Acute Infantile Liver Failure Due to Mutations in the TRMU Gene

Acute liver failure in infancy accompanied by lactic acidemia was previously shown to result from mtDNA depletion. We report on 13 unrelated infants who presented with acute liver failure and lactic acidemia with normal mtDNA content. Four died during the acute episodes, and the survivors never had a recurrence. The longest follow-up period was 14 years. Using homozygosity mapping, we identified mutations in the TRMU gene, which encodes a mitochondria-specific tRNA-modifying enzyme, tRNA 5-methylaminomethyl-2-thiouridylate methyltransferase. Accordingly, the 2-thiouridylation levels of the mitochondrial tRNAs were markedly reduced. Given that sulfur is a TRMU substrate and its availability is limited during the neonatal period, we propose that there is a window of time whereby patients with TRMU mutations are at increased risk of developing liver failure.     

Biallelic Mutations in NBAS Cause Recurrent Acute Liver Failure with Onset in Infancy

Acute liver failure (ALF) in infancy and childhood is a life-threatening emergency. Few conditions are known to cause recurrent acute liver failure (RALF), and in about 50% of cases, the underlying molecular cause remains unresolved. Exome sequencing in five unrelated individuals with fever-dependent RALF revealed biallelic mutations in NBAS. Subsequent Sanger sequencing of NBAS in 15 additional unrelated individuals with RALF or ALF identified compound heterozygous mutations in an additional six individuals from five families. Immunoblot analysis of mutant fibroblasts showed reduced protein levels of NBAS and its proposed interaction partner p31, both involved in retrograde transport between endoplasmic reticulum and Golgi. We recommend NBAS analysis in individuals with acute infantile liver failure, especially if triggered by fever.     

Mutations of the FHL1 Gene Cause Emery-Dreifuss Muscular Dystrophy

Emery-Dreifuss muscular dystrophy (EDMD) is a rare disorder characterized by early joint contractures, muscular dystrophy, and cardiac involvement with conduction defects and arrhythmias. So far, only 35% of EDMD cases are genetically elucidated and associated with EMD or LMNA gene mutations, suggesting the existence of additional major genes. By whole-genome scan, we identified linkage to the Xq26.3 locus containing the FHL1 gene in three informative families belonging to our EMD- and LMNA-negative cohort. Analysis of the FHL1 gene identified seven mutations, in the distal exons of FHL1 in these families, three additional families, and one isolated case, which differently affect the three FHL1 protein isoforms: two missense mutations affecting highly conserved cysteines, one abolishing the termination codon, and four out-of-frame insertions or deletions. The predominant phenotype was characterized by myopathy with scapulo-peroneal and/or axial distribution, as well as joint contractures, and associated with a peculiar cardiac disease characterized by conduction defects, arrhythmias, and hypertrophic cardiomyopathy in all index cases of the seven families. Heterozygous female carriers were either asymptomatic or had cardiac disease and/or mild myopathy. Interestingly, four of the FHL1-mutated male relatives had isolated cardiac disease, and an overt hypertrophic cardiomyopathy was present in two. Expression and functional studies demonstrated that the FHL1 proteins were severely reduced in all tested patients and that this was associated with a severe delay in myotube formation in the two patients for whom myoblasts were available. In conclusion, FHL1 should be considered as a gene associated with the X-linked EDMD phenotype, as well as with hypertrophic cardiomyopathy.     

Acute Liver Failure with Thyrotoxicosis Treated with Liver Transplantation

ObjectiveWe describe a young woman with previously undiagnosed thyrotoxicosis who presented with acute liver failure (ALF).MethodsWe present a case report and review the relevant literature.ResultsAn extensive evaluation excluded possible causes of ALF other than thyrotoxicosis. The management of thyrotoxicosis posed several unique challenges in the setting of ALF, particularly because we did not want to use potentially hepatotoxic thionamides. The patient was treated with prednisone and propranolol and was started on potassium iodide when she was listed for liver transplantation. She underwent an uncomplicated liver transplant and subsequent thyroidectomy and is doing well.ConclusionThis well-characterized case describes thyrotoxicosis as a possible cause of ALF after thoroughly excluding other possible causes and illustrates the challenges of simultaneously managing both disorders. To our knowledge, this is the first report of ALF possibly resulting from untreated thyrotoxicosis that was successfully treated with liver transplantation. (Endocr Pract. 2013;19:e57-e60)     

Heterozygous Germline Mutations in the CBL Tumor-Suppressor Gene Cause a Noonan Syndrome-like Phenotype

RAS signaling plays a key role in controlling appropriate cell responses to extracellular stimuli and participates in early and late developmental processes. Although enhanced flow through this pathway has been established as a major contributor to oncogenesis, recent discoveries have revealed that aberrant RAS activation causes a group of clinically related developmental disorders characterized by facial dysmorphism, a wide spectrum of cardiac disease, reduced growth, variable cognitive deficits, ectodermal and musculoskeletal anomalies, and increased risk for certain malignancies. Here, we report that heterozygous germline mutations in CBL, a tumor-suppressor gene that is mutated in myeloid malignancies and encodes a multivalent adaptor protein with E3 ubiquitin ligase activity, can underlie a phenotype with clinical features fitting or partially overlapping Noonan syndrome (NS), the most common condition of this disease family. Independent CBL mutations were identified in two sporadic cases and two families from among 365 unrelated subjects who had NS or suggestive features and were negative for mutations in previously identified disease genes. Phenotypic heterogeneity and variable expressivity were documented. Mutations were missense changes altering evolutionarily conserved residues located in the RING finger domain or the linker connecting this domain to the N-terminal tyrosine kinase binding domain, a known mutational hot spot in myeloid malignancies. Mutations were shown to affect CBL-mediated receptor ubiquitylation and dysregulate signal flow through RAS. These findings document that germline mutations in CBL alter development to cause a clinically variable condition that resembles NS and that possibly predisposes to malignancies.     

Non-Syndromic Hereditary Gingival Fibromatosis in Three Chinese Families is Not Due to SOS1 Gene Mutations

Non-syndromic hereditary gingival fibromatosis (HGF) is a rare condition, characterized by a progressive gingival hyperplasia that occurs as an isolated disease. Hitherto, only one insertion (g.126,142–126,143insC) in son-of-sevenless-1 (SOS1) gene has been associated with non-syndromic HGF in a Brazilian family. The aim of the present study was to determine if SOS1 is the causative gene of non-syndromic HGF in the Chinese population. Peripheral blood samples were collected from six affected and seven unaffected individuals from three Chinese families with history of non-syndromic HGF. Genomic DNA was extracted and SOS1 gene exons were sequenced. Neither g.126,142–126,143insC nor any other novel mutation was detected in SOS1 gene. Our results suggest that the SOS1 may not be the gene responsible for HGF in these three Chinese families and, therefore, it is possible that other genes are involved in the manifestation of HGF in these Chinese HGF families.     

Causes and Outcomes of Acute Liver Failure in China

Objectives

No extensive investigation has been performed and thus no representative data are available regarding acute liver failure (ALF) in China. This study aims to investigate the causes and outcomes of ALF in China and establish a prognostic model.

Methods

Patients diagnosed as ALF in seven hospitals in different areas of China from January 2007 to December 2012 were retrospectively selected.

Results

Of the 177 patients included in this study, 112 (63.28%) eventually died. The common causes of ALF were drug toxicity (43.50%), indeterminate etiology (29.38%) and acute viral hepatitis (11.30%). Additionally, traditional Chinese herbs predominated in the causes of drug-induced ALF (30/77). No patients in this study received liver transplantation. In the established model for predicting death in ALF, four variables were finally selected out, including age (P=0.01), the entry hepatic encephalopathy grade (P=0.04), international normalized ratio (P<0.01) and arterial blood ammonia (P=0.02). Using a threshold value of 0.5683, this model had a sensitivity of 95.24% and a specificity of 91.30%.

Conclusions

Traditional Chinese medicine was a major cause of ALF in China. The spontaneous mortality of ALF was high, whereas the rate of liver transplantation was significantly low. The established prognostic model of ALF had superior sensitivity and specificity.     

Multiple Synostoses Syndrome Is Due to a Missense Mutation in Exon 2 of FGF9 Gene

Fibroblast growth factors (FGFs) play diverse roles in several developmental processes. Mutations leading to deregulated FGF signaling can cause human skeletal dysplasias and cancer.^1,2 Here we report a missense mutation (Ser99Asp) in exon 2 of FGF9 in 12 patients with multiple synostoses syndrome (SYNS) in a large Chinese family. In vitro studies demonstrate that FGF9^S99N is expressed and secreted as efficiently as wild-type FGF9 in transfected cells. However, FGF9^S99N induces compromised chondrocyte proliferation and differentiation, which is accompanied by enhanced osteogenic differentiation and matrix mineralization of bone marrow-derived mesenchymal stem cells (BMSCs). Biochemical analysis reveals that S99N mutation in FGF9 leads to significantly impaired FGF signaling, as evidenced by diminished activity of Erk1/2 pathway and decreased β-catenin and c-Myc expression when compared with wild-type FGF9. Importantly, the binding of FGF9^S99N to its receptor is severely impaired although the dimerization ability of mutant FGF9 itself or with wild-type FGF9 is not detectably affected, providing a basis for the defective FGFR signaling. Collectively, our data demonstrate a previously uncharacterized mutation in FGF9 as one of the causes of SYNS, implicating an important role of FGF9 in normal joint development.     

糖皮质激素对小鼠急性肝衰竭的干预作用研究

摘要 目的：探讨糖皮质激素对小鼠急性肝衰竭（Acute liver failure, AHF）的干预作用。方法：通过腹腔注射D-GaIN/LPS构建AHF小鼠模型,并随机分为对照组、AHF模型组（AHF组）和糖皮质激素处理急性肝衰竭组（GCs + AHF组）。通过全自动生物化学分析仪检测小鼠血清中ALT、AST、TBA、TBIL、DBIL水平;采用ELISA试剂盒测定小鼠血清中IL-1β、TNF-α、IL-6水平;通过定量PCR检测mRNA表达水平;苏木精-伊红（H&E）染色检测肝组织病理学情况;免疫组织化学检测肝组织中SOX9表达水平;蛋白质印迹检测Hedgehog信号通路关键蛋白因子的表达水平。结果：与对照组相比,AHF组小鼠血清ALT、AST、TBA、TBIL、DBIL、MDA、TNF-α、IL-1β和IL-6水平显著升高,SOD、GSH和CAT活性水平显著降低（P＜0.01）;与AHF组相比,GCs+AHF组小鼠血清中ALT、AST、TBA、TBIL、DBIL、MDA、TNF-α、IL-1β和IL-6水平显著降低,SOD、GSH和CAT活性水平显著升高（P＜0.01）。H&E染色结果显示,对照组小鼠的肝脏切片显示正常的细胞结构,AHF组小鼠出现广泛的细胞坏死和炎性细胞浸润;与AHF组相比,GCs + AHF组小鼠中细胞坏死和炎性细胞浸润等病理变化显著减弱。免疫组织化学结果显示：与对照组相比,AHF小鼠肝脏中SOX9的表达水平显著升高（P＜0.01）;与AHF组相比,GCs+AHF组小鼠肝脏中SOX9的表达水平显著降低（P＜0.01）。Western blotting结果显示,与对照组相比,AHF组小鼠的Hedgehog通路关键蛋白因子Shh、Ptch-1和Gli-1蛋白表达水平显著升高（P<0.01）;与AHF组相比,GCs+AHF组的Shh、Ptch-1和Gli-1蛋白表达水平显著降低（P<0.01）。结论：GCs可有效改善AHF小鼠肝功能指标,降低炎性因子表达,提高急性肝衰竭小鼠抗氧化能力,并使Hedgehog通路关键蛋白因子表达恢复正常。     

Mutations in PDGFRB Cause Autosomal-Dominant Infantile Myofibromatosis

Infantile myofibromatosis (IM) is a disorder of mesenchymal proliferation characterized by the development of nonmetastasizing tumors in the skin, muscle, bone, and viscera. Occurrence within families across multiple generations is suggestive of an autosomal-dominant (AD) inheritance pattern, but autosomal-recessive (AR) modes of inheritance have also been proposed. We performed whole-exome sequencing (WES) in members of nine unrelated families clinically diagnosed with AD IM to identify the genetic origin of the disorder. In eight of the families, we identified one of two disease-causing mutations, c.1978C>A (p.Pro660Thr) and c.1681C>T (p.Arg561Cys), in PDGFRB. Intriguingly, one family did not have either of these PDGFRB mutations but all affected individuals had a c.4556T>C (p.Leu1519Pro) mutation in NOTCH3. Our studies suggest that mutations in PDGFRB are a cause of IM and highlight NOTCH3 as a candidate gene. Further studies of the crosstalk between PDGFRB and NOTCH pathways may offer new opportunities to identify mutations in other genes that result in IM and is a necessary first step toward understanding the mechanisms of both tumor growth and regression and its targeted treatment.     

Alginate Microencapsulated Hepatocytes Optimised for Transplantation in Acute Liver Failure

MethodsHuman hepatocyte microbeads (HMBs) were prepared using sterile GMP grade materials. We determined physical stability, cell viability, and hepatocyte metabolic function of HMBs using different polymerisation times and cell densities. The immune activation of peripheral blood mononuclear cells (PBMCs) after co-culture with HMBs was studied. Rats with ALF induced by galactosamine were transplanted intraperitoneally with rat hepatocyte microbeads (RMBs) produced using a similar optimised protocol. Survival rate and biochemical profiles were determined. Retrieved microbeads were evaluated for morphology and functionality.ResultsThe optimised HMBs were of uniform size (583.5±3.3 µm) and mechanically stable using 15 min polymerisation time compared to 10 min and 20 min (p<0.001). 3D confocal microscopy images demonstrated that hepatocytes with similar cell viability were evenly distributed within HMBs. Cell density of 3.5×10⁶ cells/ml provided the highest viability. HMBs incubated in human ascitic fluid showed better cell viability and function than controls. There was no significant activation of PBMCs co-cultured with empty or hepatocyte microbeads, compared to PBMCs alone. Intraperitoneal transplantation of RMBs was safe and significantly improved the severity of liver damage compared to control groups (empty microbeads and medium alone; p<0.01). Retrieved RMBs were intact and free of immune cell adherence and contained viable hepatocytes with preserved function.ConclusionAn optimised protocol to produce GMP grade alginate-encapsulated human hepatocytes has been established. Transplantation of microbeads provided effective metabolic function in ALF. These high quality HMBs should be suitable for use in clinical transplantation.