Inspissated bile syndrome in an infant with citrin deficiency and congenital anomalies of the biliary tract and esophagus: identification and pathogenicity analysis of a novel SLC25A13 mutation with incomplete penetrance.

Biallelic mutations of the SLC25A13 gene result in citrin deficiency (CD) in humans. Neonatal intrahepatic cholestasis caused by citrin deficiency (NICCD) is the major CD phenotype in pediatrics; however, knowledge on its genotypic and phenotypic characteristics remains limited. The present study aimed to explore novel molecular and clinical characteristics of CD. An infant suspected to have NICCD as well as her parents were enrolled as the research subjects. SLC25A13 mutations were investigated using various methods, including cDNA cloning and sequencing. The pathogenicity of a novel mutation was analyzed bioinformatically and functionally with a yeast model. Both the infant and her father were heterozygous for c.2T>C and c.790G>A, while the mother was only a c.2T>C carrier. The novel c.790G>A mutation proved bioinformatically and functionally pathogenic. The infant had esophageal atresia and an accessory hepatic duct, along with bile plug formation confirmed by laparoscopic surgery. However, the father seemed to be healthy thus far. The findings of the present study enrich the genotypic and phenotypic characteristics of CD patients, and provided clinical and molecular evidence suggesting the possible non-penetrance of SLC25A13 mutations and the likely involvement of this gene in primitive foregut development during early embryonic life.

Molecular analysis of SLC25A13 gene in human peripheral blood lymphocytes: Marked transcript diversity, and the feasibility of cDNA cloning as a diagnostic tool for citrin deficiency.

Human SLC25A13 gene encodes citrin, the liver-type aspartate-glutamate carrier isoform 2, and SLC25A13 mutations lead to citrin deficiency (CD). The definitive diagnosis of CD relies on SLC25A13 analysis, but conventional DNA analysis could not identify all SLC25A13 mutations. We investigated transcriptional features of SLC25A13 gene in peripheral blood lymphocytes (PBLs) from CD patients and healthy volunteers. SLC25A13 mutations were explored by PCR/LA-PCR, PCR-RFLP and direct sequencing. SLC25A13 cDNA was amplified by RT-PCR, cloned and then sequenced. All diagnoses of the CD patients were confirmed, including a heterozygote of g.2T>C and an unknown mutation yielding an aberrant transcript r.16_212dup. Twenty-eight alternative splice variants (ASVs) were identified from normal SLC25A13 alleles. Among them, r.213_328del took account for 53.7%, the normal transcript r.=, 16.6%, and the remaining 26 novel ASVs, collectively 29.3%, of all cDNA clones. Moreover, similar ASVs, all reflecting corresponsive mutations, were detected from the mutated alleles. These results indicated that the normal SLC25A13 transcript could be cloned, and the abundance of the ASV r.213_328del predicted the existence of a constructively novel protein isoform for this gene in human PBLs. And, the 26 novel ASVs, along with the novel aberrant transcript r.16_212dup and the SNP g.2T>C, enriched the transcript/variation spectrum of SLC25A13 gene in human beings. The findings in this paper, for the first time, uncovered the marked transcript diversity of SLC25A13 gene in human PBLs, and suggested that cDNA cloning analysis of this gene in human PBLs might be a feasible tool for CD molecular diagnosis.

[Antagonistic effect of baicalin on oxidative stress injury in neurons and astrocytes of rats].

OBJECTIVE: To explore the experimental conditions for H2O2 to injure astrocytes and the effect of baicalin in protecting neurons and astrocytes from oxidative stress injury. METHODS: Neurons and astrocytes from forebrain of rats were cultured in vitro and treated with H2O2, baicalin and combination of the two, respectively. The cell viability or survival rate was determined using MTT. RESULTS: Effects of H2O2 in different concentrations on survival rate of astrocytes showed significant difference (F = 28.569, P < 0.01) in a dose-dependent manner. Degrees of H2O2 injury, with the same concentration of H2O2, on cells with different seeding density were also significantly different (F = 5.439, P < 0.01), and dose-dependently. Baicalin didn't influence the survival rate of neurons and astrocytes when the concentration was within 2.5-40 mumol/L (for neurons, F = 0.49, P > 0.05; for astrocytes, F = 1.001, P > 0.05), but baicalin showed significant antagonism to the injury of oxidative stress (for neurons, F = 24.384, P < 0.01; for astrocytes, F = 5.000, P < 0.01). The higher the concentration of bainalin, the higher the cell survival rate. CONCLUSION: A model of astrocytes oxidative injury induced by H2O2 is established. Baicalin shows no toxicity on neurons and astrocytes when the concentration is within 2.5-40 mumol/L, but could antagonize the H2O2 caused oxidative injury on cells in a dose-dependent manner.