Rituximab in combination with cyclosporine and steroid pulse therapy for childhood-onset multidrug-resistant nephrotic syndrome: a multicenter single-arm clinical trial (JSKDC11 trial).

BACKGROUND: Only 80% of children with idiopathic nephrotic syndrome respond well to glucocorticoid therapy. Multidrug-resistant nephrotic syndrome (MRNS) is associated with a poor kidney prognosis. Several retrospective studies have identified rituximab as an effective treatment for MRNS; however, prospective studies are required to assess its efficacy and safety. METHODS: We conducted a multicenter, non-blinded, single-arm trial to investigate the efficacy and safety of rituximab in patients with childhood-onset MRNS who were resistant to cyclosporine and more than three courses of steroid pulse therapy. The enrolled patients received four 375 mg/m2 doses of rituximab in combination with baseline cyclosporine and steroid pulse therapy. The primary endpoint was a > 50% reduction in the urinary protein/creatinine ratio from baseline on day 169. Complete and partial remissions were also evaluated. RESULTS: Six patients with childhood-onset MRNS were enrolled. All patients were negative for pathogenic variants of podocyte-related genes. On day 169, five patients (83.3%) showed a > 50% reduction in the urinary protein/creatinine ratio, two patients showed partial remission, and two patients showed complete remission. No deaths occurred and severe adverse events occurred in two patients (infection in one patient and acute kidney injury in one patient). Three patients needed treatment for moderate-to-severe infection. CONCLUSIONS: The study treatment effectively reduced the urinary protein/creatinine ratio in patients with childhood-onset MRNS. The adverse events in this study were within the expected range; however, attention should be paid to the occurrence of infections.

Middle-term natural course after acute pancreatitis associated with pancreas divisum.

Invasive treatment might be taken into consideration at the time of initial acute pancreatitis in patients with pancreas divisum who do not complain of abdominal pain to prevent progression to chronic pancreatitis.

Hyperammonemia in a carbamoyl-phosphate synthetase 1 deficiency recipient after living-donor liver transplantation from a carrier donor: a case report.

Carbamoyl-phosphate synthetase 1 (CPS1) deficiency is an autosomal recessive congenital urea cycle disorder (UCD) characterized by hyperammonemia. The recipients of liver transplantation (LT) for UCD are often children, and the potential donors are often the parents. Hereditary congenital diseases involving UCD entail the possibility of both parents being genetically heterozygous. Herein, we describe the case of a 12-year-old girl with CPS1 deficiency receiving a liver transplant (soon after birth) from her father, who had a heterozygous CPS1 mutation. She was referred to our hospital with respiratory distress after contracting two infections (respiratory syncytial virus and human metapneumovirus) within a short period, both of which presented with hyperammonemia. Medication for hyperammonemia quickly lowered the ammonia levels. The hyperammonemia was thought to be caused by the heterozygous mutation in the donor liver; moreover, it is likely that the low enzyme activity in the patient's liver was increased due to the infections. This is the first study to report hyperammonemia in a CPS1 deficiency patient due to an infection after LT. Thus, patients with CPS1 deficiency should be aware of the development of hyperammonemia after LT.

Identification of novel OCRL isoforms associated with phenotypic differences between Dent disease-2 and Lowe syndrome.

BACKGROUND: Although Lowe syndrome and Dent disease-2 are caused by Oculocerebrorenal syndrome of Lowe (OCRL) mutations, their clinical severities differ substantially and their molecular mechanisms remain unclear. Truncating mutations in OCRL exons 1-7 lead to Dent disease-2, whereas those in exons 8-24 lead to Lowe syndrome. Herein we identified the mechanism underlying the action of novel OCRL protein isoforms. METHODS: Messenger RNA samples extracted from cultured urine-derived cells from a healthy control and a Dent disease-2 patient were examined to detect the 5' end of the OCRL isoform. For protein expression and functional analysis, vectors containing the full-length OCRL transcripts, the isoform transcripts and transcripts with truncating mutations detected in Lowe syndrome and Dent disease-2 patients were transfected into HeLa cells. RESULTS: We successfully cloned the novel isoform transcripts from OCRL exons 6-24, including the translation-initiation codons present in exon 8. In vitro protein-expression analysis detected proteins of two different sizes (105 and 80 kDa) translated from full-length OCRL, whereas only one protein (80 kDa) was found from the isoform and Dent disease-2 variants. No protein expression was observed for the Lowe syndrome variants. The isoform enzyme activity was equivalent to that of full-length OCRL; the Dent disease-2 variants retained >50% enzyme activity, whereas the Lowe syndrome variants retained <20% activity. CONCLUSIONS: We elucidated the molecular mechanism underlying the two different phenotypes in OCRL-related diseases; the functional OCRL isoform translated starting at exon 8 was associated with this mechanism.

Association between Crohn's disease and AarF domain-containing kinase 4 glomerulopathy.

Coenzyme Q10 deficiency causing gastrointestinal symptoms has not been reported. At least 15 genes are involved in CoQ10 biosynthesis, and one of the genes is AarF domain-containing kinase 4 (ADCK4). This case report first showed a patient who presented with Crohn's disease (CD) combined with ADCK4 glomerulopathy (ADCK4-GN). After approximately 2 years of infliximab treatment for CD, this patient has remained in clinical remission with no adverse effects. Moreover, two important findings were obtained: first, individuals with ADCK4-GN may present with complications, such as CD, which is a gastrointestinal disease, and infliximab may be effective for CD associated with ADCK4-GN. Second, infliximab may also be effective and safe for individuals with CD who are undergoing peritoneal dialysis. Thus, the results of the present study may be used in managing patients with such condition, and as a result, better clinical outcomes will be obtained.

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.

Chronic Stimulation of Renin Cells Leads to Vascular Pathology.

Experimental or spontaneous genomic mutations of the renin-angiotensin system or its pharmacological inhibition in early life leads to renal abnormalities, including poorly developed renal medulla, papillary atrophy, hydronephrosis, inability to concentrate the urine, polyuria, polydipsia, renal failure, and anemia. At the core of such complex phenotype is the presence of unique vascular abnormalities: the renal arterioles do not branch or elongate properly and they have disorganized, concentric hypertrophy. This lesion has been puzzling because it is often found in hypertensive individuals whereas mutant or pharmacologically inhibited animals are hypotensive. Remarkably, when renin cells are ablated with diphtheria toxin, the vascular hypertrophy does not occur, suggesting that renin cells per se may contribute to the vascular disease. To test this hypothesis, on a Ren1c-/- background, we generated mutant mice with reporter expression (Ren1c-/-;Ren1c-Cre;R26R.mTmG and Ren1c-/-;Ren1c-Cre;R26R.LacZ) to trace the fate of reninnull cells. To assess whether reninnull cells maintain their renin promoter active, we used Ren1c-/-;Ren1c-YFP mice that transcribe YFP (yellow fluorescent protein) directed by the renin promoter. We also followed the expression of Akr1b7 and miR-330-5p, markers of cells programmed for the renin phenotype. Contrary to what we expected, reninnull cells did not die or disappear. Instead, they survived, increased in number along the renal arterial tree, and maintained an active molecular memory of the myoepitheliod renin phenotype. Furthermore, null cells of the renin lineage occupied the walls of the arteries and arterioles in a chaotic, directionless pattern directly contributing to the concentric arterial hypertrophy.

Characterization of contiguous gene deletions in COL4A6 and COL4A5 in Alport syndrome-diffuse leiomyomatosis.

Alport syndrome-diffuse leiomyomatosis (AS-DL, OMIM: 308940) is a rare variant of the X-linked Alport syndrome that shows overgrowth of visceral smooth muscles in the gastrointestinal, respiratory and female reproductive tracts in addition to renal symptoms. AS-DL results from deletions that encompass the 5' ends of the COL4A5 and COL4A6 genes, but deletion breakpoints between COL4A5 and COL4A6 have been determined in only four cases. Here, we characterize deletion breakpoints in five AS-DL patients and show a contiguous COL4A6/COL4A5 deletion in each case. We also demonstrate that eight out of nine deletion alleles involved sequences homologous between COL4A5 and COL4A6. Most breakpoints took place in recognizable transposed elements, including long and short interspersed repeats, DNA transposons and long-terminal repeat retrotransposons. Because deletions involved the bidirectional promoter region in each case, we suggest that the occurrence of leiomyomatosis in AS-DL requires inactivation of both genes. Altogether, our study highlights the importance of homologous recombination involving multiple transposed elements for the development of this continuous gene syndrome and other atypical loss-of-function phenotypes.

Natural history of genetically proven autosomal recessive Alport syndrome.

BACKGROUND: Autosomal recessive Alport syndrome (ARAS) is a rare hereditary disease caused by homozygous or compound heterozygous mutations in either the COL4A3 or COL4A4 genes. Failure to diagnose ARAS cases is common, even if detailed clinical and pathological examinations are carried out. As the mutation detection rate for ARAS is unsatisfactory, we sought to develop more reliable diagnostic methods and provide a better description of the clinicopathological characteristics of this disorder. METHODS: A retrospective analysis of 30 genetically diagnosed patients with ARAS in 24 pedigrees was conducted. The mutation detection strategy comprised three steps: (1) genomic DNA analysis using polymerase chain reaction (PCR) and direct sequencing; (2) mRNA analysis using reverse transcription (RT)-PCR to detect RNA processing abnormalities; (3) semi-quantitative PCR using capillary electrophoresis to detect large heterozygous deletions. RESULTS: Using the three-step analysis, we identified homozygous or compound heterozygous mutations in all patients. Interestingly, 20% of our ARAS patients showed normal expression of α5 in kidney tissue. The median age of developing end-stage renal disease was 21 years. CONCLUSIONS: The strategy described in this study improves the diagnosis for ARAS families. Although immunohistochemical analysis of α5 can provide diagnostic information, normal distribution does not exclude the diagnosis of ARAS.

Milder clinical aspects of X-linked Alport syndrome in men positive for the collagen IV α5 chain.

X-linked Alport syndrome is caused by mutations in the COL4A5 gene encoding the type IV collagen α5 chain (α5(IV)). Complete absence of α5(IV) in the renal basal membrane is considered a pathological characteristic in male patients; however, positive α5(IV) staining has been found in over 20% of patients. We retrospectively studied 52 genetically diagnosed male X-linked Alport syndrome patients to evaluate differences in clinical characteristics and renal outcomes between 15 α5(IV)-positive and 37 α5(IV)-negative patients. Thirteen patients in the α5(IV)-positive group had non-truncating mutations consisting of nine missense mutations, three in-frame deletions, and one splice-site mutation resulting in small in-frame deletions of transcripts. The remaining two showed somatic mutations with mosaicism. Missense mutations in the α5(IV)-positive group were more likely to be located before exon 25 compared with missense mutations in the α5(IV)-negative group. Furthermore, urinary protein levels were significantly lower and the age at onset of end-stage renal disease was significantly higher in the positive group than in the negative group. These results help to clarify the milder clinical manifestations and molecular characteristics of male X-linked Alport syndrome patients expressing the α5(IV) chain.

Expression of prokineticin receptors in mouse cultured astrocytes and involvement in cell proliferation.

Effects of prokineticins (PKs), a novel family of bioactive peptides with a mitogenic action to endothelial cells of the endocrine gland and testis, on astrocytic functions were examined. Mouse cultured astrocytes expressed PK-R1 type PK receptors, while there was little expression of the PK-R2 type. PKs caused increases in astrocytic cytosolic Ca2+ levels and BrdU incorporation. Increases in Ca2+ levels by PK-2 were diminished by U73122 (a phospholipase C inhibitor). PK-induced BrdU incorporation was inhibited by U73122, GF109203 (a protein kinase C inhibitor) and PD98059 (a MEK/ERK inhibitor). These results indicate that PK receptors are expressed in astrocytes and regulate astrocytic proliferation.