Design, Synthesis and Characterization of Palladium-Functionalized Covalent Organic Framework and Its Application as Heterogeneous Catalysis for C-H Arylation of Azoles.

A novel triazine-based covalent organic framework (TFPT-Bz COF) has been constructed by the condensation of 2,4,6-tris(5-formyl-2-pyridinoxy)-1,3,5-triazine (TFPT) and benzidine (BZ) with deep eutectic solvent (DES) as the reaction medium. After the introduction of Pd ions through strong coordination to TFPT-Bz COF matrix, the constructed TFPT-Bz COF/Pd composite exhibited excellent catalytic activity for C-H arylation of azoles with aryl halides in 2-methyltetrahydrofuran. The protocol allows the arylation of a variety of substituted azoles with diverse aryl halides in high to excellent yield. Moreover, the TFPT-Bz COF/Pd catalyst can be recycled several times without significantly reducing its activity.

Alagille syndrome associated with total anomalous pulmonary venous connection and severe xanthomas: A case report.

BACKGROUND: Alagille syndrome (ALGS) is an autosomal dominant genetic disorder caused by mutations in the JAG1 or NOTCH2 gene. It is characterized by decreased intrahepatic bile ducts associated with a variety of abnormalities in many other organ systems, such as the cardiovascular, skeletal, and urinary systems. CASE SUMMARY: We report a rare case of ALGS. A 1-month-old male infant presented with sustained jaundice and had a rare congenital heart disease: Total anomalous pulmonary venous connection (TAPVC). Sustained jaundice, particularly with cardiac murmur, caught our attention. Laboratory tests revealed elevated levels of alanine aminotransferase, aspartate aminotransferase, gamma-glutamyl transpeptidase, total bilirubin, and total bile acids, indicating serious intrahepatic cholestasis. Imaging confirmed the presence of butterfly vertebra at the seventh thoracic vertebra. This suggested ALGS, which was confirmed by genetic testing with a c.3197dupC mutation in the JAG1 gene. Ursodiol was administered immediately after confirmation of the diagnosis, and cardiac surgery was performed when the patient was 1.5 month old. He recovered well after treatment and was discharged at the age of 3 mo. At the age of two years, the patient returned to our clinic because multiple cutaneous nodules with xanthomas appeared, and their size and number increased over time. CONCLUSION: We report a unique case of ALGS associated with TAPVC and severe xanthomas. This study has enriched the clinical manifestations of ALGS and emphasized the association between JAG1 gene and TAPVC.

Histone H3K4me3 modification is a transgenerational epigenetic signal for lipid metabolism in Caenorhabditis elegans.

As a major risk factor to human health, obesity presents a massive burden to people and society. Interestingly, the obese status of parents can cause progeny's lipid accumulation through epigenetic inheritance in multiple species. To date, many questions remain as to how lipid accumulation leads to signals that are transmitted across generations. In this study, we establish a nematode model of C. elegans raised on a high-fat diet (HFD) that leads to measurable lipid accumulation, which can transmit the lipid accumulation signal to their multigenerational progeny. Using this model, we find that transcription factors DAF-16/FOXO and SBP-1/SREBP, nuclear receptors NHR-49 and NHR-80, and delta-9 desaturases (fat-5, fat-6, and fat-7) are required for transgenerational lipid accumulation. Additionally, histone H3K4 trimethylation (H3K4me3) marks lipid metabolism genes and increases their transcription response to multigenerational obesogenic effects. In summary, this study establishes an interaction between a network of lipid metabolic genes and chromatin modifications, which work together to achieve transgenerational epigenetic inheritance of obesogenic effects.

Physical and neuropsychological development of children with Citrin deficiency. / Citrinç¼ºé™·ç— æ‚£å„¿ä½“æ ¼å’Œç¥žç»å¿ƒç†å‘è‚²çŠ¶å†µ.

OBJECTIVES: To study the physical and neuropsychological development of children with Citrin deficiency (CD). METHODS: A total of 93 children, aged 1.9-59.8 months, who were diagnosed with CD by SLC25A13 gene analysis in the First Affiliated Hospital of Jinan University from August 2010 to August 2015, were enrolled as subjects. A retrospective analysis was performed for their birth condition and physical growth and neuropsychological development indices. Among these children, 7 underwent physical measurement and neuropsychological development assessment within 1 year old and after 1 year old, and therefore, a total of 100 cases were included for analysis. RESULTS: For the 93 children with CD, the incidence rate of failure to thrive was 25% (23 children) and the proportion of small for gestational age was 47% (44 children). For the 100 cases of CD, the incidence rates of growth retardation, underweight, emaciation, overweight, and microcephalus were 23% (23 cases), 14% (14 cases), 4% (4 cases), 8% (8 cases), and 9% (9 cases), respectively. The incidence rate of neuropsychological developmental delay was 25% (25 cases), and the incidence rates of development delay in the five domains of adaptability, gross motor, fine motor, language, and social ability were 7% (7 cases), 15% (15 cases), 7% (7 cases), 9% (9 cases), and 7% (7 cases), respectively. CONCLUSIONS: Physical and neuropsychological developmental delay can be observed in children with CD, and physical and neuropsychological development should be regularly assessed.

Molecular epidemiologic study of citrin deficiency by screening for four reported pathogenic SLC25A13 variants in the Shaanxi and Guangdong provinces, China.

BACKGROUND: Citrin deficiency (CD) is an autosomal recessive disease resulting from biallelic mutations of the SLC25A13 gene. This study aimed to investigate the molecular epidemiological features of CD in the Guangdong and Shaanxi provinces of China. METHODS: A total of 3,409 peripheral blood samples from Guangdong and 2,746 such samples from Shaanxi province were collected. Four prevalent SLC25A13 mutations NG_012247.2 (NM_014251.3): c.852_855del, c.1638_1660dup, c.615+5G>A, and c.1751-5_1751-4ins(2684) were screened by using the conventional polymerase chain reaction (PCR)/PCR-restriction fragment length polymorphism and newly-developed multiplex PCR methods, respectively. The mutated SLC25A13 allele frequencies, carrier frequencies, and CD morbidity rates were calculated and then compared with the Chi-square and Fisher's exact tests. RESULTS: The mutations were detected in 68 out of 6,818 SLC25A13 alleles in Guangdong and 29 out of 5,492 alleles in the Shaanxi population. The carrier frequencies were subsequently calculated to be 1/51 and 1/95, while the CD morbidity rates were 1/10,053 and 1/35,865, in the 2 populations, respectively. When compared with the Shaanxi population, Guangdong exhibited a higher frequency of mutated SLC25A13 allele (68/6,818 vs. 29/5,492, χ2=8.570, P=0.003) in general, with higher c.852_855del (54/6,818 vs. 13/5,492, χ2=17.328, P=0.000) but lower c.1751-5_1751 -4ins(2684) (2/6,818 vs. 9/5,492, P=0.015) allele frequencies. The distribution of c.615+5G>A and c.1638_1660dup between the 2 provinces, as well as all 4 prevalent mutations among different geographic regions within the 2 provinces, did not differed significantly. CONCLUSIONS: Our findings depicted the CD molecular epidemiological features in Guangdong and Shaanxi populations, providing preliminary but significant laboratory evidences for the subsequent CD diagnosis and management in the 2 provinces of mainland China.

A terbium(III) lanthanide-organic framework as a selective and sensitive iodide/bromide sensor in aqueous medium.

A potent luminescent sensor for the detection of iodide ions was developed based on a terbium(iii)-based lanthanide-organic framework [Tb(cpia)(H2O)2]n·nH2O (1), which was prepared under hydrothermal conditions using the 5-(4-carboxyphenoxy)isophthalic acid (H3cpia) bridging ligand. Compound 1 exhibits superior luminescence quenching behavior towards I- with high sensitivity and selectivity among various anions and shows real-time response. Moreover, the mechanism of the selective luminescence quenching response for I- can be mainly explained by the absorption competition between 1 and I-. According to this quenching mechanism, we find that compound 1 can also detect Br- by adjusting the excitation wavelength. Significantly, this work could serve as a general guidance for the design and synthesis of pollutant sensors.

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.

Visible-Light-Mediated Oxidative Amidation of Aldehydes by Using Magnetic CdS Quantum Dots as a Photocatalyst.

A magnetic CdS quantum dot (Fe3 O4 /polydopamine (PDA)/CdS) was synthesized through a facile and convenient method from inexpensive starting materials. Characterization of the prepared catalyst was performed by means of FTIR spectroscopy, XRD, SEM, TEM, energy-dispersive X-ray spectroscopy, and vibrating-sample magnetometer techniques. Fe3 O4 /PDA/CdS was found to be a highly active photocatalyst for the amidation of aromatic aldehydes by using air as a clean oxidant under mild conditions. The photocatalyst can be recovered by magnetic separation and successfully reused for five cycles without considerable loss of its catalytic activity.

Catalyst-free Synthesis of Aminomethylphenol Derivatives in Cyclopentyl Methyl Ether via Petasis Borono-Mannich Reaction.

OBJECTIVE: Aminomethylphenol molecules have wider applications in pharmaceuticals, agrochemicals, plant protection and promising functional materials. The development of an efficient and practical method to prepare this class of compound is highly desirable from both environmental and economical points of view. MATERIALS AND METHODS: In order to establish an effective synthetic method for preparing aminomethylphenol derivatives, the Petasis borono-Mannich reaction of salicylaldehyde, phenylboronic acid and 1,2,3,4- tetrahydroisoquinoline was selected as a model reaction. A variety of reaction conditions are investigated, including solvent and temperature. The generality and limitation of the established method were also evaluated. RESULTS AND DISCUSSION: It was found that model reaction can be carried out in cyclopentyl methyl ether at 80 oC under catalyst-free conditions. This protocol, with broad substrate applicability, the reaction of various arylboronic acid, secondary amine and salicylaldehyde proceeded smoothly under optimal reaction conditions to afford various aminomethylphenol derivatives in high yields. A practical, scalable, and high-yielding synthesis of aminomethylphenol derivatives was successfully accomplished. CONCLUSION: A catalyst-free practical method for the synthesis of minomethylphenol derivatives based on Petasis borono-Mannich (PBM) reaction of various arylboronic acid, secondary amine and salicylaldehyde in cyclopentyl methyl ether has been developed. The salient features of this protocol are avoidance of any additive/catalyst and toxic organic solvents, use of cyclopentyl methyl ether as the reaction medium, clean reaction profiles, easy operation, and high to excellent yield.

Visible-Light-Initiated One-Pot, Three-Component Synthesis of 2-Amino-4H-pyran-3,5-dicarbonitrile Derivatives.

A novel approach for the visible-light-initiated synthesis of 2-amino-4H-pyran-3,5-dicarbonitrile derivatives via a one-pot, three-component reaction of aldehydes or isatins, malononitrile, and α-cyano ketones has been developed. The reaction was carried out at room temperature in ethanol/water to give the corresponding products with a wide range of functional groups in high yields. This process did not require any additives or chromatographic separation and could be applied for gram-scale synthesis.

Bioinformatic and functional analysis of promoter region of human SLC25A13 gene.

The human SLC25A13 gene encodes the liver type aspartate/glutamate carrier isoform 2 (AGC2, commonly named as citrin), which plays a key role in the main NADH-shuttle of human hepatocyte. Biallelic SLC25A13 mutations result in Citrin deficiency (CD). In order to identify the important regulatory region of SLC25A13 gene and elucidate the way how potential promoter mutations affect the citrin expression, we performed promoter deletion analysis and established the reporter constructs of luciferase gene-carrying SLC25A13 promoter containing several mutations located in putative transcription factor-binding sites. The luciferase activities of all promoter constructs were measured using a Dual-Luciferase Reporter Assay System. Bioinformatic analysis showed that the promoter of SLC25A13 gene lacks TATA box and obviously typical initiator element, but contains a CCAAT box and two GC box. Promoter deletion analysis confirmed the region from -221 to -1 upstream ATG was essential for SLC25A13 to maintain the promoter activity. We utilized dual-luciferase reporter system as function analytical model to tentatively assess the effect of artificially constructed promoter mutations on citrin expression, and our analysis revealed that mutated putative CCAAT box and GC box could significantly affect the citrin expression. Our study confirmed the important SLC25A13 promoter regions that influenced citrin expression in HL7702 cells, and constructed a function analytical model. This work may be useful to further identify the pathogenic mutations leading to CD in the promoter region.

Molecular diagnosis of citrin deficiency in an infant with intrahepatic cholestasis: identification of a 21.7kb gross deletion that completely silences the transcriptional and translational expression of the affected SLC25A13 allele.

Neonatal Intrahepatic Cholestasis caused by Citrin Deficiency (NICCD) arises from biallelic SLC25A13 mutations, and SLC25A13 analysis provides reliable evidences for NICCD definite diagnosis. However, novel large insertions/deletions in this gene could not be detected just by conventional DNA analysis. This study aimed to explore definite diagnostic evidences for an infant highly-suspected to have NICCD. Prevalent mutation screening and Sanger sequencing of SLC25A13 gene just revealed a paternally-inherited mutation c.851_854del4. Nevertheless, neither citrin protein nor SLC25A13 transcripts of maternal origin could be detected on Western blotting and cDNA cloning analysis, respectively. On this basis, the hidden maternal mutation was precisely positioned using SNP analysis and semi-quantitative PCR, and finally identified as a novel large deletion c.-3251_c.15+18443del21709bp, which involved the SLC25A13 promoter region and the entire exon 1 where locates the translation initiation codon. Hence, NICCD was definitely diagnosed in the infant. To the best of our knowledge, the novel gross deletion, which silenced the transcriptional and translational expression of the affected SLC25A13 allele, is the hitherto largest deletion in SLC25A13 mutation spectrum. The Western blotting approach using mitochondrial protein extracted from expanded peripheral blood lymphocytes, of particular note, might be a new minimally-invasive and more-feasible molecular tool for NICCD diagnosis.

SLC25A13 cDNA cloning analysis using peripheral blood lymphocytes facilitates the identification of a large deletion mutation: Molecular diagnosis of an infant with neonatal intrahepatic cholestasis caused by citrin deficiency.

Neonatal intrahepatic cholestasis caused by citrin deficiency (NICCD) is an autosomal recessive disorder resulting from biallelic mutations of the SLC25A13 gene. Due to the lack of well­recognized clinical or biochemical diagnostic criteria, the definitive diagnosis of this disease relies on the genetic analysis of SLC25A13 at present. As novel large deletion/insertion mutations of the SLC25A13 gene are difficult to detect using routine DNA analytic approaches, the timely diagnosis of patients with these types of mutations remains a challenge. The present study aimed to examine SLC25A13 mutations in an infant with a suspected diagnosis of NICCD. DNA was extracted from blood samples, and SLC25A13 mutations were examined by screening for high­frequency mutations and Sanger sequencing. Reverse transcription-polymerase chain reaction and cDNA cloning analyses were then performed using peripheral blood lymphocytes (PBLs) to identify the obscure mutation. The results demonstrated that the infant was heterozygous for a paternally­inherited mutation, c.851_854del4, and a maternally­inherited large deletion, c.1019_1177+893del, which has not been reported previously. A positive diagnosis of NICCD was made, and the infant responded favorably to a galactose­free and medium­chain triglyceride­enriched formula. The present study confirmed the effectiveness of this formula in NICCD therapy, enriched the SLC25A13 mutational spectrum and supported the feasibility of cDNA cloning analysis using PBLs as a molecular tool for facilitating the identification of large SLC25A13 deletions.

Photocontrolled Reversible Luminescent Lanthanide Molecular Switch Based on a Diarylethene-Europium Dyad.

A new europium complex coordinated between a Eu(III) ion and an unsymmetrical diarylperfluorocyclopentene yields a light-controlled diarylethene-europium dyad, DAE@TpyEu(tta)3, whose photophysical properties can be reversibly switched by optical stimuli. When DAE@TpyEu(tta)3 is exposed to 365 nm UV light, an efficient intramolecular photochromic fluorescence resonance energy transfer (pc-FRET) occurs between the emission of the Eu(3+) donor (D) and the absorption of the diarylethene acceptor (A) in closed-form DAE@TpyEu(tta)3 accompanied by luminescence quenching. However, the pc-FRET process could be effectively inhibited by visible light (λ > 600 nm) irradiation, and the lanthanide emission of DAE@TpyEu(tta)3 is rapidly recovered. Furthermore, this luminescent lanthanide molecular switch could serve as a highly reliable and sensitive "turn on" fluorescent marker in living cells irradiated by red light without any optical interference.

Molecular diagnosis of pediatric patients with citrin deficiency in China: SLC25A13 mutation spectrum and the geographic distribution.

Citrin deficiency (CD) is a Mendelian disease due to biallelic mutations of SLC25A13 gene. Neonatal intrahepatic cholestasis caused by citrin deficiency (NICCD) is the major pediatric CD phenotype, and its definite diagnosis relies on SLC25A13 genetic analysis. China is a vast country with a huge population, but the SLC25A13 genotypic features of CD patients in our country remains far from being well clarified. Via sophisticated molecular analysis, this study diagnosed 154 new CD patients in mainland China and identified 9 novel deleterious SLC25A13 mutations, i.e. c.103A > G, [c.329 - 154_c.468 + 2352del2646; c.468 + 2392_c.468 + 2393ins23], c.493C > T, c.755 - 1G > C, c.845_c.848 + 1delG, c.933_c.933 + 1insGCAG, c.1381G > T, c.1452 + 1G > A and c.1706_1707delTA. Among the 274 CD patients diagnosed by our group thus far, 41 SLC25A13 mutations/variations were detected. The 7 mutations c.775C > T, c.851_854del4, c.1078C > T, IVS11 + 1G > A, c.1364G > T, c.1399C > T and IVS16ins3kb demonstrated significantly different geographic distribution. Among the total 53 identified genotypes, only c.851_854del4/c.851_854del4 and c.851_854del4/c.1399C > T presented different geographic distribution. The northern population had a higher level of SLC25A13 allelic heterogeneity than those in the south. These findings enriched the SLC25A13 mutation spectrum and brought new insights into the geographic distribution of the variations and genotypes, providing reliable evidences for NICCD definite diagnosis and for the determination of relevant molecular targets in different Chinese areas.

[Clinical features and DGUOK mutations of an infant with mitochondrial DNA depletion syndrome].

The aim of this study was to investigate the clinical features and DGUOK gene mutations of an infant with mitochondrial DNA depletion syndrome (MDS). The patient (more than 7 months old) manifested as hepatosplenomegaly, abnormal liver function, nystagmus and psychomotor retardation. Genetic DNA was extracted from peripheral blood samples of the patient and her parents. Targeted Exome Sequencing was performed to explore the genetic causes. Sanger sequencing was carried out to confirm the detected mutations. The sequencing results showed that the patient was a compound heterozygote for c.679G>A and c.817delT in the DGUOK gene. The former was a reportedly pathogenic missense mutation of maternal origin, while the latter, a frameshift mutation from the father, has not been described yet. The findings in this study expand the mutation spectrum of DGUOK gene, and provide molecular evidence for the etiologic diagnosis of the patient as well as for the genetic counseling and prenatal diagnosis in the family.

Identification of a Large SLC25A13 Deletion via Sophisticated Molecular Analyses Using Peripheral Blood Lymphocytes in an Infant with Neonatal Intrahepatic Cholestasis Caused by Citrin Deficiency (NICCD): A Clinical and Molecular Study.

Background. Neonatal intrahepatic cholestasis caused by citrin deficiency (NICCD) is a Mendelian disorder arising from biallelic SLC25A13 mutations, and SLC25A13 genetic analysis was indispensable for its definite diagnosis. However, conventional SLC25A13 analysis could not detect all mutations, especially obscure large insertions/deletions. This paper aimed to explore the obscure SLC25A13 mutation in an NICCD infant. Methods. Genomic DNA was extracted to screen for 4 high-frequency SLC25A13 mutations, and then all 18 exons and their flanking sequences were analyzed by Sanger sequencing. Subsequently, cDNA cloning, SNP analyses, and semiquantitative PCR were performed to identify the obscure mutation. Results. A maternally inherited mutation IVS16ins3kb was screened out, and then cDNA cloning unveiled paternally inherited alternative splicing variants (ASVs) featuring exon 5 skipping. Ultimately, a large deletion c.329-1687_c.468+3865del5692bp, which has never been described in any other references, was identified via intensive study on the genomic DNA around exon 5 of SLC25A13 gene. Conclusions. An NICCD patient was definitely diagnosed as a compound heterozygote of IVS16ins3kb and c.329-1687_c.468+3865del5692bp. The large deletion enriched the SLC25A13 mutation spectrum, and its identification supported the concept that cDNA cloning analysis, along with other molecular tools such as semiquantitative PCR, could provide valuable clues, facilitating the identification of obscure SLC25A13 deletions.

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.

Screening for five prevalent mutations of SLC25A13 gene in Guangdong, China: a molecular epidemiologic survey of citrin deficiency.

Citrin is the liver-type aspartate/glutamate carrier isoform 2 (AGC2) encoded by SLC25A13 gene, playing important roles in the urea cycle and the malate-aspartate shuttle. Citrin deficiency (CD) has proven a disease entity with high prevalence in south China, including Guangdong with the largest population, but CD epidemiology in this province has not been well characterized. This study aims to screen for five prevalent SLC25A13 mutations, c.851_854del (p.R284fs286X), c.1638_1660dup (p.A554fs570X), c.615+5G>A (p.A206fs212X), IVS16ins3kb (p.A584fs585X) and c.1399C>T (p.R467X), to calculate the mutation carrier rate in Guangdong. A total of 2,428 used blood samples for health examination were collected as the research subjects, including 1,558 from 5 cities in the Pearl River Delta area and the remaining 870 from 4 peripheral cities, and the 5 mutations screened using High Resolution Melting Assay and HybProbe assay. A total of 52 carriers were detected, including 2 carriers of a novel c.1420G>A (p.V474M) mutation that impairs citrin function, as judged by the functional analysis in the yeast system. The carrier rate was higher in Pearl River Delta area than that in the peripheral cities (26/1,558 vs. 26/870, with χ(2) = 4.639 and P < 0.05). The carrier rate was around 1/47 (52/2,428), theoretically with the CD morbidity of 1/8,800 and the number of CD patients over 11,800 in Guangdong population. This study has provided primary epidemiologic data for the evaluation of CD effect in Guangdong province. Moreover, the newly identified c.1420G>A mutation that impairs AGC2 function has enriched the mutation spectrum of human SLC25A13 gene.

Clinical, molecular and functional investigation on an infant with neonatal intrahepatic cholestasis caused by citrin deficiency (NICCD).

BACKGROUND AND OBJECTIVE: SLC25A13 analysis has provided reliable evidences for the definitive diagnosis of citrin deficiency (CD) in the past decade. Meanwhile, these studies generated some issues yet to be resolved, including the pathogenicity of SLC25A13 missense mutations and the mRNA product from the mutation c.615+5G>A. This study aims to investigate the effect of a novel missense mutation on the aspartate/glutamate carrier (AGC) function of citrin protein, and to explore the aberrant transcript from c.615+5G>A in the same CD infant. METHODS AND RESULTS: By means of screening for prevalent SLC25A13 mutations and exons sequencing, the patient proved a compound heterozygote of c.615+5G>A and a novel c.1064G>A (p.Arg355Gln) mutation. An aberrant transcript with retention of the entire intron 6, r.[615+1_615+1789ins; 615+5 g>a] (GenBank accession number KJ128074), which was resulted from c.615+5G>A, was detected by RT-PCR and cDNA sequencing. After bioinformatic analyses of the novel missense mutation c.1064G>A, the growth abilities of three agc1Δ yeast strains were tested, which had been transformed with recombinant or empty vectors, respectively. Besides the bioinformatically pathogenic evidences, the growth ability of the agc1Δ strains transformed with mutant recombinant was the same as with empty vector, but significantly lower than that with normal control in functional analysis. CONCLUSIONS: A CD infant was definitely diagnosed in this paper by a genetic, transcriptional and functional analysis of SLC25A13 gene. This study provided direct laboratory evidences supporting the splice-site nature of the c.615+5G>A mutation, and the novel c.1064G>A variation, which proved a pathogenic mutation bioinformatically and functionally, enriched the SLC25A13 mutation spectrum.