Objective To discuss the application of serum hs-CRP and lipids detection and the correlations between them in type 2 diabetes mellitus.Methods The levels of serum hs-CRP,TC,TG, HDL-C,LDL-C,ApoA1,ApoB were all measured in 49 type 2 diabetic patients and 53 normal cases.Results The type 2 diabetic group had significantly higher hs-CRP, TC,TG, LDL-C, ApoB and lower HDL-C than the control group. The concentration of serum hs-CRP correlated positively with TG,ApoB and negatively with HDL-C.Conclusion Serum hs-CRP level is associated with serum lipid level and inflammatory response might play an important role in the pathogenesis of type 2 diabetes mellitus. In type 2 diabetic mellitus there is an increased risk of development of cardiovascular diseases.

Objective To discuss the clinical application of polynomial curves fitting.Methods Based on the experiments of TBIL, ALT, DBIL and Cr,the linear experimental data were polynomial fitted.Results The optimal polynomial of TBIL is y=-3.886+7.544x , and the evaluation is linear 1; The optimal polynomial of ALT is y=5.293+25.897x-0.043 x~2, and the evaluation is linear 2;The optimal polynomial of DBIL is y=-2.950+1.688x+0.011x~2, and it′s non-linear; The optimum polynomial of Cr is y=11.654+14.512x-0.010 x~2, and the evaluation is inexactitude.Conclusion The polynomial fitting curve is the perfect linear evaluation method. It guarantees both the accuracy and reliability of the experimental results, and is more suitable to clinic.

Objective To express outer membrane protein 2(Omp2) of Chlamydia trachomatis, purify expressed products and study its immunity.Methods The target gene encoding Omp2 167—434 amino acid residues was amplified by PCR from C. trachomatis template DNA. The targeted DNA fragment was cloned into expression vector pET28b(+) and introduced into competent E. coli BL21(DE3) cell. Recombinant Omp2aa_ 167 ～aa_ 434 was expressed after induction by IPTG and analyzed by SDS-PAGE and Western blot, purified with Ni-NTA-His affinity chromatography. The rOmp2aa_ 167 ～aa_ 434 was used to immune rabbits for immunogenicity assessment.Results Restriction enzymes cleavage analysis and DNA sequencing confirmed that the plasmid pET28b(+)/Omp2aa_ 167 ～aa_ 434 was correctly constructed. The 35.0?103 molecular weight pure protein, which specifically reacted with serum from C. trachomatis infected patient by Western blot, was obtained by optimizing the conditions for both expression and purification. The titer of serum antibodies was above 1∶1 280 as detected by ELISA.Conclusion The expressed product showed good immunity.

BACKGROUND: Tonifying kidney is the main method to treat aging of nervous system, which is characterized by decreasing density of nerve cells,cell aging and deposition of lipofuscin granule in the cytoplasm of nerve cell.OBJECTIVE: To investigate the effect of nourishing kidney and tonifying brain on density and ultrastructural of nerve cell in hippocampal CA1 and CA3 area of aging mouse. DESIGN: Completely randomized and controlled study.SETTING: Institute of Traditional Chinese Medicine of Hunan Province.MATERIALS: The experiment was completed at the Pharmacological Laboratory of Institute of Traditional Chinese Medicine of Hunan Province between September 2000 and November 2003. Totally 30 Kunming mice with 11-month old were selected. Solution of yishen jiannao Ⅰ (zishen prescription) was provided by Agent Laboratory of Institute of Traditional Chinese Medicine of Hunan Province (including 15 g shouwu, 15 g sangshen, 15 g gouqizi, 6 g wuweizi, 30 g danshen, 30 g gegen, 10 g honghua,10 g shichangpu, 10 g yujin, 10 g yuanzhi, 10 g shanzha and scorpion).Scorpion was crushed into power. The rest drugs were decocted with water twice, mixed together and filtered, then the powder of scropion was added.The raw material was 1.0 g/mL.METHODS: Thirty mice were randomly divided into 3 groups with 10 in each group. Mice in control group were given distilled water, those in zishen prescription group were given 20.0 g/kg zishen prescription, and in vitamin E group were given 40.0 mg/kg vitamin E. All mice were perfused with 20 mL/g, once a day for 4 weeks. After 1 hour of the last medication, mice were sacrificed at once at the drugged state. Cranium of mice was sheared and the hippocampal tissue was taken out. Numbers of nerve cells in CA1 and CA3 area were calculated under the microscope to calculate the density of nerve cell.MAIN OUTCOME MEASURES: ① Density of nerve cell in cerebral hippocampus of mice in each group; ② Ultrastructure of nerve cell in cerebral hippocamp us of mice in each group.RESULTS: Totally 30 mice entered the final analysis. ① Density of nerve cell in hippocampal CA1 and CA3 area of brain: Density in zishen prescription group and vitamin E group was higher than that in control group [CA1: (3 707±495), (3 812±281), (257±372) mm-2; CA3:(2 746±262),(2 397±366), (1 992±307) mm-2, t=2.68-8.30, P ＜ 0.05-0.01]. ② Aging of mitochondrion was decreased, deposition of lipofuscin and aging of lysosome in cytoplasm were reduced in zishen prescription group and vitamin E group.CONCLUSION: Granule of nourishing kidney and tonifying brain can defer density decrease of nerve cell and aging of cell in hippocampla area of mice' brain.

Objective To observe the clinical efficacy of acupoint thread embedding on the peripheral serum leptin and insulin levels in simple obesity patients, and to further explore the mechanism of acupoint thread embedding in treating simple obesity.Method A total of 120 patients with simple obesity were randomized into a control group, an eletroacupuncture (EA) group and a thread embedding group, 40 cases in each group. The control group was intervened by diet plus exercise intervention, the EA group by EA treatment in addition to the intervention given to the control group and the thread embedding group by acupoint thread embedding treatment in addition to the intervention given to the control group. The levels of blood serum leptin and insulin in the three groups were observed before and after 2 treatment courses and the clinical efficacies were compared among the three groups.Result The total effective rate was 87.5% in the thread embedding group, 85.0% in the EA group, and 47.5% in the control group. The total effective rates in the thread embedding group and EA group were significantly different from the rate in the control group (P<0.05). The levels of the fasting serum leptin and insulin were significantly changed in the thread embedding group and EA group after the treatment (P<0.01). After the treatment, the levels of the fasting serum leptin and insulin in the thread embedding group and EA group were significantly different from those in the control group (P<0.01,P<0.05). The insulin level in the thread embedding group was better than that in the EA group, and the difference was statistically significant (P<0.05).Conclusion Acupoint thread embedding is an effective approach for simple obesity, and it can down-regulate the fasting peripheral serum leptin and insulin levels.

Objective To construct the recombinant plasmid containing the major outer membrane protein(MOMP) gene of Chlamydia trachomatis and expres s MOMP protein in E.coli BL21. Methods The MOMP gene was amplified by polymera se chain reaction from the genome of Chlamydia trachomatis serovar D. The amplif ied fragment was directly inserted into pUCm-T vector and verified by DNA sequen cing. MOMP gene was then subcloned into the prokaryotic expression vector pET-22 b(+). The recombinant protein of MOMP was purified by Ni-NTA affinity chromatogr aphy and identified by SDS-PAGE and Western blot. Results The MOMP gene, which is about 1 200 bp, was successfully amplified and cloned. The DNA sequence of t he cloned MOMP gene was the same as that published by the GenBank. SDS-PAGE anal ysis showed that the relative molecular weight of this fusion protein was about 47 kDa which was consistent with the theoretically predicted value, and the spec ificity of this recombinant protein was confirmed by Western blot. Conclusions The MOMP gene of Chlamydia trachomatis was successfully cloned and expressed in the prokaryotic expression system, which may lay the foundation for the developm ent of Chlamydia trachomatis vaccine.

Objective To investigate the expression of eukaryotic initiation factor 4E(eIF4E) ,vascular endothelial growth factor (VEGF)‐A and VEGF‐C in gastric cancer tissues and their correlation with invasion and metastasis of gastric carcinoma .Methods The expressions of eIF4E ,VEGF‐A and VEGF‐C were detected in tissues of 58 gastric carcinomas and 25 normal gastric mucosa by using immunohistochemical method .Results The positive rate of eIF4E、VEGF‐A and VEGF‐C protein expression were 89 .7%(52/58) ,65 .5% (38/58) ,60 .3% (35/58) in gastric carcinoma ,which were higher than those in normal gastric mucosa tissues which were 4 .0% (1/25) ,12 .0% (3/25) ,8 .0% (2/25) respectively .Expressions of eIF4E ,VEGF‐A and VEGF‐C were significantly correlated with the depth of invasion and lymph node metastasis(P<0 .05) ,but not with the age ,sex of patients(P>0 .05) .Ex‐pressions of eIF4E and VEGF‐C were significantly correlated with tumor differentiation .The expression of eIF4E was being found positively correlated with VEGF‐A and VEGF‐C .Conclusion eIF4E may play certain roles in the oncogenesis and progression of gastric carcinoma .VEGF‐A and VEGF‐C may be helpful in lymph metastasis .Combined detection of eIF4E ,VEGF‐A and VEGF‐C may be helpful to assess the malignant degree and prognosis of gastric carcinoma .

Objective To construct DNA vaccine containing MOMP gene of Chlamydia trachomatis and to observe immune response in mice. Methods Mice of 4 - 6 weeks old were immunized with pcDNA3.1-MOMP or pcDNA3.1 intramuscularly at a dose of 100 μg. Booster immunizations were employed at 2-week interval for two times. Specific antibody in the sera of mice and the level of IFN-γ in murine spleen lymphocyte supernatant were detected by ELISA. The proliferation response of spleen cells was detected by MTT assay. Results Significant specific antibody titers were observed and the highest titer was 1: 1 024 in mice after three times immunization with pcDNA3.1-MOMP. The proli-feration response of spleen cells were significantly higher than that of mice injected with pc DNA3.1. IFN-γ reached(532.0 + 45.4)pg/mL in immunized mice. Conclusion Strong responses of humoral and cellular immunity can be evoked by DNA vaccine of pcDNA3.1-MOMP in mice.

Objective To evaluate the performance of the hypersensitive C-reactive protein(hs-CRP) rapid quantitative chemilu-minescent detection kit .Methods According to National Committee for Clinical Laboratory Standards (NCCLS) EP10-A2 docu-ment ,hs-CRP rapid quantitative chemiluminescent detection kit was employed to measure the CRP at low ,medium and high con-centration levels of quality control serum .Bias ,total imprecision and their slope rates ,intercepts ,carryover ,non-linearity and drift were calculated ,and its clinical acceptability was evaluated .Results Bias and total imprecision of hs-CRP rapid quantitative chemi-luminescent detection kit were within the allowable ranges ,the average values of slope rates ,intercepts ,carryover ,non-linearity and drift were 1 .005 7 ,0 .537 8 ,0 .789 6% ,0 .019 2 ,0 .036 0 ,respectively ,the differences showed no statistically significance ( P>0 .05) .Conclusion hs-CRP rapid quantitative chemiluminescent detection kit has good accuracy and precision ,stable perform-ance ,and consistent with the clinical testing requirements .

Objective To investigate the effect of heat shock protein 10 (HSP1O) of Chlamydophila pneumoniae in inducing TNF-α on THP-1 cells and the roles of TLR4 and TLR2 involved in it.Methods Purified native recombinant HSP10 from Cpn(CHSP10) were produced and inactivated the endotoxin contamination,then different concentration (0.5,1,5,10,20,30 μg/ml) of CHSP10 were used to stimulate THP-1 for different time (0,6,12,24,36,48,60 h).TNF-α were measured by using human TNF-α ELISA kit and compared among different groups.THP-1 were collected and analyzed for TLR2 and TLR4 mRNA levels and protein expression by RT-PCR and immunofluorescence.Peritoneal macrophages isolated from wide-type (C3 H/HeN) and TLR4-deficient mice (C3H/HeJ) were stimulated with endotoxin-free proteins respectively,and the TNF-α were measured.Furthermore,neutralizing anti-human TLR2/TLR4 McAb as a blocking Ab was preincubated with THP-1,after stimulation with CHSP10,ELISA was used to detect the concentration of TNF-α.Results TNF-α can be induced with CHSP10 in THP-1,while it significantly decreased with heated or deproteinized CHSP10.Both TLR2 and TLR4 mRNA and protein were detected in THP-1.Macrophages from C3H/HeN mice displayed higher TNF-α compared with it from C3H/HeJ mice after stimulation with CHSP10.The CHSP10-induced TNF-α would obviously decline when treated with antiTLR2/TLR4 McAb.Conclusion As a potential inflammation related protein,CHSP10 are involved in the pathogenesis of Cpn inducing inflammation cytokine TNF-α.TLR2 and TLR4 appear to be involved in CHSP10-mediated expression of TNF-α.