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Background & aims: We have previously shown that the hepatic vagus nerve stimulates the activation of hepatic progenitor cells (HPC), via muscarinic acetylcholine receptor type 3. Given the coproliferation of HPC and hepatic stellate cells (HSC) in acute hepatitis, we determined whether HSC proliferation is also modulated by vagal activity. Methods: We induced acute hepatitis in Wistar rats by injection of galactosamine and lipopolysaccharides. Hepatitis was preceded by hepatic branch vagotomy or sham vagotomy, by electrical stimulation or sham stimulation and by muscarinic receptor antagonist atropine, nicotinic receptor antagonist mecamylamine or saline injection. Rats were sacrificed after 12 and 48 h and HSC numbers were quantified on immunohistochemical stainings. Furthermore, we performed reverse transcriptase-polymerase chain reaction with receptor-specific primers on total RNA from isolated HSC and determined the in vitro proliferation of HSC in response to acetylcholine, atropine and mecamylamine. Results: HSC numbers were significantly lower after vagotomy than after sham vagotomy. Conversely, more HSC were seen after electrical stimulation than after sham stimulation. Atropine resulted in less HSC than saline at both time points, while mecamylamine treatment only diminished HSC after 12 h, suggesting a predominant involvement of muscarinic receptors. Moreover, HSC express muscarinic receptor type 2 mRNA and protein, as well as nicotinic receptor α1, α5, β1 and vasoactive intestinal peptide receptor 1 mRNA. Furthermore, acetylcholine enhanced the in vitro proliferation of HSC, which was inhibited by atropine, but not by mecamylamine. Conclusions: We show here that the hepatic vagus nerve stimulates HSC proliferation, most likely through binding of acetylcholine on muscarinic receptor type 2. [ABSTRACT FROM AUTHOR]

Background/Objective: Carbon monoxide (CO) produced by haem-oxygenase isoforms (HO-1 & HO-2) is involved in the regulation of systemic vascular tone. We aimed to elucidate the vasoregulatory role of CO in the microcirculation in normal and thioacetamide cirrhotic rat livers. Methods: Haem-oxygenase expression was examined by Western blot. Total HO enzymatic activity was measured spectrophotometrically. Sensitivity of hepatic stellate cells (HSCs) to CO-mediated relaxation was studied by a stress-relaxed-collagen-lattice model. To define the relative role of CO, the CO-releasing molecule CORM-2, the HO-inhibitor zinc protoporphyrin-IX and the HO-1 inducer hemin were added to an in situ liver perfusion set-up. The topography of vasoactive CO production was evaluated by applying different CO- and nitric oxide-trapping reagents in the liver perfusion set-up and by immunohistochemistry. Results: Western blot showed decreased expression of both HO isoenzymes ( P<0.036 for HO-1; P<0.001 for HO-2) in cirrhotic vs normal rat livers, confirmed by the HO-activity assay ( P=0.004). HSCs relaxed on exposure to CORM-2 ( P=0.013). The increased intrahepatic vascular resistance (IHVR) of cirrhotic rats was attenuated by perfusion with CORM-2 ( P=0.016) and pretreatment with hemin ( P<0.001). Inhibition of HO caused a dose-related increase in IHVR in normal and cirrhotic liver. In normal liver, the haemodynamically relevant CO production occurred extrasinusoidally, while intrasinusoidally HO-1 predominantly regulated the microcirculation in cirrhotic livers. Conclusion: We demonstrate a role for CO and HO in the regulation of normal and cirrhotic microcirculation. These findings are of importance in the pathophysiology of portal hypertension and establish CO/HO as novel treatment targets. [ABSTRACT FROM AUTHOR]

Aim: Hepatoblastoma (HB), the most common pediatric malignant liver tumor, is treated with chemotherapy to facilitate surgical resection. Previous studies suggest that HB acquires chemoresistance via increased expression of multidrug resistance protein 1 (MDR1, ABCC1). There is no well established evidence that this also occurs in the clinical setting and little is known about the effects of chemotherapeutic treatments on HB in situ. Methods: Clinical and histopathological features and expression patterns of ABC transporters in diagnostic needle biopsies from 7 HBs taken before chemotherapy were compared with those in surgically resected tumors. To understand the mechanism s leading to chemoresistance we also investigated the involvement of hypoxia on protein expression and functional activity of drug transporters (BCRP and MDR1) in cultures of HepG2 human HB cells. Results: We found that chemotherapeutical treatment of HBs led to an increased expression of the breast cancer resistance protein (BCRP, ABCG2) in all patients studied. There was no change in the expression pattern of MDR1 or other ABC transporters. Chemotherapy-induced specific vascular abnormalities associated with areas of necrosis and fibrosis were seen in all cases, suggesting tumor hypoxia. The observations of increased BCRP expression in hypoxic areas of three-dimensional HepG2 aggregates and the enhanced BCRP function in monolayer cultures of HepG2 cells under hypoxic conditions, support a role for hypoxia in enhanced BCRP expression. Conclusions: Chemotherapeutical treatment of HB leads to vascular alterations that modify the tumor microenvironment, and increased BCRP expression in which hypoxia might play a role. No evidence was found for upregulation of MDR1 in HBs as suggested from previous experimental studies. [ABSTRACT FROM AUTHOR]

Background and Aim: Congenital portosystemic veno-venous malformations are rare abnomalities that often remain undiagnosed. Typically they are classified by their anatomical characteristics according to Morgan (extrahepatic, Abernethy malformations type Ia,b and II) and Park (intrahepatic, types 1–4). However, their clinical presentation is less dependent on the anatomical type. Method: We reviewed the clinical characteristics of six cases drawn from our files (from 1970 to 2006). Results: One patient, a 25-year-old male, had extrahepatic shunting whereby the liver receives only arterial blood because the portal vein (PV) connects with the inferior caval vein (ICV) (Abernethy Ib); he presented with episodes of jaundice and pruritus. Three patients had extrahepatic shunting with patent intrahepatic portal veins, but with shunting of splenomesenterial blood towards the ICV (Abernethy II); these included a 66-year-old male with hepatic encephalopathy, a 17-year-old female with (porto?-)pulmonary hypertension without portal hypertension, and a 33-year-old female with epidsodes of acute pain secondary to spontaneous bleeding within a primary liver tumor. Two patients had intrahepatic shunting; these included an 8-year-old boy who was diagnosed incidentally during work-up for abnormal liver enzymes with a communication between right PV and ICV (Park type 1), and a 59-year-old male with multiple PV-ICV–shunts in several liver segments (Park, type 4) who presented with hepatic encephalopathy. Conclusion: Patients often present with signs of hepatic shunting (encephalopathy, pulmonary hypertension, hepatopulmonary syndrome, and/or hypoglycemia) with relative sparing of the synthetic liver function in the absence of portal hypertension. Some shunts present with space-occupying lesions (focal nodular hyperplasia, hepatocellular carcinoma, nodular regenerative hyperplasia, etc.) or biliary atresia. Finally, some cases are detected incidentally. [ABSTRACT FROM AUTHOR]

Background: Cystinosis is a metabolic disease characterized by accumulation of cystine in different organs and tissues, leading to potentially life-threatening organ dysfunction. Infantile cystinosis typically leads to end-stage renal disease, necessitating renal replacement therapy. Liver disease in cystinosis is rare and is mostly reported as nodular regenerative hyperplasia leading to portal hypertension. Methods: Two patients with infantile cystinosis developed cholestatic liver disease (increasing alkaline phosphatases, gamma-glutamyltransferase and mild increase in transaminases). Severe accumulation of cystine was demonstrated on liver biopsy, predominantly localized in Kupffer cells, together with morphological signs of sclerosing cholangitis on liver biopsy. One patient showed changes compatible with sclerosing cholangitis on magnetic resonance imaging. Therapy with ursodeoxycholic acid led to biochemical improvement in one and stabilization in the other patient. Conclusion: Long-term infantile nephropathic cystinosis can be associated with a form of sclerosing cholangitis, which can respond to therapy with ursodeoxycholic acid. [ABSTRACT FROM AUTHOR]

A 14-year-old girl, with no medical history except for a preterm birth at 7 months, presented with clinical and laboratory signs of pancytopenia. Radiological investigation, including abdominal ultrasound and magnetic resonance imaging revealed a focal irregular main portal vein wall and splenomegaly. Endoscopy showed major esophageal and gastric varices. Definitive diagnosis was made by direct, transhepatic portography revealing a focal stenosis associated with a dissection of the main portal vein. Percutaneous treatment including balloon angioplasty and stent placement resulted in complete re-expansion of the portal vein main branch. Clinical and radiological follow-up over 5 years showed complete disappearance of all clinical signs and normalization of laboratory data and splenic volume. [ABSTRACT FROM AUTHOR]

Background: HFE-related hereditary haemochromatosis (HH) is a common autosomal recessive disorder with clinical manifestations ranging from asymptomatic disease to possible life-threatening complications. Cirrhosis, hepatocellular carcinoma, diabetes mellitus or osteoporosis can develop in HH patients not treated or monitored optimally. The purpose of this study was to develop key-interventions (KI's) to measure and improve the quality of care delivered to patients diagnosed with HH.Methods: A RAND-Modified Delphi method was used to develop KI's. In the first round of a scoring form to prioritize the recommendations extracted from evidence-based guidelines was circulated between experts. The results of this survey were discussed in a consensus meeting, followed by a final appraisal of the selected recommendations. This resulted in a list of measurable KI's.Results: Initially, 41 key recommendations on screening, diagnosis and treatment/management were extracted from three existing guidelines on HH (European Association for the Study of the Liver, American Association for the Study of Liver Diseases and Dutch guideline on HH). Finally, a core set of 24 recommendations resulted in 15 KI's.Conclusions: This manuscript presents the results of the process to develop KI's to measure and improve the quality of care for patients with HH. [ABSTRACT FROM AUTHOR]

Objective: Stable isotope studies have shown that hepatic de novo lipogenesis (DNL) plays an important role in the pathogenesis of intrahepatic lipid (IHL) deposition. Furthermore, previous research has demonstrated that fructose 1-phosphate (F1P) not only serves as a substrate for DNL, but also acts as a signalling metabolite that stimulates DNL from glucose. The aim of this study was to elucidate the mediators of F1P-stimulated DNL, with special focus on two key regulators of intrahepatic glucose metabolism, i.e., glucokinase regulatory protein (GKRP) and carbohydrate response element binding protein (ChREBP)., Methods: Aldolase B deficient mice (Aldob -/- ), characterized by hepatocellular F1P accumulation, enhanced DNL, and hepatic steatosis, were either crossed with GKRP deficient mice (Gckr -/- ) or treated with short hairpin RNAs directed against hepatic ChREBP., Results: Aldob -/- mice showed higher rates of de novo palmitate synthesis from glucose when compared to wildtype mice (p < 0.001). Gckr knockout reduced de novo palmitate synthesis in Aldob -/- mice (p = 0.017), without affecting the hepatic mRNA expression of enzymes involved in DNL. In contrast, hepatic ChREBP knockdown normalized the hepatic mRNA expression levels of enzymes involved in DNL and reduced fractional DNL in Aldob -/- mice (p < 0.05). Of interest, despite downregulation of DNL in response to Gckr and ChREBP attenuation, no reduction in intrahepatic triglyceride levels was observed., Conclusions: Both GKRP and ChREBP mediate F1P-stimulated DNL in aldolase B deficient mice. Further studies are needed to unravel the role of GKRP and hepatic ChREBP in regulating IHL accumulation in aldolase B deficiency., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier GmbH.. All rights reserved.)

Patients with classic galactosemia (CG), an inborn error of galactose metabolism, suffer from impairments in cognition, including language processing. Potential causes are atypical brain oscillations. Recent electroencephalogram (EEG) showed differences in the P300 event-related-potential (ERP) and alterations in the alpha/theta-range during speech planning. This study investigated whether transcranial alternating current stimulation (tACS) at theta-frequency compared to sham can cause a normalization of the ERP post stimulation and improves language performance. Eleven CG patients and fourteen healthy controls participated in two tACS-sessions (theta 6.5 Hz/sham). They were engaged in an active language task, describing animated scenes at three moments, that is, pre/during/post stimulation. Pre and post stimulation, behavior (naming accuracy, voice-onset-times; VOT) and mean-amplitudes of ERP were compared, by means of a P300 time-window analysis and cluster-based-permutation testing during speech planning. The results showed that theta stimulation, not sham, significantly reduced naming error-percentage in patients, not in controls. Theta did not systematically speed up naming beyond a general learning effect, which was larger for the patients. The EEG analysis revealed a significant pre-post stimulation effect (P300/late positivity), in patients and during theta stimulation only. In conclusion, theta-tACS improved accuracy in language performance in CG patients compared to controls and altered the P300 and late positive ERP-amplitude, suggesting a lasting effect on neural oscillation and behavior., (© 2024 The Authors. Journal of Inherited Metabolic Disease published by John Wiley & Sons Ltd on behalf of SSIEM.)

Background and Aims: Acute-on-chronic liver failure (ACLF) is a complication of cirrhosis characterized by multiple organ failure and high short-term mortality. The pathophysiology of ACLF involves elevated systemic inflammation leading to organ failure, along with immune dysfunction that heightens susceptibility to bacterial infections. However, it is unclear how these aspects are associated with recovery and nonrecovery in ACLF., Approach and Results: Here, we mapped the single-cell transcriptome of circulating immune cells from patients with ACLF and acute decompensated (AD) cirrhosis and healthy individuals. We further interrogate how these findings, as well as immunometabolic and functional profiles, associate with ACLF-recovery (ACLF-R) or nonrecovery (ACLF-NR). Our analysis unveiled 2 distinct states of classical monocytes (cMons). Hereto, ACLF-R cMons were characterized by transcripts associated with immune and stress tolerance, including anti-inflammatory genes such as RETN and LGALS1 . Additional metabolomic and functional validation experiments implicated an elevated oxidative phosphorylation metabolic program as well as an impaired ACLF-R cMon functionality. Interestingly, we observed a common stress-induced tolerant state, oxidative phosphorylation program, and blunted activation among lymphoid populations in patients with ACLF-R. Conversely, ACLF-NR cMon featured elevated expression of inflammatory and stress response genes such as VIM , LGALS2 , and TREM1 , along with blunted metabolic activity and increased functionality., Conclusions: This study identifies distinct immunometabolic cellular states that contribute to disease outcomes in patients with ACLF. Our findings provide valuable insights into the pathogenesis of ACLF, shedding light on factors driving either recovery or nonrecovery phenotypes, which may be harnessed as potential therapeutic targets in the future., (Copyright © 2024 The Author(s). Published by Wolters Kluwer Health, Inc.)

Background & Aims: Wilson disease (WD) is caused by accumulation of copper primarily in the liver and brain. During maintenance therapy of WD with D-penicillamine, current guidelines recommend on-treatment ranges of urinary copper excretion (UCE) of 200-500 μg/24 h and serum non-ceruloplasmin-bound copper (NCC) of 50-150 μg/L. We compared NCC (measured by two novel assays) and UCE from patients with clinically stable WD on D-penicillamine therapy with these recommendations., Methods: This is a secondary analysis of data from the Chelate trial (NCT03539952) that enrolled physician-selected patients with clinically stable WD on D-penicillamine maintenance therapy (at an unaltered dose for at least 4 months). We analyzed laboratory samples from the first screening visit, prior to interventions. NCC was measured by either protein speciation (NCC-Sp) using anion exchange high-performance liquid chromatography protein speciation followed by copper determination with inductively coupled plasma mass spectroscopy or as exchangeable copper (NCC-Ex). NCC-Sp was also analyzed in healthy controls (n = 75)., Results: In 76 patients with WD with 21.3±14.3 average treatment-years, NCC-Sp (mean±SD: 56.6±26.2 μg/L) and NCC-Ex (mean±SD: 57.9±24.7 μg/L) were within the 50-150 μg/L target in 61% and 54% of patients, respectively. In addition, 36% and 31%, respectively, were even below the normal ranges (NCC-Sp: 46-213 μg/L, NCC-Ex: 41-71 μg/L). NCC-Ex positively correlated with NCC-Sp (r 2  = 0.66, p < 0.001) but with systematic deviation. UCE was outside the 200-500 μg/24 h target range in 58%. Only 14/69 (20%) fulfilled both the NCC-Sp and UCE targets. Clinical or biochemical signs of copper deficiency were not detected., Conclusion: Clinically stable patients with WD on maintenance D-penicillamine therapy frequently have lower NCC-Sp or higher UCE than current recommendations without signs of overtreatment. Further studies are warranted to identify appropriate target ranges of NCC-Sp, NCC-Ex and UCE in treated WD., Impact and Implications: Chelator treatment of patients with Wilson disease (WD) is currently guided by measurements of non-ceruloplasmin-bound copper (NCC) and 24 h urinary copper excretion (UCE) but validation is limited. In 76 adults with ≈21 years history of treated WD and clinically stable disease on D-penicillamine therapy, NCC was commonly found to be below normal values and recommended target ranges whether measured by protein speciation (NCC-Sp) or as exchangeable copper (NCC-Ex), while UCE values were above the recommended target range in 49%. Common wisdom would suggest overtreatment in these cases, but no clinical or biochemical signs of copper deficiency were observed. Exploratory analysis of liver enzymes suggested that NCC below levels seen in controls may be beneficial, while the relation to UCE was less clear. The data calls for critical re-evaluation of target ranges for treatment of WD, specific for drug and laboratory methodology., Clinical Trial Number: (NCT03539952)., (© 2024 The Authors.)

Classic galactosemia (CG, OMIM #230400, ORPHA: 79,239) is a hereditary disorder of galactose metabolism that, despite treatment with galactose restriction, affects brain function in 85% of the patients. Problems with cognitive function, neuropsychological/social emotional difficulties, neurological symptoms, and abnormalities in neuroimaging and electrophysiological assessments are frequently reported in this group of patients, with an enormous individual variability. In this review, we describe the role of impaired galactose metabolism on brain dysfunction based on state of the art knowledge. Several proposed disease mechanisms are discussed, as well as the time of damage and potential treatment options. Furthermore, we combine data from longitudinal, cross-sectional and retrospective studies with the observations of specialist teams treating this disease to depict the brain disease course over time. Based on current data and insights, the majority of patients do not exhibit cognitive decline. A subset of patients, often with early onset cerebral and cerebellar volume loss, can nevertheless experience neurological worsening. While a large number of patients with CG suffer from anxiety and depression, the increased complaints about memory loss, anxiety and depression at an older age are likely multifactorial in origin., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The author(s) declared that they were an editorial board member of Frontiers, at the time of submission. This had no impact on the peer review process and the final decision., (Copyright © 2024 Panis, Vos, Barić, Bosch, Brouwers, Burlina, Cassiman, Coman, Couce, Das, Demirbas, Empain, Gautschi, Grafakou, Grunewald, Kingma, Knerr, Leão-Teles, Möslinger, Murphy, Õunap, Pané, Paci, Parini, Rivera, Scholl-Bürgi, Schwartz, Sdogou, Shakerdi, Skouma, Stepien, Treacy, Waisbren, Berry and Rubio-Gozalbo.)

Background: Although patients with hereditary fructose intolerance (HFI) generally have a good prognosis on a fructose-restricted diet, relatively little is known about their quality of life. The aim of this study was to investigate the quality of life in adult patients with HFI in comparison to patients with dietary-treated, classical phenylketonuria (PKU)., Methods: Patients with HFI and patients with classical PKU were recruited from the adult metabolic centers in The Netherlands and Belgium and via social media. Patients were asked to fill out the 36-item Short Form Health survey (SF-36) and a modified PKU Quality Of Life (PKU-QoL) questionnaire., Results: Patients with HFI (n = 19) did not report any restrictions in their health-related quality of life, except for vitality and general mental health, which were scored more unfavorable compared to patients with PKU (n = 19) (p < 0.05, adjusted for level of education and country of origin). The results from the modified PKU-QoL demonstrated a statistically significantly greater impact of the disease in the social domain in HFI. A substantial proportion of both HFI and PKU patients (21%) reported a great to severe emotional impact of their disease. Finally, patients with HFI experienced statistically significantly less food temptations, less guilt if dietary restrictions not followed, and less overall difficulty following dietary restrictions., Conclusions: Although patients with HFI showed to have a generally good quality of life, they scored lower on vitality and general mental health, and reported a greater social impact of the disease. These aspects deserve further study and clinical attention., Competing Interests: Declaration of Competing Interest MCGJB received consultancy fees from Arrowhead and Editas Medicine., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Abnormal polyol metabolism is predominantly associated with diabetes, where excess glucose is converted to sorbitol by aldose reductase (AR). Recently, abnormal polyol metabolism has been implicated in phosphomannomutase 2 congenital disorder of glycosylation (PMM2-CDG) and an AR inhibitor, epalrestat, proposed as a potential therapy. Considering that the PMM2 enzyme is not directly involved in polyol metabolism, the increased polyol production and epalrestat's therapeutic mechanism in PMM2-CDG remained elusive. PMM2-CDG, caused by PMM2 deficiency, presents with depleted GDP-mannose and abnormal glycosylation. Here, we show that, apart from glycosylation abnormalities, PMM2 deficiency affects intracellular glucose flux, resulting in polyol increase. Targeting AR with epalrestat decreases polyols and increases GDP-mannose both in patient-derived fibroblasts and in pmm2 mutant zebrafish. Using tracer studies, we demonstrate that AR inhibition diverts glucose flux away from polyol production toward the synthesis of sugar nucleotides, and ultimately glycosylation. Finally, PMM2-CDG individuals treated with epalrestat show a clinical and biochemical improvement., Competing Interests: Declaration of interests Mayo Clinic and E.M. have a financial interest related to this research. This research has been reviewed by the Mayo Clinic Conflict of Interest Review Board and is being conducted in compliance with Mayo Clinic Conflict of Interest policies. E.M. has the following patents planned, issued, or pending: application title, “Methods and Materials for Treating Glycosylation Disorders”; application # 16/973,210; filing date, 12/08/2020; Mayo Case # 2018-132., (Copyright © 2023 The Author(s). Published by Elsevier Inc. All rights reserved.)

The virome remains an understudied domain of the human microbiome. The role of commensal viruses on the outcome of infections with known pathogens is not well characterized. In this study we aimed to characterize the longitudinal plasma virome dynamics in chronic hepatitis B virus (HBV) infected patients. Eighty-five longitudinal plasma samples were collected from 12 chronic HBV infected individuals that were classified in the four stages of HBV infection. The virome was characterized with an optimized viral extraction protocol and deep-sequenced on a NextSeq 2500 platform. The plasma virome was primarily composed of members of the Anello - Flavi -, and Hepadnaviridae (HBV) families. The virome structure and dynamics did not correlate with the different stages of chronic HBV infection nor with the administration of antiviral therapy. We observed a higher intrapersonal similarity of viral contigs. Genomic analysis of viruses observed in multiple timepoint demonstrated the presence of a dynamic community. This study comprehensively assessed the blood virome structure in chronic HBV infected individuals and provided insights in the longitudinal development of this viral community., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Thijssen, Tacke, Van Espen, Cassiman, Naser Aldine, Nevens, Van Ranst, Matthijnssens and Pourkarim.)

Background: Wilson disease is an inherited disorder of copper transport. Whereas penicillamine is used therapeutically to re-establish copper balance, trientine is indicated for patients with penicillamine intolerance. We aimed to compare penicillamine with trientine tetrahydrochloride (TETA4) for maintenance therapy in patients with Wilson disease., Methods: We conducted a randomised, open-label, non-inferiority, phase 3 trial at 15 health-care centres across nine countries (patients were recruited from 13 of these health-care centres across Brazil, Europe, and the USA). We enrolled patients aged 18-75 years with stable Wilson disease who were treated for at least 1 year with penicillamine. Patients entered a 12-week period to determine stability through clinical assessment by site investigators and predefined thresholds for serum non-caeruloplasmin-bound copper (NCC; by an exchangeable copper assay; 25-150 μg/L), 24 h urinary copper excretion (100-900 μg/24 h), and alanine aminotransferase (ALT; <2 × upper limit of normal). Stable patients were randomly assigned (1:1) to continue receiving the maintenance twice daily dose of oral penicillamine or switched mg-for-mg to oral TETA4 centrally with a web-based system using minimisation. The primary endpoint, assessed 24 weeks after randomisation, was NCC by speciation assay. The non-inferiority margin of mean difference in NCC by speciation assay was -50 μg/L, as estimated by a general linear model for repeated visits, adjusted for baseline values. Further data on safety and efficacy were collected during a 24-week extension period. Data were analysed using an intention-to-treat approach. Safety was assessed in all patients who received at least one dose of study treatment. This study is registered with ClinicalTrials.gov, NCT03539952 (active, not recruiting)., Findings: Between June 4, 2018, and March 10, 2020, 77 patients were screened. 53 patients were randomly assigned (27 to the penicillamine group and 26 to the TETA4 group). After 24 weeks, the mean difference in serum NCC by speciation assay between the penicillamine group and TETA4 group was -9·1 μg/L (95% CI -24·2 to 6·1), with the lower limit of the 95% CI within the defined non-inferiority margin. At 24 weeks, urinary copper excretion was lower with TETA4 than with penicillamine (mean difference 237·5 μg/24 h (99% CI 115·6 to 359·4). At 48 weeks, TETA4 remained non-inferior to penicillamine in terms of NCC by speciation assay (mean difference NCC -15·5 μg/L [95% CI -34·5 to 3·6]). Urinary copper excretion at 48 weeks remained in the expected range for well treated patients in both study groups, and the mean difference (124·8 μg/24 h [99% CI -37·6 to 287·1]) was not significantly different. At 24 weeks and 48 weeks, masked clinical adjudication of stability assessed by three independent clinicians confirmed clinical stability (100%) of all participants, in agreement with the stability seen with the NCC by speciation assay. There were no notable changes in either the Clinical Global Impression of Change or Unified Wilson Disease Rating Scale (neurological assessment) from baseline (pre-randomisation) at weeks 24 and 48. The mean change in serum total copper from baseline to 24 weeks was 17·6 μg/L (99% CI -9·5 to 44·7) with penicillamine and -6·3 μg/L (-34·7 to 22·1) with TETA4, and the mean change in serum total caeruloplasmin from baseline to 24 weeks was 1·8 mg/L (-19·2 to 22·8) with penicillamine and -2·2 mg/L (-6·1 to 1·7) with TETA4. All liver enzymes were similar at 24 weeks and 48 weeks, with the exception of elevated ALT concentration at 48 weeks for patients in the TETA4 group. Penicillamine was associated with three post-randomisation serious adverse events (leukopenia, cholangiocarcinoma, and hepatocellular cancer); none were reported for TETA4. The most common treatment-emergent adverse events were headache for penicillamine (five [19%] of 27 patients vs two [8%] of 26) and abdominal pain for TETA4 (one [4%] vs four [15%]); all treatment-emergent adverse events resolved and were mild to moderate. One patient developed a rash with TETA4 that resolved on discontinuation of therapy., Interpretation: The efficacy of TETA4 as oral maintenance therapy was non-inferior to penicillamine and well tolerated in adults with Wilson disease., Funding: Orphalan., Competing Interests: Declaration of interests MLS has received research (institutional) grants from Orphalan, Alexion, and Vivet Therapeutics and is the Chair (unpaid) of the Medical Advisory Committee of the Wilson Disease Association. DC has received research (institutional) grants, consulting fees, and speaker's fees from Orphalan. AA has received research (institutional) grants from Orphalan, Alexion, Univar, and the Wilson Disease Association; consulting fees from Alexion and Orphalan; speaker's fee from Orphalan; travel assistance to meetings from Orphalan and Univar; and advisory board payments from Alexion, Univar, and Orphalan. GD has received consulting fees from Orphalan, Alexion, and Univar, and speaker's fees from Orphalan. EC-B has received speaker's fees from Orphalan and travel assistance to meetings from Orphalan, and advisory board payments from Alexion and Orphalan. AP has received research (institutional) grants from Orphalan; consulting fees from Alexion, Orphalan, and Vivet Therapeutics; speaker's fees from Orphalan, Alexion, and Univar; and advisory board payments from Alexion, Orphalan, Univar, and Vivet Therapeutics. AC has received research (institutional) grants from Orphalan, Alexion, Public Heath Research Institute in Canada; consulting fees from Wilson Therapeutics, Alexion, Vivet Therapeutics, and Orphalan; speaker's fees from Ever Pharma; and travel assistance to meetings from Orphalan. KD'H and ND are IDDI (biostatistical CRO for the trial) employees. COFK is an Orphalan employee. PO has received research (institutional) grants from Orphalan, Alexion, Univar, and Vivex, and speaker's fees and travel assistance to meetings from Orphalan, and is convenor of the Aarhus Wilson Disease Symposium. JM has received consulting fees from Orphalan and Alexion, and speaker's fees from Shire. KHW has received research (institutional) grants from Orphalan; consulting fees from Orphalan, Univar, Pfizer, Alexion, and Vivet Therapeutics; speaker's fees from Falk, AbbVie, Alexion, and Orphalan; travel assistance to meetings from Alexion and Univar; and advisory board payments from Ultragenyx. All other authors declare no competing interests., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Peroxisome proliferator-activated receptor γ (PPARγ) is the master regulator of adipocyte differentiation, and mutations that interfere with its function cause lipodystrophy. PPARγ is a highly modular protein, and structural studies indicate that PPARγ domains engage in several intra- and inter-molecular interactions. How these interactions modulate PPARγ's ability to activate target genes in a cellular context is currently poorly understood. Here we take advantage of two previously uncharacterized lipodystrophy mutations, R212Q and E379K, that are predicted to interfere with the interaction of the hinge of PPARγ with DNA and with the interaction of PPARγ ligand binding domain (LBD) with the DNA-binding domain (DBD) of the retinoid X receptor, respectively. Using biochemical and genome-wide approaches we show that these mutations impair PPARγ function on an overlapping subset of target enhancers. The hinge region-DNA interaction appears mostly important for binding and remodelling of target enhancers in inaccessible chromatin, whereas the PPARγ-LBD:RXR-DBD interface stabilizes the PPARγ:RXR:DNA ternary complex. Our data demonstrate how in-depth analyses of lipodystrophy mutants can unravel molecular mechanisms of PPARγ function., (© 2022. The Author(s).)

Porphyrias are rare metabolic disorders of the haem synthesis. They can present with acute neurovisceral attacks, cutaneous symptoms, or a combination of both. As they present with a wide variety of clinical symptoms, diagnosis is often delayed and correct interpretation of porphyria-related tests remains a challenge for many physicians. We developed and validated two algorithms for the laboratory diagnosis of porphyrias based on presenting symptoms. Based on a literature search and clinical/laboratory expertise, we developed algorithms for acute and cutaneous porphyrias. We validated these algorithms using all porphyria related laboratory test requests between January 1st 2000 and September 30th 2020 in UZ Leuven. In addition, we also evaluated our algorithm using samples from the European porphyria network (EPNET) external quality assessment scheme (2010-2021). Sensitivity of the algorithm for acute porphyria was 100.0% [74.9%-100.0%] (13 acute intermittent porphyria (AIP) and 1 variegate porphyria [VP]) with a specificity of 98.5% [91.0%-100.0%] (65 patients). Sensitivity of the algorithm for cutaneous porphyria was 100% [95.1%-100.0%] (7 VP, 59 porphyria cutanea tarda (PCT), 23 erythropoietic protoporphyria (EPP), 2 X-linked erythropoietic protoporphyria [XLEPP]) with a specificity of 93.9% [82.9%-98.5%]. There were no diagnostic samples of other types of porphyria. The algorithms correctly identified 18 of the 19 EPNET porphyria cases. One of the two hereditary coproporphyria cases was missed. The algorithms for acute and cutaneous porphyria showed high sensitivity and specificity and can be used to aid the clinician in correctly interpreting the laboratory findings of porphyria-related tests., (© 2022 SSIEM.)

One-year data from EXPLORE Part A showed high disease burden and impaired quality of life (QOL) in patients with acute hepatic porphyria (AHP) with recurrent attacks. We report baseline data of patients who enrolled in EXPLORE Part B for up to an additional 3 years of follow-up. EXPLORE B is a long-term, prospective study evaluating disease activity, pain intensity, and QOL in patients with AHP with ≥1 attack in the 12 months before enrollment or receiving hemin or gonadotropin-releasing hormone prophylaxis. Data were evaluated in patients with more (≥3 attacks or on prophylaxis treatment) or fewer (<3 attacks and no prophylaxis treatment) attacks. Patients in the total population (N = 136), and more (n = 110) and fewer (n = 26) attack subgroups, reported a median (range) of 3 (0-52), 4 (0-52), and 1 (0-2) acute attacks, respectively, in the 12 months prior to the baseline visit. Pain, mood/sleep, digestive/bladder, and nervous system symptoms were each experienced by ≥80% of patients; most received hemin during attacks. Almost three-quarters of patients reported chronic symptoms between attacks, including 85% of patients with fewer attacks. Pain intensity was comparable among both attack subgroups; most patients required pain medication. All groups had diminished QOL on the EuroQol visual analog scale and the European Organisation for Research and Treatment of Cancer Quality-of-life Questionnaire Core 30 versus population norms. Patients with AHP with recurrent attacks, even those having fewer attacks, experience a high disease burden, as evidenced by chronic symptoms between attacks and impaired QOL., (© 2022 The Authors. Journal of Inherited Metabolic Disease published by John Wiley & Sons Ltd on behalf of SSIEM.)

Tyrosinemia type 1 (TT1) and phenylketonuria (PKU) are both inborn errors of phenylalanine-tyrosine metabolism. Neurocognitive and behavioral outcomes have always featured in PKU research but received less attention in TT1 research. This study aimed to investigate and compare neurocognitive, behavioral, and social outcomes of treated TT1 and PKU patients. We included 33 TT1 patients (mean age 11.24 years; 16 male), 31 PKU patients (mean age 10.84; 14 male), and 58 age- and gender-matched healthy controls (mean age 10.82 years; 29 male). IQ (Wechsler-subtests), executive functioning (the Behavioral Rating Inventory of Executive Functioning), mental health (the Achenbach-scales), and social functioning (the Social Skills Rating System) were assessed. Results of TT1 patients, PKU patients, and healthy controls were compared using Kruskal-Wallis tests with post-hoc Mann-Whitney U tests. TT1 patients showed a lower IQ and poorer executive functioning, mental health, and social functioning compared to healthy controls and PKU patients. PKU patients did not differ from healthy controls regarding these outcome measures. Relatively poor outcomes for TT1 patients were particularly evident for verbal IQ, BRIEF dimensions "working memory", "plan and organize" and "monitor", ASEBA dimensions "social problems" and "attention problems", and for the SSRS "assertiveness" scale (all p values <0.001). To conclude, TT1 patients showed cognitive impairments on all domains studied, and appeared to be significantly more affected than PKU patients. More attention should be paid to investigating and monitoring neurocognitive outcome in TT1 and research should focus on explaining the underlying pathophysiological mechanism., (© 2022 The Authors. Journal of Inherited Metabolic Disease published by John Wiley & Sons Ltd on behalf of SSIEM.)

Acute hepatic porphyrias (AHP) are a group of four different rare to ultra-rare, severely debilitating, and sometimes fatal diseases that significantly impact patients' lives: 5-aminolevulinic acid (ALA) dehydratase deficiency porphyria (ADP), acute intermittent porphyria (AIP), hereditary coproporphyria (HCP), and variegate porphyria (VP). Based on literature estimates, a conservative estimate of the number of AHP patients in Belgium requiring treatment, defined as patients experiencing recurrent attacks and/or chronic debilitating symptoms, is likely limited to 11-34 patients. These patients face a considerable unmet need, as there is currently no pharmaceutical treatment available that effectively prevents attacks and has an impact on other chronic symptoms of the disease.A panel consisting of the two European Porphyria Network 1 (EPNet) centers in Belgium (Center for inborn errors of metabolism of UZ Leuven and the 'Centre Belge des Porphyries' of Erasme Hospital and LHUB-ULB) participated in an advisory board on 24 January 2020. Representatives of the sponsoring pharmaceutical company, Alnylam Pharmaceuticals, organized and attended the meeting. The objective of the meeting was to obtain expert input on the state-of-the-art clinical practice of AHP in Belgium. Following this meeting, this expert consensus statement was drafted, in collaboration with and coordinated by the EPNet centers in Belgium. This statement provides an overview of the state-of-the art in AHP, by means of a concise overview of AHP pathophysiology, clinical manifestations, and burden of disease, (Belgian) epidemiology, treatments, and proposed organization of care.

Profit-driven games with the availability and prices of chenodeoxycholic acid led to the discontinuation of proper treatment for this cerebrotendinous xanthomatosis patient with disastrous consequences to his health., (© 2021 SSIEM.)

The patient, a boy born in 1991, showed pronounced polyostotic fibrous dysplasia due to McCune-Albright syndrome, as well as Gilbert syndrome and Charcot-Marie-Tooth neuropathy caused by a DNM2 mutation. In addition, the patient, his sister, mother and maternal grandfather had intermittently increased plasma arginine and lysine levels, most probably due to heterozygosity for a novel pathogenic SLC7A2 variant., (© 2021 Kristl G. Claeys et al. published by Sciendo.)

Background: Donor hepatectomy and liver implantation time reduce long-term graft and patient survival after liver transplantation. It is not known whether these surgical times influence early outcomes after liver transplantation., Methods: This single-center study evaluated the effect of donor hepatectomy and implantation time on the risk of nonanastomotic biliary strictures (NAS) occurring within 1 year and of early allograft dysfunction (EAD) after deceased-donor solitary liver transplantation, adjusting for other donors, recipient, and surgical factors., Results: Of 917 transplants performed between January 2000 and December 2016, 106 (11.56%) developed NAS and 247 (27%) developed EAD. Donor hepatectomy time (median 35 min, IQR: 26-46) was an independent risk factor of NAS [adjusted hazard ratio, 1.19; 95% CI, 1.04-1.35; P = 0.01]. Implantation time (median 80 min, IQR: 69-95) was independently associated with EAD [adjusted odds ratio (OR), 1.15; 95% CI,1.07-1.23; P < 0.0001). The risk of EAD was increased by anastomosis time of both portal vein (adjusted OR, 1.26; 95% CI, 1.12-14.42; P = 0.0001) and hepatic artery (adjusted OR, 1.13; 95% CI, 1.04-1.22; P = 0.005). The magnitude of these effects was similar in donation after circulatory death liver grafts., Conclusions: Donor hepatectomy and implantation time negatively affect short-term outcomes., Competing Interests: The authors declare no funding or conflicts of interest., (Copyright © 2020 Wolters Kluwer Health, Inc. All rights reserved.)

Cystic fibrosis (CF) is a life-threatening disorder characterised by decreased pulmonary mucociliary and pathogen clearance, and an exaggerated inflammatory response leading to progressive lung damage. CF is caused by bi-allelic pathogenic variants of the cystic fibrosis transmembrane conductance regulator (CFTR) gene, which encodes a chloride channel. CFTR is expressed in endothelial cells (ECs) and EC dysfunction has been reported in CF patients, but a role for this ion channel in ECs regarding CF disease progression is poorly described.We used an unbiased RNA sequencing approach in complementary models of CFTR silencing and blockade (by the CFTR inhibitor CFTRinh-172) in human ECs to characterise the changes upon CFTR impairment. Key findings were further validated in vitro and in vivo in CFTR-knockout mice and ex vivo in CF patient-derived ECs.Both models of CFTR impairment revealed that EC proliferation, migration and autophagy were downregulated. Remarkably though, defective CFTR function led to EC activation and a persisting pro-inflammatory state of the endothelium with increased leukocyte adhesion. Further validation in CFTR-knockout mice revealed enhanced leukocyte extravasation in lung and liver parenchyma associated with increased levels of EC activation markers. In addition, CF patient-derived ECs displayed increased EC activation markers and leukocyte adhesion, which was partially rescued by the CFTR modulators VX-770 and VX-809.Our integrated analysis thus suggests that ECs are no innocent bystanders in CF pathology, but rather may contribute to the exaggerated inflammatory phenotype, raising the question of whether normalisation of vascular inflammation might be a novel therapeutic strategy to ameliorate the disease severity of CF., Competing Interests: Conflict of interest: M. Declercq has nothing to disclose. Conflict of interest: P. de Zeeuw has nothing to disclose. Conflict of interest: N.V. Conchinha has nothing to disclose. Conflict of interest: V. Geldhof has nothing to disclose. Conflict of interest: A.S. Ramalho has nothing to disclose. Conflict of interest: M. García-Caballero has nothing to disclose. Conflict of interest: K. Brepoels has nothing to disclose. Conflict of interest: M. Ensinck has nothing to disclose. Conflict of interest: M.S. Carlon has nothing to disclose. Conflict of interest: M.J. Bird has nothing to disclose. Conflict of interest: S. Vinckier has nothing to disclose. Conflict of interest: M. Proesmans has nothing to disclose. Conflict of interest: F. Vermeulen has nothing to disclose. Conflict of interest: L. Dupont has nothing to disclose. Conflict of interest: B. Ghesquière has nothing to disclose. Conflict of interest: M. Dewerchin has nothing to disclose. Conflict of interest: P. Carmeliet has nothing to disclose. Conflict of interest: D. Cassiman has nothing to disclose. Conflict of interest: L. Treps has nothing to disclose. Conflict of interest: G. Eelen has nothing to disclose. Conflict of interest: P. Witters has nothing to disclose., (Copyright ©ERS 2021.)

Purpose: In this study we investigate the disease etiology in 12 patients with de novo variants in FAR1 all resulting in an amino acid change at position 480 (p.Arg480Cys/His/Leu)., Methods: Following next-generation sequencing and clinical phenotyping, functional characterization was performed in patients' fibroblasts using FAR1 enzyme analysis, FAR1 immunoblotting/immunofluorescence, and lipidomics., Results: All patients had spastic paraparesis and bilateral congenital/juvenile cataracts, in most combined with speech and gross motor developmental delay and truncal hypotonia. FAR1 deficiency caused by biallelic variants results in defective ether lipid synthesis and plasmalogen deficiency. In contrast, patients' fibroblasts with the de novo FAR1 variants showed elevated plasmalogen levels. Further functional studies in fibroblasts showed that these variants cause a disruption of the plasmalogen-dependent feedback regulation of FAR1 protein levels leading to uncontrolled ether lipid production., Conclusion: Heterozygous de novo variants affecting the Arg480 residue of FAR1 lead to an autosomal dominant disorder with a different disease mechanism than that of recessive FAR1 deficiency and a diametrically opposed biochemical phenotype. Our findings show that for patients with spastic paraparesis and bilateral cataracts, FAR1 should be considered as a candidate gene and added to gene panels for hereditary spastic paraplegia, cerebral palsy, and juvenile cataracts.

Purpose: Hardikar syndrome (MIM 612726) is a rare multiple congenital anomaly syndrome characterized by facial clefting, pigmentary retinopathy, biliary anomalies, and intestinal malrotation, but with preserved cognition. Only four patients have been reported previously, and none had a molecular diagnosis. Our objective was to identify the genetic basis of Hardikar syndrome (HS) and expand the phenotypic spectrum of this disorder., Methods: We performed exome sequencing on two previously reported and five unpublished female patients with a clinical diagnosis of HS. X-chromosome inactivation (XCI) studies were also performed., Results: We report clinical features of HS with previously undescribed phenotypes, including a fatal unprovoked intracranial hemorrhage at age 21. We additionally report the discovery of de novo pathogenic nonsense and frameshift variants in MED12 in these seven individuals and evidence of extremely skewed XCI in all patients with informative testing., Conclusion: Pathogenic missense variants in the X-chromosome gene MED12 have previously been associated with Opitz-Kaveggia syndrome, Lujan syndrome, Ohdo syndrome, and nonsyndromic intellectual disability, primarily in males. We propose a fifth, female-specific phenotype for MED12, and suggest that nonsense and frameshift loss-of-function MED12 variants in females cause HS. This expands the MED12-associated phenotype in females beyond intellectual disability.

Metabolic rewiring is a hallmark of cancer that supports tumor growth, survival, and chemotherapy resistance. Although normal cells often rely on extracellular serine and glycine supply, a significant subset of cancers becomes addicted to intracellular serine/glycine synthesis, offering an attractive drug target. Previously developed inhibitors of serine/glycine synthesis enzymes did not reach clinical trials due to unfavorable pharmacokinetic profiles, implying that further efforts to identify clinically applicable drugs targeting this pathway are required. In this study, we aimed to develop therapies that can rapidly enter the clinical practice by focusing on drug repurposing, as their safety and cost-effectiveness have been optimized before. Using a yeast model system, we repurposed two compounds, sertraline and thimerosal, for their selective toxicity against serine/glycine synthesis-addicted breast cancer and T-cell acute lymphoblastic leukemia cell lines. Isotope tracer metabolomics, computational docking, enzymatic assays, and drug-target interaction studies revealed that sertraline and thimerosal inhibit serine/glycine synthesis enzymes serine hydroxymethyltransferase and phosphoglycerate dehydrogenase, respectively. In addition, we demonstrated that sertraline's antiproliferative activity was further aggravated by mitochondrial inhibitors, such as the antimalarial artemether, by causing G 1 -S cell-cycle arrest. Most notably, this combination also resulted in serine-selective antitumor activity in breast cancer mouse xenografts. Collectively, this study provides molecular insights into the repurposed mode-of-action of the antidepressant sertraline and allows to delineate a hitherto unidentified group of cancers being particularly sensitive to treatment with sertraline. Furthermore, we highlight the simultaneous inhibition of serine/glycine synthesis and mitochondrial metabolism as a novel treatment strategy for serine/glycine synthesis-addicted cancers., (©2020 American Association for Cancer Research.)

Purpose: Galactokinase (GALK1) deficiency is a rare hereditary galactose metabolism disorder. Beyond cataract, the phenotypic spectrum is questionable. Data from affected patients included in the Galactosemias Network registry were collected to better characterize the phenotype., Methods: Observational study collecting medical data of 53 not previously reported GALK1 deficient patients from 17 centers in 11 countries from December 2014 to April 2020., Results: Neonatal or childhood cataract was reported in 15 and 4 patients respectively. The occurrence of neonatal hypoglycemia and infection were comparable with the general population, whereas bleeding diathesis (8.1% versus 2.17-5.9%) and encephalopathy (3.9% versus 0.3%) were reported more often. Elevated transaminases were seen in 25.5%. Cognitive delay was reported in 5 patients. Urinary galactitol was elevated in all patients at diagnosis; five showed unexpected Gal-1-P increase. Most patients showed enzyme activities ≤1%. Eleven different genotypes were described, including six unpublished variants. The majority was homozygous for NM&#95;000154.1:c.82C>A (p.Pro28Thr). Thirty-five patients were diagnosed following newborn screening, which was clearly beneficial., Conclusion: The phenotype of GALK1 deficiency may include neonatal elevation of transaminases, bleeding diathesis, and encephalopathy in addition to cataract. Potential complications beyond the neonatal period are not systematically surveyed and a better delineation is needed.

Objectives: Since 2005, a New Wilson Index (NWI) ≥11 is used as a predictor of death without transplantation in fulminant Wilson disease (WD). Plasma exchange is advocated as a new treatment modality., Methods: We present a patient with fulminant WD treated with plasma exchange. All published cases applying plasma exchange for fulminant WD were reviewed systematically., Results: A 14-year-old girl presented with hemolysis and fulminant liver failure. She had no encephalopathy; NWI was 14. As a bridge to transplantation plasma exchange was started immediately. Complete remission was achieved with plasma exchange and later chelation therapy with D-penicillamine. She is now at 3-year transplant-free survival. Literature review identified 37 patients presenting with fulminant WD and NWI ≥11 who were treated with plasma exchange. Seventeen of these patients (ie, 46%) recovered without transplantation., Conclusions: Multiple case reports and case series demonstrate transplant free survival after plasma exchange and subsequent chelation therapy, despite a NWI ≥11. Plasma exchange affects the clinical course and is a therapeutic option in children and young adults presenting with fulminant WD.

Background & Aims: Although transient elastography (TE) is used to determine liver stiffness as a surrogate to hepatic fibrosis, the normal range in children is not well defined. We performed a systematic review and individual participant data (IPD) meta-analysis to determine the range of liver stiffness in healthy children and evaluate the influence of important biological parameters., Methods: We pooled data from 10 studies that examined healthy children using TE. We divided 1702 children into two groups: ≥3 years (older group) and < 3 years of age (younger group). Univariate and multivariate linear regression models predicting liver stiffness were conducted., Results: After excluding children with obesity, diabetes, or abnormal liver tests, 652 children were analysed. Among older children, mean liver stiffness was 4.45 kPa (95% confidence interval 4.34-4.56), and increased liver stiffness was associated with age, sedation status, and S probe use. In the younger group, the mean liver stiffness was 4.79 kPa (95% confidence interval 4.46-5.12), and increased liver stiffness was associated with sedation status and Caucasian race. In a subgroup analysis, hepatic steatosis on ultrasound was significantly associated with increased liver stiffness. We define a reference range for normal liver stiffness in healthy children as 2.45-5.56 kPa., Conclusions: We have established TE-derived liver stiffness ranges for healthy children and propose an upper limit of liver stiffness in healthy children to be 5.56 kPa. We have identified increasing age, use of sedation, probe size, and presence of steatosis on ultrasound as factors that can significantly increase liver stiffness., (© 2020 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Background: Hypophosphatasia (HPP) is a rare metabolic bone disorder caused by mutations in the alkaline phosphatase (ALPL) gene, and characterized by low circulating alkaline phosphatase (ALP) levels and bone, muscle, dental and systemic manifestations. In this case series we investigate the clinical spectrum, genetic and biochemical profile of adult HPP patients from the University Hospitals Leuven, Belgium., Methodology: Adults with HPP were identified through medical record review. Inclusion criteria were: (1) age ≥ 16 yr; (2) consecutively low ALP levels not explained by secondary causes; (3) one or more of the following supporting criteria: biochemical evidence of elevated enzyme substrates; subtrochanteric fractures, metatarsal fractures or other typical clinical features; family history of HPP; a known or likely pathogenic ALPL mutation., Results: Nineteen patients met our inclusion criteria (n = 2 infantile, n = 6 childhood, n = 10 adult-onset HPP and one asymptomatic carrier). Fractures and dental abnormalities were the most reported symptoms. Fatigue was reported in n = 7/19 patients (37%), three of which had previously been misdiagnosed as having chronic fatigue syndrome and/or fibromyalgia. Empirical pyridoxine therapy in four patients (without seizures) did not provide symptomatic relief. N = 7/19 patients (37%) were inappropriately treated or planned to be treated with antiresorptive treatment. Two patients developed atypical femoral fractures following exposure to bisphosphonates and/or denosumab. Patients detected by screening were less severely affected, while patients with homozygous or compound heterozygous mutations had the most severe symptoms, significantly lower circulating ALP levels (p = 0.013) and significantly higher pyridoxal-5'-phosphate (p = 0.0018) and urinary phosphoethanolamine (p = 0.0001) concentrations., Conclusions: Screening may detect mainly less severely affected individuals, which may nevertheless avoid misdiagnosis and inappropriate antiresorptive drug exposure. Patients with biallelic mutations had more severe symptoms, significantly lower ALP and higher substrate levels. Whether the latter finding has implications for the classification and treatment of HPP should be investigated further in larger cohorts., (Copyright © 2018 The International Society for Clinical Densitometry. Published by Elsevier Inc. All rights reserved.)

Niemann-Pick type C (NPC)1 disease is a rare genetic condition in which the function of the lysosomal cholesterol transporter NPC1 protein is impaired. Consequently, sphingolipids and cholesterol accumulate in lysosomes of all tissues, triggering a cascade of pathological events that culminate in severe systemic and neurological symptoms. Lysosomal cholesterol accumulation is also a key factor in the development of atherosclerosis and NASH. In these two metabolic diseases, the administration of plant stanol esters has been shown to ameliorate cellular cholesterol accumulation and inflammation. Given the overlap of pathological mechanisms among atherosclerosis, NASH, and NPC1 disease, we sought to investigate whether dietary supplementation with plant stanol esters improves the peripheral features of NPC1 disease. To this end, we used an NPC1 murine model featuring a Npc1 -null allele ( Npc1 nih ), creating a dysfunctional NPC1 protein. Npc1 nih mice were fed a 2% or 6% plant stanol ester-enriched diet over the course of 5 weeks. During this period, hepatic and blood lipid and inflammatory profiles were assessed. Npc1 nih mice fed the plant stanol-enriched diet exhibited lower hepatic cholesterol accumulation, damage, and inflammation than regular chow-fed Npc1 nih mice. Moreover, plant stanol consumption shifted circulating T-cells and monocytes in particular toward an anti-inflammatory profile. Overall, these effects were stronger following dietary supplementation with 6% stanols, suggesting a dose-dependent effect. The findings of our study highlight the potential use of plant stanols as an affordable complementary means to ameliorate disorders in hepatic and blood lipid metabolism and reduce inflammation in NPC1 disease., (Copyright © 2020 Magro dos Reis et al.)

Objective : Previous studies have shown that patients with hereditary fructose intolerance (HFI) are characterized by a greater intrahepatic triglyceride content, despite a fructose-restricted diet. The present study aimed to examine the long-term consequences of HFI on other aldolase-B-expressing organs, i.e. the kidney and vascular endothelium. Methods : Fifteen adult HFI patients were compared to healthy control individuals matched for age, sex and body mass index. Aortic stiffness was assessed by carotid-femoral pulse wave velocity (cf-PWV) and endothelial function by peripheral arterial tonometry, skin laser doppler flowmetry and the endothelial function biomarkers soluble E -selectin [sE-selectin] and von Willebrand factor. Serum creatinine and cystatin C were measured to estimate the glomerular filtration rate (eGFR). Urinary glucose and amino acid excretion and the ratio of tubular maximum reabsorption of phosphate to GFR (TmP/GFR) were determined as measures of proximal tubular function. Results : Median systolic blood pressure was significantly higher in HFI patients (127 versus 122 mmHg, p  = .045). Pulse pressure and cf-PWV did not differ between the groups ( p  = .37 and p  = .49, respectively). Of all endothelial function markers, only sE-selectin was significantly higher in HFI patients ( p  = .004). eGFR was significantly higher in HFI patients than healthy controls (119 versus 104 ml/min/1.73m 2 , p  = .001, respectively). All measurements of proximal tubular function did not differ significantly between the groups. Conclusions : Adult HFI patients treated with a fructose-restricted diet are characterized by a higher sE-selectin level and slightly higher systolic blood pressure, which in time could contribute to a greater cardiovascular risk. The exact cause and, hence, clinical consequences of the higher eGFR in HFI patients, deserves further study., (© 2020 The Authors.)

Epilepsy, intellectual and cortical sensory deficits, and psychiatric manifestations are the most frequent manifestations of mitochondrial diseases. How mitochondrial dysfunction affects neural structure and function remains elusive, mostly because of a lack of proper in vitro neuronal model systems with mitochondrial dysfunction. Leveraging induced pluripotent stem cell technology, we differentiated excitatory cortical neurons (iNeurons) with normal (low heteroplasmy) and impaired (high heteroplasmy) mitochondrial function on an isogenic nuclear DNA background from patients with the common pathogenic m.3243A > G variant of mitochondrial encephalomyopathy, lactic acidosis, and stroke-like episodes (MELAS). iNeurons with high heteroplasmy exhibited mitochondrial dysfunction, delayed neural maturation, reduced dendritic complexity, and fewer excitatory synapses. Micro-electrode array recordings of neuronal networks displayed reduced network activity and decreased synchronous network bursting. Impaired neuronal energy metabolism and compromised structural and functional integrity of neurons and neural networks could be the primary drivers of increased susceptibility to neuropsychiatric manifestations of mitochondrial disease., Competing Interests: Declaration of Interests The authors declare no competing interests., (Copyright © 2020 The Author(s). Published by Elsevier Inc. All rights reserved.)

Predicting drug-induced liver injury in a preclinical setting remains challenging, as cultured primary human hepatocytes (PHHs), pluripotent stem cell-derived hepatocyte-like cells (HLCs), and hepatoma cells exhibit poor drug biotransformation capacity. We here demonstrate that hepatic functionality depends more on cellular metabolism and extracellular nutrients than on developmental regulators. Specifically, we demonstrate that increasing extracellular amino acids beyond the nutritional need of HLCs and HepG2 cells induces glucose independence, mitochondrial function, and the acquisition of a transcriptional profile that is closer to PHHs. Moreover, we show that these high levels of amino acids are sufficient to drive HLC and HepG2 drug biotransformation and liver-toxin sensitivity to levels similar to those in PHHs. In conclusion, we provide data indicating that extracellular nutrient levels represent a major determinant of cellular maturity and can be utilized to guide stem cell differentiation to the hepatic lineage.

Background The dietary management of methylmalonic acidaemia (MMA) is a low-protein diet providing sufficient energy to avoid catabolism and to limit production of methylmalonic acid. The goal is to achieve normal growth, good nutritional status and the maintenance of metabolic stability. Aim To describe the dietary management of patients with MMA across Europe. Methods A cross-sectional questionnaire was sent to European colleagues managing inherited metabolic disorders (IMDs) (n=53) with 27 questions about the nutritional management of organic acidaemias. Data were analysed by different age ranges (0-6 months; 7-12 months; 1-10 years; 11-16 years; >16 years). Results Questionnaires were returned from 53 centres. Twenty-five centres cared for 80 patients with MMA vitamin B12 responsive (MMAB12r) and 43 centres managed 215 patients with MMA vitamin B12 non-responsive (MMAB12nr). For MMAB12r patients, 44% of centres (n=11/25) prescribed natural protein below the World Health Organization/Food and Agriculture Organization/United Nations University (WHO/FAO/UNU) 2007 safe levels of protein intake in at least one age range. Precursor-free amino acids (PFAA) were prescribed by 40% of centres (10/25) caring for 36% (29/80) of all the patients. For MMAB12nr patients, 72% of centres (n=31/43) prescribed natural protein below the safe levels of protein intake (WHO/FAO/UNU 2007) in at least one age range. PFAA were prescribed by 77% of centres (n=33/43) managing 81% (n=174/215) of patients. In MMAB12nr patients, 90 (42%) required tube feeding: 25 via a nasogastric tube and 65 via a gastrostomy. Conclusions A high percentage of centres used PFAA in MMA patients together with a protein prescription that provided less than the safe levels of natural protein intake. However, there was inconsistent practices across Europe. Long-term efficacy studies are needed to study patient outcome when using PFAA with different severities of natural protein restrictions in patients with MMA to guide future practice.

Pompe disease is a metabolic disorder caused by a deficiency of the glycogen-hydrolyzing lysosomal enzyme acid α-glucosidase (GAA), which leads to progressive muscle wasting. This autosomal-recessive disorder is the result of disease-associated variants located in the GAA gene. In the present study, we performed extended molecular diagnostic analysis to identify novel disease-associated variants in six suspected Pompe patients from four different families for which conventional diagnostic assays were insufficient. Additional assays, such as a generic-splicing assay, minigene analysis, SNP array analysis, and targeted Sanger sequencing, allowed the identification of an exonic deletion, a promoter deletion, and a novel splicing variant located in the 5' UTR. Furthermore, we describe the diagnostic process for an infantile patient with an atypical phenotype, consisting of left ventricular hypertrophy but no signs of muscle weakness or motor problems. This led to the identification of a genetic mosaicism for a very severe GAA variant caused by a segmental uniparental isodisomy (UPD). With this study, we aim to emphasize the need for additional analyses to detect new disease-associated GAA variants and non-Mendelian genotypes in Pompe disease where conventional DNA diagnostic assays are insufficient., (© 2020 The Author(s).)