OBJECTIVE:To study the impact of amoxicillin on dysmenorrhea model mice treated with paeoniflorin.METHODS:The enrolled mice were randomized to different groups and all were given diethylstilbestrol i.g.for 12 consecutive days.From day 5 to day 12,the model control(MC)group,positive control(PC)group and 3 PA groups(different dosage of paeoniflorin plus amoxicillin)were given amoxicillin i.g.On day 12,3 P(paeoniflorin)groups and 3 PA groups were respectively given different dosage of paeoniflorin i.g.All groups were respectively given oxytocin i.p 1 hour after medication.The incubation period(TIP,seconds)between oxytocin i.p.and the first time of body twisting and the number of the body twisting(NBT)in 15 minutes of the mice were recorded.RESULTS:Compare to NC(negative control)and MC groups,the TIP was significantly prolonged and the NBT in 15min were significantly decreased in 3 P groups(P0.05).CONCLUSION:The curative effect of paeoniflorin in treating dysmenorrheal model mouse was decreased by amoxicillin suggests that the 2 drugs were unsuitable to be used concomitantly.

AIM: To amplify and analyze the differential DNA fragment between pathogenic leptospira serovar lai and nonpathagenic leptospira serovar patoc I. METHODS: The previously subtractive DNA fragment only existing in leptospira serovar Lai was amplified by cassette ligation and semi-nested PCR.The obtained gene was sequenced and searched homologically. In addition, the deduced amino acid was analyzed and the secondary stracture of protein was predicted. RESULTS: The 580 bp DNA fragment, which deposited in GenBank (AF495587), was cloned, and four overlapping open reading frames (ORF) was contained. The high homology with conserved hypothetical protein streptococcus pyogenes was found. CONCLUSION: This study lays foundation for deeply exploring biological actions of new gene and pathogenic mechanism of leptospira serovar lai.

Objective To study the molecular genetic mechanism of a patient with 17? hydroxylase (CYP17) deficiency. Methods Genomic DNA were abstracted from the blood of the patient, her parents and healthy control. The 8 exons of CYP17 gene were amplified, using 5 pairs of designed primers, with polymerase chain reaction (PCR), and the 8 exons were sequenced by the dideoxy terminator method to determined the mutation sites. The corresponding exons of the parents of the patients were also amplified and sequenced to determine the zygosity of the patient and the source of the gene variances. Results The analysis revealed that the patient (46, XY) was a compound heterozygote carrying two different inherited mutations on CYP17 gene, one from mother containing a point mutation Arg 96 (C G G)→ Gln(C A G) and the other from father containing a nine base deletion (CACTCTTTC) at amino acid position 487～489 (Asp Ser Phe) near the carboxyl terminus of P450c17. Conclusion The CYP17 gene of the patient with 17? hydroxylase deficiency is a compound heterozygous mutation. The mutation changes the amino acid sequence of P450c17 enzyme, which in turn affected the enzymatic activity. Arg 96 is essential in P450c17 enzyme activity. Deletion of Asp 487 Ser 488 Phe 489 in exon 8 may be a prevalent mutation causing P450c17 deficiency in Southeast Asia.

Objective To explore new method for enhancing the efficacy of tuberculosis DNA vaccine. Methods Two recombinant plasmids were constructed, one named as pBK GM/85A encoding mouse granulocyte macrophage colony stimulating factor(GM CSF) fused to Mycobacterium tuberculosis Ag85A antigen, the other named as pBK 85A encoding Mycobacterium tuberculosis Ag85A antigen alone. Subsequently, the two plasmids were transferred into cultured COS7 cells by using cationic liposomes. The expression products were identified by Western blotting. Then, in a murine model, we compared the immunogenicity and protective immunity of the two recombinant plasmids following genetic immunization. Results All pBK GM/85A injected mice elicited higher antibody titres than that for pBK 85A injected mice. Lymphocytes obtained from the spleen of pBK GM/85A immunized mice exhibited higher lymphocyte proliferative response、IFN ? production and CTL activity than that for pBK 85A immunized mice. The protective efficacy was also higher for pBK GM/85A immunized mice than that for pBK 85A immunized mice. However, The protective efficacy for pBK GM/85A immunized mice was lower than that for BCG immunized mice. Conclusion These results showed that DNA vaccines with GM CSF/antigen fusion constructs could greatly improve the immunogenicity of DNA vaccine against Mycobacterium tuberculosis.

Objective To study on the expression of the eukaryotic recombinant vector carrying HlyX gene of Leptospira serovar Lai in mammalian cell and explore the humoral immune response in BALB/c mice immunized with the recombinant plasmid. Methods The HlyX gene was amplified from Leptospira serovar Lai genomic DNA by PCR and inserted into pcDNA3.1 vector. After transformed into E. coli DH5α,the recombinant plasmid was assayed for identification by PCR analysis,restriction nuclease enzyme digestion and sequencing. The recombinant plasmid was transfected into COS-7 cells,then RT-PCR and Western blot were performed to test the expression of the target gene. The recombinant plasmid was injected intramuscularly into BALB/c mice for three times at intervals of two weeks,and the antibody titer was measured by ELISA. Results PCR showed the full length HlyX gene was about 1100 bp. PCR analysis,restriction nuclease enzyme digestion and sequencing indicated the recombinant vector was constructed successfully. After the plasmid Was transfected into COS-7 cells,a fragment about 1100 bp was found by RT-PCR and a specific band relative molecular mass(Mr)about 40×103,which was consistent with the expected size of the target proteins was showed by Western blot. ELISA showed the antibody titer in BALB/c mice immunized by the HlyX gene of Leptospira serovar Lai can elicit high-titer antibody in BALB/c mice,which has laid the foundation for the application of the DNA vaccine.

As the dominant antigens, early secreted antigenic target 6 (ESAT6, E6) and culture filtrate protein 10 (CFP10, C10) had once been the focus of tuberculosis (TB) vaccine due to their capability of inducing strong cell immune response in the host. They are also endowed with promising future of prevention against and diagnosis of TB. In this review, we systematically introduce recent research progress of E6 and C10, especially in structure-function, biological characteristics, protein expression and secretion, host immunity and vaccine development, and the prospects of their application are also discussed.
Antigens, Bacterial ; chemistry ; genetics ; immunology ; Bacterial Proteins ; chemistry ; genetics ; immunology ; Humans ; Immunodominant Epitopes ; immunology ; Molecular Biology ; Peptide Fragments ; chemistry ; genetics ; immunology ; Tuberculosis Vaccines ; genetics ; immunology ; Vaccines, DNA ; immunology

Objective To observe the clinical efficacy of Jin's three-needling therapy on children autism with different TCM syndrome types.Methods A total of 202 autism children was enrolled into the study.Autism children were divided into two groups:the treatment group(N=118)received Jin's three-needling therapy,the acupoints mainly consisted of Sishen needling,Dingshen needling,temporal triple needling,supertemporal triple needling,brain triple needling,mental triple needling,Xingshen needling,hand needling for improving mental,foot needling for improving mental and tongue triple needling,and some other acupoints were selected according to the syndrome patterns;the control group(N=84)received special training.The treatment was done one time per day,six times one week,and 4 months constituted one treatment course.After treatment,the therapeutic effect was evaluated by Children Autism Rating Scale(CARS).Results After one treatment course,the total curative effective rate was 88.1% in the treatment group and 65.5% in the control group(P

In order to provide substantial scientific information for exploring the mechanism of porcine liver injury caused by Taenia asiatica (T.asiatica),the expression of Cysteine aspartyl proteinase 3 (Caspase-3) from liver tissues of porkets that were experimentally infected by T.asiatica was examined.The T.asiatica adults were collected from the taeniasis patients in Duyun,Guizhou Province and identified biologically.The eggs were harvested from gravid proglottids and prepared by repeated washing and centrifugation.Twelve 20-days old Yorkshire and Seghers hybrid porkets were randomly divided into experimental and control groups as six pigs per group.The experimental group was orally administrated with 1.5 × 106 eggs per porket at day 0 post-infection.The porkets of both groups were sacrificed on the day 15 and day 75 post-infection (three pigs per time point) respectively,and liver samples were collected for further experiments.Quantitative real-time polymerase chain reaction method was employed to detect the mRNA levels of Caspase-3,and western blotting and immunohistochemistry methods were performed to detect the level of Caspase-3 expression in both groups.At the day 15 post-infection,the mRNA level and expression level of Caspase-3 of the experimental group were significantly decreased,comparison with the control group (P =0.011,P=0.008 and P=0.004 respectively).It was positive with Caspase-3 when yellow or brown signal appeared in the cytoplasm of liver cells by immunohistochemistry.However,at the day 75 post-infection,the mRNA level and expression level of Caspase-3 of the experimental group were dramatically similar to the control group.Furthermore,in the experimental group,the mRNA level and expression level of Caspase-3 were significantly increased at day 75 post-infection than day 15 post-infection (P--0.018,P=0.003 and P=0.002 respectively).These results suggested that Caspase-3 might be involved into the regulation of the damage of porcine liver induced by T.asiatica challenge at the early infection stage and have on effect to the hepatic injury because of the dramatic recovery of Caspase-3 at the consequent infection stage.

AIM: Construction of an eukaryote- E. coli shuttle expressing recombinant plasmid which expresses OmpL1 envelope protein of pathogenic Leptospira, serovar Lai strain 017. METHODS: The OmpL 1 gene was amplified by PCR from the leptospiral genome. Then it was cut with restriction enzymes and ligated to the plasmid pBK-CMV. The correct recombinant plasmid was screened out with analysis of restriction enzymes and PCR. After inducing the E. coli baring recombinant plasmid with IPTG,the complete protein of the bacteria was extracted for SDS-PAGE. At the same time, OD600 of the host bacteria was examined at different time after inducing or uninducing with IPTG. RESULTS: Five strains E. coli containing proper recombinant plasmids were screened out. Four strains E. coli expressed a new protein with a weight of 37 kD among them. With the expression of the heterogenous protein,the OD600 of the host bacteria decreased. CONCLUSION: The shuttle expressing plasmid of the OmpL 1 gene of strong virulent Leptospira strain 017 was successfully constructed. Furthermore,the recombinant plasmid expressed the expected OmpL 1 fusion protein in E. coli and the expression of the heterogenous protein had toxic effect on the host bacteria. This work was important for the future research of OmpL1 protein which relates to the diagnosis,new vaccine preparing and the pathogenic mechanism of leptospirosis.

AIM:To detect the drug-resistance mycobacterium tuberculosis by using DNA microarray hybridization.METHODS: DNA microarray for detecting drug-resistance of mycobacterium tuberculosis was prepared; Clinical isolated strains were cultivated and their drug-resistance sensitivity was detected. The genome DNA of mycobacterium tuberculosis was prepared and the drug-resistance genes of the mycobacterium tuberculosis were amplified by PCR. Then the gene chip was hybridized, washed, detected and analyzed. RESULTS: Results of cultivating mycobacterium tuberculosis and detecting the drug-resistance sensitivity: one strain was drug-sensitive; four strains were multi-drug-resistant; The detecting results of the drug-resistance was consistent with the results of diagnosis therapy of the 5 clinical patients. The detecting results of gene chip confirmed the above facts. CONCLUSION: Detecting drug-resistance mycobacterium tuberculosis by the gene chip is precise, fast and highly-efficient.