BACKGROUNDNevoid basal cell carcinoma syndrome (NBCCS) is a rare autosomal dominant disease characterized by a combination of development anomalies and a predisposition to tumour formation. Mutation of patched gene (PTCH), considered the molecular defect of NBCCS, in a Chinese NBCCS family was investigated in this study.

METHODSGenomic DNA was isolated from blood samples of all 12 members of this family. The mutated PTCH gene was screened by polymerase chain reaction amplification and direct sequencing.

RESULTSA new mutation of 3 bp (GAT deletion) was found in all seven affected members of this family. This mutation caused one aspartate deletion in the fourth transmembrane domain of the PTCH protein located within the sterol sensing domain (SSD). This deletion was not found in any unaffected members of this family nor in 200 control samples.

CONCLUSIONSOur findings suggest that one 3-bp deletion in PTCH gene was the cause of nevoid basal cell carcinoma in a Chinese family through affecting the conformation and function of PTCH protein.

Basal Cell Nevus Syndrome ; genetics ; Humans ; Mutation ; Patched Receptors ; Patched-1 Receptor ; Receptors, Cell Surface ; genetics

OBJECTIVE: To explore the genetic etiology of a fetus with autosomal recessive polycystic kidney disease (ARPKD). METHODS: Prenatal ultrasonography has revealed oligohydramnios and abnormal structure of fetal kidneys. After careful counseling, the couple opted induced abortion. With informed consent, genomic DNA was extracted from the muscle sample of the abortus and peripheral blood samples of the couple. High throughput whole exome sequencing was carried out to detect potential variants in relation with the disease. Suspected variants were verified by Sanger sequencing. RESULTS: Prenatal ultrasound revealed increased size of fetal kidneys, with multiple hyperechos from the right kidney, and multiple hyperechos with anechoic masses within the left kidney. DNA sequencing revealed that the fetus has carried heterozygous variants of the PKHD1 gene, including c.7994T>C inherited from its father, and two heterozygous variants of the PKHD1 gene c.5681G>A from its mother. CONCLUSION The compound heterozygous c.7994T>C and c.5681G>A variants of the PKHD1 gene probably underlay the pathogenesis of ARPKD in this fetus. Above results can provide guidance for subsequent pregnancies of the couple.
Female ; Fetus ; Genetic Testing ; Humans ; Mutation ; Polycystic Kidney, Autosomal Recessive/genetics* ; Pregnancy ; Receptors, Cell Surface/genetics*

OBJECTIVETo investigate whether leptin receptor Lys109Arg polymorphism influences non-alcoholic fatty liver disease.

METHODSGenomic DNA samples were extracted from blood of subjects who had received a physical examination. Genotyping was performed using oligonucleotide microarray and these fluorescence labeled PCR-amplified fragments were hybridized to allele-specific oligonucleotide probes. The relevant mutation was confirmed by sequencing analysis.

RESULTSA total of 180 subjects (109 males and 71 females) were included in the study, 117 of them had fatty liver disease and the other 63 had no liver problems and served as healthy controls. There were 144 (80%) subjects with GG genotype (Arg109Arg), 33 (18.3%) with GA genotype (Lys109Arg) and 3 (1.7%) with AA genotype (Lys109Lys). The distribution of leptin receptor Lys109Arg polymorphism had no significant difference (P > 0.05) between the fatty liver disease patients (95GG, 21GA and 1AA) and the healthy control subjects (49GG, 12GA and 2AA). The abdominal wall fat was significantly thicker in AA genotype subjects (4.1+/-0.4) cm than that in GA (2.8+/-0.6) cm and GG genotype subjects (2.7+/-0.7) cm (F = 5.197, P = 0.006). The serum cholesterol levels in AA genotype subjects (5.1+/-0.4) mmol/L was significantly lower than that in AG (25.5+/-6.9) mmol/L and GG genotype (27.2+/-8.4) mmol/L subjects (F = 8.164, P = 0.005). There were no significant differences in age, body mass index, hip circumference, waist circumference, blood pressure (BP), percentage of body fat, blood protein, triglyceride, HDL and fasting blood glucose between AA, GG and GA genotype subjects.

CONCLUSIONLeptin receptor Lys109Arg polymorphism may be involved in the regulation of distribution of abdominal wall fat thickness and cholesterol metabolism. Whether leptin receptor Lys109Arg polymorphism is in any way related to fatty liver disease is still not known.

Adult ; Arginine ; chemistry ; genetics ; Fatty Liver ; etiology ; genetics ; Female ; Genotype ; Humans ; Lysine ; chemistry ; genetics ; Male ; Middle Aged ; Polymorphism, Genetic ; genetics ; Receptors, Cell Surface ; genetics ; Receptors, Leptin

OBJECTIVETo screen rare blood groups Fy(a-), s-, k-, Di(b-) and Js(b-) in an ethnic Zhuang population.

METHODSSequence-specific primers were designed based on single nucleotide polymorphism (SNP) sites of blood group antigens Fy(b) and s. A specific multiplex PCR system I was established. Multiplex PCR system II was applied to detect alleles antigens Di(b), k, Js(b)1910 and Js(b) 2019 at the same time. The two systems was were used to screen for rare blood group antigens in 4490 randomly selected healthy donors of Guangxi Zhuang ethnic origin.

RESULTSWe successfully made the multiplex PCR system I. We detected the rare blood group antigens using the two PCR system. There are five Fy(a-), three s(-), two Di(b-) in 4490 Guangxi zhuang random samples. The multiplex PCR system I has achieved good accuracy and stability. With multiplex PCR systems I and II, 4490 samples were screened. Five Fy(a-), three s(-) and two Di(b-) samples were discovered.

CONCLUSIONMultiplex PCR is an effective methods, which can be used for high throughput screening of rare blood groups. The rare blood types of Guangxi Zhuang ethnic origin obtained through the screening can provide valuable information for compatible blood transfusion. Through screening we obtained precious rare blood type materials which can be used to improve the capability of compatible infusion and reduce the transfusion reactions.

Blood Group Antigens ; genetics ; Duffy Blood-Group System ; genetics ; Genotype ; Humans ; Multiplex Polymerase Chain Reaction ; methods ; Receptors, Cell Surface ; genetics

OBJECTIVETo construct Epithelia Membrane Protein 1 gene-deficient in human fetal nucleus pulposus model by lentivirus-mediated RNA interference for building a platform for illustrating the biomechanisms role of EMP-1 during human intervertebral disc degeneration.

METHODSThe lentivirus vector with shRNA targeting EMP-1 mRNA was transected into 293FT cells by liposome. Then the lentivirus supernatant was obtained and used for infecting human fetal nucleus pulposus. The expression of GFP was observed under fluorescence microscope after 48 h. The viral particles were collected at 72 h after transfection. The efficacy of gene interference was tested by Western blot and Real-time RT-PCR. Analysis the results of the fluorescent microscope scenes and get the average values of EMP-1/GAPDH by detected the interference efficiency of various interference DNA sequences with western blot and semi quantitative RT-PCR methods.

RESULTSThe lentivirns with high titer were obtained and the EMP-1 gene deficient cell strains were obtained. Semi quantitative RT-PCR and Western blot proved the average values of EMP-1/GAPDH decreased from 0.46 to 0.32 and 0.5 to 0.25 (P < 0.01).

CONCLUSIONLentivirus packaging technology can be mastered skillfully. EMP-1 gene-deficient cell models are successfully established.

Fetus ; HEK293 Cells ; Humans ; Intervertebral Disc ; metabolism ; Lentivirus ; genetics ; Neoplasm Proteins ; genetics ; RNA Interference ; Receptors, Cell Surface ; genetics ; Transfection

BACKGROUNDAutosomal recessive polycystic kidney disease (ARPKD) is a rare inherited disease, which is a disorder with multiple organ involvement, mainly the kidney and liver. It is caused by mutations in the PKHD1 gene. Here, we reported the clinical characteristics of a case with ARPKD and analyze the genetic features of this patient as well as of his father using targeted exome sequencing and Sanger sequencing.

METHODSGenomic DNA was extracted from peripheral blood leukocytes obtained from a patient with ARPKD. The mutations were identified using exome sequencing and confirmed by Sanger sequencing.

RESULTSThe patient was diagnosed as ARPKD based on ultrasonography and abdominal computed tomography which showed polycystic changes, multiple calcinosis of both kidneys, and multiple dilated bile ducts of the liver. Compound heterozygous PKHD1 gene mutations A979G and G5935A, which lead to substitution of an asparagine for an aspartate at amino acid 327 (N327D) and a glycine for an arginine at amino acid 1979 (G1979R) respectively, were identified using targeted exome sequencing and confirmed by Sanger sequencing for the patient. In addition, the father of the patient was identified to be a carrier of heterozygous A979G mutation of this gene.

CONCLUSIONSWe identified that the compound heterozygous PKHD1 gene mutations are the molecular basis of the patient with ARPKD. Targeted exome sequencing is suitable for genetic diagnosis of single-gene inherited diseases like ARPKD in which the pathogenic gene is a large.

Adolescent ; Exome ; genetics ; Genetic Predisposition to Disease ; Humans ; Male ; Mutation ; Polycystic Kidney, Autosomal Recessive ; genetics ; Receptors, Cell Surface ; genetics

OBJECTIVETo study the molecular genetic etiology of a Chinese pedigree with basal cell nevus syndrome.

METHODSThe proband and his affected mother and a unaffected individual in the pedigree were chosen and peripheral blood was collected from them for DNA. Direct sequencing was performed to detect the mutations of PTCH gene. In order to further confirm the results of sequence analysis, all available family members were analyzed with genetic linkage analysis using 3 highly polymorphic microsatellite DNA markers in the region of 9q22.3-q31.

RESULTSNo mutations of PTCH gene was detected in the proband's mother, one synonymous mutation was detected in the proband. Linkage analysis showed that the Lod scores of the 3 markers were: D9S283, Z = -2.11 (theta = 0.00); D9S1690, Z = -2.95 (theta = 0.00); D9S1677, Z = -5.94 (theta = 0.00).

CONCLUSIONSIn this pedigree, mutation of PTCH gene is not related to the underlying pathogenesis of the syndrome.

Asian Continental Ancestry Group ; genetics ; Basal Cell Nevus Syndrome ; genetics ; Female ; Genetic Linkage ; Humans ; Male ; Mutation ; Patched Receptors ; Patched-1 Receptor ; Pedigree ; Receptors, Cell Surface ; genetics

OBJECTIVE: To evaluate the effects of CLEC5A expression level on cell proliferation, migration and invasion and epithelial-mesenchymal transition (EMT) in hepatocellular carcinoma (HCC) and explore the role of CLEC5A in the tumorigenesis and progression of HCC. METHODS: The expression level of CLEC5A was detected in 50 pairs of HCC and adjacent tissues using immunohistochemical staining, and its association with clinicopathological parameters of HCC patients was analyzed. Cultured HCC cell line SK-HEP-1 was transfected with a lentiviral vector overexpressing CLEC5A, and the transfection efficiency was verified using real-time fluorescence quantitative PCR and Western blotting. The changes in proliferation, migration and invasion abilities of the transfected cells were analyzed using CCK-8, 5-ethynyl-29-deoxyuridine (EdU) and Transwell assays, and EMT of the cells was determined using Western blotting. RESULTS: The protein expression level of CLEC5A was significantly lower in HCC tissues than in the adjacent tissues (P < 0.001). The expression level of CLEC5A was significantly correlated with tumor size (P=0.008), tumor number (P=0.010), histological differentiation (P=0.016), microvascular invasion (P=0.024) and BCLC stage (P=0.040). In SK-HEP-1 cells, overexpression of CLEC5A obviously inhibited the cell proliferation, migration and invasion and reversed EMT phenotype of the cells. CONCLUSION CLEC5A is a potential HCC suppressor gene and may serve as a promising therapeutic target for HCC.
Humans ; Carcinoma, Hepatocellular/genetics* ; Epithelial-Mesenchymal Transition ; Liver Neoplasms/genetics* ; Cell Proliferation ; Cell Differentiation ; Receptors, Cell Surface/genetics* ; Lectins, C-Type/genetics*

Toll-like receptor (TLR) 3 is a member of the TLR family that confers innate immunity by recognizing viral pathogens. Herein, we report that the TLR3 isoform is expressed on human primary cells and cell lines. This isoform has 2, 520 bp cDNAs compared to the 2, 712 bp of full cDNA, is produced by deletion of an intron-like sequence within exon 4 and is co-expressed with wild type TLR3 in primary human astrocytes and glioblastoma cell lines. This finding suggests the TLR3 isoform in astrocytes may have a different immunological role for binding ligands during the immune response in brain.
Astrocytes/*physiology ; Cloning, Molecular ; Human ; Isomerism ; Membrane Glycoproteins/chemistry/*genetics ; Receptors, Cell Surface/chemistry/*genetics ; Support, Non-U.S. Gov't

OBJECTIVE: To explore the genetic basis for a fetus with renal abnormalities through whole exome sequencing and imaging examination. METHODS: Clinical data and result of medical imaging of the fetus was collected. Amniotic fluid sample was collected for the extraction of fetal DNA. Whole exome sequencing was carried out. Candidate variants were verified by Sanger sequencing. RESULTS: Prenatal ultrasonography showed that the fetus had bilateral enlargement of the kidneys with hyperechogenicity and diffuse renal cysts. Whole exome sequencing revealed that the fetus carried compound heterozygous variants of the PKHD1 gene, namely c.5137G>T and c.2335_2336delCA, which were derived from its mother and father, respectively. CONCLUSION The fetus was diagnosed with autosomal recessive polycystic kidney disease through combined prenatal ultrasonography and whole exome sequencing. The compound heterozygous variants of the PKHD1 gene probably underlay the pathogenesis in the fetus. The results have enabled prenatal diagnosis and genetic counseling for its parents.
Female ; Genetic Testing ; Humans ; Polycystic Kidney, Autosomal Recessive/genetics* ; Pregnancy ; Prenatal Diagnosis ; Receptors, Cell Surface/genetics* ; Whole Exome Sequencing