Objective To prenatally diagnose HA fetus with different clinical backgrounds. Methods Genetic tests were performed on 15 gravidas subjected for prenatal diagnosis of HA and different methods were employed for diagnosis according to the gestational weeks and clinical data. Amniotic fluid were taken from pregnant women within 23 gestational weeks for direct genotyping and indirect linkage analysis, since these women had probands with clear-cut mutations. Cordocentesis was performed for linkage analysis in pregnant women over 23 gestational weeks with probands whose types of mutation were unknown, while the FⅧ activity tests were carried out simultaneously. For the pregnant women over 23 gestational weeks without proband, cordocentesis was operated for measurement of FⅧ activity and karyotyping, but carriers of hemophilia A could not be detected in these cases. The introns 22 and 1 inversion of F8 gene were identified by long distance-polymerase chain reaction. Nucleotide sequencing was employed if the gene inversion could not be found and linkage analysis of 7 polymorphic markers, including DXS1108, F8Civs13, INTRON22,DXS1073,DXS9901, DXS15, DXS8069 and sex site (Amelo) were applied eventually. Identification of maternal blood contamination must be done before the tests. Results Fifteen samples were identified without maternal blood contamination. Five fetuses were diagnosed with hemophilia A. Meanwhile there were three pregnant women whose cord blood FⅧ activities were less than 1%. One of them was accompanied by trisomy 21; another had inversion mutation in introns 22 of F8 gene; the remaining one was identified with missense mutation in exon 23 (p. Arg2182Cys) of F8 gene. Conclusions Diverse methods should be applied in prenatal diagnosis of hemophilia A with different clinical backgrounds, for the sake of birth defects prevention.

HIV/STIs remain a major global public health problem. One of the global strategies for the prevention and control of HIV/STIs is to interrupt their transmission, which requires the public health methods based on scientific evidence and cost-effectiveness. The scale-up of male circumcision services in the priority countries of the HIV-prevention project in sub-Saharan Africa has been hampered by the scarcity of trained providers and relative technical difficulty of male circumcision techniques recommended by WHO and UNAIDS. Shang Ring is an innovative and disposable device for male circumcision, which has been safely used for over 600 000 males in China since 2006. Clinical studies of more than 3 000 cases of Shang Ring circumcision in China, Kenya, Zambia, and Uganda have demonstrated its safety, effectiveness, acceptability and ease of use. The most obvious advantages of Shang Ring include short procedure time (3-6 min), excellent postoperative cosmesis, low rate of complications, high acceptance by clients and providers, ease of use, and standardization for reliable performance. As an innovative technique, Shang Ring has a great potential for facilitating the safe and effective scale-up of circumcision services. This article comprehensively reviews the clinical studies of Shang Ring male circumcision in China and Africa.
Africa ; China ; Circumcision, Male ; instrumentation ; methods ; HIV Infections ; prevention & control ; Humans ; Male

To analyze killer immunoglobulin (Ig)-like receptor (KIR) gene content and allelic polymorphism in Zhejiang Han population, samples were genotyped by polymerase chain reaction sequence-specific primers (PCR-SSP). The results demonstrated that all 17 KIR genes could be observed in the population. All individuals contained 2DL4, 3DL2 and 3DL3 genes. The frequencies of these genes was 1.00. 2DL1, 2DL3, 2DP1, 3DP1*003, 3DL1, 2DS4*001/002 loci were more common, their frequencies were 0.902, 0.902, 0.902, 0.902, 0.7598, 0.5615 respectively, while the frequencies of 2DL2, 2DL5A, 2DL5B, 2DS1, 2DS2, 2DS3, 2DS4*003, 2DS5, 3DS1 and 3DP1*001/002 were relatively lower. The A KIR haplotype was the most prevalent (74.7%) in Zhejiang Han population and there were 12 different KIR haplotypes in total, the most common was 2 (53.0%). Twenty six different genotypes have been found in the population, AJ (2, 2) and AF (1, 2) showed higher frequencies, followed by AH (2, 5), NN2 (2, 6), AI (1, 5) and AG (1, 1). Fifteen of these genotypes have not been found in Caucasians so far and four new KIR profiles could not be assigned to the haplotypes according to standard assign method. In conclusion, there are distinctive frequencies of KIR gene content, haplotype as well as genotype in Zhejiang Han population.
China ; Gene Frequency ; Genotype ; Haplotypes ; Humans ; Polymerase Chain Reaction ; methods ; Polymorphism, Genetic ; Receptors, KIR ; genetics

The purpose of this study was to establish beta block matching technique. DNA was extracted from whole blood by salting-out method, beta block matching was performed by PCR and GeneScan technique. The results showed that the length of fragments amplificated in 100 samples was different and the range of them was 91-197 bp. Amplification fragments could be divided into four regions: 91-93, 105-113, 125-139 and 177-197 bp respectively. 91 bp DNA fragments could be found in all of samples. The numbers of DNA fragments with different length have been shown high polymorphism and they focused on the range of seven to twenty four. In conclusion, the beta block matching technique is reliable and applicable to the selection of hematopoietic stem cell transplantation donors.
DNA ; genetics ; HLA-B Antigens ; genetics ; HLA-C Antigens ; genetics ; HLA-DR Antigens ; genetics ; HLA-DRB1 Chains ; Hematopoietic Stem Cell Transplantation ; Histocompatibility Testing ; methods ; Humans ; Polymerase Chain Reaction ; methods

OBJECTIVETo establish a method of indirect immunofluorescence (IIF) to measure DNA (mDNA)-associated autoantibodies to cell membrane, and to evaluate diagnostic value of the anti-mDNA antibodies in patients with juvenile systemic lupus erythematosus (SLE) in comparison with anti-dsDNA antibody.

METHODSForty-four children with SLE were enrolled in this study. As a control group, 30 children with other rheumatic diseases were also enrolled. Anti-mDNA and anti-dsDNA antibodies were measured by IIF. Anti-smooth muscle (Sm) antibodies were measured by immuno-double diffusion (ID) and IIF.

RESULTSOut of 44 juvenile SLE patients, 34 (77.27%) were seropositive for anti-mDNA, which was significantly higher than that of patients with other rheumatic diseases (20.00%, P<0.05). The sensitivity and specificity of anti-mDNA for juvenile SLE diagnosis were 77.27% and 80.00%, respectively. The positive predictive value and negative predictive value were 85.00% and 70.59%, respectively. The positive rate of anti-mDNA in SLE lacking of anti-dsDNA and anti-Sm antibodies were 68.00% (17/25) and 79.49% (31/39), respectively.

CONCLUSIONThe detection of anti-mDNA antibodies is useful for diagnosis of juvenile SLE, especially in patients who are negative for anti-dsDNA antibodies and anti-Sm antibodies.

Adolescent ; Antibodies, Antinuclear ; analysis ; Case-Control Studies ; Cell Membrane ; immunology ; Child ; Female ; Fluorescent Antibody Technique, Indirect ; methods ; Humans ; Lupus Erythematosus, Systemic ; immunology ; Male ; Predictive Value of Tests ; Sensitivity and Specificity

To establish delta block HLA-matching technique, DNA was extracted from whole blood by salting-out method, delta block was amplified by polymerase chain reaction (PCR), and PCR product was detected by GeneScan. The results showed that delta block had polymorphism in 104 samples without sibship of the Han people from Zhejiang province. The range of DNA fragment length was 81-393 bp and could be divided into 4 groups: 81-118 bp, 140-175 bp, 217-301 bp, 340-393 bp. The numbers of DNA fragments were 6-32. It is concluded that the method of delta block matching is reliable and can be applied to select donors for the patients to be transplanted. It is the first time to get delta block data of the Han people in China.
HLA-A Antigens ; genetics ; immunology ; HLA-B Antigens ; genetics ; immunology ; HLA-DQ Antigens ; genetics ; immunology ; HLA-DR Antigens ; genetics ; immunology ; HLA-DRB1 Chains ; Hematopoietic Stem Cell Transplantation ; Histocompatibility Testing ; methods ; Humans

The aim of this study was aimed to investigate the molecular genetic basis for a novel HLA allele, HLA-B*4061, in Chinese population. DNA was extracted from whole blood by salting-out method. The HLA-B exons 1 - 8 of the proband was amplified and the amplified product was cloned using TOPO TA cloning sequencing kit to split the two alleles apart. Both strands of exons 2, 3 and 4 of chosen colonies were sequencing. The PCR-SSP was performed to confirm the mutations detected by sequencing. The sequencing results showed HLA-B alleles of the proband as B*4601 and the novel allele. The sequences of the novel allele have been submitted to GenBank (DQ089628, DQ089629, DQ089630). After HLA blast analysis, the novel allele showed a single nucleotide mismatch with B*400101 in exon 2 at position 272 C-->A, as the results, changing amino acid from Ser to Tyr at codon 67. It is concluded that this allele is a novel one and has been officially named B*4061 by the WHO Nomenclature Committee.
Alleles ; Amino Acid Substitution ; Asian Continental Ancestry Group ; genetics ; Base Sequence ; China ; HLA-B Antigens ; genetics ; immunology ; Histocompatibility Testing ; Humans ; Molecular Sequence Data ; Sequence Analysis, DNA ; Sequence Homology, Nucleic Acid

OBJECTIVEThe aim of this study was to explore the possibility of creating a toxin, C-CPE-ETA', by fusing C-terminal high affinity binding domain of CPE (C-CPE) with a truncated form of Pseudomonas aeruginosa exotoxin A (ETA') and to examine whether C-CPE-ETA' could specifically target CLDN-3, 4 molecule and the targeted toxin was cytotoxic against CLDN-3,4-overexpressing ovarian cancer.

METHODSCLDN-3 and CLDN-4 expressions were analyzed at the mRNA level in three ovarian cancer cell lines and epithelial ovarian cancer tissues from 20 patients. After transforming an expression plasmid of C-CPE-ETA' into E. coli BL21 (DE3) plysS strain, the recombinant protein was purified using His-Bind resin chromatography column and analyzed by Western blot and Coomassie blue staining. The specific binding, proapoptotic and cytolytic activities were evaluated by flow cytometry, fluorescence microscopy with the JC-1 probe and MTT assay in CLDN-3,4-overexpressing ovarian cancer cells.

RESULTSQuantitive RT-PCR results showed there existed high levels of CLDN-3 and CLDN-4 in ovarian cancer cells, CAOV3, OVCAR3 and SKOV3. Moreover, high expressions of CLDN-3 and CLDN-4 were observed in 90.0% (18/20) and 60.0% (12/20) of ovarian cancer tissues, with an expression level 10-fold higher than that in the normal ovarian tissue. A 58 000 recombinant protein C-CPE-ETA' was demonstrated by Western blot and Coomassie blue staining. Purified and recombinant C-CPE-ETA' was bound with high affinity to CLDN-3,4-overexpressing ovarian cancer cells, CAOV3, OVCAR3 and SKOV3 cells. C-CPE-ETA' was strongly proapoptotic and cytotoxic towards the CLDN-3,4-overexpressing ovarian cancer cells. The concentration of IC(50) was 7.364 ng/ml for CAOV3 cells, 8.110 ng/ml for OVCAR3 cells and 22.340 ng/ml for SKOV3 cells, respectively. However, control CLDN-3,4-deficient cell line HUVEC was not susceptible to the recombinant C-CPE-ETA' at a concentration up to 10 µg/ml.

CONCLUSIONSThe C-CPE-ETA' protein exhibits remarkably specific cytotoxicity for CLDN-3,4-overexpressing ovarian cancer cells. Its therapeutic potential warrants further development for ovarian cancer molecular targeted therapy.

ADP Ribose Transferases ; metabolism ; physiology ; Apoptosis ; Bacterial Toxins ; metabolism ; Cell Line, Tumor ; Claudin-3 ; Claudin-4 ; Claudins ; genetics ; metabolism ; Enterotoxins ; metabolism ; physiology ; Exotoxins ; metabolism ; physiology ; Female ; Humans ; Immunotoxins ; metabolism ; Ovarian Neoplasms ; metabolism ; pathology ; RNA, Messenger ; metabolism ; Recombinant Fusion Proteins ; metabolism ; physiology ; Virulence Factors ; metabolism ; physiology

Objective: This research aimed to explore the therapeutic effect and its mechanism of pirfenidone in liver cirrhosis induced by carbon tetrachloride in mice. Methods: Sixty male C57 BL/6 mice were randomly divided into the control group, model group and different doses of pirfenidone group, twelve rats in each group. Mice were intraperitoneally injected with 20％ CCl4 soybean oil solution ( 5 ml/kg), twice a week for 7 weeks. And these mice were free to drink 20％ ethanol solution in the third week after building the model. The low, medium and high dose groups were respectively given 50, 100 and 200 mg/kg of pirfenidone solution according to the body weights, while the model group and control group were given equal volume of blank solvent after building the model, once a day for 2 weeks. The serum level of ALT and AST, liver index, spleen index, the gene or protein expression level of TGF-β1 and Smad3 were analyzed before and after the treatment of pirfenidone. Results: The serum level of ALT, AST increased significantly in the model group ( P＜0. 05), while decreased significantly in different doses of pirfenidone group ( P＜0. 05). The liver and spleen index in the model group was significantly higher than that in the control group ( P＜0. 05). However, after treating with pirfenidone, the liver and spleen index were significantly lower than that in the model group ( P＜0. 05). The number of TGF-β1 positive cells in the model group was significantly more than that in the control group, but it was significantly decreased in the pirfenidone group. The gene expression level of Smad3 in the model group was significantly higher than that in the control group ( P＜0. 05). The gene expression level of TGF-β1 and Smad3 in different doses of pirfenidone group were significantly lower than that in the model group ( P＜ 0. 05). Meanwhile, the protein level of TGF-β1 and Smad3 were significantly increased in the model group, while decreased in the pirfenidone group. Conclusion: Pirfenidone relieves liver cirrhosis caused by carbon tetrachloride in mice by inhibiting the TGF-β1/Smad3 signaling pathway.

Objective:To purpose of this study is to introduce how peripheral blood neutrophil/lymphocyte ratio (NLR) before operations influences the prognosis of patients with breast cancer.Methods:Review of systems were used to analyze patients who suffered from breast cancer and accepted modified radical mastectomy of breast cancer according to the clinical data of 180 cases of Shenyang Military Region General Hospital between January 2002 and January 2005.All the patients were classified into two groups according to the NLR with the critical value at 6.0.2 statistics were used to evaluate the relationship between NLR of two groups and clinical pathological characteristic.Kaplan-Meier survival analysis and Cox's proportional hazards regression model were used to analyze the relationship among NLR of two groups,other clinical pathologic characteristic and prognosis of patients.Results:The high level of NLR is related with the size of patients' tumor,lymph node metastasis and TNM stages (P＜0.05).Kaplan-Meier survival curves indicated the group of high level of NLR's overall survival (OS) and disease-free survival (DFS) was significantly lower than the low level NLR group (P＜0.05).Single factor and multivariate cox's proportional hazards regression model indicated the high level of NLR before operations,the size of tumor,lymph node metastasis and TNM stages were significantly related with the OS and DFS (P＜0.05).Conclusion:The high level of NLR before operations is an independent risk factor to influence the OS and DFS of the patients who accepted modified radical mastectomy of breast cancer.