largest proportion of hospitalizations and of total cost in patients suffering from kidney transplant complications. Improved immunosuppression regimens have resulted in longer allograft survival. We speculate that the overlap between infection and rejection is compounding and contributing to graft injury and thus, it is important to try to prevent and/or properly identify those episodes as well in order to improve graft survival.

Thousands of chemical compounds produced by industry are dispersed in the human environment widely enough to reach the world population, and the introduction of new chemicals constantly occurs. As new synthetic molecules emerge, rapid analytical workflows for screening possible presence of exogenous compounds in biofluids can be useful as a first pass analysis to detect chemical exposure and guide the development and application of more elaborate LC-MS/MS methods for quantification. In this study, a suspect screening workflow using the multiple reaction monitoring (MRM) profiling method is proposed as a first pass exploratory technique to survey selected exogenous molecules in human urine samples. The workflow was applied to investigate 12 human urine samples using 310 MRMs related to the chemical functionalities of 87 exogenous compounds present in the METLIN database and reported in the literature. A total of 11 MRMs associated with five different compounds were detected in the samples. Product ion scans for the precursor ions of the selected MRMs were acquired as a further identification step for these chemicals. The suspect screening results suggested the presence of five exogenous compounds in the human urine samples analyzed, namely metformin, metoprolol, acetaminophen, paraxanthine and acrylamide. LC-MS/MS was applied as a last step to confirm these results, and the presence of four out of the five targets selected by MRM profiling were corroborated, indicating that this workflow can support the selection of suspect compounds to screen complex samples and guide more time-consuming and specific quantification analyses.

Urinary tract infections (UTI), associated with vesicoureteral reflux (VUR), can lead to chronic kidney disease. Genetic alterations in the innate immune defenses contribute to UTI risk. We investigated a novel gene, Dachsous Cadherin-Related 1 ( DCHS1), in children with UTI. We determined absolute DNA copy number (CN) of DCHS1 in children with UTI. In this case-control study, we utilized multiple complementary methods to determine the genomic CN of DCHS1. Children with ( n = 370) and without ( n = 71) VUR from two well-phenotyped clinical trials of UTI were copy-typed and compared to 491 healthy controls with no known history of VUR or UTI. Less than 1% of controls had a single copy of DCHS1, while 31% of children with UTI and no VUR and 7% of children with UTI and VUR had a single copy of the DCHS1 gene. Using immunostaining, we localized expression postnatally to the bladder and renal epithelia. Mice were also challenged with two uropathogenic Escherichia coli strains, and Dchs1 mRNA was quantified. This study represents the first report of DCHS1 in association with pediatric UTI. We hypothesize that its role in innate immunity is critical to lower urinary tract defense. Further investigation is required to determine the role of DCHS1 in innate immunity.

Background: The 2011 American Academy of Pediatrics clinical practice guideline recommends when to obtain renal and bladder ultrasound (RBUS) and voiding cystourethrography (VCUG) following febrile urinary tract infection (UTI) for children age 2–24 months. However, there is not consensus about when to obtain imaging in neonates. The objective of this study is to evaluate UTI diagnostic criteria along with RBUS and VCUG in neonates admitted to the NICU in the first 3 months of life. Methods: A retrospective electronic medical record review was performed of neonates admitted to Nationwide Children's Hospital system NICUs between January 2010 and December 2014 with UTI as a primary or secondary diagnosis. Urine culture results were evaluated versus established UTI criteria and renal US and VCUG results were compared. Results: Of 81 patients with a straight catheterized urine culture obtained, 28 patients met laboratory criteria for diagnosis of UTI and all but 4 had a RBUS. Urine cultures had an equal distribution of Enterobacter cloacae, Escherichia coli, Klebsiella pneumoniae, and Coagulase negative staphylococcus. RBUS showed dilation of the collecting system in 37.5% of patients with UTI compared to 41.3% without UTI. VCUG showed vesicourteral reflux (VUR) on 41.7% of those with UTI compared to 34.8% without UTI. For patients with UTI, the sensitivity of RBUS for VUR on VCUG was 60% with CI [0.17, 0.93] and specificity was 43% with CI [0.12, 0.80]. In patients without UTI, sensitivity of RBUS for VUR on VCUG was 63% with CI [0.26, 0.90] and specificity was 71% with CI [0.42, 0.90]. Conclusions: Fewer than half of neonates that were diagnosed clinically with UTI met laboratory criteria for a UTI. Positive urine cultures grew a wide variety of organisms. The sensitivity of renal ultrasound for VUR is only about 60%. Key Words: cystitis, cystourethrogram, ultrasound, urinalysis

Acute pyelonephritis is frequently associated with metabolic acidosis. We previously reported that metabolic acidosis stimulates expression of hypoxia-inducible factor (HIF)-1α-induced target genes such as stromal derived factor-1 and cathelicidin, an antimicrobial peptide. Since the collecting duct (CD) plays a pivotal role in regulating acid-base homeostasis and is the first nephron segment encountered by an ascending microbial infection, we examined the contribution of HIF-1α to innate immune responses elicited by acid loading of an M-1 immortalized mouse CD cell line. Acid loading of confluent M-1 cells was achieved by culture in pH 6.8 medium supplemented with 5-( N-ethyl- N-isopropyl)-amiloride to block Na+/H+ exchange activity for 24 h. Acid loading induced antimicrobial peptide [cathelicidin and β-defensin (Defb2 and Defb26)] mRNA expression and M-1 cell resistance to uropathogenic Escherichia coli infection to an extent similar to that obtained by inhibition of HIF prolyl hydroxylases, which promote HIF-1α protein degradation. The effect of acid loading on M-1 cell resistance to uropathogenic E. coli infection was reduced by inhibition of HIF-1α (PX-478), and, in combination with prolyl hydroxylase inhibitors, acidosis did not confer additional resistance. Thus, metabolic stress of acidosis triggers HIF-1α-dependent innate immune responses in CD (M-1) cells. Whether pharmacological stabilization of HIF prevents or ameliorates pyelonephritis in vivo warrants further investigation.

Ribonuclease 7 (RNase 7) is an antimicrobial peptide that prevents urinary tract infections (UTI); however, it is yet unknown how RNASE7 genetic variations affect its antimicrobial activity and its mitigation of UTI risk. This study determined whether the RNASE7 SNP rs1263872 is more prevalent in children with UTI and defined how rs1263872 affects RNase 7’s antimicrobial activity against uropathogenic E. coli (UPEC). We performed genotyping for rs1263872 in 2 national UTI cohorts, including children enrolled in the Randomized Intervention for Children with Vesicoureteral Reflux trial or the Careful Urinary Tract Infection Evaluation study. Genotypes from these cohorts were compared with those of female controls with no UTI. To assess whether rs1263872 affects RNase 7’s antimicrobial activity, we generated RNase 7 peptides and genetically modified urothelial cultures encoding wild-type RNase 7 and its variant. Compared with controls, girls in both UTI cohorts had an increased prevalence of the RNASE7 variant. Compared with the missense variant, wild-type RNase 7 peptide showed greater bactericidal activity against UPEC. Wild-type RNase 7 overexpression in human urothelial cultures reduced UPEC invasive infection compared with mutant overexpression. These results show that children with UTI have an increased prevalence of RNASE7 rs1263872, which may increase UTI susceptibility by suppressing RNase 7’s antibacterial activity.

UropathogenicWe studied the effects of UPEC infection using single cell RNA sequencing (scRNAseq) in zebrafish. Bulk RNA sequencing has historically been used to evaluate gene expression patterns, but scRNAseq allows gene expression to be evaluated at the single cell level and is optimal for evaluating heterogeneity within cell types and rare cell types. Zebrafish cohorts were injected with either saline or UPEC, and scRNAseq and canonical pathway analyses were performed.Canonical pathway analysis of scRNAseq data provided key information regarding innate immune pathways in the cells determined to be thymus cells, ionocytes, macrophages/monocytes, and pronephros cells. Pathways activated in thymus cells included interleukin 6 (IL-6) signaling and production of reactive oxygen species. Fc receptor-mediated phagocytosis was a leading canonical pathway in the pronephros and macrophages. Genes that were downregulated in UPEC vs saline exposed embryos involved the cellular response to the Gram-negative endotoxin lipopolysaccharide (LPS) and included Forkhead Box O1aBecause 4-day post fertilization zebrafish embryos only have innate immune systems, the scRNAseq provides insights into pathways and genes that cell types utilize in the bacterial response. Based on our analysis, we have identified genes and pathways that might serve as genetic targets for treatment and further investigation in UPEC infections at the single cell level.

Historically, the role of bacteria in urinary stone disease (USD) has been limited to urease-producing bacteria associated with struvite stone formation. However, growing evidence has revealed bacteria associated with stones of non-struvite composition. These bacteria may be derived from either urine or from the stones themselves. Using 16S rRNA gene sequencing and an enhanced culture technique (EQUC), we identified the urine and stone microbiota of USD patients and then determined if bacteria were statistically enriched in the stones relative to the urine. From 52 patients, bladder urine and urinary stones were collected intraoperatively during ureteroscopy. Stone homogenate and urine specimens were subjected to 16S rRNA gene sequencing and EQUC. Standard Chi-squared tests were applied to determine if the relative abundance of any bacterial taxon was significantly enriched in urinary stones compared to urine. Stones were primarily calcium-based. 29/52 (55.8%) stones had bacteria detected by 16S rRNA gene sequencing. Of these, dominant bacterial taxa were enriched from 12 stones. Bacterial taxa isolated by EQUC include members of the genera Staphylococcus, Enterobacter, Escherichia, Corynebacterium, and Lactobacillus. Dominant bacterial genera were enriched compared to paired bladder urine. Differences between the stone and urine microbiota may indicate that certain bacteria contribute to USD pathophysiology. Further investigation is warranted.

People with diabetes mellitus have increased infection risk. With diabetes, urinary tract infection (UTI) is more common and has worse outcomes. Here, we investigate how diabetes and insulin resistance impact the kidney’s innate defenses and urine sterility. We report that type 2 diabetic mice have increased UTI risk. Moreover, insulin-resistant prediabetic mice have increased UTI susceptibility, independent of hyperglycemia or glucosuria. To identify how insulin resistance affects renal antimicrobial defenses, we genetically deleted the insulin receptor in the kidney’s collecting tubules and intercalated cells. Intercalated cells, located within collecting tubules, contribute to epithelial defenses by acidifying the urine and secreting antimicrobial peptides (AMPs) into the urinary stream. Collecting duct and intercalated cell–specific insulin receptor deletion did not impact urine acidification, suppressed downstream insulin-mediated targets and AMP expression, and increased UTI susceptibility. Specifically, insulin receptor–mediated signaling regulates AMPs, including lipocalin 2 and ribonuclease 4, via phosphatidylinositol-3-kinase signaling. These data suggest that insulin signaling plays a critical role in renal antibacterial defenses.

The urinary tract is usually culture negative despite its close proximity to microbial flora. The precise mechanism by which the kidneys and urinary tract defends against infection is not well understood. The initial kidney cells to encounter ascending pathogens are the collecting tubule cells that consist of principal cells (PCs) that express aquaporin 2 (AQP2) and intercalated cells (ICs) that express vacuolar H+-ATPase (V-ATPase, B1 subunit). We have previously shown that ICs are involved with the human renal innate immune defense. Here we generated two reporter mice, VATPase B1-cre+tdT+mice to fluorescently label ICs and AQP2-cre+tdT+mice to fluorescently label PCs, and then performed flow sorting to enrich PCs and ICs for analysis. Isolated ICs and PCs along with proximal tubular cells were used to measure antimicrobial peptide (AMP) mRNA expression. ICs and PCs were significantly enriched for AMPs. Isolated ICs responded to uropathogenic Escherichia coli (UPEC) challenge in vitro and had higher RNase4 gene expression than control while both ICs and PCs responded to UPEC challenge in vivo by upregulating Defb1 mRNA expression. To our knowledge, this is the first report of isolating murine collecting tubule cells and performing targeted analysis for multiple classes of AMPs.

Background: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and its associated disease COVID-19 (coronavirus disease 2019) has presented graduate medical education (GME) training programs with a unique set of challenges. One of the most pressing is how should hospital systems that rely on graduate medical residents provide appropriate care for patients while protecting trainees. This question is of particular concern as healthcare workers are at high risk of SARS-CoV-2 exposure.Objective: This cross-sectional study sought to assess the impact of hospital COVID-19 patient placement on pediatric graduate medical residents by comparing rates of SARS-CoV-2 seroconversion rates of residents who worked on designated COVID-19 teams and those who did not.Methods: Forty-four pediatric and medicine–pediatric residents at Riley Children's Hospital (Indianapolis, IN) were tested for SARS-CoV-2 immunoglobulin M (IgM) and IgG seroconversion in May 2020 using enzyme-linked immunosorbent assays (Abnova catalog no. KA5826), 2 months after the first known COVID-19 case in Indiana. These residents were divided into two groups: those residents who worked on designated COVID-19 teams, and those who did not. Groups were compared using χ2 or Fisher exact test for categorical variables, and continuous variables were compared using Student t testing.Results: Forty-four of 104 eligible residents participated in this study. Despite high rates of seroconversion, there was no difference in the risk of SARS-CoV-2 seroconversion between residents who worked on designated COVID-19 teams (26% or 8/31) and those who did not (31% or 4/13). Eleven of 44 residents (25%) tested positive for SARS-CoV-2 IgG, whereas only 5/44 (11.4%) tested positive for SARS-CoV-2 IgM, without a detectable difference between exposure groups.Conclusion: We did not observe a difference in SARS-CoV-2 seroconversion between different exposure groups. These data are consistent with growing evidence supporting the efficacy of personal protective equipment. Further population-based research on the role of children in transmitting the SARS-CoV-2 virus is needed to allow for a more evidence-based approach toward managing the COVID-19 pandemic.

Background:Acute kidney injury (AKI) due to urinary stone disease (USD) is rare in adults; AKI rates in children with USD may be higher, and emerging data links stones to chronic kidney disease (CKD) development in adults.Methods:This study is a retrospective analysis of USD patients at a single pediatric hospital system's emergency department (ED). Patients were initially identified by USD ICD codes; USD was then confirmed by imaging or physician documentation; patients had to have baseline creatinine (Cr) and Cr in the ED for comparison to be included. AKI was defined by Kidney Disease: Improving Global Outcomes (KDIGO), Acute Kidney Injury Network (AKIN), and Pediatric Risk, Injury, Failure, Loss, End Stage (pRIFLE).Results:Of the 589 total visits, 264/589 (45%) had data to evaluate for AKI, 23% were AKI(+) and 77% were AKI(–). pRIFLE was most common (82%) and 18% were only positive by AKIN/KDIGO. AKI(+) were more likely to be younger (16.7 vs. 17.4 years,p= 0.046) and more likely to present with vomiting {odds ratio [OR] [95% confidence interval (CI)]: 2.4 [1.4–4.3],p= 0.002}; also, the proportion of AKI(+) was significantly higher in p= 0.032, OR (95% CI): 2.0 (1.1–3.9)]. Urinary tract infection (UTI) and obstruction rates were similar between groups. AKI(+) patients had a significant OR p= 0.001).Conclusion:We have demonstrated a novel association between USD-induced renal colic and AKI in a group of young adults and children. AKI(+) patients were younger and were more likely to present with vomiting. AKI(+) patients did not have higher rates of obstruction or UTI, and 51% of AKI(+) received NSAIDs.

BACKGROUND: The occurrence of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and the associated coronavirus disease 2019 (COVID-19) have profoundly affected adult kidney disease patients. In contrast, pediatric solid organ transplant recipients, including pediatric kidney transplant (KT) recipients, do not seem to be at particularly higher risk for SARS-CoV-2 infection or for severe COVID-19 disease. This patient population might be protected by certain mechanisms, such as the immunosuppressive medications with their anti-inflammatory properties or simply being well-versed in self-protection techniques. Assessing SARS-CoV-2 antibody serologies could potentially help understand why this patient population is apparently spared from severe SARS-CoV-2 clinical courses. OBJECTIVE: To examine SARS-CoV-2 serologic status in a cohort of pediatric KT recipients. METHODS: SARS-CoV-2 anti-spike IgG and IgM antibodies were measured by three different methods in pediatric KT recipients coming for routine clinic visits immediately post-confinement in May-June of 2020. The patients were considered seroconverted if SARS-CoV-2 antibodies were positive by 2/3 methods and weak positive/indeterminate if positive by 1/3. RESULTS: Thirty-one patients were evaluated (about 1/3 of our institution's pediatric KT population). One patient seroconverted, while three were considered weak positive/indeterminate. None were symptomatic and none had nasopharyngeal PCR confirmed SARS-CoV-2 disease. CONCLUSIONS: Seroconversion to SARS-CoV-2 was rare in this population and likely reflects the social distancing practiced by these patients. The results will serve as a foundation for a future longitudinal study to evaluate the long-term emergence and persistence of antibodies in this population and may inform studies of response to a future vaccine.

Dyspnea is the second leading cause of US emergency department (ED) visits and an independent predictor of morbidity and mortality1 in older adult patients aged ≥65 years. Unfortunately, the diagnosis of the cause of dyspnea presents diagnostic challenges to emergency physicians2-4 that disproportionately affects older adults.5 One in 5 dyspneic older adults experience missed diagnosis in the ED2 and 21% are treated for ≥1 pneumonia, acute exacerbation of chronic obstructive pulmonary disease [COPD], and acute exacerbation of heart failure [HF].5 Importantly, some may have multiple causes of their dyspnea but accurate diagnosis remains critical.

Coronavirus disease 2019 (COVID‐19) is a highly infectious disease caused by the severe acute respiratory syndrome coronavirus 2 virus (SARS‐CoV‐2). While children appear to experience less severe disease than adults, those with underlying conditions such as kidney disease may be more susceptible to infection. Limited data are present for children with kidney disease, and there are limited prior reports of pediatric hemodialysis patients with COVID‐19. This report describes the mild clinical disease course of COVID‐19 in two pediatric patients with chronic kidney disease, one on hemodialysis and both on chronic immunosuppression. We review treatment in these patients, as well as our measures to reduce transmission among our hemodialysis patients and staff.

Urinary tract infections (UTI), associated with vesicoureteral reflux (VUR), can lead to chronic kidney disease. Genetic alterations in the innate immune defenses contribute to UTI risk. We investigated a novel gene, Dachsous Cadherin-Related 1 ( DCHS1 ), in children with UTI. We determined absolute DNA copy number (CN) of DCHS1 in children with UTI. In this case-control study, we utilized multiple complementary methods to determine the genomic CN of DCHS1 . Children with ( n = 370) and without ( n = 71) VUR from two well-phenotyped clinical trials of UTI were copy-typed and compared to 491 healthy controls with no known history of VUR or UTI. Less than 1% of controls had a single copy of DCHS1 , while 31% of children with UTI and no VUR and 7% of children with UTI and VUR had a single copy of the DCHS1 gene. Using immunostaining, we localized expression postnatally to the bladder and renal epithelia. Mice were also challenged with two uropathogenic Escherichia coli strains, and Dchs1 mRNA was quantified. This study represents the first report of DCHS1 in association with pediatric UTI. We hypothesize that its role in innate immunity is critical to lower urinary tract defense. Further investigation is required to determine the role of DCHS1 in innate immunity.

Background Acute kidney injury (AKI) is increasingly recognized as a consequential clinical complication in children with severe malaria. However, approaches to estimate baseline creatinine (bSCr) are not standardized in this unique patient population. Prior to wide-spread utilization, bSCr estimation methods need to be evaluated in many populations, particularly in children from low-income countries. Methods We evaluated six methods to estimate bSCr in Ugandan children aged 6 months to 12 years of age in two cohorts of children with severe malaria (n = 1078) and healthy community children (n = 289). Using isotope dilution mass spectrometry (IDMS)-traceable creatinine measures from community children, we evaluated the bias, accuracy and precision of estimating bSCr using height-dependent and height-independent estimated glomerular filtration (eGFR) equations to back-calculate bSCr or estimating bSCr directly using published or population-specific norms. Results We compared methods to estimate bSCr in healthy community children against the IDMS-traceable SCr measure. The Pottel-age based equation, assuming a normal GFR of 120 mL/min per 1.73m2, was the more accurate method with minimal bias when compared to the Schwartz height-based equation. Using the different bSCr estimates, we demonstrated the prevalence of KDIGO-defined AKI in children with severe malaria ranged from 15.6–43.4%. The lowest estimate was derived using population upper levels of normal and the highest estimate was derived using the mean GFR of the community children (137 mL/min per 1.73m2) to back-calculate the bSCr. Irrespective of approach, AKI was strongly associated with mortality with a step-wise increase in mortality across AKI stages (p Conclusions We recommend using height-independent age-based approaches to estimate bSCr in hospitalized children in sub-Saharan Africa due to challenges in accurate height measurements and undernutrition which may impact bSCr estimates. In this population the Pottel-age based GFR estimating equation obtained comparable bSCr estimates to population-based estimates in healthy children.

Carbonic anhydrase II knockout ( Car2−/−) mice have depleted numbers of renal intercalated cells, which are increasingly recognized to be innate immune effectors. We compared pyelonephritis susceptibility following reciprocal renal transplantations between Car2−/−and wild-type mice. We examined the effect of pharmacological CA suppression using acetazolamide in an experimental murine model of urinary tract infection. Car2−/−versus wild-type mice were compared for differences in renal innate immunity. In our transplant scheme, mice lacking CA-II in the kidney had increased pyelonephritis risk. Mice treated with acetazolamide had lower kidney bacterial burdens at 6 h postinfection, which appeared to be due to tubular flow from diuresis because comparable results were obtained when furosemide was substituted for acetazolamide. Isolated Car2−/−kidney cells enriched for intercalated cells demonstrated altered intercalated cell innate immune gene expression, notably increased calgizzarin and insulin receptor expression. Intercalated cell number and function along with renal tubular flow are determinants of pyelonephritis risk.

More than 10% of the global human population is now afflicted with kidney stones, which are commonly associated with other significant health problems including diabetes, hypertension and obesity. Nearly 70% of these stones are primarily composed of calcium oxalate, a mineral previously assumed to be effectively insoluble within the kidney. This has limited currently available treatment options to painful passage and/or invasive surgical procedures. We analyze kidney stone thin sections with a combination of optical techniques, which include bright field, polarization, confocal and super-resolution nanometer-scale auto-fluorescence microscopy. Here we demonstrate using interdisciplinary geology and biology (geobiology) approaches that calcium oxalate stones undergo multiple events of dissolution as they crystallize and grow within the kidney. These observations open a fundamentally new paradigm for clinical approaches that include in vivo stone dissolution and identify high-frequency layering of organic matter and minerals as a template for biomineralization in natural and engineered settings.

Acute pyelonephritis is frequently associated with metabolic acidosis. We previously reported that metabolic acidosis stimulates expression of hypoxia-inducible factor (HIF)-1α-induced target genes such as stromal derived factor-1 and cathelicidin, an antimicrobial peptide. Since the collecting duct (CD) plays a pivotal role in regulating acid-base homeostasis and is the first nephron segment encountered by an ascending microbial infection, we examined the contribution of HIF-1α to innate immune responses elicited by acid loading of an M-1 immortalized mouse CD cell line. Acid loading of confluent M-1 cells was achieved by culture in pH 6.8 medium supplemented with 5-(N-ethyl-N-isopropyl)-amiloride to block Na(+)/H(+) exchange activity for 24 h. Acid loading induced antimicrobial peptide [cathelicidin and β-defensin (Defb2 and Defb26)] mRNA expression and M-1 cell resistance to uropathogenic Escherichia coli infection to an extent similar to that obtained by inhibition of HIF prolyl hydroxylases, which promote HIF-1α protein degradation. The effect of acid loading on M-1 cell resistance to uropathogenic E. coli infection was reduced by inhibition of HIF-1α (PX-478), and, in combination with prolyl hydroxylase inhibitors, acidosis did not confer additional resistance. Thus, metabolic stress of acidosis triggers HIF-1α-dependent innate immune responses in CD (M-1) cells. Whether pharmacological stabilization of HIF prevents or ameliorates pyelonephritis in vivo warrants further investigation.

Urinary stone disease (USD) is affecting a greater number of children and low bone mineral density (BMD) and increased skeletal fractures have been demonstrated in stone patients; however, the mechanism(s) driving bone disease remain unclear. This pilot study was undertaken to assess an adolescent kidney stone cohort’s BMD and evaluate for an inverse correlation between BMD and urine concentration of lithogenic minerals and/or inflammatory levels. Prospective case–control study was carried out at a large pediatric center. 15 participants with USD (12–18 years of age, 8 female) were matched by age, sex, and body mass index to 15 controls. Lumbar and total body BMD z-score did not differ between groups. When stone formers were separated by sex, there was a significant difference between male stone formers vs. controls total body BMD z-score (Fig. 1). BMD z-score did not significantly correlate with urine calcium, oxalate, citrate or magnesium. Higher urine IL-13 did significantly correlate with higher total body BMD z-score (r = 0.677, p = 0.018). Total body BMD z-score did significantly correlate with body mass index (BMI) as expected for the control group (r = 0.6321, p = 0.0133). However, this relationship was not present in the USD group (r = − 0.1629, p = 0.5619). This is a small but hypothesis-generating study which demonstrates novel evidence of male-specific low BMD in adolescent stone formers. Furthermore, we demonstrated a positive association between urine IL-13 and total body BMD z-score USD patients as well as a lack of a positive BMD and BMI correlations in stone formers.

ObjectiveCurrent urinary tract infection (UTI) diagnostic strategies that rely on leukocyte esterase have limited accuracy. We performed an aptamer-based proteomics pilot study to identify urine protein levels that could differentiate a culture proven UTI from culture negative samples, regardless of pyuria status.MethodsWe analyzed urine from 16 children with UTIs, 8 children with culture negative pyuria and 8 children with negative urine culture and no pyuria. The urine levels of 1,310 proteins were quantified using the Somascan™ platform and normalized to urine creatinine. Machine learning with support vector machine (SVM)-based feature selection was performed to determine the combination of urine biomarkers that optimized diagnostic accuracy.ResultsEight candidate urine protein biomarkers met filtering criteria. B-cell lymphoma protein, C-X-C motif chemokine 6, C-X-C motif chemokine 13, cathepsin S, heat shock 70kDA protein 1A, mitogen activated protein kinase, protein E7 HPV18 and transgelin. AUCs ranged from 0.91 to 0.95. The best prediction was achieved by the SVMs with radial basis function kernel.ConclusionsBiomarkers panel can be identified by the emerging technologies of aptamer-based proteomics and machine learning that offer the potential to increase UTI diagnostic accuracy, thereby limiting unneeded antibiotics.

Autoantibodies targeting host antigens contribute to autoimmune disorders, frequently occur during and after infections and have been proposed to contribute to malaria-induced anemia. We measured anti-phosphatidylserine (PS) and anti-DNA antibody levels in 382 Ugandan children prospectively recruited in a study of severe malaria (SM). High antibody levels were defined as antibody levels greater than the mean plus 3 standard deviations of community children (CC). We observed increases in median levels of anti-PS and anti-DNA antibodies in children with SM compared to CC (p

The incidence of nephrolithiasis continues to rise. Previously, we showed that a monogenic cause could be detected in 11.4% of individuals with adult-onset nephrolithiasis or nephrocalcinosis and in 16.7-20.8% of individuals with onset before 18 years of age, using gene panel sequencing of 30 genes known to cause nephrolithiasis/nephrocalcinosis. To overcome the limitations of panel sequencing, we utilized whole exome sequencing in 51 families, who presented before age 25 years with at least one renal stone or with a renal ultrasound finding of nephrocalcinosis to identify the underlying molecular genetic cause of disease. In 15 of 51 families, we detected a monogenic causative mutation by whole exome sequencing. A mutation in seven recessive genes ( AGXT, ATP6V1B1, CLDN16, CLDN19, GRHPR, SLC3A1, SLC12A1 ), in one dominant gene ( SLC9A3R1 ), and in one gene ( SLC34A1 ) with both recessive and dominant inheritance was detected. Seven of the 19 different mutations were not previously described as disease-causing. In one family, a causative mutation in one of 117 genes that may represent phenocopies of nephrolithiasis-causing genes was detected. In nine of 15 families, the genetic diagnosis may have specific implications for stone management and prevention. Several factors that correlated with the higher detection rate in our cohort were younger age at onset of nephrolithiasis/nephrocalcinosis, presence of multiple affected members in a family, and presence of consanguinity. Thus, we established whole exome sequencing as an efficient approach toward a molecular genetic diagnosis in individuals with nephrolithiasis/nephrocalcinosis who manifest before age 25 years.

Urinary stone disease (USD) is increasing in adult and pediatric populations. Adult and pediatric studies have demonstrated decreased bone mineral density and increased fracture rates. USD has also been independently linked to increased rates of myocardial infarction and cerebral vascular accidents. Although USD is a multisystem disorder involving the kidneys, bone, and vasculature, the molecular mechanisms linking these three organs remain unknown. Calcium oxalate nephropathy was induced in C57BL/6J mice with intra-peritoneal (ip) injection of sodium glyoxolate. Half of each kidney underwent Pizzalato staining and half was snap frozen for RNA extraction. RT2 Profiler Mouse Atherosclerosis, Osteoporosis, and Calcium Signaling PCR Arrays (Qiagen) were performed. Only results that passed quality checks in PCR array reproducibility and genomic DNA contamination were included. Genes had to show at least fourfold differential expression and P 10-fold increase. All 10 have P ≤ 0.003. The calcium signaling array showed significant fourfold upregulation of 10 genes, four of which were ≥10-fold. All 10 have P ≤ 0.03. We have demonstrated that calcium oxalate nephropathy can induce upregulation of atherosclerotic, metabolic bone, and calcium homeostasis genes in a murine model. This may be and initial step in identifying the molecular mechanisms linking stone, bone, and cardiovascular disease. J. Cell. Biochem. 118: 2744–2751, 2017. © 2017 Wiley Periodicals, Inc.

Kidney stones are increasing in the pediatric and adult populations; similarly osteoporosis is increasingly recognized in children. While kidney stone formers are known to suffer from low bone density, metabolic bone patients have not been considered a high risk population for kidney stones. Retrospective chart review of Nationwide Children’s Hospital Metabolic Bone Clinic patients from October 2009–2013. Patients were identified by ICD 9 codes for osteoporosis, osteopenia, low bone density and kidney stones. Only patients with radiologic evidence of both diseases were included.Twenty-six of 889 patients met criteria; this is equivalent to an incidence of 30 per 100,000 patients. Osteoporosis was the most frequent bone diagnosis. Males were the majority (68%). Most common secondary diagnoses: seizure (52%) and cerebral palsy (44%). Treatment: calcium (48%), vitamin D (40%), bisphosphonates (48%). The majority (75%) were non-ambulatory. Most frequent lithogenic medications: Topiramate (42%) and corticosteroids (27%). This is one of the first studies to consider metabolic bone patients as high risk for urinary stone disease. We found a higher rate of kidney stones in pediatric metabolic bone patients compared to data available for the general pediatric kidney stone population. The most common risk factor for bone and stone disease was nonambulatory status. Males were more frequently affected than females; this is the reverse of general adolescent kidney stone population. The predominance of cerebral palsy and seizure patients can be attributed to their frequency of non-ambulatory status and lithogenic medications such as Topiramate.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Vesicoureteral reflux (VUR) is a complex, heritable disorder. Genome-wide linkage analyses of families affected by VUR have revealed multiple genomic loci linked to VUR. These loci normally harbor a number of genes whose biologically functional variant is yet to be identified. DNA copy number variations (CNVs) have not been extensively studied at high resolution in VUR patients. In this study, we performed array comparative genomic hybridization (aCGH) on a cohort of patients with a history of both VUR and urinary tract infection (UTI) with the objective of identifying genetic variations responsible for VUR and/or UTI susceptibility. UTI/VUR-associated CNVs were identified by aCGH results from the 192 Randomized Intervention for Children With Vesicoureteral Reflux (RIVUR) patients compared to 683 controls. Rare, large CNVs that are likely pathogenic and lead to VUR development were identified using stringent analysis criteria. Because UTI is a common affliction with multiple risk factors, we utilized standard analysis to identify potential disease-modifying CNVs that can contribute to UTI risk. Gene ontology analysis identified that CNVs in innate immunity and development genes were enriched in RIVUR patients. CNVs affecting innate immune genes may contribute to UTI susceptibility in VUR patients and may provide the first step in assisting clinical medicine in determining adverse outcome risk in children with VUR.

Background: Urinary stones are affecting more children, and pediatric stone formers have unique pathophysiology compared to adults. While adult stone formers are most frequently male, children have an age dependent sex prevalence. Under 10 years, a majority of stone formers are boys; adolescent stone formers are mostly female. Previous adult studies have shown that stone composition is influenced by the sex and age of the stone former. Thus, we hypothesize that female and male stone forming children will also have sex and age specific stone phenotypes.Methods: Retrospective chart review of a large pediatric center's stone forming children 6/1/2009 to 6/1/2016. Patients were identified by ICD 9 codes: N20, N20.1, and N20.9. Charts were reviewed for radiographic evidence of stones or documented visualized stone passage.Results: One hundred and thirty six subjects: 54 males and 82 females. Females were older, median age 14 years [interquartile range (IQR): 11, 15] vs. males' median age 12 years (IQR: 11, 14) (p < 0.01). Females had lower height z-scores, median 0.2 (IQR: −0.8, 0.8) vs. males' median 0.8 (IQR: −0.2, 1.8) (p < 0.01). Presenting symptoms were similar except flank pain affecting 39% of females vs. 22% of males (p = 0.04). Leukocyte esterase was positive in more females than males (33 vs. 4%) (p < 0.001). Males had a higher BUN/Cr ratio, mean ± standard deviation of 19.8 ± 6.3 vs. 16.6 ± 6.5 in females (p = 0.01). Glomerular hyperfiltration was present in 9% of patients while 35% of patients had estimated glomerular filtration rate (eGFR) < 90 ml/min/1.73 m2. Treatment strategies and clinical course were similar except females were told to increase dietary citrate more frequently than males (21 vs. 4%) (p < 0.01).Conclusion: We have provided a novel analysis and demonstrated that low height z-score and pyuria are more common in female stone formers. We have also shown that 9% of pediatric stone formers have labs consistent with hyperfiltration. Whether high protein intake and/or chronic dehydration are associated with hyperfiltration and long-term renal function in children with kidney stones will be an area for future research.