Biallelic loss-of-function ZFYVE19 mutations are associated with congenital hepatic fibrosis, sclerosing cholangiopathy and high-GGT cholestasis.

BACKGROUND: For many children with intrahepatic cholestasis and high-serum gamma-glutamyl transferase (GGT) activity, a genetic aetiology of hepatobiliary disease remains undefined. We sought to identify novel genes mutated in children with idiopathic high-GGT intrahepatic cholestasis, with clinical, histopathological and functional correlations. METHODS: We assembled a cohort of 25 children with undiagnosed high-GGT cholestasis and without clinical features of biliary-tract infection or radiological features of choledochal malformation, sclerosing cholangitis or cholelithiasis. Mutations were identified through whole-exome sequencing and targeted Sanger sequencing. We reviewed histopathological findings and assessed phenotypical effects of ZFYVE19 deficiency in cultured cells by immunofluorescence microscopy. RESULTS: Nine Han Chinese children harboured biallelic, predictedly complete loss-of-function pathogenic mutations in ZFYVE19 (c.314C>G, p.S105X; c.379C>T, p.Q127X; c.514C>T, p.R172X; c.547C>T, p.R183X; c.226A>G, p.M76V). All had portal hypertension and, at liver biopsy, histopathological features of the ductal plate malformation (DPM)/congenital hepatic fibrosis (CHF). Four children required liver transplantation for recurrent gastrointestinal haemorrhage. DPM/CHF was confirmed at hepatectomy, with sclerosing small-duct cholangitis. Immunostaining for two primary-cilium axonemal proteins found expression that was deficient intraluminally and ectopic within cholangiocyte cytoplasm. ZFYVE19 depletion in cultured cells yielded abnormalities of centriole and axoneme. CONCLUSION: Biallelic ZFYVE19 mutations can lead to high-GGT cholestasis and DPM/CHF in vivo. In vitro, they can lead to centriolar and axonemal abnormalities. These observations indicate that mutation in ZFYVE19 results, through as yet undefined mechanisms, in a ciliopathy.

Successful treatment of infantile oxysterol 7α-hydroxylase deficiency with oral chenodeoxycholic acid.

BACKGROUND: Deficiency of oxysterol 7α-hydroxylase, encoded by CYP7B1, is associated with fatal infantile progressive intrahepatic cholestasis and hereditary spastic paraplegia type 5. Most reported patients with CYP7B1 mutations presenting with liver disease in infancy have died of liver failure. However, it was recently reported that two patients treated with chenodeoxycholic acid survived. Correlations between the phenotype and genotype of CYP7B1 deficiency have not been clearly established. CASE PRESENTATION: A 5-month-7-day-old Chinese baby from non-consanguineous parents was referred for progressive cholestasis and prolonged prothrombin time from one month of age. Genetic testing revealed compound heterozygous mutations c.187C > T(p.R63X)/c.334C > T(p.R112X) in CYP7B1, and fast atom bombardment mass spectrometry analysis of the urinary bile acid confirmed the presence of atypical hepatotoxic 3ß-hydroxy-Δ5-bile acids. While awaiting liver transplantation she was orally administered chenodeoxycholic acid. Her liver function rapidly improved, urine atypical bile acids normalized, and she thrived well until the last follow-up at 23 months of age. Her 15-year-old brother, with no history of infantile cholestasis but harboring the same mutations in CYP7B1, had gait abnormality from 13 years of age. Neurological examination revealed hyper-reflexia and spasticity of the lower limbs. Brain MRI revealed enlarged perivascular space in the bilateral basal ganglia and white matter of frontal parietal. CONCLUSIONS: In summary, these findings highlight that the phenotype of CYP7B1 deficiency varies widely, even in siblings and that early administration of chenodeoxycholic acid may improve prognosis.

TJP2 hepatobiliary disorders: Novel variants and clinical diversity.

To assess the spectrum of pediatric clinical phenotypes in TJP2 disease, we reviewed records of our seven patients in whom intrahepatic cholestasis was associated with biallelic TJP2 variants (13; 12 novel) and correlated clinical manifestations with mutation type. The effect of a splicing variant was analyzed with a minigene assay. The effects of three missense variants were analyzed with protein expression in vitro. Our patients had both remitting and persistent cholestasis. Three exhibited growth retardation. Six responded to treatment with cholestyramine, ursodeoxycholic acid, or both. Two had cholecystolithiasis. None required liver transplantation or developed hepatocellular or cholangiocellular malignancy. None manifested extrahepatic disease not attributable to effects of cholestasis. The variant c.2180-5T>G resulted in exon 15 skipping with in-frame deletion of 32 amino acid residues in TJP2. The three missense variants decreased but did not abolish TJP2 expression. Patients with truncating or canonical splice-site variants had clinically more severe disease. TJP2 disease in children includes a full clinical spectrum of severity, with mild or intermittent forms as well as the severe and minimal forms hitherto described. Biallelic TJP2 variants must be considered in children with clinically intermittent or resolved intrahepatic cholestasis.

Low-GGT intrahepatic cholestasis associated with biallelic USP53 variants: Clinical, histological and ultrastructural characterization.

BACKGROUND & AIMS: In about 20% of children with cholestasis and normal or low serum gamma-glutamyltransferase (GGT) activity, no aetiology is identified. We sought new genes implicated in paediatric hepatobiliary disease. METHODS: We conducted whole-exome sequencing in 69 children evaluated at our centre from 2011 to 2018 who had low-GGT cholestasis and in whom homozygous/compound heterozygous predictedly pathogenic variants (PPVs) in ATP8B1, ABCB11, NR1H4, MYO5B or TJP2 were not found. Clinical records and findings on light microscopy and transmission electron microscopy of liver biopsy materials were reviewed. RESULTS: In seven patients from seven unrelated families, biallelic PPVs (10 in total) were found in USP53, recently associated with intrahepatic cholestasis. Seven variants were classified as pathogenic: one canonical splicing, c.569 + 2T > C, and six nonsense or frameshifting: c.169C > T (p.Arg57Ter), c.581delA (p.Arg195GlufsTer38), c.831_832insAG (p.Val279GlufsTer16), c.1012C > T (p.Arg338Ter), c.1426C > T (p.Arg476Ter) and c.1558C > T (p.Arg520Ter). Three were likely pathogenic: c.297G > T (p.Arg99Ser), c.395A > G (p.His132Arg) and c.878G > T (p.Gly293Val). In all patients, jaundice began at age <7 months. Cholestasis was transient, with documented resolution of hyperbilirubinaemia in all (oldest patient now aged 5 years) except one, who was lost to follow-up. Light microscopy identified intralobular cholestasis, giant-cell change of hepatocytes and perisinusoidal-perihepatocytic and portal-tract fibrosis. Ultrastructural study revealed elongated hepatocyte-hepatocyte tight junctions. One patient was deaf. CONCLUSION: USP53 interacts with the tight junction constituent TJP2. TJP2 mutation can cause low-GGT intrahepatic cholestasis, with elongated hepatocyte-hepatocyte tight junctions, as well as deafness. Our findings extend a preliminary report of USP53 disease and indicate that USP53 mutation may generate a partial phenocopy of TJP2 disease.

Molecular findings in children with inherited intrahepatic cholestasis.

BACKGROUND: Genetic defects account for a substantial proportion of pediatric cholestasis. This study explored the molecular findings in a large cohort of Chinese patients with inherited cholestasis. METHODS: Between January 2012 and June 2016, 809 Chinese pediatric patients with suspected inherited intrahepatic cholestasis were evaluated by Sanger sequencing and/or panel sequencing. RESULTS: Of the 809 patients, 273 (33.7%) obtained a genetic diagnosis. The rate of positive genetic diagnosis in patients with disease onset at 0-3 month of age was higher than that in patients with disease onset at 4 month of age or later. There were 17 distinct genetic defects diagnosed. The top 4 resulted from mutations in SLC25A13 (44.3%), JAG1 (24.5%), ABCB11 (11.0%), and ATP8B1 (5.9%). All 17 genetic disorders were diagnosed in patients with disease onset at 0-3 months of age; but only 5 were diagnosed in patients with disease onset beyond 4 months of age. A total of 217 distinct pathogenic variants, including 41 novel variants, were identified. Ten recurrent mutations were detected in SLC25A13, ATP8B1, and CYP27A1. They accounted for 48.2% of the total 477 mutant alleles. CONCLUSIONS: There were 17 distinct genetic disorders diagnosed in Chinese pediatric patients with inherited cholestasis.

Abnormal Bilirubin Metabolism in Patients With Sodium Taurocholate Cotransporting Polypeptide Deficiency.

OBJECTIVES: The aim of the study was to explore the significance of sodium taurocholate cotransporting polypeptide (NTCP) deficiency and its clinical features in Chinese children presenting with isolated persistent hypercholanemia. METHODS: The exon and adjacent regions of SLC10A1, the gene encoding NTCP, were sequenced in 33 Chinese children presenting with isolated hypercholanemia. Clinical history and medical data were reviewed. Growth milestones were compared with the national standard. The serum direct bilirubin concentration at last follow-up was compared with age- and sex-matched controls. RESULTS: A variant, c.800C>T, p. S267F of SLC10A1 was detected in all subjects; 30 patients were homozygotes and 3 were compound heterozygotes. Nine patients presented with transient neonatal cholestasis, and 1 with a persistent mild conjugated hyperbilirubinemia. The serum direct bilirubin level in NTCP-deficient patients was significantly higher than age- and sex-matched controls even after the neonatal cholestasis stage (2.85 ±â€Š1.50 vs 1.49 ±â€Š0.70 µmol/L, P = 0.00008). No growth delay or other severe long-term clinical consequences were observed. CONCLUSIONS: NTCP deficiency is the exclusive or major cause of isolated hypercholanemia in Han Chinese children, with c.800C>T the major contributing genetic variation. The defect may affect bilirubin metabolism and present as transient neonatal cholestasis and/or persistent mild conjugated hyperbilirubinmia, but with no apparent long-term clinical consequences.

Novel missense mutation in VPS33B is associated with isolated low gamma-glutamyltransferase cholestasis: Attenuated, incomplete phenotype of arthrogryposis, renal dysfunction, and cholestasis syndrome.

The typical phenotype of arthrogryposis, renal dysfunction, and cholestasis (ARC) syndrome involves three cardinal symptoms as the name describes, harboring biallelic mutations on VPS33B or VIPAS39. Except for ARC syndrome, low gamma-glutamyltransferase (GGT) cholestasis often implies hereditary hepatopathy of different severity; however, some remain undiagnosed. Several monogenic defects typically with multiorgan manifestations may only present liver dysfunction at times, such as DGUOK defect and AGL defect. Previously, four VPS33B mutated cases were reported without arthrogryposis, or with less severe symptoms and longer lifespan, indicating the possibility of incomplete ARC phenotype of isolated hepatopathy. So we retrospectively reviewed all patients with confirmed VPS33B/VIPARS39 defect in our center and identified three presenting isolated low-GGT cholestasis with intractable pruritus. Distinguished from others with typical ARC phenotype, these patients did not suffer the other two typical characteristics, survived much longer, and shared a novel missense VPS33B variation c.1726T>C, p.Cys576Arg, causing declined protein expression and abolished interaction with VIPAS39 in-vitro. Serum bile acid profiles of our VPS33B/VIPAS39 mutated patients revealed similar changes to primary defect of bile salt export pump, among which those with isolated cholestasis phenotype had a higher level of total secondary bile acids than that with typical ARC phenotype, indicating the partial residual function of VPS33B.

Anemia following zinc treatment for Wilson's disease: a case report and literature review.

BACKGROUND: Zinc therapy is considered an effective and safe treatment for Wilson's disease. Hypocupremia-related anemia is rarely reported after long-term zinc administration or combination therapy with copper-chelating agent. CASE PRESENTATION: We herein report a 12-year-old girl with pre-symptomatic Wilson's disease diagnosed 5 years ago who presented with severe anemia after high-dose oral zinc for 4 years and 4 months. Her hemoglobin was gradually restored to the normal range after the adjustment of zinc dose and diet therapy for 4 months. A review of the literature revealed eight patients with hypocupremia-associated anemia following zinc therapy for Wilson's disease, including 7 adults and 1 child. The only child patient was a 16-year-old boy, in whom the zinc therapy was succession to penicillamine administration. CONCLUSIONS: This is the first report worldwide that a child developed severe anemia following high-dose single zinc administration for Wilson's disease. It highlights the importance of regular follow-up during zinc treatment and the involvement of specialists in the long-term management of Wilson's disease. We hope that this will alert pediatricians the issue of zinc over-treatment.

Defects in myosin VB are associated with a spectrum of previously undiagnosed low γ-glutamyltransferase cholestasis.

Hereditary cholestasis in childhood and infancy with normal serum gamma-glutamyltransferase (GGT) activity is linked to several genes. Many patients, however, remain genetically undiagnosed. Defects in myosin VB (MYO5B; encoded by MYO5B) cause microvillus inclusion disease (MVID; MIM251850) with recurrent watery diarrhea. Cholestasis, reported as an atypical presentation in MVID, has been considered a side effect of parenteral alimentation. Here, however, we report on 10 patients who experienced cholestasis associated with biallelic, or suspected biallelic, mutations in MYO5B and who had neither recurrent diarrhea nor received parenteral alimentation. Seven of them are from two study cohorts, together comprising 31 undiagnosed low-GGT cholestasis patients; 3 are sporadic. Cholestasis in 2 patients was progressive, in 3 recurrent, in 2 transient, and in 3 uncategorized because of insufficient follow-up. Liver biopsy specimens revealed giant-cell change of hepatocytes and intralobular cholestasis with abnormal distribution of bile salt export pump (BSEP) at canaliculi, as well as coarse granular dislocation of MYO5B. Mass spectrometry of plasma demonstrated increased total bile acids, primary bile acids, and conjugated bile acids, with decreased free bile acids, similar to changes in BSEP-deficient patients. Literature review revealed that patients with biallelic mutations predicted to eliminate MYO5B expression were more frequent in typical MVID than in isolated-cholestasis patients (11 of 38 vs. 0 of 13). CONCLUSION: MYO5B deficiency may underlie 20% of previously undiagnosed low-GGT cholestasis. MYO5B deficiency appears to impair targeting of BSEP to the canalicular membrane with hampered bile acid excretion, resulting in a spectrum of cholestasis without diarrhea. (Hepatology 2017;65:1655-1669).

Comprehensive bile acid profiling in hereditary intrahepatic cholestasis: Genetic and clinical correlations.

BACKGROUND & AIMS: Genetic defects causing dysfunction in bile salt export pump (BSEP/ABCB11) lead to liver diseases. ABCB11 mutations alter the bile acid metabolome. We asked whether profiling plasma bile acids could reveal compensatory mechanisms and track genetic and clinical status. METHODS: We compared plasma bile acids in 17 ABCB11-mutated patients, 35 healthy controls and 12 genetically undiagnosed cholestasis patients by ultra-high-performance liquid chromatography/multiple-reaction monitoring-mass spectrometry (UPLC/MRM-MS). We developed an index to rank bile acid hydrophobicity, and thus toxicity, based on LC retention times. We recruited 42 genetically diagnosed hereditary cholestasis patients, of whom 12 were presumed to have impaired BSEP function but carried mutations in genes other than ABCB11, and 8 healthy controls, for further verification. RESULTS: The overall hydrophobicity indices of total bile acids in both the ABCB11-mutated group (11.89 ± 1.07 min) and the undiagnosed cholestasis group (11.46 ± 1.07 min) were lower than those of healthy controls (13.69 ± 0.77 min) (both p < 0.005). This was owing to increased bile acid modifications. Secondary bile acids were detected in patients without BSEP expression, suggesting biliary bile acid secretion through alternative routes. A diagnostic panel comprising lithocholic acid (LCA), tauro-LCA, glyco-LCA and hyocholic acid was identified that could differentiate the ABCB11-mutated cohort from healthy controls and undiagnosed cholestasis patients (AUC=0.946, p < 0.0001) and, in non-ABCB11-mutated cholestasis patients, could distinguish BSEP dysfunction from normal BSEP function (9/12 vs 0/38, p < 0.0000001). CONCLUSIONS: Profiling of plasma bile acids has provided insights into cholestasis alleviation and may be useful for the clinical management of cholestatic diseases.

Novel NBAS mutations and fever-related recurrent acute liver failure in Chinese children: a retrospective study.

BACKGROUND: Underlying causes in Chinese children with recurrent acute liver failure (RALF), including liver crises less than full acute liver failure, are incompletely understood. We sought to address this by searching for genes mutated in such children. METHODS: Five unrelated Chinese boys presenting between 2012 and 2015 with RALF of unexplained etiology were studied. Results of whole exome sequencing were screened for mutations in candidate genes. Mutations were verified in patients and their family members by Sanger sequencing. All 5 boys underwent liver biopsy. RESULTS: NBAS was the only candidate gene mutated in more than one patient (biallelic mutations, 3 of 5 patients; 5 separate mutations). All NBAS mutations were novel and predictedly pathogenic (frameshift insertion mutation c.6611_6612insCA, missense mutations c.2407G > A and c.3596G > A, nonsense mutation c.586C > T, and splicing-site mutation c.5389 + 1G > T). Of these mutations, 3 lay in distal (C-terminal) regions of NBAS, a novel distribution. Unlike the 2 patients without NBAS mutations, the 3 patients with confirmed NBAS mutations all suffered from a febrile illness before each episode of liver crisis (fever-related RALF), with markedly elevated alanine aminotransferase and aspartate aminotransferase activities 24-72 h after elevation of body temperature, succeeded by severe coagulopathy and mild to moderate jaundice. CONCLUSIONS: As in other countries, so too in China; NBAS disease is a major cause of fever-related RALF in children. The mutation spectrum of NBAS in Chinese children seems different from that described in other populations.

Severe Neonatal Cholestasis in Cerebrotendinous Xanthomatosis: Genetics, Immunostaining, Mass Spectrometry.

OBJECTIVES: Cerebrotendinous xanthomatosis (CTX) is caused by defects in sterol 27-hydroxylase (CYP27A1, encoded by CYP27A1), a key enzyme in the bile acid synthesis pathway. CTX usually presents as neurologic disease in adults or older children. The rare reports of CTX manifest as neonatal cholestasis assess the cholestasis as transient, with patient survival. Our experience differs. METHODS: Homozygous or compound heterozygous CYP27A1 mutations were detected in 8 neonatal cholestasis patients by whole exome sequencing, panel sequencing, or Sanger sequencing. Their clinical and biochemical data were retrospectively reviewed. Immunostaining for CYP27A1 was conducted in liver of 4 patients. Mass spectrometry was used to analyze patients' urine samples. RESULTS: All 8 infants had severe cholestasis. Five died from, or were transplanted for, liver failure; 3 cleared their jaundice eventually. Marking for CYP27A1 was weak or absent in 3 of the 4 patient specimens. Mass spectrometry of urine revealed a predominance of sulfated and doubly conjugated (sulfated-glucuronidated) bile alcohols. No patient harbored a putatively pathogenic mutation in genes other than CYP27A1 that have been implicated in cholestatic liver disease. CONCLUSIONS: CTX manifest as neonatal cholestasis has a bile acid profile different from CTX manifest in later life, and thus may be overlooked. Immunostaining, mass spectrometry of urine, and genetic studies can support one another in making the diagnosis. A substantial proportion of CTX patients with severe neonatal cholestasis may die or need liver transplantation. CTX manifest in infancy as severe cholestasis warrants further investigation of biochemical diagnostic criteria and best management.

Fibrinogen storage disease in a Chinese boy with de novo fibrinogen Aguadilla mutation: Incomplete response to carbamazepine and ursodeoxycholic acid.

BACKGROUND: Fibrinogen storage disease (FSD) is a rare autosomal-dominant disorder caused by mutation in FGG, encoding the fibrinogen gamma chain. Here we report the first Han Chinese patient with FSD, caused by de novo fibrinogen Aguadilla mutation, and his response to pharmacologic management. CASE PRESENTATION: Epistaxis and persistent clinical-biochemistry test-result abnormalities prompted liver biopsy in a boy, with molecular study of FGG in him and his parents. He was treated with the autophagy enhancer carbamazepine, reportedly effective in FSD, and with ursodeoxycholic acid thereafter. Inclusion bodies in hepatocellular cytoplasm stained immune-histochemically for fibrinogen. Selective analysis of FGG found the heterozygous mutation c.1201C > T (p.Arg401Trp), absent in both parents. Over more than one year's follow-up, transaminase and gamma-glutamyl transpeptidase activities have lessened but not normalized. CONCLUSION: This report expands the epidemiology of FSD and demonstrates idiosyncrasy in response to oral carbamazepine and/or ursodeoxycholic acid in FSD.

Efficacy of rituximab-containing regimens used as first-line and rescue therapy for giant cell hepatitis with autoimmune hemolytic anemia a retrospective study.

OBJECTIVE: To evaluate the efficacy of rituximab (RTX)-containing therapy as first-line as well as rescue treatment for giant cell hepatitis with autoimmune hemolytic anemia (GCH-AHA). METHODS: This retrospective study recruited patients diagnosed with GCH-AHA and treated with conventional immunosuppressor regimens consisting of prednisone or RTX-containing regimes consisting of RTX and prednisone, with or without another immunosuppressor. The primary outcomes were the complete remission (CR) rate and time-period required for CR. The secondary outcomes included relapses and adverse events. RESULTS: Twenty patients (8 females and 12 males; age range 1-26 months), 15 receiving conventional regimens and 5 receiving RTX-containing regimens, were included. The CR rates were 73.3 % (11/15) and 100 % (5/5) in the conventional and RTX-containing groups, respectively. The time-period required for CR was significantly shorter in the RTX-containing group than in the conventional group (6 (3-8) versus 14 (5-25) months, P = 0.015). Relapses occurred in 30.8 % (4/13) of patients in the conventional group; all achieved CR after adding RTX. Relapses occurred in 40.0 % (2/5) of patients in the RTX-containing group; both achieved CR after adding intravenous immune globulins or tacrolimus. Transient low immunoglobulin and infections were recorded in both groups. Treatment withdrawal was achieved in 73.3 % (11/15) and 60.0 % (3/5) of patients receiving conventional and RTX-containing regimens after 36 (2-101) and 22 (4-41) months, respectively. Two patients in conventional group died of disease progression and infection. CONCLUSIONS: RTX-containing first-line therapy achieves CR of GCH-AHA more quickly than the conventional therapy. RTX is efficacious when added to rescue therapy.

Poly-hydroxylated bile acids and their prognostic roles in Alagille syndrome.

BACKGROUND: The liver manifestations of Alagille syndrome (ALGS) are highly variable, and factors affecting its prognosis are poorly understood. We asked whether the composition of bile acids in ALGS patients with good clinical outcomes differs from that in patients with poor outcomes and whether bile acids could be used as prognostic biomarkers. METHODS: Blood for bile acid profiling was collected from genetically confirmed JAG1-associated ALGS patients before one year of age. A good prognosis was defined as survival with native liver and total bilirubin (TB) < 85.5 µmol/L, while a poor prognosis was defined as either liver transplantation, death from liver failure, or TB ≥ 85.5 µmol/L at the last follow-up. RESULTS: We found that the concentrations of two poly-hydroxylated bile acids, tauro-2ß,3α,7α,12α-tetrahydroxylated bile acid (THBA) and glyco-hyocholic acid (GHCA), were significantly increased in patients with good prognosis compared to those with poor prognosis [area under curve (AUC) = 0.836 and 0.782, respectively] in the discovery cohort. The same trend was also observed in the molar ratios of GHCA to glyco- chenodeoxycholic acid (GCDCA) and tetrahydroxylated bile acid (THCA) to tauro-chenodeoxycholic acid (TCDCA) (both AUC = 0.836). A validation cohort confirmed these findings. Notably, tauro-2ß,3α,7α,12α-THBA achieved the highest prediction accuracy of 88.00% (92.31% sensitivity and 83.33% specificity); GHCA at > 607.69 nmol/L was associated with native liver survival [hazard ratio: 13.03, 95% confidence interval (CI): (2.662-63.753), P = 0.002]. CONCLUSIONS: We identified two poly-hydroxylated bile acids as liver prognostic biomarkers of ALGS patients. Enhanced hydroxylation of bile acids may result in better clinical outcomes.

Effect of continuous veno-venous hemodiafiltration on endotoxin-induced acute lung injury of the piglets.

OBJECTIVE: To evaluate the effect of continuous veno-venous hemodialysis filtration (CVVHDF) on cardiopulmonary function and clearance of inflammatory mediators in piglets with endotoxin-induced acute lung injury. DESIGN: A randomized controlled trial. SETTING: An animal laboratory in a tertiary care pediatric center. SUBJECTS: : Eighteen piglets, weighing 6-8 kg. INTERVENTIONS: The piglets were anesthetized, ventilated, and received an intravenous infusion of 150 µg/kg of endotoxin. They were then randomly divided into three groups: control group (n = 6) received Ringer's lactate solution; the heparin group (n = 6) received heparin infusion plus Ringer's lactate solution; and the CVVHDF group (n = 6) received CVVHDF plus heparin infusion and Ringer's lactate solution. MEASUREMENTS AND MAIN RESULTS: Parameters measured simultaneously were: heart rate, mean arterial blood pressure, central venous pressure, pulse contour cardiac index, cardiac function index, left ventricular contractile index, and systemic vascular resistance index; extra vascular lung water index, respiratory rate, dynamic pulmonary compliance, airway resistance, Pao2/Fio2 ratio, serum tumor necrosis factor-α, and soluble tumor necrosis factor receptor. Lung tissue was obtained for pathologic lung injury scoring and wet/dry weight ratio. Endotoxin challenge decreased oxygenation and pulmonary mechanics, suppressed cardiac function, increased extravascular lung water, and elevated serum inflammatory mediators (tumor necrosis factor-α and soluble tumor necrosis factor receptor). After CVVHDF, pulmonary function and oxygenation improved (Pao2/Fio2, 291.5 ± 75.9 vs. 217.2 ± 45.4, respectively, p < .05); arterial blood pressure and cardiac function were restored (pulse contour cardiac index, 3.95 ± 0.52 L/min/m(2) vs. 2.69 ± 0.49 L/min/m(2), respectively, p < .05); extravascular lung water decreased, and serum inflammatory markers also decreased. Lung injury score improved and wet/dry weight ratio decreased. CONCLUSION: Early CVVHDF has a beneficial effect on acute lung injury in piglets and is associated with a reduction in inflammatory cytokines, improving pulmonary function and hemodynamics and decreasing extravascular lung water and lung damage.

Whole-Genome Sequencing Reveals Large ATP8B1 Deletion/Duplications as Second Mutations Missed by Exome-Based Sequencing.

Progressive familial intrahepatic cholestasis type 1 (PFIC1) results from biallelic pathogenic variants in ATP8B1. This study sought second pathogenic variants in ATP8B1 by whole-genome sequencing (WGS) in four unrelated low γ-glutamyl transpeptidase cholestasis patients in whom clinical suspicion of PFIC1 was high and gene-panel or Sanger sequencing had identified only one pathogenic variant in ATP8B1. Sanger sequencing confirmed WGS findings and determined the origin of each variant. Novel nonrecurrent structural variants in three patients (patient 1 to patient 3) were identified in trans: g.55396652_55403080del (6427-bp deletion), g.55335906_55346620dup (10,715-bp duplication), and g.55362063_55364293dup (2231-bp duplication). One synonymous variant in patient 4 was recognized in trans (c.1029G>A, p. Thr343Thr) and demonstrated as deleterious. In conclusion, WGS improves genetic diagnostic yield in PFIC1. These findings expand the gene-variant spectrum associated with familiar intrahepatic cholestasis 1 (FIC1) disease and for the first time report tandem duplication in ATP8B1 associated with cholestasis.

Recurrent acute liver failure associated with novel SCYL1 mutation: A case report.

BACKGROUND: Pediatric recurrent acute liver failure (RALF) with recovery between episodes is rare. Causes include autoimmune disease, which may flare and subside; intermittent exposure to toxins, as with ingestions; and metabolic disorders, among them the fever-associated crises ascribed to biallelic mutations in SCYL1, with RALF beginning in infancy. SCYL1 disease manifest with RALF, as known to date, includes central and peripheral neurologic and muscular morbidity (hepatocerebellar neuropathy syndrome). Primary ventilatory and skeletal diseases also have been noted in some reports. CASE SUMMARY: We describe a Han Chinese boy in whom fever-associated RALF began at age 14 mo. Bilateral femoral head abnormalities and mild impairment of neurologic function were first noted aged 8 years 6 mo. Liver biopsy after the third RALF episode (7 years) and during resolution of the fourth RALF episode (8 years 6 mo) found abnormal architecture and hepatic fibrosis, respectively. Whole-exome sequencing revealed homozygosity for the novel frameshift mutation c.92_93insGGGCCCT, p.(H32Gfs*20) in SCYL1 (parental heterozygosity confirmed). CONCLUSION: Our findings expand the mutational and clinical spectrum of SCYL1 disease. In our patient a substantial neurologic component was lacking and skeletal disease was identified relatively late.

Δ4-3-oxosteroid-5ß-reductase deficiency: Responses to oral bile acid therapy and long-term outcomes.

BACKGROUND: Disorders of primary bile acid synthesis may be life-threatening if undiagnosed, or not treated with primary bile acid replacement therapy. To date, there are few reports on the management and follow-up of patients with Δ4-3-oxosteroid 5ß-reductase (AKR1D1) deficiency. We hypothesized that a retrospective analysis of the responses to oral bile acid replacement therapy with chenodeoxycholic acid (CDCA) in patients with this bile acid synthesis disorder will increase our understanding of the disease progression and permit evaluation of this treatment regimen as an alternative to the Food and Drug Administration (FDA) approved drug cholic acid, which is currently unavailable in China. AIM: To evaluate the therapeutic responses of patients with AKR1D1 deficiency to oral bile acid therapy, specifically CDCA. METHODS: Twelve patients with AKR1D1 deficiency, confirmed by fast atom bombardment ionization-mass spectrometry analysis of urine and by gene sequencing for mutations in AKR1D1, were treated with differing doses of CDCA or ursodeoxycholic acid (UDCA). The clinical and biochemical responses to therapy were monitored over a period ranging 0.5-6.4 years. Dose adjustment, to optimize the therapeutic dose, was based on changes in serum biochemistry parameters, notably liver function tests, and suppression of the urinary levels of atypical hepatotoxic 3-oxo-Δ4-bile acids measured by mass spectrometry. RESULTS: Physical examination, serum biochemistry parameters, and sonographic findings improved in all 12 patients during bile acid therapy, except one who underwent liver transplantation. Urine bile acid analysis confirmed a significant reduction in atypical hepatotoxic 3-oxo-Δ4 bile acids concomitant with clinical and biochemical improvements in those patients treated with CDCA. UDCA was ineffective in down-regulating endogenous bile acid synthesis as evidenced from the inability to suppress the urinary excretion of atypical 3-oxo-Δ4-bile acids. The dose of CDCA required for optimal clinical and biochemical responses varied from 5.5-10 mg/kg per day among patients based on maximum suppression of the atypical bile acids and improvement in serum biochemistry parameters, and careful titration of the dose was necessary to avoid side effects from CDCA. CONCLUSION: The primary bile acid CDCA is effective in treating AKR1D1 deficiency but the therapeutic dose requires individualized optimization. UDCA is not recommended for long-term management.

UGT1A1 genotypes and unconjugated hyperbilirubinemia phenotypes in post-neonatal Chinese children: A retrospective analysis and quantitative correlation.

To retrospectively analyze and quantitatively correlate UGT1A1 (bilirubin UDP- glucuronosyltransferase gene) genotypes and unconjugated hyperbilirubinemia (UCH) phenotypes among Chinese children.We retrospectively reviewed UCH patients, quantitatively analyzed genotype-phenotype correlation by comparing with healthy controls. Pfam database, SWISS-model, and Pymol were used for UGT1A1 protein domain analysis and protein modeling for assessing the effect of novel missense variants on protein structure.Seventy four cases, including 21 prolonged unconjugated hyperbilirubinemia (PUCH), 30 Gilbert syndrome (GS), 22 Crigler-Najjar syndrome type II (CNS-II), and 1 Crigler-Najjar syndrome type I (CNS-I) phenotypes were analyzed. Total of 21 variants, including 7 novel variants (c.764T>A/p.L255Q, c.1112C>T/p.T371I, c.1028C>A/p.S343X, c.1047delG/p.I350YfsX16, c.996 + 5G>C/g.6923G>C, c.287G>A/p.G96E, and c.1142G>A/p.S381N) were found. In the multiple regression model, heterozygous A(TA)7TAA, G71R/P364L, and Y486D/other mutations were significantly associated with increased risk of GS, PUCH, and CNS-II, respectively. Total allele number is significantly associated with GS and CNS-II, with each increase in total allele number, the odds ratio (OR) of having GS and CNS-II increased by 1.46 and 4.47 fold, respectively. Having only functional polymorphisms in UGT1A1 gene is associated with increased risk of PUCH, and GS with OR values of 5.67 (95% CI: 1.52-21.13), and 3.88 (95% CI: 1.02-14.78), respectively. Having only mutation is associated with significantly increased risk of having GS phenotype (OR: 34.00, 95% CI: 4.65-248.37), but not CNS-II. Polymorphism plus mutation had the strongest association with CNS-II with OR value of 64.80 (95% CI: 7.68-546.41), followed by GS (OR: 4.53, 95% CI: 1.08-19.08).We detected 7 novel variants, and quantitatively calculated risks of having specific phenotypes using genetic data. Among Chinese children, G71R and P364L is independently associated with PUCH, A(TA)7TAA is associated with GS, and Y486D or other disease-causing mutations were associated with CNS-II. Multiple alleles were associated with more severe phenotypes. Combined variant of G71R+Y486D is a common occurrence among Chinese children with UCH.