ADH1B, the adipocyte-enriched alcohol dehydrogenase, plays an essential, cell-autonomous role in human adipogenesis.

Alcohol dehydrogenase 1B (ADH1B) is a primate-specific enzyme which, uniquely among the ADH class 1 family, is highly expressed both in adipose tissue and liver. Its expression in adipose tissue is reduced in obesity and increased by insulin stimulation. Interference with ADH1B expression has also been reported to impair adipocyte function. To better understand the role of ADH1B in adipocytes, we used CRISPR/Cas9 to delete ADH1B in human adipose stem cells (ASC). Cells lacking ADH1B failed to differentiate into mature adipocytes manifested by minimal triglyceride accumulation and a marked reduction in expression of established adipocyte markers. As ADH1B is capable of converting retinol to retinoic acid (RA), we conducted rescue experiments. Incubation of ADH1B-deficient preadipocytes with 9-cis-RA, but not with all-transretinol, significantly rescued their ability to accumulate lipids and express markers of adipocyte differentiation. A homozygous missense variant in ADH1B (p.Arg313Cys) was found in a patient with congenital lipodystrophy of unknown cause. This variant significantly impaired the protein's dimerization, enzymatic activity, and its ability to rescue differentiation in ADH1B-deficient ASC. The allele frequency of this variant in the Middle Eastern population suggests that it is unlikely to be a fully penetrant cause of severe lipodystrophy. In conclusion, ADH1B appears to play an unexpected, crucial and cell-autonomous role in human adipocyte differentiation by serving as a necessary source of endogenous retinoic acid.

Safety and efficacy of combined sofosbuvir/daclatasvir treatment of children and adolescents with chronic hepatitis C Genotype 4.

Direct-acting antivirals have become available for treating chronic HCV (hepatitis C virus) infection in adults and, recently, in children at least 12 years old. Our aim was to investigate the safety and efficacy of combined sofosbuvir (SOF)/daclatasvir (DCV) for HCV Genotype 4 in children aged 8 to 18 years or weighing 17 kg or more. A total of 40 chronic HCV-infected, treatment-naïve children with well compensated livers were recruited from two sites. Patients received combined therapy of SOF (400 mg/d for patients weighing greater than 45 kg; 200 mg/d for patients weighing 17 to 45 kg) and DCV (60 mg/d for patients weighing greater than 45 kg; 30 mg/d for patients weighing 17 to 45 kg) for 12 weeks. They were followed up regularly by clinical examination and laboratory tests during treatment (weekly in the first month then monthly to the end of treatment), every 3 months for 6 months post-treatment, and at 48 weeks post-treatment. In our cohort, which included 45% of children below the age of 12 years (72.5% genotype 4 and 27.5% mixed genotype 4 and 1), end of treatment response (ETR) was 97.5%. Sustained virologic response for weeks 12 and 24 post-treatment (SVR12 and SVR24) were 97.5% and 95%, respectively, on an intention to treat basis, and 100% and 100% for those who completed the study protocol. Observed side effects were mild and none required drug cessation. Combined SOF/DCV was found to be effective and safe for treating HCV Genotype 4-infected children, 8 years of age and above.

Phenotypic and genetic characterization of a cohort of pediatric Wilson disease patients.

BACKGROUND: In Egypt, Wilson disease seems to be under diagnosed and clinical data on large cohorts are limited. The aim of this study is to highlight the clinical, laboratory and genetic characteristics of this disease in our pediatric population as well as to report our experience with both treatment options and outcome. METHODS: The study included 77 patients from 50 unrelated families (62 were followed up for a mean period of 58.9 ± 6.4 months and 27 were asymptomatic siblings). Data were collected retrospectively by record analysis and patient interviews. Diagnosis was confirmed by sequencing of the ATP7B gene in 64 patients. RESULTS: Our patients had unique characteristics compared to other populations. They had a younger age of onset (median: 10 years), higher prevalence of Kayser-Fleischer rings (97.6% in the symptomatic patients), low ceruloplasmin (93.5%), high rate of parental consanguinity (78.9%) as well as a more severe course. 71.42% of those on long term D-penicillamine improved or were stable during the follow up with severe side effects occurring in only 11.5%. Preemptive treatment with zinc monotherapy was an effective non-toxic alternative to D-penicillamine. Homozygous mutations were found in 85.7%, yet limited by the large number of mutations detected, it was difficult to find genotype-phenotype correlations. Missense mutations were the most common while protein-truncating mutations resulted in a more severe course with higher incidence of acute liver failure and neurological symptoms. CONCLUSIONS: Egyptian children with Wilson disease present with early Kayser-Fleischer rings and early onset of liver and neurological disease. The mutational spectrum identified differs from that observed in other countries. The high rate of homozygous mutations (reflecting the high rate of consanguinity) may potentially offer further insights on genotype-phenotype correlation.

Factor V G1691A (Leiden) is a major etiological factor in Egyptian Budd-Chiari syndrome patients.

OBJECTIVE: Budd-Chiari syndrome is a multifactorial disease in which several prothrombotic disorders may predispose patients to the development of thrombosis at this uncommon location (hepatic veins). The aim of this study was to determine the prevalence and characteristics of inherited thrombophilia in Egyptian Budd-Chiari syndrome patients. METHODS: The study included 47 Budd-Chiari syndrome patients (20 children and 27 adults). Genotyping of Factor V G1691A (Leiden), prothrombin G20210A (PT), and methylenetetrahydrofolate reductase C677T were performed using real-time PCR and fluorescence melting curve detection analysis. RESULTS: Factor V Leiden was observed in 29 patients (61.7%). It is the only factor that caused Budd-Chiari syndrome in 18 of the patients and in 5 of the patients with inferior vena cava involvement. Myeloproliferative disease was noted in 12 (25.5%) patients, antiphospholipid syndrome in 5 (10.6%), and Behcet's disease in 3 (6.4%). Interestingly, 3 of the children with Budd-Chiari syndrome had lipid storage disease. CONCLUSION: Factor V Leiden was a major etiological factor in Egyptian Budd-Chiari syndrome patients, which may have been related to the high frequency of this mutation in the study region. Factor V Leiden was also a strong thrombophilic factor and the leading cause of inferior vena cava thrombosis in these patients. Lipid storage disease should be included as a risk factor for Budd-Chiari syndrome.

MPI-CDG from a hepatic perspective: Report of two Egyptian cases and review of literature.

MPI-CDG is a rare congenital disorder of glycosylation (CDG) which presents with hepato-gastrointestinal symptoms and hypoglycemia. We report on hepatic evaluation of two pediatric patients who presented to us with gastrointestinal symptoms. Analysis of carbohydrate deficient transferrin (CDT) showed a Type 1 pattern and molecular analysis confirmed the diagnosis of MPI-CDG. Oral mannose therapy was markedly effective in one patient but was only partially effective in the other who showed progressive portal hypertension.

Hepatitis C genotype 4: The past, present, and future.

Hepatitis C virus (HCV) genotype (GT) 4 represents 12%-15% (15-18 million) of total global HCV infection. It is prevalent in Northern and Equatorial Africa and the Middle East, and is also present in some countries in Europe. GT-4 (and subtype 4a in particular) dominates the HCV epidemic in Egypt. In underdeveloped countries, risk factors associated with HCV infection may be due to unsafe medical practices or other factors such as familial transmission, mother's HCV status, or illiteracy. HCV prevention and control programs should include health education, increased community awareness towards the disease, controlling infection distribution in health-care centers, proper sterilization of medical and dental instruments, and ensuring safe supply of blood and blood-products. Response rates to a 48-wk combined pegylated-interferon (PEG-IFN) and ribavirin (RBV) treatment range from 40%-69%, and HCV-GT-4 has been considered better than GT-1 but worse than GT-2 and GT-3 in treatment with PEG-IFN/RBV. However, with the introduction of the HCV-GT-1 effective protease inhibitors boceprevir and telaprevir in 2011, HCV-GT-4 became the "most difficult (GT) to treat". Recently, the direct-acting antivirals (DAAs) with pan- genotypic activities simeprevir, sofosbuvir, and daclatasvir have been recommended in triple regimens with PEG-IFN/RBV for the treatment of HCV-GT-4. An IFN-free regimen will be available for treatment of all genotypes of HCV in the near future. To date, several DAAs have been developed and are currently being evaluated in various combinations in clinical trials. As new regimens and new agents are being approved by the Food and Drug Administration, we can expect the guidelines for HCV treatment to be changed. The availability of shorter, simpler, and more tolerable treatment regimens can reduce the morbidity and mortality associated with HCV infection. With such a large number of therapeutic agents available, we can end up with a range of choices that we can select from to treat patients.

Serological markers of autoimmunity in children with hepatitis A: relation to acute and fulminant presentation.

OBJECTIVES: Hepatitis A virus (HAV) infection tends to be a self-limiting disease without serious sequelae, but fulminant hepatitis, with a high mortality, develops in 0.1-0.2% of the cases. Sometimes, HAV infection precipitates autoimmune hepatitis (AIH). We aimed to assess the frequency and clinical significance of serologic markers of autoimmunity during hepatitis A infection with an acute or fulminant presentation compared with those in AIH. METHODS: The study included 126 children: 46 with HAV infection (33 with acute and 13 with fulminant presentation), 53 with AIH, and 27 healthy controls. In all, we measured autoantibodies titer (antinuclear antibody, antismooth muscle antibody, and liver kidney microsomal antibody-1) and serum gammaglobulins. RESULTS: Autoantibodies were detected in the majority of HAV (63.1%) and AIH (79.2%) groups, but in none of the controls. Gammaglobulins were significantly higher in the HAV group (1.93±0.57 g/dl) than in the controls (1.32±0.29 g/dl), but lower than that in the AIH group (2.93±1.2 g/dl) (P<0.0001 for all). In the HAV group, gammaglobulins were significantly higher in those with fulminant (2.21±0.46 g/dl) than in those with acute presentation (1.82±0.57 g/dl) (P=0.019), but comparable with that in AIH (P=0.095). Gammaglobulins correlated significantly with disease severity in both HAV and AIH groups. CONCLUSION: Hypergammaglobulinemia and a high occurrence of autoantibodies are encountered in HAV infection. This may support the immunological basis of its pathogenesis. Moreover, the higher gammaglobulins in fulminant HAV, with an insignificant difference from that in AIH, suggest that a more aggressive immunological reaction is related to this presentation.

Safety and efficacy of Hansenula-derived PEGylated-interferon alpha-2a and ribavirin combination in chronic hepatitis C Egyptian children.

AIM: To investigate the safety and efficacy of a Hansenula-derived PEGylated (polyethylene glycol) interferon (IFN)-alpha-2a (Reiferon Retard) plus ribavirin customized regimen in treatment-naïve and previously treated (non-responders and relapsers) Egyptian children with chronic hepatitis C infection. METHODS: Forty-six children with chronic hepatitis C virus (HCV) infection were selected from three tertiary pediatric hepatology centers. Clinical and laboratory evaluations were undertaken. Quantitative polymerase chain reaction (PCR) for HCV-RNA was performed before starting treatment, and again at 4, 12, 24, 48, 72 wk during treatment and 6 mo after treatment cessation. All patients were assigned to receive a weekly subcutaneous injection of PEG-IFN-alpha-2a plus daily oral ribavirin for 12 wk. Thirty-four patients were treatment-naïve and 12 had a previous treatment trial. Patients were then divided according to PCR results into two groups. Group I included patients who continued treatment on a weekly basis (7-d schedule), while group II included patients who continued treatment on a 5-d schedule. Patients from either group who were PCR-negative at week 48, but had at least one PCR-positive test during therapy, were assigned to have an extended treatment course up to 72 wk. The occurrence of adverse effects was assessed during treatment and follow up. The study was registered at www.ClinicalTrials.gov (NCT02027493). RESULTS: Only 11 out of 46 (23.9%) patients showed a sustained virological response (SVR), two patients were responders at the end of treatment; however, they were lost to follow up at 6 mo post treatment. Breakthrough was seen in 18 (39.1%) patients, one patient (2.17%) showed relapse and 14 (30.4%) were non-responders. Male gender, short duration of infection, low viral load, mild activity, and mild fibrosis were the factors related to a better response. On the other hand, patients with high viral load and absence of fibrosis failed to respond to treatment. Before treatment, liver transaminases were elevated. After commencing treatment, they were normalized in all patients at week 4 and were maintained normal in responders till the end of treatment, while they increased again significantly in non-responders (P = 0.007 and 0.003 at week 24 and 72 respectively). The 5-d schedule did not affect the response rate (1/17 had SVR). Treatment duration (whether 48 wk or extended course to 72 wk) gave similar response rates (9/36 vs 2/8 respectively; P = 0.49). Type of previous treatment (short acting IFN vs PEG-IFN) did not affect the response to retreatment. On the other hand, SVR was significantly higher in previous relapsers than in previous non-responders (P = 0.039). Only mild reversible adverse effects were observed and children tolerated the treatment well. CONCLUSION: Reiferon Retard plus ribavirin combined therapy was safe. Our customized regimen did not influence SVR rates. Further trials on larger numbers of patients are warranted.

Cholestasis in patients with Cockayne syndrome and suggested modified criteria for clinical diagnosis.

BACKGROUND: Cockayne syndrome is a rare autosomal recessive neurodegenerative disease characterized by low-to-normal birth weight; growth failure; brain dysmyelination with calcium deposits, cutaneous photosensitivity; pigmentary retinopathy, cataract, and sensorineural hearing loss. To the best of our knowledge, cholestatic liver disease was not previously reported in these patients. AIM: To highlight the presence of cholestasis and liver dysfunction in this group of patients and to suggest modified criteria for clinical diagnosis. METHODS: The study included nine patients with Cockayne from four different families (five males and four females) in which Cockayne was suspected clinically. In all patients chromosomal breakage studies revealed mild (45%) to moderate (60%) increase in frequency of chromatid and chromosome gaps and breaks versus 25% in normal controls. Diagnosis was confirmed by DNA repair assay. RESULTS: During routine follow up of these patients, seven of them had evident liver affection ranging from mild elevation in liver enzymes to cholestatic liver disease and liver cell failure. The attacks were recurrent in two patients and were sometimes preceded by infection. The attack may lead to deterioration of neurological and/or liver condition. It may end in liver cell failure that either recovers completely or may lead to death. CONCLUSIONS: liver disease could be considered common in Egyptian patients with Cockayne with the cholestatic form being the most evident. The syndrome should be included in the list of causes of cholestatic liver disease. Chromosomal breakage study and positive family history should be included as major criteria for clinical diagnosis of Cockayne especially in a population like ours where consanguineous marriage is very high and molecular testing and UV sensitivity tests are considered unaffordable.

Clinical Benefits of Biochemical Markers of Fibrosis in Egyptian Children With Chronic Liver Diseases.

BACKGROUND: The need for repetition of liver biopsy, especially in assessing the degree of fibrosis and follow-up of treatment protocols, justifies an intensive search for non-invasive alternatives. We attempted to investigate the clinical usefulness of serum fibrogenesis markers in pediatric chronic liver diseases. METHODS: We measured serum levels of TGF-ß1, collagen IV, laminin, MMP-2 and EGF-R, in 50 children with chronic liver disease (HBV, HCV and Bilharziasis) and 30 healthy controls, and determined their relationship to frequently used liver function tests and liver biopsy findings in patients. RESULTS: TGF-ß1, collagen IV, laminin and MMP-2, but not EGF-R, were significantly higher in patients than in controls (P < 0.01). None of these markers correlated with the histological fibrosis stage, whereas laminin correlated with necroinflammatory activity (P < 0.01). TGF-ß1, collagen IV, laminin and MMP-2 had the ability to discriminate patients with significant fibrosis, while only collagen IV and laminin were able to discriminate those with cirrhosis. Among these markers, collagen IV had the best predictive accuracy for significant fibrosis (AUROC 0.94; PPV 91.5%) and cirrhosis (AUROC 0.85; PPV 80%). CONCLUSIONS: In conclusion, these markers may be useful in reducing but not replacing the need for liver biopsy in the monitoring of disease progression and treatment effectiveness and might be an inseparable part of assessment of chronic hepatopathies.