Abnormal Development of Microbiota May Be a Risk Factor for Febrile Urinary Tract Infection in Infancy.

Febrile urinary tract infection (fUTI) is common in infants, but specific risk factors for developing it remain unclear. As most fUTIs are caused by ascending infections of intestinal bacteria, dysbiosis-an imbalance in gut microbial communities-may increase fUTI risk. This study was conducted to test the hypothesis that abnormal development of gut microbiota during infancy increases the risk of developing fUTI. Stool samples were collected from 28 infants aged 3-11 months with first-onset fUTI (fUTI group) and 51 healthy infants of the same age (HC group). After bacterial DNA extraction, 16S rRNA expression was measured and the diversity of gut microbiota and constituent bacteria were compared between the two groups. The alpha diversity of gut microbiota (median Shannon index and Chao index) was significantly lower in the fUTI group (3.0 and 42.5) than in the HC group (3.7 and 97.0; p < 0.001). The beta diversity also formed different clusters between the two groups (p < 0.001), suggesting differences in their microbial composition. The linear discriminant analysis effect size showed that the fUTI group proportionally featured significantly more Escherichia-Shigella in the gut microbiota (9.5%) than the HC group (3.1%; p < 0.001). In summary, abnormal gut microbiota development during infancy may increase the risk of fUTI.

Accuracy of diagnosing acute kidney injury by assessing urine output within the first week of life in extremely preterm infants.

BACKGROUND: Neonatal acute kidney injury (AKI) is associated with increased mortality and is often assessed with the neonatal modified Kidney Disease: improving Global Outcomes (KDIGO) classification, which uses changes in serum creatinine levels. However, because this classification has many drawbacks, a novel method, the neonatal Risk, Injury, Failure, Loss, and End-Stage Kidney Disease (nRIFLE) classification for diagnosing neonatal AKI according to urine output (UO), was recently proposed. To date, no data on the incidence of AKI according to nRIFLE are available for extremely preterm infants (born at gestational age less than 28 weeks). This study was conducted to clarify the association between incidence of AKI and in-hospital mortality in extremely preterm infants. METHODS: Of 171 extremely preterm infants hospitalized from 2006 to 2020, 84 in whom indwelling bladder catheters were placed for UO measurements within 24 h of life were included. The incidence of AKI was assessed using the nRIFLE classification. In-hospital mortality was compared between patients with AKI and those without it. RESULTS: The incidence of AKI during the first week of life was 56% and that of in-hospital mortality was significantly higher in patients with AKI (25.5%) than in those without it (2.8%). The odds ratio was 12.3 with 95% confidence interval ranging from 1.5 to 100.0. CONCLUSION: The incidence of AKI according to nRIFLE was higher than reported in most previous studies using the neonatal modified KDIGO classification, suggesting that assessment by nRIFLE criteria using UO may improve diagnostic accuracy of AKI in extremely preterm infants.

Association of Neonatal Jaundice with Gut Dysbiosis Characterized by Decreased Bifidobacteriales.

Neonatal jaundice, caused by excess serum bilirubin levels, is a common condition in neonates. Imbalance in the gut microbiota is believed to play a role in the development of neonatal jaundice. Thus, we aimed to reveal the gut microbiota characteristics in neonates with jaundice. 16S rRNA gene sequencing was performed on stool samples collected on day 4 from 26 neonates with jaundice (serum total bilirubin > 15.0 mg/dL) and 17 neonates without jaundice (total serum bilirubin < 10.0 mg/dL). All neonates were born full term, with normal weight, by vaginal delivery, and were breastfed. Neonates who were administered antibiotics, had serum direct bilirubin levels above 1 mg/dL, or had conditions possibly leading to hemolytic anemia were excluded. The median serum bilirubin was 16.0 mg/dL (interquartile range: 15.5-16.8) and 7.4 mg/dL (interquartile range: 6.8-8.3) for the jaundice and non-jaundice groups, respectively. There was no difference in the alpha diversity indices. Meanwhile, in the jaundice group, linear discriminant analysis effect size revealed that Bifidobacteriales were decreased at the order level, while Enterococcaceae were increased and Bifidobacteriaceae were decreased at the family level. Bifidobacteriaceae may act preventatively because of their suppressive effect on beta-glucuronidase, leading to accelerated deconjugation of conjugated bilirubin in the intestine. In summary, neonates with jaundice had dysbiosis characterized by a decreased abundance of Bifidobacteriales.

Clinical Significance of Probiotics for Children with Idiopathic Nephrotic Syndrome.

We previously reported that a decrease in butyrate-producing bacteria in the gut is a potential cause of regulatory T cell (Treg) abnormalities in children with idiopathic nephrotic syndrome (INS). Therefore, we hypothesized that administration of butyrate-producing bacteria might reduce INS relapse and the need for immunosuppressants in these patients. Twenty patients in remission from INS (median age 5.3 years, 15 boys) were enrolled in the study and assigned to receive either daily oral treatment with a preparation of 3 g Clostridium butyricum or no probiotic treatment. The number of relapses and requirement for immunosuppressive agents were compared between the two groups. In the probiotic treatment group, analyses of the gut microbiota and Treg measurements were also performed. Probiotic-treated patients experienced fewer INS relapses per year compared with non-probiotic-treated patients (p = 0.016). Further, administration of rituximab in the probiotic treatment group was significantly less frequent compared with the non-probiotic-treated group (p = 0.025). In the probiotic treatment group, analyses before and after probiotic treatment revealed the significant increases in the relative abundance of butyrate-producing bacteria (p = 0.017) and blood Treg counts (p = 0.0065). Thus, oral administration of butyrate-producing bacteria during INS remission may reduce the frequency of relapse and the need for immunosuppressive agents.

Reduced urinary excretion of neutrophil gelatinase-associated lipocalin as a risk factor for recurrence of febrile urinary tract infection in children.

BACKGROUND: This study aimed to test the hypothesis that reduced urinary excretions of neutrophil gelatinase-associated lipocalin (NGAL) predispose children to recurrence of febrile urinary tract infection (fUTI). METHODS: Subjects were 38 children diagnosed with fUTI. To examine risk factors for recurrence of fUTI, the subjects were divided into a non-recurrent group and a recurrent group according to the presence or absence of fUTI over 3 years since the first episode. We measured the urinary NGAL levels in patients with fUTI at the non-infected stage in addition to age-matched healthy control children. RESULTS: In a multiple logistic regression analysis, significant differences between the groups were not observed for age, sex, the prevalence of kidney scarring and bladder bowel dysfunction, urinary ß2-microglobulin/creatinine (Cr) level, and serum levels of Cr and Cystatin C, while the recurrent group had significantly more cases with grade III or higher vesicoureteral reflux (p < 0.01). Furthermore, the urinary NGAL/Cr in the recurrent group (median, 3.60 µg/gCr) was significantly lower than that in the non-recurrent group (median, 16.47 µg/gCr; p < 0.01), and age-matched healthy control children (median, 14.14 µg/gCr; p < 0.05). The area under the receiver operating characteristic curve of NGAL/Cr was 0.86 for predicting recurrence of fUTI. A cut-off value of 11.59 µg/gCr had the best accuracy to predict recurrent fUTI yielding a specificity of 78% and a sensitivity of 93%. CONCLUSIONS: Reduced levels of urinary NGAL, which protects against urinary infection, are a risk factor for recurrence of fUTI and could serve as a biomarker.

Idiopathic nephrotic syndrome in children: role of regulatory T cells and gut microbiota.

BACKGROUND: We investigated whether an association exists between regulatory T cells (Tregs) during initial presentation in children with idiopathic nephrotic syndrome (INS) and later development of frequently relapsing INS. METHODS: Blood samples were obtained at onset and at remission from 25 patients (median age, 4.0 years) with INS; eight did not show relapse after initial response (non-relapsing [NR]), whereas 17 showed frequent relapses (frequently relapsing [FR]). Tregs were measured by flow cytometry; increases were compared between groups. Fecal samples were obtained at onset from 20 patients with INS, as well as from 20 age-matched healthy children. Gut microbiota composition was assessed using 16S ribosomal RNA (rRNA) sequencing (ion PGM). RESULTS: The rate of increase in Tregs from onset to remission was significantly lower in the FR group (124.78%) than in the NR group (879.16%; P < 0.001). Additionally, 16S rRNA sequencing of gut microbiota showed that the proportion of butyric acid-producing bacteria was significantly lower in the FR group (7.08%) than in the healthy children (17.45%; P < 0.001). CONCLUSIONS: In children with INS, small increases in Tregs in response to steroid treatment were associated with subsequent increased risk of frequent relapses. In addition, the FR group had a greater degree of dysbiosis at onset. IMPACT: A low rate of Tregs increase is associated with subsequent frequent relapses of INS. The increase in Tregs in response to steroid treatment was small when dysbiosis was present in patients with INS, particularly when the proportion of butyrate-producing bacteria was considerably reduced We presume that improvement of dysbiosis by administration of probiotics and prebiotics may enhance the rate of Tregs' increase, thus preventing frequent relapse.

Nephrotic syndrome-associated hypercoagulopathy is alleviated by both pioglitazone and glucocorticoid which target two different nuclear receptors.

BACKGROUND: Thrombosis is a potentially life-threatening nephrotic syndrome (NS) complication. We have previously demonstrated that hypercoagulopathy is proportional to NS severity in rat models and that pioglitazone (Pio) reduces proteinuria both independently and in combination with methylprednisolone (MP), a glucocorticoid (GC). However, the effect of these treatments on NS-associated hypercoagulopathy remains unknown. We thus sought to determine the ability of Pio and GC to alleviate NS-associated hypercoagulopathy. METHODS: Puromycin aminonucleoside-induced rat NS was treated with sham, Low- or High-dose MP, Pio, or combination (Pio + Low-MP) and plasma was collected at day 11. Plasma samples were collected from children with steroid-sensitive NS (SSNS) and steroid-resistant NS (SRNS) upon presentation and after 7 weeks of GC therapy. Plasma endogenous thrombin potential (ETP), antithrombin (AT) activity, and albumin (Alb) were measured using thrombin generation, amidolytic, and colorimetric assays, respectively. RESULTS: In a rat model of NS, both High-MP and Pio improved proteinuria and corrected hypoalbuminemia, ETP and AT activity (p < .05). Proteinuria (p = .005) and hypoalbuminemia (p < .001) were correlated with ETP. In childhood NS, while ETP was not different at presentation, GC therapy improved proteinuria, hypoalbuminemia, and ETP in children with SSNS (p < .001) but not SRNS (p = .330). CONCLUSIONS: Both Pio and GC diminish proteinuria and significantly alleviate hypercoagulopathy. Both Pio and MP improved hypercoagulopathy in rats, and successful GC therapy (SSNS) also improved hypercoagulopathy in childhood NS. These data suggest that even a partial reduction in proteinuria may reduce NS-associated thrombotic risk.

Predicting and Defining Steroid Resistance in Pediatric Nephrotic Syndrome Using Plasma Proteomics.

INTRODUCTION: Nephrotic syndrome (NS) is a characterized by massive proteinuria, edema, hypoalbuminemia, and dyslipidemia. Glucocorticoids (GCs), the primary therapy for >60 years, are ineffective in approximately 50% of adults and approximately 20% of children. Unfortunately, there are no validated biomarkers able to predict steroid-resistant NS (SRNS) or to define the pathways regulating SRNS. METHODS: We performed proteomic analyses on paired pediatric NS patient plasma samples obtained both at disease presentation before glucocorticoid initiation and after approximately 7 weeks of GC therapy to identify candidate biomarkers able to either predict steroid resistance before treatment or define critical molecular pathways/targets regulating steroid resistance. RESULTS: Proteomic analyses of 15 paired NS patient samples identified 215 prevalent proteins, including 13 candidate biomarkers that predicted SRNS before GC treatment, and 66 candidate biomarkers that mechanistically differentiated steroid-sensitive NS (SSNS) from SRNS. Ingenuity Pathway Analyses and protein networking pathways approaches further identified proteins and pathways associated with SRNS. Validation using 37 NS patient samples (24 SSNS/13 SRNS) confirmed vitamin D binding protein (VDB) and APOL1 as strong predictive candidate biomarkers for SRNS, and VDB, hemopexin (HPX), adiponectin (ADIPOQ), sex hormone-binding globulin (SHBG), and APOL1 as strong candidate biomarkers to mechanistically distinguish SRNS from SSNS. Logistic regression analysis identified a candidate biomarker panel (VDB, ADIPOQ, and matrix metalloproteinase 2 [MMP-2]) with significant ability to predict SRNS at disease presentation (P = 0.003; area under the receiver operating characteristic curve = 0.78). CONCLUSION: Plasma proteomic analyses and immunoblotting of serial samples in childhood NS identified a candidate biomarker panel able to predict SRNS at disease presentation, as well as candidate molecular targets/pathways associated with clinical steroid resistance.

High daily salt intake had a negative impact on how well nocturnal enuresis treatment worked on children aged 7-10 years.

AIM: We investigated whether the daily salt intake of children with nocturnal enuresis influenced their response to 1-desamino-8-D-arginine vasopressin therapy. METHODS: This study comprised 129 children (67.4% boys) with a median age of 9.2 years (range 7.2-10.4) with monosymptomatic nocturnal enuresis who were seen at Kansai Medical University Hospital, Osaka, Japan, from 2013 to 2017. Urinary sodium concentrations were determined using a spot urine test, and the children were divided into appropriate (n = 55) and excessive salt intake (n = 74) groups based on Japanese Government guidelines. After a month of therapy, the treatment responses were compared for 39 and 50 children, respectively. RESULTS: There were no significant differences in the urea nitrogen-to-creatinine or calcium-to-creatinine ratios in the two groups. However, the excessive salt intake group showed a significantly reduced treatment response to the appropriate salt intake group. In addition, the excessive and appropriate salt intake groups showed median efficacy ratios of 8.2% and 21.8%, respectively, based on intention-to-treat analysis (P = 0.029) and 12.0% and 30.8% based on per-protocol analysis (P = 0.029). CONCLUSION: High daily salt intake significantly reduced the efficacy of ddavp therapy for nocturnal enuresis and consumption should be controlled during treatment.

A Calcium-Deficient Diet in Dams during Gestation Increases Insulin Resistance in Male Offspring.

Calcium (Ca) plays an important role in the pathogenesis of insulin resistance syndrome. Osteocalcin (OC), a bone formation biomarker, acts directly on ß-cells and increases insulin secretion. We determined the effects of Ca deficiency during pregnancy and/or lactation on insulin resistance in offspring. Female Wistar rats consumed either a Ca-deficient or control diet ad libitum from three weeks preconception to 21 days postparturition. Pups were allowed to nurse their original mothers until weaning. The offspring were fed a control diet beginning at weaning and were killed on day 180. Serum carboxylated OC (Gla-OC) and undercarboxylated OC (Glu-OC), insulin and adipokines in offspring were measured. In males, mean levels of insulin, glucose, and HOMA-IR were higher in the Ca-deficient group than in the control group. In addition, ionized Ca (iCa) was inversely associated with serum Glu-OC and adiponectin in males. In females, mean levels of Glu-OC and Gla-OC in the Ca-deficient group were higher than in the control group. In all offspring, serum leptin levels were correlated with serum insulin levels, and inversely correlated with iCa. In conclusion, maternal Ca restriction during pregnancy and/or lactation influences postnatal offspring Ca metabolism and insulin resistance in a sex-specific manner.

Gut Microbiota Dysbiosis in Children with Relapsing Idiopathic Nephrotic Syndrome.

BACKGROUND: While the etiology of idiopathic nephrotic syndrome (idiopathic nephrotic syndrome [INS]; characterized by repeated relapses and comorbid allergic conditions) remains unknown, recent evidence suggests that dysfunction in regulatory T cells (Tregs) plays an important role in the development of INS as well as allergic diseases. We hypothesized that dysbiosis involving decreased butyric acid-producing gut microbiota leads to defective induction and differentiation of peripherally induced Tregs, resulting in INS relapse. METHODS: Study subjects were 12 children with INS, 8 classified as relapsing (R group; median age: 3.0 years) and 4 as non-relapsing (NR group; median age: 4.3 years), and 11 healthy children (HC group; median age: 5.1 years) serving as normal controls. Measurement of microbiota was performed using 16S ribosomal RNA metagenomic analysis, and fecal butyric acid was measured using high performance liquid chromatography. Flow-cytometric analysis of Tregs and CD4-positive (CD4+) cells in peripheral blood was also performed. RESULTS: Metagenomic analysis of gut microbiota using feces showed that the proportion of butyric acid-producing bacteria was significantly lower in R (median 6.36%) than HC (median 18.84%; p = 0.0013), but no different between NR (median 16.71%) and HC (p = 0.29). Fecal organic acid analysis revealed significantly lower butyric acid quantities in R than HC (medians: 0.48 vs. 0.99 mg/g, p = 0.042). Circulating Tregs as a proportion of CD4+ cells were decreased in 75% of R and NR. CONCLUSION: Pediatric relapsing INS patients show gut microbiota dysbiosis, characterized by a decreased proportion of butyric acid-producing bacteria and lower fecal butyric acid quantities, concomitant with reduced circulatory Tregs.

Antiproteinuric effect of an endothelin-1 receptor antagonist in puromycin aminonucleoside-induced nephrosis in rat.

BackgroundThe pathogenesis of idiopathic nephrotic syndrome (INS) remains unclear, although recent studies suggest endothelin 1 (ET-1) and CD80 of podocytes are involved. We investigated the potential of antagonist to ET-1 receptor type A (ETRA) as therapeutic agent through the suppression of CD80 in a rat model of INS.MethodsPuromycin aminonucleoside (PAN) was injected to Wister rats to induce proteinuria: some were treated with ETRA antagonist and others were treated with 0.5% methylcellulose. Blood and tissue samples were collected. Quantitative PCR was used to determine the expression of Toll-like receptor-3 (TLR-3), nuclear factor-κB (NF-κB), CD80, talin, ETRA, and ET-1 in the kidney. To confirm the level of CD80 protein expression, immunofluorescence staining and western blot analysis of the renal tissue were performed.ResultsAmount of proteinuria in the treatment group was significantly lower than the other groups. The same-day body weight, serum creatinine values, and blood pressure were not significantly different. ETRA antagonist restores podocyte foot process effacement as well as the aberrant expression of TLR-3, nuclear factor-κB (NF-κB), and CD80 in PAN-injured kidneys.ConclusionsThe ETRA antagonist may be promising drug for INS as it showed an antiproteinuric effect. Its action was considered to be through suppression of CD80 expression on podocytes.

The Effect of Family Assistance to Wake Children with Monosymptomatic Enuresis in Alarm Therapy: A Pilot Study.

PURPOSE: Alarm therapy is widely used as first line treatment for nocturnal enuresis. However, some children do not wake when nocturnal urination activates the alarm. It is currently unclear whether waking the child when the alarm is activated improves the efficacy of alarm therapy. In this study we investigated the efficacy of alarm therapy for nocturnal enuresis when children do not wake in response to the sound and their parents do not wake them. MATERIALS AND METHODS: Detailed information regarding incontinence was retrospectively obtained from 78 of 112 patients who underwent alarm therapy between 2006 and 2016, and completed a questionnaire and a 14-day bladder diary. The enrolled patients were divided into 2 groups. In the family assisted group (44) the children were awakened by family members when the alarm sounded. In the alarm control group (34) the children were self-responsible for waking to the alarm. The groups were compared to investigate differences at 16 weeks after alarm therapy began. The efficacy rate was calculated using the International Children's Continence Society criteria. RESULTS: The efficacy was similar between the groups. Full response and partial response were observed in 36.4% and 20.5% of patients in the family assisted group, and 26.5% and 29.4% of patients in the alarm control group (p = 1.00), respectively. There was no significant difference in the percentage of children who woke spontaneously to the alarm in the 2 groups (56.7% and 64.0%, respectively). CONCLUSIONS: Family assisted alarm therapy and self-responsible alarm therapy are equally efficacious in the treatment of childhood nocturnal enuresis.

Urinary C-megalin for screening of renal scarring in children after febrile urinary tract infection.

BackgroundFebrile urinary tract infection (fUTI) in children may cause renal scarring. This study aimed to investigate the usefulness of urinary biomarkers for diagnosing renal scarring after fUTI.MethodsThirty-seven children (median age: 1.36 years, range: 0.52-12.17 years, 25 boys) with a history of fUTI, who underwent renal scintigraphy for 4 months or longer after the last episode of fUTI, were analyzed. A spot urine sample was obtained on the day of renal scintigraphy to measure levels of total protein, N-acetyl-ß-D-glucosaminidase (NAG), ß2-microglobulin (BMG), neutrophil gelatinase-associated lipocalin (NGAL), liver-type fatty acid binding protein (L-FABP), and C-megalin (full-length megalin). Results were corrected for urinary creatinine (Cr) and compared between the group with renal scarring (n=23) and that without scarring (n=14). Urinary levels of C-megalin were also measured in healthy control subjects.ResultsNo significant differences in total protein, NGAL, L-FABP, NAG, and BMG levels were found between the groups. However, C-megalin levels were significantly higher in the renal scarring group than in the non-renal scarring group and healthy controls (P<0.001). A cutoff value of 6.5 pmol/nmol of urinary C-megalin/Cr yielded 73.9% of specificity and 92.9% of sensitivity.ConclusionUrinary C-megalin is useful for diagnosing renal scarring caused by fUTI.

Effect of cesarean section on relapse of childhood idiopathic nephrotic syndrome.

In parallel with the increase in the prevalence of childhood chronic diseases, the rate of cesarean delivery has risen during the past decades. This study tested the hypothesis that children delivered by cesarean section (CS) have a higher risk of relapse of idiopathic nephrotic syndrome (INS). Fifty-six children with INS were categorized into three groups. Group A consisted of patients with INS who had no relapses after the onset of INS; group B consisted of patients with INS who had infrequent relapse; and group C consisted of patients with INS who had frequent relapse. The number of enrolled patients in groups A, B, and C was 10, 14, and 32, respectively. The ratio of neonates delivered via CS was significantly higher in group C (37.5%, P < 0.001) than in groups A (0%) and B (7.1%). This study shows that CS is associated with an increased risk of relapse of childhood INS.

Nephron development and extrarenal features in a child with congenital nephrotic syndrome caused by null LAMB2 mutations.

BACKGROUND: Congenital nephrotic syndrome (CNS) is a rare disorder caused by various structural and developmental defects of glomeruli. It occurs typically as an isolated kidney disorder but associates sometimes with other systemic, extrarenal manifestations. CASE PRESENTATIONS: An infant presented with severe CNS, which progressed rapidly to renal failure at age of 3 months and death at 27 months. The clinical phenotypes and genetic causes were studied, including the renal pathology at autopsy. Besides the CNS, the affected child had remarkable right-side predominant eye-ball hypoplasia with bilateral anterior chamber dysgenesis (microcoria). Brain MRI revealed grossly normal development in the cerebrum, cerebellum, and brain stem. Auditory brainstem responses were bilaterally blunted, suggesting a defective auditory system. At autopsy, both kidneys were mildly atrophied with persistent fetal lobulation. Microscopic examination showed a diffuse global sclerosis. However, despite of the smaller size of glomeruli, the nephron number remained similar to that of the age-matched control. Whole-exome sequencing revealed that the affected child was compound heterozygous for novel truncating LAMB2 mutations: a 4-bp insertion (p.Gly1693Alafs*8) and a splicing donor-site substitution (c.1225 + 1G > A), presumably deleting the coiled-coil domains that form the laminin 5-2-1 heterotrimer complex. CONCLUSIONS: Our case represents a variation of Pierson syndrome that accompanies CNS with unilateral ocular hypoplasia. The average number but smaller glomeruli could reflect either mal-development or glomerulosclerosis. Heterogeneous clinical expression of LAMB2 defects may associate with the difference in fetal ß1 subtype compensation among affected tissues. Further study is necessary to evaluate incidence and features of auditory defect under LAMB2 deficiency.

Regulatory T cells and CTLA-4 in idiopathic nephrotic syndrome.

The pathogenesis of idiopathic nephrotic syndrome (INS) remains unknown. Recently, it was postulated that suppression of regulatory T cells (Treg) leads to massive proteinuria in INS, although there is some controversy. Considering the important role of cytotoxic T-lymphocyte-associated protein 4 (CTLA-4) in Treg-mediated immune suppression, the aim of this study was therefore to clarify the involvement of Treg and CTLA-4 in the pathogenesis of INS. Fifteen patients with INS were enrolled. Their blood was sampled twice, once at onset and once at remission induced by glucocorticoid. Although median Treg number was significantly lower at onset than in healthy children, it increased at remission. Similarly, serum CTLA-4 concentration significantly increased at remission compared with onset. Furthermore, a positive significant correlation was observed between Treg number and serum CTLA-4 level. This suggests that Treg and CTLA-4 are involved in the induction of remission in INS.