Adeno-Associated Virus 2 and Human Adenovirus F41 in Wastewater during Outbreak of Severe Acute Hepatitis in Children, Ireland.

During April-July 2022, outbreaks of severe acute hepatitis of unknown etiology (SAHUE) were reported in 35 countries. Five percent of cases required liver transplantation, and 22 patients died. Viral metagenomic studies of clinical samples from SAHUE cases showed a correlation with human adenovirus F type 41 (HAdV-F41) and adeno-associated virus type 2 (AAV2). To explore the association between those DNA viruses and SAHUE in children in Ireland, we quantified HAdV-F41 and AAV2 in samples collected from a wastewater treatment plant serving 40% of Ireland's population. We noted a high correlation between HAdV-F41 and AAV2 circulation in the community and SAHUE clinical cases. Next-generation sequencing of the adenovirus hexon in wastewater demonstrated HAdV-F41 was the predominant HAdV type circulating. Our environmental analysis showed increased HAdV-F41 and AAV2 prevalence in the community during the SAHUE outbreak. Our findings highlight how wastewater sampling could aid in surveillance for respiratory adenovirus species.

Acute Hepatitis of Unknown Etiology Among Young Children: Research Agenda by the ESPGHAN Hepatology Committee.

In April 2022, an increased incidence of acute hepatitis cases of unknown etiology among previously healthy children across the United Kingdom was described. Since, more than 270 cases from the United Kingdom and hundreds more from all across the world have been reported. The majority of affected children were younger than 6 years of age. The clinical presentation was nonspecific with diarrhea and vomiting usually preceding the appearance of jaundice, abdominal pain, nausea, and malaise. Approximately 5% have required liver transplantation. An infectious etiology has been considered likely given the epidemiological and clinical features of the reported cases. Between 50 and 60% of the children tested were diagnosed with adenovirus infection although a clear etiological connection has still to be demonstrated. No link with SARS-CoV-2 infection and COVID-19 vaccine was found. What is not clear to date is whether the high number of acute hepatitis cases reported is related to a true increase in incidence or heightened awareness following on from the initial reports from the United Kingdom. The Hepatology Committee of the European Society of Pediatric Gastroenterology, Hepatology and Nutrition (ESPGHAN) developed a paper on the current outbreak of acute hepatitis of unknown etiology recognizing its importance and the need of approaching the current situation with a scientifically rigorous approach. The aims of the article are to summarize the current knowledge and to identify the most pertinent issues regarding the diagnosis and management of this condition and the research questions raised.

Emerging clinical perspectives in cystic fibrosis liver disease.

PURPOSE OF REVIEW: Liver disease (CFLD) as a complication of cystic fibrosis is recognized as a more severe disease phenotype in both children and adults. We review recent advances in understanding the disease mechanism and consider the implications of new strategies for the diagnosis and management of cystic fibrosis in those with evidence of clinically significant liver disease. RECENT FINDINGS: Evidence suggests that the prevalence of CFLD has not declined with the introduction of newborn screening. Furthermore, children with CFLD, who have been diagnosed with cystic fibrosis following newborn screening continue to have a much higher mortality rate compared with those with no liver disease. There is further data suggesting noncirrhotic obliterative portal venopathy as the predominant pathological mechanism in the majority of children and young adults receiving a liver transplantation. Little progress has been made in developing an accurate noninvasive test for early diagnosis or monitoring disease progression in CFLD. The benefit of new modulator therapies is not well understood in those with established CFLD, whereas the risk of hepatotoxicity as a complication of treatment must be carefully monitored. SUMMARY: Better understanding of the pathophysiology of CFLD would allow a standardized approach to diagnosis, with the potential to improve outcomes for those with CFLD.

Alginate microencapsulated human hepatocytes for the treatment of acute liver failure in children.

BACKGROUND & AIMS: Liver transplantation (LT) is the most effective treatment for patients with acute liver failure (ALF), but is limited by surgical risks and the need for life-long immunosuppression. Transplantation of microencapsulated human hepatocytes in alginate is an attractive option over whole liver replacement. The safety and efficacy of hepatocyte microbead transplantation have been shown in animal models. We report our experience of this therapy in children with ALF treated on a named-patient basis. METHODS: Clinical grade human hepatocyte microbeads (HMBs) and empty microbeads were tested in immunocompetent healthy rats. Subsequently, 8 children with ALF, who were awaiting a suitable allograft for LT, received intraperitoneal transplantation of HMBs. We monitored complications of the procedure, assessing the host immune response and residual function of the retrieved HMBs, either after spontaneous native liver regeneration or at the time of LT. RESULTS: Intraperitoneal transplantation of HMBs in healthy rats was safe and preserved synthetic and detoxification functions, without the need for immunosuppression. Subsequently, 8 children with ALF received HMBs (4 neonatal haemochromatosis, 2 viral infections and 2 children with unknown cause at time of infusion) at a median age of 14.5 days, range 1 day to 6 years. The procedure was well tolerated without complications. Of the 8 children, 4 avoided LT while 3 were successfully bridged to LT following the intervention. HMBs retrieved after infusions (at the time of LT) were structurally intact, free of host cell adherence and contained viable hepatocytes with preserved functions. CONCLUSION: The results demonstrate the feasibility and safety of an HMB infusion in children with ALF. LAY SUMMARY: Acute liver failure in children is a rare but devastating condition. Liver transplantation is the most effective treatment, but it has several important limitations. Liver cell (hepatocyte) transplantation is an attractive option, as many patients only require short-term liver support while their own liver recovers. Human hepatocytes encapsulated in alginate beads can perform the functions of the liver while alginate coating protects the cells from immune attack. Herein, we demonstrated that transplantation of these beads was safe and feasible in children with acute liver failure.

Repeatability of transient elastography in children.

BACKGROUND: Poorly performing diagnostic tests can impact patient safety. Clinical investigations must have good precision and diagnostic accuracy before widespread use in clinical practice. Transient elastography (TE) measures liver stiffness, a surrogate marker of liver fibrosis in adults and children. Studies to evaluate its repeatability and reproducibility (precision) in children are limited. Our aim was to determine (i) the normal range of TE measurements and (ii) the repeatability and reproducibility of TE in healthy children. METHODS: TE was performed in 257 healthy children, of whom 235 (91%, mean age 11.7 years, standard deviation (SD) 2.51, 107 were males (45.5%)) had two valid TE measurements performed, at least 24 h apart, by two operators under similar circumstances. High-quality TE images were obtained for each examination. RESULTS: The normal range of TE was 2.88-6.52 kPa. The mean difference between paired measurements was 0.044 (SD 0.4). The 95% limits of agreement ranged from -0.8 to +0.76 kPa for repeat measurements. There was a difference of >1 kPa between measurements in 61/235 (25.9%) children. The lack of precision was similar across all age groups. CONCLUSIONS: This study demonstrates that TE does not have acceptable precision in healthy children, because random measurement variation results in the lack of agreement between paired measurements. IMPACT: The precision and diagnostic accuracy of a new technology must be determined before it is deployed in children in order to ensure that appropriate clinical decisions are made, and healthcare resources are not wasted. TE is widely used to diagnose liver disease in children without adequate evaluation of the precision (repeatability) of TE either in healthy children or children with liver disease. This study demonstrates that TE does not have adequate precision in children. This study was performed in accordance with methods previously published for children. Refinements to the test protocol, such as duration of fasting or probe size, will have to be evaluated for their impact on precision and accuracy before the test is deployed in research studies or clinical practice.

Paediatric fatty liver disease (PeFLD): All is not NAFLD - Pathophysiological insights and approach to management.

The recognition of a pattern of steatotic liver injury where histology mimicked alcoholic liver disease, but alcohol consumption was denied, led to the identification of non-alcoholic fatty liver disease (NAFLD). Non-alcoholic fatty liver disease has since become the most common chronic liver disease in adults owing to the global epidemic of obesity. However, in paediatrics, the term NAFLD seems incongruous: alcohol consumption is largely not a factor and inherited metabolic disorders can mimic or co-exist with a diagnosis of NAFLD. The term paediatric fatty liver disease may be more appropriate. In this article, we summarise the known causes of steatosis in children according to their typical, clinical presentation: i) acute liver failure; ii) neonatal or infantile jaundice; iii) hepatomegaly, splenomegaly or hepatosplenomegaly; iv) developmental delay/psychomotor retardation and perhaps most commonly; v) the asymptomatic child with incidental discovery of abnormal liver enzymes. We offer this model as a means to provide pathophysiological insights and an approach to management of the ever more complex subject of fatty liver.

Cryopreserved neonatal hepatocytes may be a source for transplantation: Evaluation of functionality toward clinical use.

Neonatal livers are a potential source of good-quality hepatocytes for clinical transplantation. We compared viability and function of neonatal hepatocytes (NHs) and adult hepatocytes (AHs) and report their clinical use both intraportally and in alginate microbeads. Following isolation from donor livers, hepatocyte function was assessed using albumin, alpha-1-antitrypsin, and factor VII. Metabolic function was investigated by measuring resorufin conjugation, ammonia metabolism, uridine diphosphate glucuronosyltransferase enzyme activity, and cytochrome P450 (CYP) function following induction. Activation of the instant blood-mediated inflammatory reaction by NHs and AHs was investigated using an in vitro blood perfusion model, and tissue factor expression was analyzed using real-time polymerase chain reaction (RT-PCR). Clinical hepatocyte transplantation (HT) was undertaken using standard protocols. Hepatocytes were isolated from 14 neonatal livers, with an average viability of 89.4% ± 1.8% (mean ± standard error of the mean) and average yield of 9.3 × 106 ± 2.0 × 106 cells/g. Hepatocytes were isolated from 14 adult livers with an average viability of 78.6% ± 2.4% and yield 2.2 × 106 ± 0.5 × 105 cells/g. NHs had significantly higher viability after cryopreservation than AHs, with better attachment efficiency and less plasma membrane leakage. There were no differences in albumin, alpha-1-antitrypsin, and factor VII synthesis between NHs and AHs (P > 0.05). Neonatal cells had inducible phase 1 enzymes as assessed by CYP function and functional phase 2 enzymes, in which activity was comparable to AHs. In an in vitro blood perfusion model, AHs elicited increased thrombus formation with a greater consumption of platelets and white cells compared with NHs (28.3 × 109 versus 118.7 × 109 and 3.3 × 109 versus 6.6 × 109 ; P < 0.01). Intraportal transplantation and intraperitoneal transplantation of alginate encapsulated hepatocytes was safe, and preliminary data suggest the cells may activate the immune response to a lesser degree than adult cells. In conclusion, we have shown NHs have excellent cell viability, function, and drug metabolism making them a suitable alternative source for clinical HT. Liver Transplantation 24 394-406 2018 AASLD.

Correction to: Proteomic identification and characterization of hepatic glyoxalase 1 dysregulation in non-alcoholic fatty liver disease.

[This corrects the article DOI: 10.1186/s12953-018-0131-y.].

Proteomic identification and characterization of hepatic glyoxalase 1 dysregulation in non-alcoholic fatty liver disease.

Background: Non-alcoholic fatty liver disease (NAFLD) is the most common liver disease worldwide. However, its molecular pathogenesis is incompletely characterized and clinical biomarkers remain scarce. The aims of these experiments were to identify and characterize liver protein alterations in an animal model of early, diet-related, liver injury and to assess novel candidate biomarkers in NAFLD patients. Methods: Liver membrane and cytosolic protein fractions from high fat fed apolipoprotein E knockout (ApoE-/-) animals were analyzed by quantitative proteomics, utilizing isobaric tags for relative and absolute quantitation (iTRAQ) combined with nano-liquid chromatography and tandem mass spectrometry (nLC-MS/MS). Differential protein expression was confirmed independently by immunoblotting and immunohistochemistry in both murine tissue and biopsies from paediatric NAFLD patients. Candidate biomarkers were analyzed by enzyme-linked immunosorbent assay in serum from adult NAFLD patients. Results: Through proteomic profiling, we identified decreased expression of hepatic glyoxalase 1 (GLO1) in a murine model. GLO1 protein expression was also found altered in tissue biopsies from paediatric NAFLD patients. In vitro experiments demonstrated that, in response to lipid loading in hepatocytes, GLO1 is first hyperacetylated then ubiquitinated and degraded, leading to an increase in reactive methylglyoxal. In a cohort of 59 biopsy-confirmed adult NAFLD patients, increased serum levels of the primary methylglyoxal-derived advanced glycation endproduct, hydroimidazolone (MG-H1) were significantly correlated with body mass index (r = 0.520, p < 0.0001). Conclusion: Collectively these results demonstrate the dysregulation of GLO1 in NAFLD and implicate the acetylation-ubquitination degradation pathway as the functional mechanism. Further investigation of the role of GLO1 in the molecular pathogenesis of NAFLD is warranted.

Transient Elastography Measurements of Spleen Stiffness as a Predictor of Clinically Significant Varices in Children.

OBJECTIVES: Investigate the use of spleen stiffness measurements (SSMs), measured by transient elastography (TE), for the prediction of clinically significant varices (CSV) in children with portal hypertension. METHODS: This observational cohort study included children selected for endoscopy, as per department protocol, between September 2015 and June 2016. Those included underwent single TE FibroScan for liver stiffness measurements and SSM. Clinical and laboratory data were collected and variceal prediction scores were calculated at time of elastography. RESULTS: In total 67 children (32 boys) underwent TE. Fifty-two children (25 boys) had chronic liver disease (CLD), 15 (7 boys) portal vein thrombosis (PVT). In all children SSM was the best predictor of CSV+ve, with an optimal cut-off value of 38.0 kPa (area under the receiver operator curve [AUROC] = 0.92, sensitivity = 89%, specificity = 82%, P < 0.01). In the CLD group SSM was also the best predictor, with an optimal cut-off value of 38.05 kPa (AUROC = 0.90, sensitivity = 84%, specificity = 87%, P < 0.01). In the PVT group only SSM was predictive of CSV+ve, with an optimal cut-off value of 16.8 kPa (AUROC = 1.00, sensitivity = 100%, specificity = 100%, P < 0.001). For the prediction of GI bleeding (n = 6), liver stiffness measurement performed the best, with an optimal cut-off value of 34.3 kPa (AUROC = 0.84, sensitivity of 80%, specificity of 88%, P = 0.01). CONCLUSIONS: SSM was the greatest predictor of CSV+ve in the whole cohort, and individual CLD and PVT groups. SSM could be used as a noninvasive screening tool for children with portal hypertension to stratify the risk of having CSV.

Fifteen-minute consultation: liver disease in children.

Liver disease in children can present in many ways from the frequently encountered prolonged neonatal jaundice to the comparatively rare acute liver failure. In this article, we will discuss 'red flags' of liver disease, the initial investigations required and when to refer to a specialist liver centre. Across all presentations, the degree of elevation of alanine aminotransferase or aspartate aminotransferase provides only little diagnostic information. Measurement of clotting is vital, and coagulopathy should be followed by a trial of intravenous vitamin K before being repeated.

Interim assessment of liver damage in patients with sickle cell disease using new non-invasive techniques.

We explored transient elastography (TE) and enhanced liver fibrosis (ELF™ ) score with standard markers of liver function to assess liver damage in 193 well patients with sickle cell disease (SCD). Patients with HbSS or HbSß0 thalassaemia (sickle cell anaemia, SCA; N = 134), had significantly higher TE results and ELF scores than those with HbSC (N = 49) disease (TE, 6·8 vs. 5·3, P < 0·0001 and ELF, 9·2 vs. 8·6 P < 0·0001). In SCA patients, TE and ELF correlated significantly with age and all serum liver function tests (LFTs). Additionally, (weak) positive correlation was found with lactate dehydrogenase (TE: r = 0·24, P = 0·004; ELF: r = 0·26 P = 0·002), and (weak) negative correlation with haemoglobin (TE: r = -0·25, P = 0·002; ELF: r = -0·25 P = 0·004). In HbSC patients, correlations were weaker or not significant between TE or ELF, and serum LFTs. All markers of iron loading correlated with TE values when corrected for sickle genotype (serum ferritin, ß = 0·25, P < 0·0001, total blood transfusion units, ß = 0·25, P < 0·0001 and LIC ß = 0·32, P = 0·046). The exploratory study suggests that, while TE could have a role, the utility of ELF score in monitoring liver damage in SCD, needs further longitudinal studies.

Autoimmune Liver Disease in Children with Sickle Cell Disease.

OBJECTIVE: To assess the incidence, clinical features, and outcome of autoimmune liver disease (AILD) in patients with sickle cell disease (SCD). STUDY DESIGN: Single center retrospective review of patients with SCD with AILD referred between 1999 and 2015. RESULTS: Thirteen of 77 (17%) patients with SCD with hepatic dysfunction were diagnosed with AILD (median age 11, range, 3.4-16 years) with a female preponderance (77%). Acute hepatitis and insidious onset were the commonest presentations. Two patients (15%) presented with acute liver failure. In 2 patients (15%), parvovirus B19-induced transient red cell aplasia preceded the diagnosis of AILD. All patients were positive for antinuclear and/or smooth muscle autoantibodies. Six of 12 patients (50%) had cholangiopathy on cholangiogram suggesting autoimmune sclerosing cholangitis (ASC). Liver biopsy, performed in 11 patients without complications, showed interface hepatitis in 90%. Patients with AILD were treated with standard immunosuppression. After a median follow-up of 3.8 years (range, 0.2-14.3), 10 patients are alive (1 was transplanted 6.4 years after diagnosis); 2 are lost to follow-up; 1 died of subdural hemorrhage before starting treatment for AILD. Five (42%) achieved full and 4 (33%) partial biochemical remission. Ulcerative colitis, present in 4 patients (2 male patients, 3 with ASC) was diagnosed in 2 patients before and in 2 patients after the diagnosis of AILD. CONCLUSIONS: AILD is not uncommon in patients with SCD, affecting mainly female patients and responding satisfactorily to immunosuppressive treatment. Liver biopsy is helpful in confirming the diagnosis and can be safely performed in the absence of acute vaso-occlusive sickling episodes. Ulcerative colitis is common in the presence of ASC.

Systematic Review: Nutrition and Physical Activity in the Management of Paediatric Nonalcoholic Fatty Liver Disease.

OBJECTIVES: The aim of the study was to evaluate efficacy of nutrition and physical activity interventions in the clinical management of paediatric nonalcoholic fatty liver disease. The prevalence of paediatric nonalcoholic fatty liver disease continues to rise alongside childhood obesity. Weight loss through lifestyle modification is currently first-line treatment, although supplementation of specific dietary components may be beneficial. METHODS: Medline, CINAHL, EMBASE, Scopus, and Cochrane Libraries were systematically searched to identify randomized controlled trials assessing nutritional and physical activity interventions. Primary outcome measures were changes to liver biomarkers assessed by imaging, histology, or serum liver function tests. Study quality was evaluated using the American Dietetic Association Quality Criteria Checklist. RESULTS: Fifteen articles met eligibility criteria investigating nutritional supplementation (vitamin E [n = 6], probiotics [n = 2], omega-3 fatty acids [n = 5]), dietary modification (low glycaemic load [n = 1] and reducing fructose intake [n = 1]). No randomized controlled trials examining physical activity interventions were identified. Vitamin E was ineffective at improving alanine transaminase levels, whereas omega-3 fatty acids decreased hepatic fat content. Probiotics gave mixed results, whereas reduced fructose consumption did not improve primary outcome measures. A low glycaemic load diet and a low-fat diet appeared equally effective in decreasing hepatic fat content and transaminases. Most studies were deemed neutral as assessed by the American Dietetic Association Quality Criteria Checklist. CONCLUSIONS: The limited evidence base inhibits the prescription of specific dietary and/or lifestyle strategies for clinical practice. General healthy eating and physical activity guidelines, promoting weight loss, should remain first-line treatment until high-quality evidence emerges that support specific interventions that offer additional clinical benefit.

AST-to-platelet ratio index in non-invasive assessment of long-term graft fibrosis following pediatric liver transplantation.

Long-term graft fibrosis occurs in the majority of pediatric liver transplant recipients. Serial biopsies to monitor graft health are impractical and invasive. The APRI has been evaluated in pediatric liver disease, but not in the context of post-transplantation fibrosis. We aimed to investigate the validity of APRI as a predictor of long-term graft fibrosis in pediatric liver transplant recipients. This was a retrospective, observational study of a cohort of children who underwent liver transplantation at King's College Hospital between 1989 and 2003, with a relevant dataset available. Protocol liver biopsies were performed at 10-yr follow-up and fibrosis was graded using the Ishak scoring system, with S3-6 denoting "significant fibrosis." APRI was calculated concurrently with biopsy. A total of 39 asymptomatic patients (20 males; median age at transplant, 1.43 yr) underwent protocol liver biopsies at a median of 10.39 yr post-transplantation. APRI was associated with significant fibrosis (p = 0.012). AUROC for APRI as a predictor of significant fibrosis was 0.74 (p = 0.013). The optimal cutoff APRI value for significant fibrosis was 0.45 (sensitivity = 0.67; specificity = 0.79; PPV = 0.67; NPV = 0.79). APRI appears to be a useful non-invasive adjunct in the assessment of significant graft fibrosis in the long-term follow-up of pediatric liver transplant survivors.

Systemic Complications Secondary to Chronic Liver Disease.

The systemic sequelae of chronic liver disease (CLD) may be due to portal hypertension and shunting, malnutrition, and/or a low grade inflammatory state. This article will focus on the consequences of chronic liver disease affecting extrahepatic organs. Portal hypertension underlies many systemic complications of CLD. Aside from varices and ascites, portal hypertension may cause both hepatopulmonary syndrome and portopulmonary hypertension leading to respiratory compromise. Cardiomyopathy may also occur secondary to end stage liver disease. Hepatorenal syndrome is also well recognised and hepatic encephalopathy is a consequence of the effect of liver dysfunction on the brain. Compromise of the immune system is well described in end-stage liver disease leading to sepsis and its consequences. Bony disease including osteoporosis and hepatic arthropathy may both be seen in children with CLD. CLD may be asymptomatic initially but then complications may present as the disease progresses. Furthermore, systemic effects of end stage liver disease may complicate liver transplant. These complications often present insidiously or at the time of acute decompensation. Thus, it is important that healthcare providers are vigilant when caring for children with CLD. This article outlines the secondary complications of CLD with an overview of the definition and diagnosis, pathophysiology, management and prognosis of each.

Management of cholestatic pruritus in paediatric patients with alagille syndrome: the King's College Hospital experience.

OBJECTIVES: The aims of the study were to perform a retrospective observational review of the present management and outcome of cholestatic pruritus in children with Alagille syndrome (AGS) at King's College Hospital and to use results to inform appropriate guidelines. METHODS: A retrospective review of 62 patients diagnosed as having AGS from January 1995 to November 2010 treated at King's College Hospital was performed. The departmental database of the Paediatric Liver Centre was searched to identify all patients and the clinical records were then analysed. RESULTS: Fifty-one (82.3%) patients experienced pruritus and 50 (80.6%) received antipruritic medication. Ursodeoxycholic acid was the most prescribed drug (n = 40). Other drugs prescribed were rifampicin (n = 39), cholestyramine (n = 18), naltrexone (n = 14), alimemazine (n = 13), nonsedating antihistamine agents (n = 7), ondansetron (n = 5), and phenobarbitone (n = 1). Albumin dialysis using the molecular adsorbent recirculation system was used in 1 patient. Sixteen patients (25.8%) were listed for liver transplantation, and 11 had undergone transplantation by November 2010. Patient survival was high at 95.2%. Pruritus resolved permanently in 39.2% (n = 20) of patients. Fifty-five percent (n = 11) of such patients had undergone liver transplantation. Pruritus was controlled by medication in 41.2% (n = 21). Itching remained a significant problem, affecting quality of life in 19.6% of patients (n = 10). CONCLUSIONS: The management of cholestatic pruritus in AGS is difficult and often suboptimal. Pruritus may remain intractable even with combination medical treatment, and at this stage, surgery or liver transplantation is indicated. At our centre, pruritus was successfully treated in 80.4% of patients with medical and surgical management.