Objective To characterize and quantify the CD4 +CD25 + regulatory T (Treg) cell population in peripheral blood of patients with ankylosing spondylitis (AS) and to determine the influence of treatment with tumor necrosis factor (TNF)-a inhibitors on them.Methods Peripheral blood mononuclear cells (PBMC) were isolated from 25 patients with active AS,in which 10 patients were treated with 12 weeks of etanercept,and 21 healthy subjects.CD4+CD25high T cells were analyzed using flow cytometry,and mRNA expression of FOXP3 was determined by real-time polymerase chain reaction (PCR).Proliferation of T cells to PHA was measured by WST-1 assay using depleted CD25+ cells by immunomagnetic sorting.Results There was no significant difference in the percentage of CD4+CD25high cells in peripheral blood between patients with active AS and controls (P＞0.05).However,PBMC from patients with active AS expressed reduced levels of FOXP3 mRNA (P＜0.01) which were inversely correlated with C-reactive protein (CRP)(P＜0.01).CD4+CD25+ cells in peripheral blood of both active AS patients and controls exhibited suppressive capacity on the proliferation of effector T cells in vitro (both P＜0.01).Treatment with etanereept increased significantly CD4+CD25high cells and FOXP3 mRNA expression (both P＜0.01),with negative correlations between these increases and decrease in CRP levels (P＜0.05 and P＜0.01,respectively).Conclusion In AS patients,peripheral FOXP3-expressing CD4 +CD25 + Treg cells are abnormal,and are up-regulated by etanercept treatment.This suggests a possible pathogenesis of AS and a potential mechanism for clinical efficacy of TNF-α inhibitors.

Objective To provide the experimental basis for the further research of the interacting proteins with Stathmin ,the Stathmin gene Pichia pastoris expression system was constructed ,the expressed Stathmin product was purified and identified .Meth‐ods Stathmin gene was amplified from tumor cell line of SKBR3 by PCR method and cloned into the yeast expression vector pPIC3 .5K .The recombinant vector pPIC3 .5K‐Stathmin was constructed and transformed into Pichia pastoris GS115 .The positive clones were screened by YPD medium containing Geneticin 600 μg/mL .Expression was induced with 0 .5% methanol and expres‐sion products were identified by SDS‐PAGE and Western Blotting .Results DNA sequencing result showed that the gene fragment was consistent with Stathmin gene sequence .pPIC3 .5K‐Stathmin was selected from YPD culture medium containing Geneticin ,and the positive clones were identified by PCR .SDS‐PAGE showed that a 37 × 103 protein band could be seen on the PAGE gel after Coomassie Blue staining ,which was further confirmed and identified as Stathmin protein by Western Blotting .Conclusion Stathmin yeast expression vector is successfully constructed and expressed in Pichia pastoris ,which laid the foundation for the study of inter‐acting proteins with Stathmin ,and for the preparation of the biological treatment drugs of Stahtmin target .

Objective To explore the relationship between the Stathmin gene expression in breast cancer cells MDA-MB-231, MCF-7 and the biological behaviours such as cell growth,adhesion and invasion,and provide experimental basis of breast cancer metastasis for further study.Methods Used RT-PCR and Western Blot methods to detect the Stathmin gene expres-sion levels in MDA-MB-231 and MCF-7 cells,and in the mean while to test the MDA-MB-231 and MCF-7 cell growth,adhe-sion,invasion ability by CCK-8 cell proliferation experiments,cell adhesion experiments,cell invasion experiments,then, analyed the relationship of Stathmin gene expression and cell growth,adhesion,invasion ability.Results Over-expression levels of Stathmin gene were observed both in the MDA-MB-231 and MCF-7 cells (F=10.173,P<0.05),and furthermore, the expression levels of Stathmin gene in MDA-MB-231 cells was higher than in MCF-7 cells (t=4.562,P<0.05).While, the growth,adhesion and invasion ability of the MDA-MB-231 cells was higher than that of MCF-7 cells(P<0.05).Conclu-sion The higher level of Stathmin gene expression,the stronger breast cancer cells had ability of growth,invasion,and ad-hesive.The Stathmin gene expression levels was closely correlated with breast cancer cell invasive.

Objective To explore the potential role of high levels of adiponectin (AD) in the inflammatory joint of rheumatoid arthritis (RA). Methods ELISA was used to measure the levels of AD, IL-Iβ,IL-6, IL-8, TNF-α, MCP-1 and MMP-9 in the synovial fluids of RA and osteroarthritis (OA), the levels of these cytokines were tested after the synovial fibroblasts (SFLs) were stimulated with AD. Doublelabeling immunohistochemistry was used to analyze the expression of AD in RA synovium. Cytokines were measured by ELISA after SFLs were stimulated with AD. The expression of RANKL was detected by real-time PCR after MH7A were treated with AD and IL-6 ANOVA, Student's t-test, Mann-Whitney U-tese, Spearman's-test were used for statistical analysis. Results High levels of AD in RA synovial fluids were correlated with IL-6 levels. Double labeling immunohistochemistry showed that AD was localized in fibroblasts. MCP-1 and IL-6 were dramatically increased in human synovial fibroblasts following incubation with recombinant AD for 24 h. RANKL mRNA was significantly increased in MH7A after treated with AD and IL-6. Conclusion High levels of AD in the inflammatory joints of RA are likely to contribute to the high expression of IL-6, MCP-1 and RANKL, which may play an important role in the chronic inflammation, osteoclasts activation and bone erosion in RA.

Objective To study the effect of methylprednisolone on serum interleukin-1(IL-1),8,10 during immature myocardial ischemia reperfusion injury.Methods A total of 70 rats were randomly divided into 3 groups:control group(n=10),model group(n=30)and drug group(n=30).And the later 2 groups were observed at 0.5 h after ischemia and 2,4,8,12,24 h after reperfusion with 5 rats in each group and the control group were observed only at 12 h.The rat model of myocardial ischemia reperfusion injury(MIRI)was established.Morphology changes were observed and the levels of serum IL-1,8,10 were examined.Results Compared with model group,inflammatory response in drug group weaken clearly,IL-1,8 were lower(Pa

A bacillus strain was isolated from the dejecta of the Giant Panda " Xiangxiang" returned to wild, afforded by China Giant Panda Protection and Research Center, Wolong Sichuan. By primary research, the strain was identified as Serratia and called as Serratia JF-1116. The 16S rDNA gene of the bacteria strain, was amplified , cloned and sequenced. BLAST of the sequence in GenBank indicated that the species with close similarity to JF-1116 were from genus Enterobacter only. The phylogenetic tree was producted from 16S rDNA of JF-1116 and other 18 Enterobacter species with the high similarity. JF-1116 was clustered with 3 strains of Serratia.

Objective: To compare tea polysaccharides(TPS) characteristics and their role in scavenging free radicals and reducing blood glucose(BG) in diabetic mice(DM). Methods: TPS was extracted from green,Oolong and black tea which were made from the same fresh leaves from Hubei,Fujian and Yunnan. Then the recovery rate of TPS, contents of neutral sugar, uronic acid and protein were analysed, and scavenging rate of -2Oand 稯H in vitro and hypoglycemic effect were also determined. Results: 1. The yield and contents of neutral sugar, uronic acid and protein of green tea TPS were the highest, and those of black tea TPS were the lowest. Oolong tea TPS acted the best in scavenging-2O and 稯H . 2. The hypoglycemic effect of TPS from Hubei tea was the best . The effect of TPS extracted from semi-fermented Oolong tea and fermented black tea was better than that of non-fermented green tea. 3. There were obvious differences in yield, free radical scavenging rate and effect of reducing BG among TPS extracted from tea in different regions. TPS extracted from Fujian tea had the best effect in reducing BG,but that from Yunnan tea had not. Conclusion: There was remarkable effect of region and process on physico-chemical characteristics,effect of scavenging radical and reducing blood sugar TSP.

Objective To detect mycoplasma pneumoniae antibody in children having the upper respiratory tract infection.And then investigate mycoplasma pneumoniae infection status of different season different age children.Methods In 5 403 cases of suspected pneumonia mycoplasma infection of 0 to 14 years old children using the method of passive particle agglutination determination of mycoplasma pneumoniae antibody,and analysis of the statistical results.Results The positive rate was 67.8％ in the groups of children.The rates of infection was biggest during 2 to 3 years old children and 3-4 years old children,14.9％ and 18.4％,respectively.In addition,we found that the highest rate of mycoplasma pneumoniae infection arised from October to January every year of the following year.Conclusion The infection of mycoplasma pneumoniae is on the rise,and children aged 0 to 6 years old are the main population.

Objective To investigate the mechanism of klotho reversing the resistance of breast cancer to paclitaxel in MCF-7/PTX cells.Methods The Klotho expression in MCF-7 and MCF-7/PTX cells was detected by Western blot.The effects of Klotho on paclitaxel resistance in MCF-7/PTX cells was measured by MTT assay.The effects of Klotho and 3-methyladenine (3-MA) on proliferation and expression of Beclin1 in MCF-7/PTX cells were detected by MTT and Western blot assay,respectively.Results The expression of Klotho in MCF-7/PTX cells was decreased compared with MCF-7 cells.Klotho could sensitize MCF-7/PTX cells to paclitaxel.The expression of Beclin1 in MCF-7/PTX cells was higher than that in MCF-7 cells.Klotho and 3-MA could decrease the expression of Beclin1 in MCF-7/PTX cells,and the effects of Klotho on paclitaxel resistance in MCF-7/PTX cells was similar to that of 3-MA.Conclusion Paclitaxel resistance in breast cancer cells is related to expression of the Klotho which can reverse the resistance of breast cancer to paclitaxel by inhibiting autophagy.

Objective To investigate the inhibitory effect of metformin on high glucose-induced lipolysis and further elucidate the underlying mechanisms,for better understanding of metformin.Methods Adipocytes were isolated from epididymal fat pads of Sprague-Dawley rats.After isolation and digestion,packed adipocytes were divided into four groups according to the glucose and metformin in the media,control A group containing 5 mmol · L-1 glucose,control B group containing 25 mmol · L-1 glucose,test A group containing 500 μmol · L-1 metformin with 5 mmol · L-1 glucose and test B group containing 25 mmol · L-1 glucose with 500 μmol · L-1 metformin or at the concentrations as planned.After incubation,glycerol accumulated or released into the media in 30 minutes was determined by the use of a colorimetric assay and served as an index of lipolysis.The expressions of phosphorylated and total perilipin as well as adipose triglyceride lipase (ATGL) were examined by Western Blot.Adipose lipases activity was assayed by using an enzymatic method.Results The glycerol release in the control B and test B groups were (1.61 ± 0.08) and (0.50 ± 0.06) μmol · mL-1 packed cell volume,respectively,suggesting that metformin significantly inhibited the lipolytic action of high glucose (P ＜0.001).Such an inhibitory effect lasted from 16 h to 24 h after incubation.The adipose lipases activity in the control B and test B groups were (344.28 ± 65.98) and (200.44 ± 64.25)μmol glycerol · mg protein-1 · h-1,respectively,and thus metformin caused a 41.78％ (P ＜0.05) reduction of adipose lipases activity elevated by high glucose.Compared with control B group,metformin in test B group significantly attenuated the phosphorylation of perilipin induced by high glucose (P ＜ 0.01).However,the protein amount of total ATGL was not changed in control B group by high glucose or in test group (A and B) by metformin compared with that of control A group (all P ＞ 0.05).Conclusion Metformin inhibits high glucose-induced lipolysis through eliminating perilipin phosphorylation and suppressing lipolytic lipases activity.We suggest that such antilipolytic effect of metformin in hyperglycemia could be a molecular basis by which metformin reduces the release of free fatty acids from adipose tissue to bloodstream and thus ameliorates insulin resistance in diabetes.