Supplementary Material for: A Cross-Sectional Growth Reference and Chart of Stretched Penile Length for Japanese Boys Aged 0-7 Years

<b><i>Background/Aim:</i></b> Reference values for penile length have not been established for Japanese boys. We aimed to develop percentiles and means (and standard deviations) of stretched penile length (SPL) in Japanese boys. <b><i>Methods:</i></b> A cross-sectional study was performed in 1,628 Japanese boys aged <9 years from 2007 through 2014. The LMS method was used to develop a growth reference and chart for SPL in boys aged 0-7 years. Interobserver variation in SPLs was assessed in 32 boys (aged 0-11 years, median 3 years) using the Bland-Altman plot. The correlation between SPL and stature, weight or body mass index (BMI) was analyzed by the Pearson test. <b><i>Results:</i></b> SPL increases continuously during the prepubertal period, and most rapidly in the first 4 months of life. No significant fixed or proportional bias was found for interobserver variation (p = 0.5; r<i> = </i>0.33, p = 0.06). There was no significant correlation between SPL z-score and stature z-score (r<i> = </i>0.14, p < 0.001), weight z-score (r<i> = </i>0.09, p < 0.001) or BMI z-score (r = 0.01, p = 0.71). <b><i>Conclusion:</i></b> These data serve as an updated growth reference for SPL in Japanese boys aged 0-7 years

Supplementary Material for: Anemia and Long-Term Renal Prognosis in Patients with Post-Renal Acute Kidney Injury of Nonmalignant Cause

<p><b><i>Background/Aims:</i></b> The renal prognosis of post-renal acute kidney injury (PoR-AKI) has not been verified so far. The objective of this study was to assess the association of baseline anemia with long-term renal prognosis in patients with PoR-AKI. <b><i>Methods:</i></b> We performed a multicenter retrospective cohort study. Consecutive adult patients from December 2006 to February 2010, who met the requirements as mentioned in the definition of PoR-AKI, were included. Patients without data on baseline renal function and at 6 months after PoR-AKI were excluded. We set baseline hemoglobin (Hb) level (g/dl) as the main exposure to be tested. The main outcome measure was long-term renal prognosis as determined by the difference between proximate estimated glomerular filtration rate (eGFR) at 6 months after diagnosis of PoR-AKI and baseline eGFR prior to the occurrence of the present PoR-AKI (ΔeGFR after 6 months) using the general linear model. <b><i>Results:</i></b> We included 136 patients with PoR-AKI. The most frequent cause of PoR-AKI was malignancy, accounting for 39.0% (n = 53) of cases. Multivariate analysis adjusted for possible confounders showed that ΔeGFR after 6 months significantly changed by -4.28 ml/min/1.73 m² for every 1 g/dl lower Hb at diagnosis (95% CI 1.86-6.69, p < 0.01). An additional multivariate analysis that was stratified by the presence or absence of malignancy as the cause of PoR-AKI yielded the same significant result only in the stratum of the nonmalignant cause of PoR-AKI. <b><i>Conclusion:</i></b> Patients with a nonmalignant cause of PoR-AKI who have baseline anemia may have poor long-term renal prognosis. In these cases, close observation of renal function after renal recovery may be required.</p

Supplementary Material for: Discordant Genotype-Phenotype Correlation in Familial Hyperaldosteronism Type III with KCNJ5 Gene Mutation: A Patient Report and Review of the Literature

<b><i>Background:</i></b> Familial hyperaldosteronism type III (FH-III) is a rare autosomal dominant disease for which five missense mutations in <i>KCNJ5</i> have been identified. FH-III has a wide phenotypic variability from spironolactone-responsive hyperaldosteronism to massive adrenal hypertrophy with drug-resistant hypertension. This variation has mainly been attributed to genotype, because, in contrast to other genotypes (G151R, T158A, I157S, and Y152C), (1) FH-III patients with G151E have shown milder phenotype, and (2) G151E-harboring cells were found to have rapid lethality due to much larger sodium conductance of the encoded channel (Kir3.4), which prevents adrenal hypertrophy. <b><i>Methods:</i></b> Here we describe the clinical course of a sporadic case of FH-III, with de novo G151R mutation. <b><i>Results:</i></b> The patient developed polyuria at around 1.5 years of age and developed hypertension and hypokalemia by 4 years of age. Thereafter, spironolactone treatment successfully ameliorated hyperaldosteronism for 7 years with no discernible adrenal enlargement. <b><i>Conclusion:</i></b> Diverse clinical severity in FH-III cannot be defined solely by <i>KCNJ5</i> genotype

Supplementary Material for: Population Pharmacokinetics of Diazoxide in Children with Hyperinsulinemic Hypoglycemia

<p><b><i>Background:</i></b> Diazoxide is the first-line treatment for pediatric hyperinsulinemic hypoglycemia (HI). This study aimed to elucidate the pharmacokinetics of diazoxide in children with HI. <b><i>Methods:</i></b> We obtained 81 blood samples from 22 children with HI. Measured serum diazoxide concentrations were used for population pharmacokinetic analysis. Patient factors influencing pharmacokinetics were estimated using nonlinear mixed-effects model analysis. Relationships between drug exposure and adverse drug reactions were also investigated. <b><i>Results:</i></b> Diazoxide disposition in the body was described by a 1-compartment model. Oral clearance (CL/F) and the volume of distribution were proportional to body weight (WT), as expressed by CL/F in males (liters/h) = 0.0358 + 0.00374 × WT (kg). CL/F in females was 39% greater than that in males. Steady-state concentrations of diazoxide were similar following twice- and 3 times-daily dosing when the total daily doses were comparable. A patient whose serum diazoxide concentration exceeded 100 μg/mL over a 4-month period developed hyperglycemia. No significant correlation was observed between severity of hirsutism and diazoxide concentration. <b><i>Conclusion:</i></b> We have proposed for the first time a population pharmacokinetic model for diazoxide in children with HI. The potential risk of diabetes mellitus and/or hyperglycemia increases when serum concentrations of diazoxide exceed 100 μg/mL.</p

Supplementary Material for: DHX37 Variant is One of Common Genetic Causes in Japanese Patients with Testicular Regression Syndrome / Partial Gonadal Dysgenesis without Müllerian Derivatives

Introduction: The testicular regression syndrome (TRS) is a form of differences of sex development (DSD) in which the testes differentiate and function during early embryonic development, but subsequently regress. The clinical phenotype of TRS often overlaps with that of partial gonadal dysgenesis (PGD). Previous studies have demonstrated a causal association between TRS/PGD and heterozygous missense variants of DHX37. Methods: We enrolled 11 Japanese 46,XY individuals (from 10 families) with TRS/PGD who exhibited undetected or hypoplastic testes, Müllerian duct regression, and low serum testosterone or anti-Müllerian hormone levels. The subjects underwent targeted sequencing of 36 known causative genes for DSD, PCR-based Sanger sequencing of DHX37, or whole exome sequencing. Results: Previously described pathogenic variants or novel nonsense variants (SRY, NR5A1, and DMRT1) were observed in four out of 10 families. Additionally, we identified two heterozygous rare variants of DHX37 in four families: a previously reported pathogenic variant (c.923G>A, p.Arg308Gln) in three and a novel likely pathogenic variant (c.1882A>C, p.Thr628Pro) in one. The external genitalia of patients with the DHX37 variants varied from female-type to male-type without micropenis. Eighty percent of Japanese patients with TRS/PGD had monogenic disorders including DHX37 variant being the most commonly identified (40%). The external or internal genital phenotype of TRS/PGD overlaps between DHX37 variant carriers and others. Conclusions: DHX37 variant is one of common genetic causes in Japanese patients with TRS/PGD without Müllerian derivatives. Genetic test is helpful in detecting DHX37-related TRS/PGD, because of the phenotypic diversity of the external genitalia in this disorder

Supplementary Material for: The cancer cachexia index can be used to prognostically predict patients with gastric cancer undergoing gastrectomy

Objective. Cancer cachexia occurs in cancer patients more frequently as the cancer progresses, with a negative impact on treatment outcomes. In this study, we sought to clarify the clinical impact of a cancer cachexia index CXI in patients with gastric cancer (GC) undergoing gastrectomy. Methods. Between January 2013 and December 2018, we reviewed data from 556 patients treated for GC at our hospital. CXI was calculated using skeletal muscle index (SMI), serum albumin, and neutrophil-lymphocyte ratios (NLR). Patients were divided into high (n=414) or low CXI (n=142) groups. We investigated the clinical impact of CXI in patients with GC undergoing gastrectomy. Results. Multivariate analyses of 5-year overall survival (OS) and cancer specific survival (CSS) rates indicated that a low CXI was independently associated with unfavorable outcomes for patients with GC. In multivariate analyses, SMI was independent predictor of OS but not CSS. NLR was not an independent predictor of either OS or CSS. Complication incidences [≥ Clavien Dindo (CD) 3] were non-significantly higher in the low (vs. high) CXI group. Conclusion. CXI was a more valuable prognostic biomarker when compared with SMI or NLR in GC patients undergoing gastrectomy. We suggest that patients with low CXI values should be given more comprehensive treatment, including exercise and nutritional therapy to improve clinical outcomes

PowerPoint Slides for: Erythropoietin Hyporesponsiveness in Dialysis Patients: Possible Role of Statins

<p><b><i>Background:</i></b> Hypothesizing that statins may be useful as adjuvant treatment for renal anemia, we examined the association between statin prescription (Rx) and erythropoiesis-stimulating agent (ESA) hyporesponsiveness in Japanese hemodialysis (HD) patients prescribed ESAs. <b><i>Methods:</i></b> We examined 3,602 patients in 60 HD facilities dialyzed 3 times/week for ≥4 months from the Japan Dialysis Outcomes and Practice Patterns Study phases 3-5 (2005-2015). Statin Rx was reported at the end of a 4-month interval (baseline) for each patient. ESA hyporesponsiveness in the subsequent 4 months was then defined as a binary indicator (mean hemoglobin [Hgb] level <10 g/dL and mean ESA dose >6,000 units/week) and separately as the ESA resistance index (ERI; mean ESA dose/[dry weight × mean Hgb]). We used adjusted logistic and linear regressions to evaluate the associations between statin Rx and ESA hyporesponsiveness. <b><i>Results:</i></b> At baseline, 16.2% of patients reported statin Rx; 12.8% were classified as having ESA hyporesponsiveness during 4 months of follow-up. Compared to patients without statin Rx, patients with statin Rx had lower odds of ESA hyporesponsiveness (OR 0.87; 95% CI 0.66-1.15). Similarly, the ERI was lower for those with statin Rx than without (ratio of means, 0.94; 95% CI 0.89-0.99) after adjustment for possible confounders. <b><i>Conclusions:</i></b> Our results suggest that statins may slightly reduce ESA hyporesponsiveness in HD patients. However, any causal inference is limited by the observational study design and unmeasured compliance with statin Rx.</p

Supplementary Material for: The Effects of All-Trans Retinoic Acid on the Induction of Oral Tolerance in a Murine Model of Bronchial Asthma

<b><i>Background:</i></b> Active suppression induced by regulatory T (Treg) cells is reported to be one of the mechanisms involved in oral tolerance. All-trans retinoic acid (ATRA) has been reported to affect Treg cell differentiation. The present study examined the effects of ATRA on the induction of oral tolerance in a murine model of bronchial asthma. <b><i>Methods:</i></b> BALB/c mice were sensitized to and challenged with ovalbumin (OVA) through feeding followed by OVA challenges. In some study groups ATRA was orally administered concomitantly with OVA feeding either in the presence or absence of the retinoic acid receptor antagonist LE135. Lung CD4⁺ T cells were isolated from mice exposed to ATRA and/or OVA, and transferred to control mice. Airway hyperresponsiveness (AHR), cell counts and cytokine levels in bronchoalveolar lavage (BAL) fluid, and lung histology were assessed. <b><i>Results:</i></b> Concomitant administration of ATRA with OVA ameliorated AHR, airway eosinophilia, elevation of cytokines in BAL fluid and goblet cell metaplasia. The proportion of Treg cells in the lungs was increased in mice treated with OVA and ATRA, as compared to those treated with OVA only. Transfer of lung CD4⁺ T cells from mice treated with OVA and ATRA induced suppression of AHR and airway inflammation. LE135 completely reversed the effects of ATRA on AHR, airway allergic inflammation and the number of Treg cells in the lungs. <b><i>Conclusion:</i></b> These data suggested that oral administration of ATRA with OVA had the potential to enhance oral tolerance in this murine model of bronchial asthma. These effects were mediated, at least in part, by Treg cell expansion

Supplementary Material for: Significance of the Lysyl Oxidase Members Lysyl Oxidase Like 1, 3, and 4 in Gastric Cancer

<b><i>Background/Aims:</i></b> Lysyl oxidase (LOX) family members play a key role in modifying the primary tumor microenvironment by crosslinking collagens and elastin in the extracellular matrix. The aim of this study was to analyze the LOX-like (LOXL)1, LOXL3, and LOXL4 expressions in gastric cancer tissue by immunohistochemical staining. <b><i>Methods:</i></b> The correlations between the clinicopathological features of 597 primary gastric carcinomas and LOX family members – LOXL1, LOXL3, and LOXL4 – were investigated by immunohistochemical studies. The effect of the transforming growth ­factor β1 (TGFβ1) on the expressions of <i>LOXL1, LOXL3,</i> and <i>LOXL4</i> in gastric cancer was examined using diffuse-type gastric cancer cell lines in vitro. <b><i>Results:</i></b> The expressions of LOXL1, LOXL3, and LOXL4 were correlated with T invasion, lymph node metastasis, and lymphatic and venous invasion. LOXL1 expression was associated with histological intestinal-type and expanding growth patterns. The overall survival of patients with LOXL1-, LOXL3-, or LOXL4-positive cancer was poorer than those with negative cancer. <i>LOXL3</i> and <i>LOXL4</i> mRNA expressions were significantly high in diffuse-type gastric cancer cells with high invasion ability. TGFβ decreased the LOXL1 expression and increased LOXL3 and LOXL4 expression. <b><i>Conclusion:</i></b> LOXL1, LOXL3, and LOXL4 expressions are associated with distant metastasis of gastric cancer

Supplementary Material for: The association between high-dose allopurinol and erythropoietin hyporesponsiveness in advanced chronic kidney disease. JOINT-KD study

Introduction: To explore the association between urate-lowering agents and reduced response to erythropoietin-stimulating agents in patients suffering from chronic kidney disease G5. Methods: We conducted a cross-sectional, multicenter study in Japan between April and June 2013, enrolling patients aged 20 years or older with an estimated glomerular filtration rate of ≤15 mL/min/1.73 m2. Exclusion criteria encompassed patients with a history of hemodialysis, peritoneal dialysis, or organ transplantation. The patients were categorized into four groups based on the use of urate-lowering drugs: high-dose allopurinol (>50 mg/day), low-dose allopurinol (≤50 mg/day), febuxostat, and no-treatment groups. We used a multivariable logistic regression model, adjusted for covariates, to determine the odds ratio (OR) for erythropoietin hyporesponsiveness, defined by an erythropoietin resistance index (ERI) of ≥10, associated with urate-lowering drugs. Results: A total of 542 patients were included in the analysis, with 105, 36, 165, and 236 patients in the high-dose allopurinol, low-dose allopurinol, febuxostat, and no-treatment groups, respectively. The median and quartiles of ERIs were 6.3 (0, 12.2), 3.8 (0, 11.2), 3.4 (0, 9.8), and 4.8 (0, 11.2) in the high-dose allopurinol, low-dose allopurinol, febuxostat, and no-treatment groups, respectively. The multivariate regression model showed a statistically significant association between the high-dose allopurinol group and erythropoietin hyporesponsiveness, compared to the no-treatment group (OR=1.98, 95% confidence interval: 1.10–3.57). Conclusions: Our study suggests that the use of high-dose allopurinol exceeding the optimal dose may lead to hyporesponsiveness to erythropoiesis-stimulating agents