Objective To study the joint effect of small dosages of Aspirin and compound Danshen dropping pills on type 2 diabetes(T2DM) patients'and impaired glucose tolerance(IGT) patients' hypercoagulation blood.Methods The study followed diagnostic standards for T2DM by World Health Organization (WHO) in 1999,and randomly selected two groups of clinical patients:one had 63 patients with IGT and the other contained 68 patients with T2DM.Both groups took compound Danshen dropping pills with the total dosage of 10 pills each time,3 times per day,for continuum of 3 months.After that both groups stopped medication for 7 days,then began taking Aspirin after meal,with the dosage of 100 mg per day,for a continuum of 3 month.After that both groups stopped the medication for 7 days,then both group continued to take same dosage of ASA with addition of compound Danshen dropping pills of total amount of 10 pills per each time,3 times per day,for acontinuum of 3 months.Each medication period (ev ery 3 months) was monitored for platelet count (PET),prothrombin time (PT),kaolin partial thromboplastin time (APPT),The effective rate of anticoagulation for both groups were then calculated.Results The difference of PT、APTT between before and after taking single dosage of ASA for patients with impaired glucose tolerance (IGT) was statistically significant (P＜0.05),while patients with T2DM had no significant difference before and after taking same single dosage of ASA.Similarly, both groups with single dosage of compound Danshen dropping pills had no significant difference between before and after taking the medication.However,the difference of PT、APTT between before and after taking the combination of Aspirin and Danshen dropping pills of both groups were statistically significant (P＜0.05).IGT group with single dosage of ASA had an effective rate of 71.4％ for anti coagulation,while T2DM group with the same dosage reached 23.8％.The IGT group with combinatory dosage of ASA and Danshen dropping pills reached an effective rate of 100％ for anticoagulation,while T2DM group with the same dosage was 86.7％.The difference between those effective rates of anticoagulation was statistically significant in different medicine takeways of two groups (P＜0.05).Conclusion The combinatory use of small dosage of Aspirin with compound Danshen dropping pills had evident effect to improve IGT and hypercoagulation blood of T2DM patients who had good control of their blood glucose.

In order to study the analgesic effect of Shaoyao Gancao Decoction, this paper discussed material basis and mechanism from the perspective of macromolecules in traditional Chinese medicine. Inspired by the phenomenon of turbidity after boiling Chinese medicine, this experiment took Shaoyao Gancao Decoction as the research object to study the formation process of precipitation during boiling. The results showed that aggregates with a certain shape were formed in the solvent during the boiling process, and the precipitate was obtained by standing and centrifuging. Analysis found that the precipitation was mainly composed of small molecules such as paeoniflorin, albiflorin, liquiritin, glycyrrhizic acid, isoliquiritin and gallic acid, and macromolecules such as protein and polysaccharide. The composition of precipitate was consistent with that of Shaoyao Gancao Decoction, but the analgesic effect of Shaoyao Gancao Decoction after removing the precipitate was significantly reduced. Based on these results, we isolated small molecular compounds, polysaccharides and protein from Shaoyao Gancao Decoction and their contents are 60.4, 700.7 and 207.2 mg·g^-1 respectively. We get the ratio, polysaccharide: small molecule = 11.6∶1, protein: small molecule = 3.4∶1, the precipitate is prepared in the state of boiling. The characterization results showed that the particle size of the precipitate will change significantly after co-heating, and the content determination results showed that the content of the six small molecular compounds which was free in solvent was significantly reduced after the formation of the precipitate. The acetic acid writhing experiment proved that the precipitate has a good analgesic effect, and effectively reduced the levels of inflammatory factors prostaglandin E2 and nitric oxide, and increased the level of anti-inflammatory factor interleukin-10. These results proved that the precipitate in Shaoyao Gancao Decoction is an important material basis for analgesic effect, and macromolecules such as protein and polysaccharide are the main components of the precipitate. The study of macromolecules in the precipitate of Shaoyao Gancao Decoction not only provides new ideas and methods for elucidating the pharmacodynamic material basis of Shaoyao Gancao Decoction, but also provides a reference for analyzing the scientificity of traditional decoction.

Traditional clinical students training model is teacher-centered this education model has some defects.Whether an education model is advanced or not is relative to whether we can cultivate creative students with high quality.We use“one center, two auxiliarys,three stages”in our clinical education and fully utilize advanced education methods,combine theory with practice and cultivate creative students wiht high quality.

Objective To investigate the relationship of Maternal-fetal interface local cytokine GM-CSF and its receptor with the occurrence of early spontaneous abortion. Methods From August 2009 to December 2011,we collected 30 villi tissue samples with artificial abortion and 30 villi tissue samples with spontaneous abortion. At the same time, we collected 30 villi tissue samples with artificial abortion and 30 decidua tissue samples with spontaneous abortion,and 30 decidua tissue samples with spontaneous abortion. The human chorionic gonadotropin ( HCG) was detected by radioimmunoassay in every group. The expressions of granulocyte-macrophage colony-stimulating factor (GM-CSF) and granulocyte-macrophage colony-stimulating factor receptor (GM-CSFR ) were detected by Immunohistochemical staining and Western Blot in the villi tissue and decidua tissue in every group. Results The concentration of HCG in the spontaneous abortion group was lower than that in the artificial abortion group ( <0.05) . The protein expressions of GM-CSF and GM-CSFR were found in villus and deciduas tissues in both groups. The protein expression levels of GM-CSFR in the villus tissues were higher in spontaneous abortion group than those in artificial abortion group ( <0.05), the protein expression of GM-CSF was upregulated, but there was no statistically significant difference between two groups. In deciduas tissues, the protein expressions of GM-CSF and GM-CSFR were upregualted in spontaneous abortion group ( <0.05) . Conclusions The suitable concentrations of GM-CSF and GM-CSFR in decidua tissue maintain the pregnancy continued. However,the higher concentrations of GM-CSF and GM-CSFR in the decidua tissue may be one of reasons of spontaneous abortion.

Objective To clone ,construction ,express and purify Tp47 of Treponema pallidum (Tp) ,and assess the immunoreac‐tivity by Western‐Blot .Methods Tp47 was amplified by polymerase chain reaction ,and then cloned to the vector pGEX‐6P‐1 .The correct sequence of the recombinant plasmids pGEX‐6P1‐Tp47 was transformed into Escherichia coli BL21 (DE3) and induced .The expression product was analyzed by sodium dodecyl sulfate polyacrylamide gel electropheresis and Western‐Blot .The expression protein was purified .Serum of different clinical stages of syphilis was used as the antibody to detect the immunoreactivity of the protein by Western‐Blot .Results A fusion protein with molecular weight about 71 × 103 was attained .Western‐Blot proved that the recombinant protein can react with Tp IgG positive sera .And the specificities and sensitivities of the diagnostic reagent detected by sera were 100% .Conclusion The recombinant protein Tp47 was expressed and purified with good antigen activity ,which could provide the basis of theory and practice for the development of early diagnostic kit applying to detect Tp infection .

?AIM: To study the association of the single nucleotide polymorphism ( SNP) rs1157699 in the calcitonin receptor-like receptor ( CRLR ) gene with primary angle closure ( PAC) in a Han Chinese population.?METHODS: All samples, involved 232 PAC cases and 306 controls, were obtained from an epidemiologic survey conducted in Funing, Jiangsu Province, China. Genotyping were carried out by TaqMan-MGB probe using the real time quantitative polymerase chain reaction system to study the relationship between SNP of rs1157699 in CRLR gene and PAC.?RESULTS: The prevalence of CRLRrs1157699 genotype was 67.4%, 30.0%, 2.6% for CC, CT, TT in cases, and 71.3%, 27.0%, 1.7% in controls respectively.There was no difference between the two groups in the distribution of genotype and allele frequencies of rs1157699 (P>0.05).?CONCLUSION:Our results do not support a significant role for rs1157699 in CRLR with PAC.

Objective To clone and express of Rv0091 encoding protein in Mycobacterium tuberculosis,identify and characterize of the enzyme activities.Methods Construct the Rv0091 prokaryotic expression plasmid,the vector was transformed into E.coli strain BL21trxB.After induced by IPTG,recombinant protein was purified by Ni2+-NTA chromatography and analyzed for purity by SDS-PAGE gels stained with Coomassie Blue.Immunological activity was identified by Western blot.The recombinant protein molecular weight was identified by Mass spectrometry.The enzyme-coupled assay detectes enzyme activity.Results The expression plasmid pET32a-Rv0091 was constructed and expressed in E.coli.BL21trxB,and the optimum expression system was conformed.The purity of the recombinant protein was more than 95％.Western blot analysis confirmed that recombinant protein was one of Mycobacterium tuberculosis proteins.Mass spectrometry identified the relative molecular weight and theoretical molecular weight was basically the same.Enzyme assay showed the recombinant protein able to catalyze the substrate MTA.Enzymatic properties showed that the optimal buffer for the phosphate and Hepes buffer,the poor thermal stability of the enzyme,the optimal temperature of 37℃,optimal pH10-12,when the pH ≤7,the protein denaturation and loss of some vitality.Conclusion The recombinant protein methylthioadenosine nucleosidase(MTAN) was obtained and enzyme activity was detected and plays a key role in the metabolism of Mycobacterium tuberculosis.

Background Researches showed that triterpenoids,with a similar structure to lanosterol,has therapeutical effect on many systemic diseases,and lanosterol was determined to have a therapeutical effect on cataract recently.However,how the lanosterol plays effects on other eye diseases is still unelucidated.Understanding the distribution of lanosterol in ocular tissue is helpful for us to elucidate the relationship of lanosterol with eye diseases.Objective This study attempted to investigate the distribution of lanosterol synthase (LSS) and lanosterol in cornea,lens and retina tissue of rats and offer a basis for the targeting treatment of eye diseases.Methods Fifteen SPF male SD rats were sacrificed by excessive anesthesia to obtain the eyeballs.The relative expressions of LSS protein and gene in the cornea,lens and retina tissue of the rats were detected by Western blot and reverse transcription (RT)-PCR,respectively.Immunofluorescence staining technology was used to locate the distribution of LSS in cornea,lens and retina tissue.The contents of lanosterol in the cornea,lens and retina tissue were analyzed by liquid chromatograph mass spectrometer (LC-MS).Results No LSS protein and mRNA was expressed in the retinal tissue in normal rats.The mean relative expression of LSS protein in the lens and cornea was 0.43±0.05 and 0.25±0.03,respectively,showing a significant difference between them (t =-5.35,P＜ 0.01).The relative expression of LSS mRNA was 0.51 ±0.04 and 0.29 ±0.02 in the lens and cornea,respectively,with a stronger expression in the lens in comparison with the cornea (t =-8.34,P＜0.01).Immunofluorescence staining showed that LSS primarily located in corneal epithelial layer,stromal layer and endothelial layer as well as lens epithelial cells and shallow cortex layer and hardly expressed in retina,and no co-expression of LSS with the neuron marked by NeuN and the Müller cell marked by glutamine synthetase (GS) in retinal tissue.LC-MS analysis revealed that the contents of lanosterol in lens and cornea was (24.37 ±2.91) ng/mg and (5.31 ±0.58) ng/mg,respectively,with a significant difference between them (t =-11.13,P＜0.01).Conclusions LSS and lanosterol extensively distribute in cornea and lens of normal rats,but not in retina tissue.These results offer new strategies for the target treatment of relevant eye diseases.

Objective To investigate the etfect of microenvironment simulated by colon carcinoma homogenate supernatant on the differentiation and development of human dendritic ceils (DCs), and to investigate the function of vascular endothelial growth factor A (VEGF-A) during this process . Methods Fresh colon carcinoma and peri-cancer tissues were collected to prepare homogenate supernatant. The pe-ripberal blood mononuclear cells were isolated and cultured with 1640 medium including rhGM-CSF and rhIL-4. Then the colon carcinoma homogenate supernatant, peri--carcinoma homogenate supernatant and VEGF-A were added to the cultures at day 2. Antigen of colon carcinoma cell line SW620 was added at day 4 and lipopolysaccharide (LPS) was added at day 6. DCs were collected at day 8 for further study. The con-tent of VEGF-A was tested by ELISA. The morphology and the immunopbenotype of DCs were checked by microscope and flow cytometry, respectively. The expression of CDIa was tested by RT-PCR, and the prolif-eration and killing rate of T cell was measured by CCK-8. Results The content of VEGF-A in the homoge-nate supernatant of colon carcinoma was significantly higher than that of the peri-carcinoma (P ＜ 0. 05). Compared with normal DCs, the cell morphology of colon carcinoma homogenate aupernatant group was in-hibited, and the cell number was decreased. Besides, the positive expression rate of DC surface markers de-creased (P ＜ 0.01). The capacity of mixed lymphocyte reaction (MLR) and killing capacity of T cells de-creased(P ＜0.01). However, there was almost no difference between VEGF-A group and normal DCs on the cell morphology and cell number, and VEGF-A had no obvious inhibition on the expression of DCs sur-face markers (P ＞ 0.05). But VEGF-A group had significantly inhibitory effect on the MLR and T cells kill-ing. Conclusion The tumor microenvironment simulated by the colon carcinoma homogenate supernatant obviously has inhibitory effect on the differentiation and function of DCs, and VEGF-A has the inhibitory effect on DC function, but the inhibitory effect is not through the inhibition of the expression of DC costimu-lators.

Objective To observe the sequence of fat deposit and its relationship with insulin resistance in SD rats fed by high fat diet.Methods Normal 8-week-old male SD rats were randomly divided into normal chow (NC,n=40)and high fat diet(HF,n=40)groups.Triglyceride(TG)in serum,liver and muscle were measured;glucose infusion rate(GIR)and the mRNA level of genes related to lipid metabolism in liver and muscle were determined in different stages.GIR was detected by eugiyeemic-hyperinsulinemic clamp for evaluating the insulin sensitivity.Gene expression was determined by real-time PCR.Results(1)As compared with NC group,serum TG was not increased after high fat feeding for4 and 8 weeks,it began to increase after 12 weeks [0.52(0.15-1.00) mmol/L vs O.31(0.09-0.53)retool/L, P0.05)in skeletal muscle.After 8 weeks,the expression of ACC1 in liver in HF group was increased by 20.6%,CPT-1 was decreased by 27.1%(P