Biallelic mutations in DCDC2 cause neonatal sclerosing cholangitis in a Chinese family.

Background: Neonatal sclerosing cholangitis (NSC) is a severe cholestatic liver disease, which often develops into end-stage liver disease in childhood and requires liver transplantation. Mutations in CLDN1 and DCDC2 are confirmed to be the main pathogenic mechanism of NSC.

Methods: Whole exon sequencing (WES) was performed to find the possible disease-causing mutations of this family.

Splicing of exon 9a in FMR1 transcripts results in a truncated FMRP with altered subcellular distribution.

FMRP is an RNA-binding protein, loss of which causes fragile X syndrome (FXS). FMRP has several isoforms resulted from alternative splicing (AS) of fragile X mental retardation 1 (FMR1) gene, but their biological functions are still poorly understood. In the analysis of alternatively spliced FMR1 transcripts in the blood cells from a patient with FXS-like phenotypes (normal CGG repeats and no mutation in coding sequence of FMR1), we identified three novel FMR1 transcripts that include a previously unidentified microexon (46 bp), terming the exon 9a.

Hybrid minigene splicing assay verifies the pathogenicity of a novel splice site variant in the COL1A1 gene of a chinese patient with osteogenesis imperfecta type I.

Background: Osteogenesis imperfecta (OI) is a rare genetic bone disease associated with brittle bones and fractures. Among all known types, OI type I is the most common type and characterized by increased bone fragility, low bone mass, distinctly blue-gray sclera, and susceptibility to conductive hearing loss beginning in adolescence. Mutations in genes encoding type I collagen (COL1A1 and COL1A2) contribute to the main pathogenic mechanism of OI.

Whole-exome sequencing identifies compound heterozygous mutations in ARSA of two siblings presented with atypical onset of metachromatic leukodystrophy from a Chinese pedigree.

Background: Metachromatic leukodystrophy (MLD) is a rare inherited lysosomal storage disorder caused mainly by variants in arylsulfatase A (ARSA) gene. MLD can be divided into three major clinical forms according to the age of onset: late infantile, juvenile, and adult. We report two siblings of late infantile MLD presenting with cerebellar ataxia as the only first clinical symptom.

The Value of Preoperative Neutrophil to Lymphocyte Ratio in Indicating Lymph Node Metastasis in Patients with Resectable T2 Stage Gastric Adenocarcinoma.

Background: Lymph node metastasis (LNM) is closely associated with poor prognosis in patients with resectable T2 stage gastric adenocarcinoma (RT2-GA). Preoperative blood neutrophil to lymphocyte ratio (NLR) has been identified to be a very valuable predictor for prognosis in patients with diverse cancers. The aim of this investigation was to assess the relationship between NLR and LNM in RT2-GA.

Improvement and Evaluation of a Staining Method for Measuring Sperm Lactate Dehydrogenase C4 Activity.

Background: This study sought to improve and evaluate a 2-hydroxyvaleric acid based staining method for detection of lactate dehydrogenase C4 (LDH-C4) activity in human spermatozoa.

Methods: A staining method for measuring sperm LDH-C4 activity with the substrate 2-hydroxyvaleric acid was improved. Expression level of LDH-C4 was assessed by Western blotting.

Hybrid minigene splicing assay verified the pathogenicity of a novel splice site variant in the dystrophin gene of a Chinese patient with typical Duchenne muscular dystrophy phenotype.

Background: Duchenne muscular dystrophy (DMD) is typically caused by disrupting the reading frame of the dystrophin gene: approximately 70%-80% of mutational events are represented by deletions or duplications of one or more exons in the dystrophin gene, and the remaining cases by subtle mutations, including point mutations, small indels, small inversions, and complex small rearrangements. The dystrophin gene is the largest known gene with one of the highest known rates of new mutations.

Methods: Deletions and duplications were detected in the DMD gene of the proband by using multiple ligation-dependent probe amplification (MLPA).

Alternatively spliced products lacking exon 12 dominate the expression of fragile X mental retardation 1 gene in human tissues.

Fragile X mental retardation 1 gene (FMR1) expression is associated with fragile X syndrome (FXS) and exhibits several splicing products. However, the proportion of spliced isoforms that are expressed in different tissues remains unclear. In the present study, long-chain reverse transcription-polymerase chain reaction with a T cloning-sequencing method was conducted in order to analyze the entire coding region of the FMR1 gene in human tissues.

[A novel mutation of NTRK1 gene in a family with congenital insensitivity to pain with anhidrosis].

Objective: To screen for mutations in the neurotrophic tyrosine kinase receptor type 1 (NTRK1) gene in a Chinese family affected with congenital insensitivity to pain with anhidrosis (CIPA).

Methods: With informed consent obtained, peripheral blood samples were obtained from the patient and his family members. Seventeen coding exons and intron-exon boundaries of the NTRK1 gene were amplified with PCR and analyzed by direct sequencing.

[Analysis of small supernumerary marker chromosome 15q11 in four infertile males].

Objective: To delineate the origins of small supernumerary marker chromosomes (sSMCs) identified in 4 infertile males.

Methods: The sSMCs were analyzed with combined G-banding, N-banding, multiplex ligation-dependent probe amplification (MLPA), fluorescence in situ hybridization (FISH) and single nucleotide polymorphisms array (SNP-array) techniques.

Results: G-banding analysis has suggested a 46,X,-Y,+mar karyotype in all of the 4 cases.

[Delineation of three structural Y chromosome aberrations combined molecular techniques].

Objective: To delineate the structure of Y chromosome aberrations and recombinant mechanisms for three patients.

Methods: Karyotype analysis, multiplex ligation dependent probe amplification (MLPA), fluorescence in situ hybridization (FISH), Y chromosome sequence tagged sites (STS) analysis, human whole genome-wide SNP array were used.

Results: The karyotypes of the three patients were 46, X, +mar.

A rapid and sensitive protocol for prenatal molecular diagnosis of X-linked adrenoleukodystrophy.

Background: X-linked adrenoleukodystrophy (X-ALD) is a neurodegenerative genetic disease characterized by progressive demylination of the brain, adrenal insufficiency and elevated VLCFA level. ABCD1gene is the disease gene and more than 500 unique mutations in the ABCD1gene have been recorded in the database, approximately 60% of which are noncurrent ones. Although great progress has been made in the treatment of X-ALD, prenatal diagnosis is still badly needed by X-ALD-stricken families.

Evaluation of an in-house protocol for prenatal molecular diagnosis of SMA in Chinese.

Background: Spinal muscular atrophy (SMA) is a common autosomal recessive neuromuscular disorder characterized by degeneration of the anterior horn of the spinal cord, leading to symmetric muscle weakness and atrophy. About 95% of SMA patients have homozygous loss of SMN1 which can be detected by conventional PCR-RFLP testing. However, the method cannot distinguish heterozygous healthy carriers.

Increased expression of hLRH-1 in human gastric cancer and its implication in tumorigenesis.

Altered signaling pathways or deregulated transcription factors represent an important category of molecular events leading to aberrant gene regulation in gastric cancer, among which the role of WNT/beta-catenin pathway remains unclear. LRH-1 is a critical transcription factor in controlling cell proliferation via crosstalk with the beta-catenin signaling pathway. In order to gain a knowledge of the expression of hLRH-1v1 and hLRH-1 in gastric cancer, a Q-PCR analysis was carried out.

Microarray analysis of gene-expression profile in hepatocellular carcinoma cell, BEL-7402, with stable suppression of hLRH-1 via a DNA vector-based RNA interference.

To establish a cell line with a permanent suppression of hLRH-1 in this study, a stable RNAi vector (pSineohLRH-1) targeting hLRH-1 was constructed and introduced into hepatocellular carcinoma cell, BEL-7402. By semiquantitative RT-PCR analysis, the expression of hLRH-1 in BEL-7402 cells carrying pSineohLRH-1 was shown to be significantly suppressed by up to approximately 60%. In addition, microarray analysis was carried out to assess the extent of altered gene expression in BEL-7402 cells with stable knockdown of hLRH-1.

[Expression and characterization of subunit C of mouse lactate dehydrogenase in Escherichia coli].

Objectives: To construct a prokaryotic recombinant vector for mouse lactate dehydrogenase-C and to detect its expression in BL21.

Methods: The coding sequence of mouse lactate dehydrogenase subunit C was amplified from mouse testis RNA with specific primers, and cloned into pGEX-2T after the restriction digestion with BamH I and EcoR I. GST fusion protein was expressed after induction with IPTG.