Clinical characterization of NTCP deficiency in paediatric patients : A case-control study based on SLC10A1 genotyping analysis.

Na+ -taurocholate cotransporting polypeptide deficiency (NTCPD) is a newly described disorder arising from biallelic mutations of the SLC10A1 gene. As a result of a lack of compelling evidence from case-control studies, its genotypic and phenotypic features remain open for in-depth investigation. This study aimed to explore the genotypic and clinical phenotypic characteristics of paediatric patients with NTCPD. The SLC10A1 genotypes of all NTCPD patients were confirmed by screening for the prevalent variant c.800C>T and Sanger sequencing when necessary. The clinical presentations and laboratory changes were collected, reviewed and analysed, and then qualitatively and quantitatively compared with the relevant controls. A total of 113 paediatric NTCPD patients were diagnosed while c.374dupG and c.682_683delCT were detected as two novel pathogenic mutations. Hypercholanemia was observed in 99.12% of the patients. Indirect hyperbilirubinemia in affected neonates exhibited higher positive rates in comparison to controls. Moreover, transient cholestatic jaundice, elevated liver enzymes and 25-hydroxyvitamin D (Vit D) deficiency during early infancy were more commonly observed in patients than in controls. All NTCPD patients exhibited favourable clinical outcomes as a result of symptomatic and supportive treatment. The findings enriched the SLC10A1 mutation spectrum and provided comprehensive insights into the phenotypic characteristics of NTCPD. NTCPD should be considered and SLC10A1 gene should be analysed in patients with above age-dependent clinical features. Furthermore, over investigation and intervention should be avoided in the management of NTCPD patients.

Neonatal Intrahepatic Cholestasis caused by Citrin Deficiency: In vivo and in vitro studies of the aberrant transcription arising from two novel splice-site variants in SLC25A13.

Neonatal Intrahepatic Cholestasis caused by Citrin Deficiency (NICCD) is an autosomal recessive disease resulting from biallelic SLC25A13 mutations, and its diagnosis relies on genetic analysis. This study aimed to characterize the pathogenicity of 2 novel splice-site variants of SLC25A13 gene. Two patients (C0476 and C0556) suspected to have NICCD, their family members and 9 healthy volunteers were recruited as the research subjects. The SLC25A13 genotypes NG_012247.2(NM_014251.3): c.[852_855del]; [69+5G > A] in patient C0476 and c.[1453-1G > A]; [1751-5_1751-4ins (2684)] in patient C0556 were identified by means of polymerase chain reaction, long and accurate polymerase chain reaction, as well as Sanger sequencing. The 2 splice-site variants were absent in control databases and predicted to be pathogenic by computational analysis. The alternative splice variants in monocyte-derived macrophages from patient C0476 demonstrated exon 2 skipping [r.16_69del; p.(Val6_Lys23del)] in vivo, while minigene analysis revealed both exon 2-skipping and retained products from c.69+5G > A in vitro. In the patient C0556, an aberrant transcript [r.1453del; p.(Gly485Valfs*22)] resulting from c.1453-1G > A was detected on minigene splicing study. Thus, c.69+5G > A and c.1453-1G > A were both proved to be pathogenic. The 2 novel splice-site variants expanded the SLC25A13 mutation spectrum and provided reliable molecular markers for the definite diagnosis and genetic counseling of NICCD in the affected families.

[DGUOK-related mitochondrial DNA depletion syndrome: a case report and literature review].

A boy, aged 4 months, had the major clinical manifestations of prolonged jaundice and hepatomegaly. Multiple biochemical tests revealed abnormal liver function along with elevated alpha-fetoprotein and lactate. Genetic analysis confirmed that the boy had the mutations of c.589C>T(p.Gln197Ter) and c.687G>C(p.Trp229Cys) in the DGUOK gene, both of which were novel mutations and were determined to be pathogenic and likely pathogenic respectively, by a variety of bioinformatics tools and the ACMG standard. Therefore, the boy was confirmed to have DGUOK-related mitochondrial DNA depletion syndrome. Literature review showed that onset of liver disease in infancy was the main clinical feature of this disease, and some children presented with nervous system manifestations. Abnormal laboratory results included abnormal liver function, increases in blood lactate, serum ferritin and alpha-fetoprotein, and hypoglycemia. Such children had marked heterogeneity of DGUOK gene mutations, with missense mutations as the most common type. This disease tended to have a poor prognosis, and 79.6% of the children died before the age of 3 years.

Sodium Taurocholate Cotransporting Polypeptide (NTCP) Deficiency Hidden Behind Citrin Deficiency in Early Infancy: A Report of Three Cases.

Sodium taurocholate cotransporting polypeptide (NTCP), a carrier protein encoded by the gene SLC10A1, is expressed in the basolateral membrane of the hepatocyte to uptake bile acids from plasma. As a new inborn error of bile acid metabolism, NTCP deficiency remains far from being well understood in terms of the clinical and molecular features. Citrin deficiency is a well-known autosomal recessive disease arising from SLC25A13 mutations, and in neonates or infants, this condition presents as transient intrahepatic cholestasis which usually resolves before 1 year of age. All the three patients in this paper exhibited cholestatic jaundice and elevated total bile acids in their early infancy, which were attributed to citrin deficiency by SLC25A13 genetic analysis. In response to feeding with lactose-free and medium-chain triglycerides-enrich formula, their clinical and laboratory presentations disappeared gradually while the hypercholanemia persisted, even beyond 1 year of age. On subsequent SLC10A1 analysis, they were all homozygous for the well-known pathogenic variant c.800C > T (p.Ser267Phe), and NTCP deficiency was thus definitely diagnosed. The findings in this paper indicated that NTCP deficiency could be covered up by citrin deficiency during early infancy; however, in citrin-deficient patients with intractable hypercholanemia following resolved cholestatic jaundice, NTCP deficiency should be taken into consideration.

Clinical and molecular characterization of four patients with NTCP deficiency from two unrelated families harboring the novel SLC10A1 variant c.595A>C (p.Ser199Arg).

Sodium taurocholate cotransporting polypeptide (NTCP), a carrier protein encoded by solute carrier family 10 member 1 (SLC10A1), is expressed in the basolateral membrane of hepatocytes, where it is responsible for the uptake of bile acids from plasma into hepatocytes. The first patient with NTCP deficiency was described in 2015. A limited number of such patients have been reported in the literature and their genotypic and phenotypic features require further investigation. The current study investigated 4 patients with NTCP deficiency from two unrelated families. The patients were subjected to SLC10A1 genetic analysis and it was revealed that all patients were compound heterozygous for the c.800C>T (p.Ser267Phe) and c.595A>C (p.Ser199Arg) SLC10A1 variants. To the best of the authors' knowledge, the latter variant had not been previously reported. Further analysis in 50 healthy individuals did not identify carriers. The c.595A>C (p.Ser199Arg) variant exhibited co­segregation with hypercholanemia and exhibited a relatively conserved amino acid when compared with homologous peptides. Moreover, SWISS­MODEL prediction revealed that the mutation affected the conformation of the NTCP molecule. The 4 patients demonstrated varying degrees of hypercholanemia while a downward trend in the plasma levels of total bile acids (TBA) in 2 pediatric patients and occasionally normal TBA level in an adult case were observed. The results indicated an autosomal recessive trait for NTCP deficiency, supported the primary role of NTCP in the uptake of bile acids from plasma and suggested that hepatic uptake of bile acids may occur by means other than NTCP uptake. Moreover, the novel missense variant c.595A>C(p.Ser199Arg) enriched the SLC10A1 mutation spectrum and may serve as a new genetic marker for the molecular diagnosis and genetic counseling of NTCP deficiency.

Intrahepatic Cholestasis of Pregnancy as a Clinical Manifestation of Sodium-Taurocholate Cotransporting Polypeptide Deficiency.

Intrahepatic cholestasis of pregnancy (ICP) is the most common pregnancy-related liver disorder. Although the etiology of ICP is not fully understood thus far, some genetic factors might contribute to the development of this condition. Sodium-taurocholate cotransporting polypeptide (NTCP), the protein encoded by the gene Solute Carrier Family 10, Member 1 (SLC10A1), is the primary transporter expressed in the basolateral membrane of the hepatocyte to uptake conjugated bile salts from the plasma. NTCP deficiency arises from biallelic SLC10A1 mutations which impair the NTCP function and cause intractably elevated levels of total bile acids (TBA) in the plasma (hypercholanemia). In this study, all the SLC10A1 exons and their flanking sequences were analyzed by Sanger sequencing to investigate the etiology for hypercholanemia in two male infants aged 2 and 20 months, respectively, from two unrelated families. As a result, both patients are homozygous for the reported pathogenic variant c.800C>T (p.Ser267Phe) that could impair the NTCP function to uptake bile acids, and the diagnosis of NTCP deficiency was thus made. Their mothers are also homozygotes of the same variant and both had been diagnosed to have ICP in the third trimester, with one of them undergoing cesarean section. The father of the first patient in this paper has the same SLC10A1 genotype c.800C>T/c.800C>T, also exhibiting slight hypercholanemia with a plasma TBA level of 21.5 µmol/L. In conclusion, we suggest that with hypercholanemia being a common laboratory change, NTCP deficiency may be a genetic factor leading to ICP and even cesarean section in clinical practice.

[Clinical features and ABCC2 genotypic analysis of an infant with Dubin-Johnson syndrome].

Dubin-Johnson syndrome (DJS) is an autosomal recessive disorder resulting from biallelic mutations of ABCC2 gene, with long-term or intermittent conjugated hyperbilirubinemia being the main clinical manifestation. This paper aims to report the clinical features and ABCC2 genotypes of an infant with DJS. A 9.5-month-old male infant was referred to the hospital due to abnormal liver function discovered over 9 months. The major clinical presentation was prolonged jaundice since neonatal period. A series of biochemistry analysis revealed markedly elevated total bilirubin, conjugated bilirubin and total bile acids. The patient had been managed in different hospitals, but the therapeutic effects were unsatisfactory due to undetermined etiology. Physical examination revealed jaundiced skin and sclera, and a palpable liver 3 cm below the right subcostal margin with medium texture. The spleen was not enlarged. Genetic analysis revealed a splice-site variant c.3988-2A>T and a nonsense variant c.3825C>G (p.Y1275X) in the ABCC2 gene of the infant, which were inherited from his mother and father respectively. The former had not been previously reported. Then ursodeoxycholic acid and phenobarbital were given orally. Half a month later, as a result, his jaundice disappeared and the biochemistry indices improved. However, the long-term outcome needs to be observed. Literature review revealed that neonates/infants with DJS presented with cholestatic jaundice soon after birth as the major clinical feature, and the ABCC2 variants exhibited marked heterogeneity.

[Identification and pathogenicity prediction of a novel GLB1 variant c.101T>C (p.Ile34Thr) in an infant with GM1 gangliosidosis].

GM1 gangliosidosis is an autosomal recessive disorder caused by galactosidase beta1 (GLB1) gene variants which affect the activity of ß-galactosidase (GLB). GLB dysfunction causes abnormalities in the degradation of GM1 and its accumulation in lysosome. This article reports the clinical and genetic features of a child with GM1 gangliosidosis. The girl, aged 2 years and 5 months, was referred to the hospital due to motor developmental regression for more than one year. Physical examination showed binocular deflection and horizontal nystagmus, but no abnormality was found on fundoscopy. The girl had increased muscular tone of the extremities, limitation of motion of the elbow, knee, and ankle joints, and hyperactive patellar tendon reflex. Blood biochemical examination showed a significant increase in aspartate aminotransferase. The 24-hour electroencephalographic monitoring detected frequent seizure attacks and diffuse θ wave activity, especially in the right hemisphere. Head magnetic resonance imaging showed thinner white matter in the periventricular region and diffuse high T2WI signal with unclear boundary. Three-dimensional reconstruction of white matter fiber tracts by diffusion tensor imaging showed smaller and thinner white matter fiber tracts, especially in the right hemisphere. Genetic analysis showed that the girl had compound heterozygous mutations of c.446C>T (p.Ser149Phe) and c.101T>C (p.Ile34Thr) in the GLB1 gene from her parents, among which c.101T>C (p.Ile34Thr) had not been reported in the literatures. The girl was finally diagnosed with GM1 gangliosidosis. Her conditions were not improved after antiepileptic treatment and rehabilitation training for 2 months.

Monozygotic Twins Suffering From Sodium Taurocholate Cotransporting Polypeptide Deficiency: A Case Report.

Sodium taurocholate cotransporting polypeptide (NTCP) is a carrier protein encoded by the human SLC10A1 gene, acting as the principal transporter of conjugated bile salts from the plasma into hepatocytes. Although NTCP was cloned as early as in 1994 and its function has been studied extensively, clinical description of NTCP deficiency remains rather limited thus far. The patients in this paper were 2 female monozygotic twins, who were referred to our hospital at the age 2 years with the complaint of persistently-raised total bile acids (TBA) for 21 months. At age 3 months, they were both diagnosed to have cholestatic liver disease due to raised serum TBA and direct bilirubin (DBIL) with the fraction >20% of the elevated total bilirubin (TBIL). Thereafter, their jaundice subsided and the DBIL levels recovered gradually, while serum TBA remained raised persistently. In view of their refractory hypercholanemia but negative symptoms and signs, SLC10A1 genetic analysis was performed for all family members to evaluate the possibility of NTCP deficiency. As a result, the twins were both homozygotes, while the parents, carriers, of the reportedly pathogenic variant c.800C>T (p.Ser267Phe). These findings suggested that NTCP deficiency may be a unique genetic factor causing transient cholestasis in early infancy, as well as, persistent hypercholanemia in pediatric patients.

[Clinical and genetic analysis of an infant with progressive familial intrahepatic cholestasis type II].

Progressive familial intrahepatic cholestasis type II (PFIC-2) is an autosomal recessive disorder caused by biallelic variants of ABCB11 gene. This paper reports the clinical and laboratory features of a pediatric patient with PFIC-2. The patient was a 2.4-month-old male infant with jaundice and hepatomegaly as the main clinical manifestations. The serum levels of total bilirubin, direct bilirubin and total bile acids were increased, while the serum γ-glutamyl transpeptidase (GGT) level was normal. Next generation sequencing revealed two missense variants, c.1493T>C(p.Ile498Thr) and c.1502T>G(p.Val501Gly), in the ABCB11 gene of the patient, which were inherited from his father and mother, respectively. The latter was a novel variant which was predicted to be pathogenic by using a variety of bioinformatic tools, and the affected p.Val501 residue was highly conserved in 112 homologous peptides.

[Clinical and genetic analysis of a pediatric patient with sodium taurocholate cotransporting polypeptide deficiency].

Sodium taurocholate cotransporting polypeptide (NTCP) deficiency is an inborn error of bile acid metabolism caused by mutations of SLC10A1 gene. This paper reports the clinical and genetic features of a patient with this disease. A 3.3-month-old male infant was referred to the hospital with the complaint of jaundiced skin and sclera over 3 months. Physical examination revealed moderate jaundice of the skin and sclera. The liver was palpable 3.5 cm below the right subcostal margin with a medium texture. Serum biochemistry analysis revealed markedly elevated bilirubin (predominantly direct bilirubin) and total bile acids (TBA), as well as decreased 25-OH-VitD level. On pathological analysis of the biopsied liver tissue, hepatocyte ballooning and cholestatic multinucleate giant cells were noted. The lobular architecture was distorted. Infiltration of inflammatory cells, predominantly lymphocytes, was seen in the portal tracts. In response to the anti-inflammatory and liver protective drugs as well as fat-soluble vitamins over 2 months, the bilirubin and transaminases levels were improved markedly while the TBA kept elevated. Because of persisting hypercholanemia on the follow-up, SLC10A1 gene analysis was performed at his age of 17.2 months. The child proved to be a homozygote of the reportedly pathogenic variant c.800C>T (p. Ser267Phe), while the parents were both carriers. NTCP deficiency was thus diagnosed. The infant was followed up until 34.3 months old. He developed well in terms of the anthropometric indices and neurobehavioral milestones. The jaundice disappeared completely. The liver size, texture and function indices all recovered. However, the hypercholanemia persisted, and the long-term outcome needs to be observed.

Sodium taurocholate cotransporting polypeptide (NTCP) deficiency: Identification of a novel SLC10A1 mutation in two unrelated infants presenting with neonatal indirect hyperbilirubinemia and remarkable hypercholanemia.

Sodium taurocholate cotransporting polypeptide (NTCP) is encoded by the gene SLC10A1 and expressed in the basolateral membrane of the hepatocyte, functioning to uptake bile acids from plasma. Although SLC10A1 has been cloned and NTCP function studied intensively for years, clinical description of NTCP deficiency remains rather limited. This study reported the genotypic and phenotypic features of two neonatal patients with NTCP deficiency. They both presented with neonatal indirect hyperbilirubinemia and remarkable hypercholanemia, and harbored the SLC10A1 variants c.800C>T (p.S267F) and c.263T>C (p.I88T). On genetic analysis of the two family trios, the latter missense variant was detected in trans with the former, a reported loss-of-function variant. Having not been reported in any databases, the c.263T>C (p.I88T) variant demonstrated an allele frequency of 0.67% (1/150) in healthy controls. Moreover, this variant involved a relatively conservative amino acid, and was predicted to be pathogenic or deleterious by changing the conformation of the NTCP molecule. In conclusion, the novel variant c.263T>C (p.I88T) in this study enriched the SLC10A1 mutation spectrum; the clinical findings lent support to the primary role of NTCP in hepatic bile acid clearance, and suggested that NTCP deficiency might be a contributing factor for the development of neonatal indirect hyperbilirubinemia.

[Role of p38MAPK in apoptosis of human paratumor cirrhosis hepatocellular cell line QSG-7701 induced by cisplatin].

OBJECTIVE: To investigate the roles of p38 MAPK in apoptosis of the normal liver cell, the paratumor cirrhosis hepatocellular cell and the hepatocellular carcinoma cell. METHODS: Three cell lines were adopted (the normal liver cell line HL-7702, the paratumor cirrhosis hepatocellular cell line QSG-7701 and the hepatocellular carcinoma cell line QGY-7703) and treated with Diamminedichloroplatin (DDP, cisplatin) and p38MAPK inhibitor SB203580. The apoptosis and cell cycles were detected by flow cytometry and electromicroscopy. The expressions of p38MAPK, CDC25B, p34cdc2 and cyclinB1 were detected by immunocytochemical staining , confocal microscopy and western blot. RESULTS: The apoptotic rates in all three cell lines pretreated with DDP increased obviously and the rates in normal liver cells and HCC cells increased continuously even after SB203580 treatment, whereas in paratumor cirrhosis cells the rate decreased and the cell cycle stopped at S phase. CONCLUSION: Cisplatin induces apoptosis in the paratumor cirrhosis hepatocellular cell line QSG-7701 via activation of p38MAPK pathway and it differs in the normal liver cells from the hepatocellular carcinoma cells.