Objectives and Study

Background. Adjuvant therapy post-Kasai Portoenterostomy (KPE), has not reduced liver transplantation (LT) in Biliary Atresia (BA). Increasingly, a gut microbiota(GM)-host interplay, in chronic liver disease, is being described.

Aims. To characterise GM, stool short chain fatty acids (SCFAs) and intestinal barrier (IB) markers, in BA, and association with clinical outcomes.

Methods

Stool, plasma, were prospectively collected, in BA infants (n=55) pre-KPE, and 6weeks-,3months-,6months-post-KPE. Age-matched healthy (HC;n=19) and cholestatic control (CC;n=21) stool was collected. Stool 16S rRNA sequencing (GM) and gas chromatography (SCFAs), were performed. IB markers included plasma Claudin-3 and stool calprotectin (SC). Clinical outcomes: 6month-JC (jaundice clearance), 1year-LT, cholangitis by 6month-post-KPE, fibrosis biomarkers [Aspartate Aminotransferase-to-platelet ratio index (APRi); Liver Stiffness Measurement (LSM)]

Results

Pre-KPE, beta-diversity revealed differences in genus-level-clustering, between BA vs HC [p<0.03], not CC [p=0.5] (Fig1). BA incorporated ↓Bifidobacterium [p=0.01], ↑Enterococcus [p=0.01], ↑Clostridium [p=0.04], compared to HC. 6weeks-post-KPE, increased alpha-diversity [p<0.001], increased genus-level-clustering variation [BA:↓Bifidobacterium,p<0.001; ↑Enterococcus,p=0.02], and reduced acetate [p<0.001], was shown in BA vs HC. Pre-KPE, increased alpha-diversity [p=0.04], and reduced acetate [p=0.004], was associated with 1year-LT. Increased propionate was associated with poor 6month-JC [p=0.02]; Clostridium positively correlated with propionate [rs=0.5;p=0.01]. Early-post-KPE, increased alpha-diversity [p=0.01], ↑Streptococcus [p<0.001], ↓Blautia [p=0.03], were associated with poor 6month-JC. APRi positively and negatively correlated with Streptococcus [rs=0.4;p=0.03] and Bifidobacterium [rs=-0.5;p=0.01]. ↓Bifidobacterium, 6weeks-post-KPE, was associated with cholangitis development [p=0.03]. Butyrate, 6weeks-post-KPE, inversely correlated with total bilirubin [rs=-0.3;p=0.02]. Increased Claudin-3, early-post-KPE, was associated with poor 6month-JC [p=0.01] and 1year-LT [p=0.001]. 6months-post-KPE, ↓Blautia [p=0.01], ↓SCFAs [p=0.04], ↑SC [p=0.05], were associated with 1year-LT; Bifidobacterium inversely correlated with LSM [rs=-0.5;p=0.02].

Conclusions

Dysbiosis, characterised by pathobiont enrichment and ‘probiotic’ (Blautia, Bifidobacterium) deficiency, and subsequent SCFA dysregulation, in pre- and early-post-KPE BA, are associated with non-favourable clinical outcomes. Increased intestinal permeability early-post-KPE, is a likely a key factor in GM-BA-pathogenesis. Early microbiota-modulatory therapy may reduce LT in BA.

Objectives and Study

Altered gut microbiota (GM; dysbiosis), have been implicated in the pathogenesis of chronic liver diseases, including Biliary Atresia (BA), and are a major source of endogenous ethanol production. Plasma ethanol concentration was increased in NASH vs obese/healthy controls, and we have previously described an increase in potential ethanol-producing GM in BA.

Aim: To characterise ethanol production in BA, and its association with clinical outcomes, liver disease severity, fibrosis, bacterial translocation (BT) and intestinal permeability (IP).

Methods

Stool and plasma samples, were prospectively collected, in BA infants (n=55) pre-Kasai Portoenterostomy (KPE), 6weeks-, 3months-, and 6months-post-KPE. Plasma ethanol, BT [Lipopolysaccharide-Binding Protein (LBP), D-lactate] and IP [Claudin-3] biomarkers, were measured. Stool 16S-rRNA sequencing was performed for GM genus abundance. Outcomes: 6month jaundice clearance vs 6month jaundice [BA-JC vs BA-J], 1year-liver transplant (LT) status [BA-native liver survivor (NLS) vs BA-LT], Paediatric End Stage Liver Disease (PELD) score, fibrosis [Liver Stiffness Measurement (LSM)].

Results

Plasma ethanol concentration pre-KPE, 6weeks-post-KPE, were not associated with 6month-JC or 1year-LT outcome groups. At 3months-post-KPE, increased ethanol was associated with 1year-LT [BA-NLS, 2.4 mg/L (0, 12.3) vs BA-LT 7 mg/L (1, 140, p=0.02) and predicted the need for LT at 1year-post-KPE, with AUROC of 0.74. At 6months-post-KPE, ethanol was not associated with clinical outcomes. At 3months-post-KPE, ethanol positively correlated with disease severity [PELD,rs=0.4,p=0.02] and fibrosis [LSM,rs=0.5,p=0.01]. Regarding BT, ethanol positively correlated with D-lactate at 6weeks-post-KPE [rs=0.4,p=0.02] and 3months-post-KPE [rs=0.5,p=0.01], but inversely with LBP [3months-post-KPE; rs=-0.3,p=0.05; 6months-post-KPE, rs=-0.5,p=0.01]. No correlation between ethanol and individual genus abundance or IP (Claudin-3), was identified.

Conclusions

Ethanol, at 3months-post-KPE, is associated with LT in BA. Ethanol and D-lactate production are linked, suggesting a role for GM-ethanol-BA pathogenesis. The inverse correlation between LBP and ethanol may reflect a protective role for LBP in early endotoxemia. Mechanistic pathways for ethanol-BA pathogenesis, warrant further investigation.

Objectives and Study

Biliary atresia (BA) is a fibro-obliterative disease of the extrahepatic bile ducts affecting newborns, and is the leading indication for pediatric liver transplant. The etiology remains unknown and there is no effective treatment. Previously, using a toxic model of BA we found that the toxin biliatresone decreases reduced glutathione, causes cell polarity changes and increased epithelial permeability and involves genes in the WNT, Hippo and Notch signaling pathways. Here we wanted to determine if this was applicable to human disease via human cholangiocyte organoids (HCOs) form BA patients.

Methods

We propagated HCOs from BA extrahepatic bile ducts of BA as well as non-BA controls. Bulk RNAseq analysis was performed and enrichment analysis compared between the two groups.

Results

BA-derived organoids demonstrated deformed shapes and poor proliferation capability compared to normal HCOs. There was a mild correlation between mice and human expressed genes. Gene set enrichment analysis looking at gene ontology biological processes showed that biological adhesion, cell junction organization, cell junction assembly were significantly different in BA HOCs from normal controls. And energy derivation by oxidation of organic compounds expression was decreased in BA patients. Though there were some genes in the Wnt, Hippo and Notch signaling pathways that were different between the two groups, they were not identical to the previous genes we and others described in the biliatresone mouse model.

Conclusions

RNAseq from human HOCs display a different pattern than normal human HOCs. Gene ontology biological processes show some similarities to biliatresone induced in murine cholangiocyte, specifically cell polarity and oxidative stress.

Objectives and Study

To describe disease course, treatment, and outcome in a multicentre prospective registry of paediatric autoimmune liver disease (p-AILD) patients, developed within ERN Rare-Liver.

Methods

New cases of AILD in children <18 years-old were enrolled in a prospective European database from 2017-2021. Patients were included in the present study if they had ≥ one year of follow-up. Biochemical data and data on medication were collected at diagnosis and after 12 months. Biochemical remission at 1 year was defined as ALT <45 UI/L and IgG <13.5 g/L.

Chi-square test was used to compare frequencies. One-way ANOVA was used to compare continuously variables.

Results

Eighty-seven patients were enrolled from 6 centres. 43 (49%) were females. Diagnosis was 61 (69%) AIH1, 10 (11%) AIH2 and 16 (18%) ASC. AIH2 were younger (p<0.001) at diagnosis. At presentation, neither ALT nor IgG differ between pAILDs. INR was<1.5 in 77 (87,5%) at presentation.

Initial treatment was mainly prednisolone with or without azathioprine. Almost all p-AILD received prednisolone (86 [98%]) and 70 [80%] received azathioprine. One patient had mycophenolate and one had tacrolimus as first choice. During the first year of treatment 15 (17%) changed medication to mycophenolate (8), budesonide (2), tacrolimus (2), everolimus (1), or biological treatment (2).

At one year of follow up, 75% had achieved normal ALT. AIH2 tended to have a higher rate of ALT lower than 45 U/l (64% AIH1, 90% AIH2, 81% ASC p=0.07). IgG below 13.5 g/l was observed in 67%, with the highest rate among AIH2 (90%) compared to the other p-AILDs (p=0.05). Only 52% had normalised both IgG and ALT.

Conclusions

In this large prospective, multicentre study, we describe the distribution of p-AILD. We found a high degree of normal ALT after one year (75%), which is in line with previous studies.

Objectives and Study

An increased risk for medical complications and morbidity surrounding transfer from paediatric to adult hepatology care is well documented. Health care transition (HCT) requires a continuous dynamic process between paediatric and adult providers and ideally should continue throughout early adulthood. On behalf of the European Reference Network (ERN) RARE-Liver, the Transition Working group conducted a survey within Europe exploring current practice.

Methods

A questionnaire was developed and circulated electronically via ERN members and other national/international professional bodies.

Results

86 responses (56% paediatric) from 66 centres in 27 countries were received with 16 centres providing information from paediatrics and adults members. A liver HCT programme was available in 56% (n=37) centres organised by paediatrics in 32%, adults in 11% and jointly in 54%. 46% of HCT services see >100 patients/year and 32% <20/year. HCT <18yrs is rare (8%) with 70% transferring young people by their 18^th/19^th birthday and 22% ‘whenever they are ready’. Not having a liver HCT programme is mostly due to lack of resource (52%). Overall, evaluation regarding readiness to transition is determined by age (30%), followed by multidisciplinary team decision (26%), transition checklist (5%) and a combination of different factors in 22%. Only 29% of professionals have received training in adolescent care; 80% with paediatric background. The barriers to adequate HCT are illustrated in the figure with the main concern the patient/family’s dependence on the paediatric provider and inadequate communication between teams with 76.7% and 68.6% responding strongly agree/agree respectively. Only 14% of respondents feel that Health authorities in their country see liver HCT as a priority .

Conclusions

The current provision of liver HCT programmes in Europe is variable with regard to size and set-up. Lack of resource is the main reason for absence of HCT programmes and these programmes are not seen as a priority for local Health authorities.

Objectives and Study

Paediatric acute liver failure (PALF) is a severe, potentially fatal condition of variable and often unknown etiology with the need for liver transplantation (pLTx) as only curative treatment in many cases. PALF mostly arises from complete health and impairs physical and mental health, and as existing data are scarce, we aimed to assess long-term outcome and quality of life in PALF patients.

Methods

Patients treated with liver failure in our centre (1988-2021) were evaluated (n=429), cases with chronic liver disease and multi-organ failure of non-hepatic origin were excluded and PALF-SG criteria were applied leading to a cohort of 130 patients. Data were collected retrospectively from patient files and our pLTx database. Of the 79 survivors, 52 could be contacted, and 22 patients/caregivers (1988-2005:14.8%; 2006-2022:34.7%; p=0.05) agreed to QoL assessment. Peds-QL-score-tests and multidimensional fatique scale were used and compared to published controls.

Results

Etiology of PALF in 130 included patients was metabolic (25%), infectious (10%) and in 46% unknown. Most patients had hepatic encephalopathy (72%). 81 patients received pLTx, 22 needed re-transplantation. Mortality was 39.2 % (30% <1 year after PALF), and shows significant improvement over time (1988-2005:54%; 2006–2021:30%; p=0.008). QoL was assessed in 22 patients (13f, 13 pLTx) on average 11.5 (1.8-33) years after PALF. QoL was reduced in child self-report/parent proxy-report after pLTx for PALF (71.9/67.9), especially regarding emotional/school functioning, compared to healthy controls (85.6/85.4), but also to children after pLTx for heterogeneous indications (76-77/73-77). Children after PALF without liver transplantation show higher QoL-values (85.4/84.0). Fatique also affects children after transplantation (64.2/59.0) more than patients after PALF without pLTx (77.7/84.3) and healthy controls (81.8/88.2).

Conclusions

Despite improved outcome PALF remains a disease entity with high mortality even after pLTx. PALF seems to impair long-term quality of life and fatique only if pLTx is performed, whereas patients without liver transplantation achieve near-normal values.