Identification of clinical factors related to antibody-mediated immune response to the subfornical organ.

OBJECTIVE: We recently reported cases of adipsic hypernatremia caused by autoantibodies against the subfornical organ in patients with hypothalamic-pituitary lesions. This study aimed to clarify the clinical features of newly identified patients with adipsic hypernatremia whose sera displayed immunoreactivity to the mouse subfornical organ. DESIGN: Observational cohort study of patients diagnosed with adipsic hypernatremia in Japan, United States, and Europe. METHODS: The study included 22 patients with adipsic hypernatremia but without overt structural changes in the hypothalamic-pituitary region and congenital disease. Antibody response to the mouse subfornical organ was determined using immunohistochemistry. The clinical characteristics were compared between the patients with positive and negative antibody responses. RESULTS: Antibody response to the mouse subfornical organ was detected in the sera of 16 patients (72.7%, female/male ratio, 1:1, 12 pediatric and 4 adult patients). The prolactin levels at the time of diagnosis were significantly higher in patients with positive subfornical organ (SFO) immunoreactivity than in those with negative SFO immunoreactivity (58.9 ± 33.5 vs. 22.9 ± 13.9 ng/ml, p < .05). Hypothalamic disorders were found in 37.5% of the patients with positive SFO immunoreactivity. Moreover, six patients were diagnosed with rapid-onset obesity with hypothalamic dysfunction, hypoventilation, and autonomic dysregulation/neural tumor syndrome after the diagnosis of adipsic hypernatremia. Plasma renin activity levels were significantly higher in patients with serum immunoreactivity to the Nax channel. CONCLUSIONS: The patients with serum immunoreactivity to the SFO had higher prolactin levels and hypothalamic disorders compared to those without the immunoreactivity. The clinical characteristics of patients with serum immunoreactivity to the subfornical organ included higher prolactin levels and hypothalamic disorders, which were frequently associated with central hypothyroidism and the presence of retroperitoneal tumors.

Child Neurology: Pathologically Confirmed Thrombotic Microangiopathy Caused by Onasemnogene Abeparvovec Treatment for SMA.

Onasemnogene abeparvovec is an adeno-associated virus vector-based gene therapy for spinal muscular atrophy (SMA). Although several cases of drug-induced thrombotic microangiopathy due to onasemnogene abeparvovec have been reported, none has been confirmed pathologically. Here, we present renal histopathologic findings of TMA due to onasemnogene abeparvovec. On day 5 after receiving onasemnogene abeparvovec, a 23-month-old girl with SMA type 1 developed thrombocytopenia, microangiopathic hemolytic anemia, liver dysfunction, acute kidney injury, and hypertension. She was diagnosed with TMA and received an increased dose of prednisolone, antihypertensives, diuretics, packed red blood cell and platelet transfusion, a single dose of eculizumab, 4 cycles of plasmapheresis, and intermittent and continuous hemodialysis. Her TMA resolved by day 30. On day 49, renal biopsy was performed. Light microscopy revealed proliferation of glomerular mesangial cells and matrix, with mesangiolysis, endothelial cell swelling, and partial double contours of the glomerular basement membrane. Electron microscopy showed endothelial injury, with edematous changes of the subendothelial spaces and neoformation of the basement membrane, without electron-dense depositions. These findings are compatible with the recovery phase of TMA. One year after drug administration, her motor function is improved. She can hold her posture against gravity and has neither dysphagia nor respiratory disturbance, but mild hypertension persists. Physicians should be vigilant regarding TMA as a severe side effect of onasemnogene abeparvovec treatment, especially when thrombocytopenia, hemolytic anemia, increased lactate dehydrogenase, or acute kidney injury is present.

Glomerular galactose-deficient IgA1 expression analysis in pediatric patients with glomerular diseases.

Galactose-deficient IgA1 (Gd-IgA1) is important in the pathogenesis of IgA nephropathy (IgAN). A Gd-IgA1-specific monoclonal antibody (KM55) has revealed glomerular Gd-IgA1 deposition solely in patients with IgAN and IgA vasculitis with nephritis (IgAV-N). However, this specificity is controversial and has not been demonstrated in pediatric patients. Here, we conducted double-immunofluorescence staining of IgA and Gd-IgA1 in 60 pediatric patients with various glomerular diseases. We divided patients into four groups: (1) patients with IgAN and IgAV-N (n = 23); (2) patients with immunocomplex-mediated glomerulonephritis accompanied by IgA deposition, including lupus nephritis, membranoproliferative glomerulonephritis, and membranous nephropathy (n = 14); (3) patients with other glomerular diseases involving IgA deposition, including idiopathic nephrotic syndrome (INS), oligomeganephronia, Alport syndrome, dense deposit disease, and crescentic glomerulonephritis (n = 11); and (4) patients with IgA-negative diseases including INS, membranoproliferative glomerulonephritis, membranous nephropathy, oligomeganephronia, Alport syndrome, C3 glomerulonephritis, poststreptococcal acute glomerulonephritis, and hemolytic uremic syndrome (n = 12). KM55 staining revealed Gd-IgA1-positive findings in 23/23 patients in Group 1 and 13/14 patients in Group 2, but not in patients in Groups 3 or 4. Therefore, KM55 may detect incidental IgA deposition in pediatric patients. Gd-IgA1 may be involved in the pathogenesis of these immune-related diseases; alternatively, KM55 may recognize IgA-related immunocomplexes in a non-specific manner.

Common risk variants in NPHS1 and TNFSF15 are associated with childhood steroid-sensitive nephrotic syndrome.

To understand the genetics of steroid-sensitive nephrotic syndrome (SSNS), we conducted a genome-wide association study in 987 childhood SSNS patients and 3,206 healthy controls with Japanese ancestry. Beyond known associations in the HLA-DR/DQ region, common variants in NPHS1-KIRREL2 (rs56117924, P=4.94E-20, odds ratio (OR) =1.90) and TNFSF15 (rs6478109, P=2.54E-8, OR=0.72) regions achieved genome-wide significance and were replicated in Korean, South Asian and African populations. Trans-ethnic meta-analyses including Japanese, Korean, South Asian, African, European, Hispanic and Maghrebian populations confirmed the significant associations of variants in NPHS1-KIRREL2 (Pmeta=6.71E-28, OR=1.88) and TNFSF15 (Pmeta=5.40E-11, OR=1.33) loci. Analysis of the NPHS1 risk alleles with glomerular NPHS1 mRNA expression from the same person revealed allele specific expression with significantly lower expression of the transcript derived from the risk haplotype (Wilcox test p=9.3E-4). Because rare pathogenic variants in NPHS1 cause congenital nephrotic syndrome of the Finnish type (CNSF), the present study provides further evidence that variation along the allele frequency spectrum in the same gene can cause or contribute to both a rare monogenic disease (CNSF) and a more complex, polygenic disease (SSNS).

A review of clinical characteristics and genetic backgrounds in Alport syndrome.

Alport syndrome (AS) is a progressive hereditary renal disease that is characterized by sensorineural hearing loss and ocular abnormalities. It is divided into three modes of inheritance, namely, X-linked Alport syndrome (XLAS), autosomal recessive AS (ARAS), and autosomal dominant AS (ADAS). XLAS is caused by pathogenic variants in COL4A5, while ADAS and ARAS are caused by those in COL4A3/COL4A4. Diagnosis is conventionally made pathologically, but recent advances in comprehensive genetic analysis have enabled genetic testing to be performed for the diagnosis of AS as first-line diagnosis. Because of these advances, substantial information about the genetics of AS has been obtained and the genetic background of this disease has been revealed, including genotype-phenotype correlations and mechanisms of onset in some male XLAS cases that lead to milder phenotypes of late-onset end-stage renal disease (ESRD). There is currently no radical therapy for AS and treatment is only performed to delay progression to ESRD using nephron-protective drugs. Angiotensin-converting enzyme inhibitors can remarkably delay the development of ESRD. Recently, some new drugs for this disease have entered clinical trials or been developed in laboratories. In this article, we review the diagnostic strategy, genotype-phenotype correlation, mechanisms of onset of milder phenotypes, and treatment of AS, among others.

Strong Association of the HLA-DR/DQ Locus with Childhood Steroid-Sensitive Nephrotic Syndrome in the Japanese Population.

Background Nephrotic syndrome is the most common cause of chronic glomerular disease in children. Most of these patients develop steroid-sensitive nephrotic syndrome (SSNS), but the loci conferring susceptibility to childhood SSNS are mainly unknown.Methods We conducted a genome-wide association study (GWAS) in the Japanese population; 224 patients with childhood SSNS and 419 adult healthy controls were genotyped using the Affymetrix Japonica Array in the discovery stage. Imputation for six HLA genes (HLA-A, -C, -B, -DRB1, -DQB1, and -DPB1) was conducted on the basis of Japanese-specific references. We performed genotyping for HLA-DRB1/-DQB1 using a sequence-specific oligonucleotide-probing method on a Luminex platform. Whole-genome imputation was conducted using a phased reference panel of 2049 healthy Japanese individuals. Replication was performed in an independent Japanese sample set including 216 patients and 719 healthy controls. We genotyped candidate single-nucleotide polymorphisms using the DigiTag2 assay.Results The most significant association was detected in the HLA-DR/DQ region and replicated (rs4642516 [minor allele G], combined Pallelic=7.84×10-23; odds ratio [OR], 0.33; 95% confidence interval [95% CI], 0.26 to 0.41; rs3134996 [minor allele A], combined Pallelic=1.72×10-25; OR, 0.29; 95% CI, 0.23 to 0.37). HLA-DRB1*08:02 (Pc=1.82×10-9; OR, 2.62; 95% CI, 1.94 to 3.54) and HLA-DQB1*06:04 (Pc=2.09×10-12; OR, 0.10; 95% CI, 0.05 to 0.21) were considered primary HLA alleles associated with childhood SSNS. HLA-DRB1*08:02-DQB1*03:02 (Pc=7.01×10-11; OR, 3.60; 95% CI, 2.46 to 5.29) was identified as the most significant genetic susceptibility factor.Conclusions The most significant association with childhood SSNS was detected in the HLA-DR/DQ region. Further HLA allele/haplotype analyses should enhance our understanding of molecular mechanisms underlying SSNS.

Detection of Splicing Abnormalities and Genotype-Phenotype Correlation in X-linked Alport Syndrome.

BACKGROUND: X-linked Alport syndrome (XLAS) is a progressive hereditary nephropathy caused by mutations in the COL4A5 gene. Genotype-phenotype correlation in male XLAS is relatively well established; relative to truncating mutations, nontruncating mutations exhibit milder phenotypes. However, transcript comparison between XLAS cases with splicing abnormalities that result in a premature stop codon and those with nontruncating splicing abnormalities has not been reported, mainly because transcript analysis is not routinely conducted in patients with XLAS. METHODS: We examined transcript expression for all patients with suspected splicing abnormalities who were treated at one hospital between January of 2006 and July of 2017. Additionally, we recruited 46 males from 29 families with splicing abnormalities to examine genotype-phenotype correlation in patients with truncating (n=21, from 14 families) and nontruncating (n=25, from 15 families) mutations at the transcript level. RESULTS: We detected 41 XLAS families with abnormal splicing patterns and described novel XLAS atypical splicing patterns (n=14) other than exon skipping caused by point mutations in the splice consensus sequence. The median age for developing ESRD was 20 years (95% confidence interval, 14 to 23 years) among patients with truncating mutations and 29 years (95% confidence interval, 25 to 40 years) among patients with nontruncating mutations (P=0.001). CONCLUSIONS: We report unpredictable atypical splicing in the COL4A5 gene in male patients with XLAS and reveal that renal prognosis differs significantly for patients with truncating versus nontruncating splicing abnormalities. Our results suggest that splicing modulation should be explored as a therapy for XLAS with truncating mutations.

Functional splicing analysis in an infantile case of atypical hemolytic uremic syndrome caused by digenic mutations in C3 and MCP genes.

Pathogenic variants in specific complement-related genes lead to atypical hemolytic uremic syndrome (aHUS). Some reports have indicated that patients with digenic variants in these genes might present severer phenotypes. Upon detecting novel intronic variants, transcriptional analysis is necessary to prove pathogenicity; however, when intronic variants are located in intron 1 and, as a result, no transcript is produced, no appropriate method had been established to reveal the pathogenicity. Recently, the minigene assay was used to assess the pathogenicity of intronic variants. Here, we report an infantile case of aHUS caused by digenic mutations in two different complement-related genes, C3 and MCP. Targeted sequencing detected a known variant in C3 and a novel variant in the intron 1 splicing donor site of MCP. To assess the pathogenicity of this intronic variant, we conducted functional splicing assay using a minigene construct and quantitative PCR analysis of the MCP transcript, revealing the pathogenicity of the intronic variant. In conclusion, the minigene assay revealed the pathogenicity of the intron 1 splicing donor site variant for the first time. This case showed a severe phenotype of infantile-onset aHUS associated with digenic variants in two complement-related genes.

Clinical features in a series of 258 Japanese pediatric patients with thrombotic microangiopathy.

BACKGROUND: Thrombotic microangiopathy (TMA) includes hemolytic uremic syndrome (HUS) and thrombotic thrombocytopenic purpura (TTP). This study examined the epidemiological characteristics of pediatric patients with TMA classified according to etiology. METHODS: The survey evaluated 258 Japanese pediatric patients diagnosed with TMA between 2012 and 2015. RESULTS: The primary diseases responsible for TMA were categorized as TTP (15 cases), Shiga toxin-producing Escherichia coli-associated HUS (STEC-HUS) (166 cases), atypical HUS (aHUS) (40 cases), and secondary TMA (27 cases). The remaining 10 cases were unable to be classified to one of the four categories of the primary disease. Renal replacement therapy was required in the acute phase in 103 patients with TMA, including 65 with STEC-HUS, 22 with aHUS, two with TTP, 10 with secondary TMA, and four unclassified cases. The last observational findings were normal renal function in 95 patients and chronic kidney disease (CKD) stage 1 in 62. For 31 patients, chronic renal insufficiency (CKD stage 2-5) persisted, including four patients with end-stage kidney disease (CKD stage 5). Seventeen patients suffered recurrence of TMA, and eight patients died. CONCLUSION: This study clarified differences in the relative proportions of primary diseases between patients from Japan and North America and Europe. The difference may be attributable to the lower estimated incidence of STEC-HUS in Japan.

An in vitro splicing assay reveals the pathogenicity of a novel intronic variant in ATP6V0A4 for autosomal recessive distal renal tubular acidosis.

BACKGROUND: Autosomal recessive distal renal tubular acidosis (dRTA) is a rare hereditary disease caused by pathogenic variants in the ATP6V0A4 gene or ATP6V1B1 gene, and characterized by hyperchloremic metabolic acidosis with normal anion gap, hypokalemia, hypercalciuria, hypocitraturia and nephrocalcinosis. Although several intronic nucleotide variants in these genes have been detected, all of them fell in the apparent splice consensus sequence. In general, transcriptional analysis is necessary to determine the effect on function of the novel intronic variants located out of splicing consensus sequences. In recent years, functional splicing analysis using minigene construction was used to assess the pathogenicity of novel intoronic variant in various field. METHODS: We investigated a sporadic case of dRTA with a compound heterozygous mutation in the ATP6V0A4 gene, revealed by next generation sequencing. One variant was already reported as pathogenic; however, the other was a novel variant in intron 11 (c.1029 + 5G > A) falling outside of the apparent splicing consensus sequence. Expression of ATP6V0A4 was not detected in peripheral leukocytes by RT-PCR analysis. Therefore, an in vitro functional splicing study using minigene construction was conducted to analyze the splicing pattern of the novel variant. RESULTS: A minigene assay revealed that the novel intronic variant leads to a 104 bp insertion immediately following exon 11. In addition, this result was confirmed using RNA extracted from the patient's cultured leukocytes. CONCLUSION: These results proved the pathogenicity of a novel intronic variant in our patient. We concluded that the minigene assay is a useful, non-invasive method for functional splicing analysis of inherited kidney disease, even if standard transcriptional analysis could not detect abnormal mRNA.

Clinical guides for atypical hemolytic uremic syndrome in Japan.

Atypical hemolytic uremic syndrome (aHUS) is a rare disease characterized by the triad of microangiopathic hemolytic anemia, thrombocytopenia, and acute kidney injury. In 2013, we developed diagnostic criteria to enable early diagnosis and timely initiation of appropriate treatment for aHUS. Recent clinical and molecular findings have resulted in several proposed classifications and definitions of thrombotic microangiopathy and aHUS. Based on recent advances in this field and the emerging international consensus to exclude secondary TMAs from the definition of aHUS, we have redefined aHUS and proposed diagnostic algorithms, differential diagnosis, and therapeutic strategies for aHUS.

Clinical guides for atypical hemolytic uremic syndrome in Japan.

Atypical hemolytic uremic syndrome (aHUS) is a rare disease characterized by the triad of microangiopathic hemolytic anemia, thrombocytopenia, and acute kidney injury. In 2013, we developed diagnostic criteria to enable early diagnosis and timely initiation of appropriate treatment for aHUS. Recent clinical and molecular findings have resulted in several proposed classifications and definitions of thrombotic microangiopathy and aHUS. Based on recent advances in this field and the emerging international consensus to exclude secondary TMAs from the definition of aHUS, we have redefined aHUS and proposed diagnostic algorithms, differential diagnosis, and therapeutic strategies for aHUS.

Focal form of congenital hyperinsulinism clearly detectable by contrast-enhanced computed tomography imaging.

The focal form of congenital hyperinsulinism (CHI) is characterized by a cluster of abnormal insulin-oversecreting ß cells within a restricted area of the pancreas. Although identification of the focal lesion is very important in the management of CHI, it has been reported that imaging studies, including computed tomography (CT), magnetic resonance imaging (MRI) scans, or angiography, are not helpful in identifying the focal lesion. Currently, fluorine-18-L-dihydroxyphenylalanine positron emission tomography ((18)F-DOPA PET) is believed to be the only imaging modality that can identify the focal lesions. In this report, however, we present a case of a 7-month-old girl with the focal form of CHI, caused by a loss-of-function mutation in the ABCC8 gene, whose lesion was clearly visible as a hyperenhancing nodule on contrast-enhanced CT and dynamic MRI imaging.

Diagnostic criteria for atypical hemolytic uremic syndrome proposed by the Joint Committee of the Japanese Society of Nephrology and the Japan Pediatric Society.

Atypical hemolytic uremic syndrome (aHUS) is rare and comprises the triad of microangiopathic hemolytic anemia, thrombocytopenia, and acute kidney injury. Recently, abnormalities in the mechanisms underlying complement regulation have been focused upon as causes of aHUS. The prognosis for patients who present with aHUS is very poor, with the first aHUS attack being associated with a mortality rate of approximately 25%, and with approximately 50% of cases resulting in end-stage renal disease requiring dialysis. If treatment is delayed, there is a high risk of this syndrome progressing to renal failure. Therefore, we have developed diagnostic criteria for aHUS to enable its early diagnosis and to facilitate the timely initiation of appropriate treatment. We hope these diagnostic criteria will be disseminated to as many clinicians as possible and that they will be used widely.

Relapsing 6q24-related transient neonatal diabetes mellitus successfully treated with a dipeptidyl peptidase-4 inhibitor: a case report.

The most common form of transient neonatal diabetes mellitus (TNDM) is 6q24-related TNDM. Patients are treated with insulin during the neonatal period until spontaneous remission. However, diabetes often recurs in adolescence, and there is no standard therapy for patients with a relapse. A paternal duplication at the 6q24 critical region spanning the pleiomorphic adenoma gene-like 1 PLAGL1 gene was found in a Japanese patient with TNDM relapse. The patient was treated with a dipeptidyl peptidase-4 (DPP4) inhibitor, alogliptin, at a dose of 25 mg per day. Immediately after treatment initiation, his hemoglobin A1c (HbA1c) levels dropped from 7.0-7.5% (52-58 mmol/mol) to 6.0-6.5% (41-47 mmol/mol) and remained stable for over a year. We reported the successful treatment of relapsed 6q24-related TNDM with a DPP4 inhibitor. Although insulin has been the conventional treatment for such patients, treatments targeting the GLP1 pathway can be a useful alternative because these patients retain the ß cell mass and responsiveness through G protein-coupled pathways.

Diagnostic criteria for atypical hemolytic uremic syndrome proposed by the Joint Committee of the Japanese Society of Nephrology and the Japan Pediatric Society.

Atypical hemolytic uremic syndrome (aHUS) is rare and comprises the triad of microangiopathic hemolytic anemia, thrombocytopenia, and acute kidney injury. Recently, abnormalities in the mechanisms underlying complement regulation have been focused upon as causes of aHUS. The prognosis for patients who present with aHUS is very poor, with the first aHUS attack being associated with a mortality rate of ~25 %, and with ~50 % of cases resulting in end-stage renal disease requiring dialysis. If treatment is delayed, there is a high risk of this syndrome progressing to renal failure. Therefore, we have developed diagnostic criteria for aHUS to enable its early diagnosis and to facilitate the timely initiation of appropriate treatment. We hope these diagnostic criteria will be disseminated to as many clinicians as possible and that they will be used widely.

Childhood pheochromocytoma in a survivor of central primitive neuroectodermal tumor.

Pheochromocytoma and central nervous system primitive neuroectodermal tumor are both neural crest-derived tumors. The former is usually benign and develops mainly in adulthood and the latter brain tumor mainly occurs in childhood and has a poor prognosis. We report a case of a 15-year-old boy who developed pheochromocytoma after more than 10 years of complete remission of central primitive neuroectodermal tumor. Thus far, there have been no reports of childhood cancer survivors who developed pheochromocytoma. This quite rare occurrence of two tumors in a single patient may imply some unidentified linkage or common genetic background.

Analysis of genetic and predisposing factors in Japanese patients with atypical hemolytic uremic syndrome.

Hemolytic uremic syndrome (HUS) is characterized by microangiopathic hemolytic anemia, thrombocytopenia, and renal impairment. Approximately 10% of cases are classified as atypical due to the absence of Shiga toxin-producing bacteria as a trigger. Uncontrolled activation of the complement system plays a role in the pathogenesis of atypical HUS (aHUS). Although many genetic studies on aHUS have been published in recent years, only limited data has been gathered in Asian countries. We analyzed the genetic variants of 6 candidate genes and the gene deletion in complement factor H (CFH) and CFH-related genes, examined the prevalence of CFH autoantibodies and evaluated the genotype-phenotype relationship in 10 Japanese patients with aHUS. We identified 7 causative or potentially causative mutations in CFH (p.R1215Q), C3 (p.R425C, p.S562L, and p.I1157T), membrane cofactor protein (p.Y189D and p.A359V) and thrombomodulin (p.T500M) in 8 out of 10 patients. All 7 of the mutations were heterozygous and four of them were novel. Two patients carried CFH p.R1215Q and 3 other patients carried C3 p.I1157T. One patient had 2 causative mutations in different genes. One patient was a compound heterozygote of the 2 MCP mutations. The patients carrying mutations in CFH or C3 had a high frequency of relapse and a worse prognosis. One patient had CFH autoantibodies. The present study identified the cause of aHUS in 9 out of 10 Japanese patients. Since the phenotype-genotype correlation of aHUS has clinical significance in predicting renal recovery and transplant outcome, a comprehensively accurate assessment of molecular variation would be necessary for the proper management of aHUS patients in Japan.