Protein Z (PZ) is a vitamin K-dependent protein. As a cofactor for the protein Z-dependent protease inhibitor (ZPI), it inhibits coagulation factor X under the existence of phospholipid and calcium ion, and increases the ZPI activity by nearly 1000-fold, thus it plays a role in the process of thrombosis. ZPI inhibits coagulation factor Ⅺ a alone. ZPI activity is also consumed in the process of inhabiting factor Ⅹa and Ⅺa. This article reviews the biological characteristics of PZ and ZPI and their association with stroke.

Objective To construct prokaryotic recombinant expression plasmid carrying Pneumocystis carinii Mr 55 000 antigen(p55) gene fragment and express the recombinant protein. Methods P. carinii pneumonia(PcP) rat models were established by subcutaneous injection of dexamethasone for 14 weeks. Total RNA was extracted from lung of P. carinii rat and p55 antigen gene fragment was cloned by RT-PCR,which was identified by sequencing. The 690 bp fragment was cloned to pGEX-4T-1,the recombinant plasmid was screened and identified by restriction analysis and PCR. The recombinant plasmid was finally induced with IPTG to express a new fusion protein,and the products were analyzed by SDS-PAGE and Western blot. Results A fragment of 690 bp was obtained by RT-PCR. The recombinant pGEX-4T-1/690 was constructed. SDS-PAGE revealed that the molecular weight of the recombinant protein was approximately Mr 62 000,the maximum amount of the fusion protein produced was 11.6% of the total protein. The recombinant protein can be recognized by GST antibody and by the sera from P. carinii infected rats using Western blotting. Conclusion Prokaryotic expression plasmid pGEX-4T-1/690 has been constructed and the recombinant fusion protein shows antigenicity.

Objective To evaluate immunogenicity of the recombinant protein GST-p55/570 of Pneumocystis carinii.Methods The fusion protein GST-p55/570 was expressed from the prokaryotic expression plasmid pGEX-570,and purified by using glutathione-agarose.The expressed product was analyzed by SDS-PAGE.Thirty-three mice were randomly divided into three groups,immunized with GST-p55/570,GST and PBS,respectively.Each group was immunized for four times at 2 week intervals.At the 7th day after final immunization,spleen was removed to obtain single cell suspension.Proliferation ability of lymphocytes was determined by MTT.Serum samples were collected at pre-immunizaton and two weeks after each immunization.Antibody level in sera of mice was determined by ELISA.The immune response to the recombinant GST-p55/570 recognized by sera of immunized mice was examined by Western blotting.Results The expressed fusion protein GST-p55/570 showed a Mr 47 000.Compared with GST group(1.134 5?0.073 5) or PBS group(1.124 8?0.041 6),a higher stimulation index(2.063 0?0.160 2) was revealed in GST-p55/570-immunized mice(P

Objective To investigate the effects of methylprednisolone on neurocyte apoptosis in rats with severe acute pancreatitis(SAP).Methods Thirty-six SD rats were divided into sham operation group,SAP group and methylprednisolone group(12 rats in each group).SAP model was constructed by injecting 5%sodium taurodeoxycholate into biliary-pancreatic duct.Serum amylase,interleukin-6(IL-6),tumor necrosis factor α (TNF-α),volume of aseites and histopathological changes of pancreas were determined.The mRNA expressions of Bcl-2 and Bax in brain tissue were analyzed by RT-PCR.and neuroeyte apoptosis was detected by TUNEL method.Results The levels of serum IL-6 and TNF-α were significantly increased:the expression of Bcl-2 mRNA in brain tissue was down-regulated;the expression of Bax mRNA was up-regulated;the Bcl-2/Bax ratio Was decreased:the apoptosis of the neurocytes was increased in SAP group.Compared with SAP group,the levels of serum IL-6 and TNF-α were significantly decreased;the expression of Bcl-2 mRNA was unchanged but the expression of Bax mRNA was down-regulated in brain tissue,so the Bcl-2/Bax ratio was elevated significantly;the rate of the ueurocyte apoptosis in brain tissue were reduced in methylprednisolone group.Conclusions The apoptosis of neurocytes in brain tissue may be one of the factors causing pancreatic encephalopathy.Methylprednisoione can inhibit the release of IL-6 and TNF-α.improve the balance of Bcl-2 and Bax expression and decrease the apoptosis of neurocytes in brain tissue.

To construct the eukaryotic expression plasmid containing the p55 gene fragment of Pneumocystis and to investigate the efficient expression in COS-7 cells, the gene fragment conaining the whole length of p55 gene was used as template to amplify this fragment with PCR and the amplified fragment was then cloned to vector pGEM-T. After enzyme digestion, p55 gene was cloned to the eukaryotic expression vector pcDNA3.1(+) to construct the plasmid pcDNA3.1(+)-582. This plasmid was then transfected to the eukaryotic expression cells COS-7 and PCR and SDS-PAGE assays were used to confirm the presence of target protein in these cells. In these ways, the eukaryotic expression vector for the p55 gene of Pneumocystis of rats was successfully constructed and expressed in COS-7 cells, thus providing the basis for further studies on the nucleic acid vaccine.

Objective To observe the effects of GABA on proliferation, cell cycle and expression of MMP-2, MMP-9 of pancreatic cancer cell line SW1990. Methods The effects of different concentration of GABA (0 ～ 320 μmol/L) on proliferation and cell cycle of pancreatic cancer cell line SW1990 was investigated by MTT assay and flow cytometry analysis, respectively. Expressions of MMP-2 and MMP-9 proteins were evaluated by Western blot analysis. Results GABA could promote the proliferation of SW1990 cells and influence the distribution of cell cycle, which made less cells of G0/G1 phase and more cells of S and G2/M phase. The value of A570 after GABA pretreatment at a dose of 320 μmol/L was 1. 11 ± 0.03, which was significantly higher than that in the control group (0. 56 ± 0.01, P < 0. 01 ), the cells of G0/G1 phase was (46.18 ± 1.12 )% ,which was significantly lower than (87.29 ± 1.34)% in the control group (P < 0. 01 ) ;the expressions of MMP-2 mRNA, MMP-9 mRNA and their proteins were 8.6, 6.8, 10.5, 8.4, respectively, which were significantly higher than those in the groups of the doses of 0 ～ 40 μmol/L ( P < 0. 05 ). Conclusions GABA could influence the proliferation and expression of MMP of SW1990 cells.

Objective To investigate the association of Mycoplasma pneumoniae(MP) infection with disease activity of ankylosing spondylitis. Methods A total of 158 subjects in our hospital were enrolled in this study, including patients with ankylosing spondylitis(AS, n=66), rheumatoid arthritis (RA, n=31),osteoarthritis(OA, n=25) and normal controls(NC, n=36). MP infection was defined as anti-MP IgM antibody positive. Anti-MP IgM antibodies were determined by a mycoplasma pneumoniae(Mac strain)membrane-based agglutination test. AS patients were divided into two groups: MP infection group and non-MP infection group. T-test was used for statistical analysis of age, blood white cells, ESR, CRP, immunoglobulin, BASDAI index, global assessment on VAS scale, Schober test and chest expansion reflecting spinal mobility.χ2-test was used to compare the positive rate of MP infection in different groups. Gender difference and prevalence of clinical infection in past four weeks between MP infection and MP-free group in AS patients was also compared. Ridit analysis was used to analyze the association of MP infection with degree of sacroiliac damage on CT. Results The prevalence of MP infection in AS (52%, 34/66) was much higher than that in rheumatoid arthritis (RA, 6%, P＜0.01 ), osteoarthritis(OA, 4%, P＜0.01 ) and normal controls (NC, 11%, P＜0.01) . Compared with the non-MP infection group, the MP infection group had more active disease in term of BASDAI(4.0±1.1 vs 3.0±1.9, P=0.017), ESR[(44±32) mm/1h vs (28±23) mm/1h, P=0.029], CRP [(40±38) mg/L vs (22±21) mg/L, P=0.025] serum total IgG level [(18±3) g/L vs (16±5) g/L, P=0.027],but not in serum total IgA and IgM. Regarding to the sacroiliac joint and spinal mobility, MP infection group did not exhibit any association with the sacroiliac grading on CT, Schober test and expansion. In AS patients with MP infection, only 44.1%(15/34) was complicated by clinical manifestations of upper respiratory tract in the past 4 weeks. However, a higher prevalence of MP infection was found in AS patients with clinical manifestation of upper respiratory tract, compared with those with negative clinical manifestation(71% vs 42%,P=0.027). Conclusion Mycoplasma pneumoniae is the most common reported pathogen in ankylosing spondylitis and relates to the disease activity of AS. MP infection is probably a principal triggering factor in the pathogenesis of AS.

Objective To evaluate the efficacy of pegylated interferon ( PegIFN ) α-2a plus ribavirin ( RBV) therapy for chronic hepatitis C ( CHC) in non-responders, and to investigate the related influencing factors.Methods A prospective, open, multicenter and randomized study was conducted.A total of 81 CHC non-responders were recruited from 10 clinical centers during February 2009 to November 2011.Patients were randomly assigned into two groups:group A (n=37) was given PegIFNα-2a plus RBV treatment for 72 weeks and group B (n=44) was given PegIFNα-2a plus RBV treatment for 96 weeks.Both groups were followed up for 24 weeks after treatment.Virological responses in two groups were observed, and treatment efficacies among patients with different genotypes, and among those with different previous treatment were compared.SAS software was used for statistical analysis.Results Fifty-two patients ( 28 from group A and 24 from group B) completed the study in total.The rates of rapid virological response ( RVR) , complete early virological response ( cEVR ) , end of treatment viral response ( ETVR ) and sustained virological response (SVR) in group A were 25.0% (7/28), 60.7% (17/28), 67.9%(19/28) and 60.7%(17/28), respectively; while those in the group B were 41.7% (10/24), 70.8%(17/24), 70.8%(17/24) and 70.8% (17/24), respectively; and there were no significant differences between two groups (P>0.05).SVR was observed in 82.9%(29/35) of patients with CC genotype of IL-28B, which was higher than that in patients with other genotypes ( 3/13 ) , and the difference was of statistical significance (P<0.01).There was no significant difference in viral responses between patients previously treated with IFN plus RBV and those treated by IFN only (P>0.05).The rates of RVR, cEVR, ETVR and SVR in patients who were previously treated with IFN were 36.4%(12/33), 81.8%(27/33), 81.8%(27/33) and 75.8%(25/33), and the rates of cEVR, ETVR and SVR were higher than those in patients who were previously treated with PegIFN (P<0.05), but no significant difference was observed in RVR (P>0.05).Adverse events occurred in 38 patients (46.9%), but no severe ones were observed. Conclusion The efficacy of PegIFNα-2a plus RBV therapy for CHC in non-responders is satisfactory, which may influenced by IL-28B genotypes and previous treatment.

Objective To investigate the epidemiological and clinical features of brucellosis in Foshan.Methods The epidemiological history,clinical manifestations,laboratory tests,treatment and outcomes of 41 patients with brucellosis from 2013 to 2016 in the First People's Hospital of Foshan were retrospectively analyzed.Results Brucellosis onsets occurred mainly from February to June [85.4％ (35/41)],and 58.5％ (24/41) of them had positive epidemic history.Irregular fever,rachialgia/arthralgia,fatigue and hepatosplenomegaly occurred in 29 (70.7％),20 (48.8％),19 (46.3％) and 16 (39.0％) of the patients,respectively.The C-reactive protein (hs-CRP) of patients infected with Brucella only was lower than that in patients infected also with other bacteria (26.72 vs 50.87 mg/L,Z =-2.300,P ＜ 0.05),but no significant difference of white blood cell counts (5.77 × 109/L vs 5.83 × 109/L),neutrophil (3.50 × 109/L vs 3.84 × 109/L) and procalcitonin (PCT,0.10 vs 0.14 μg/L) between the two groups were observed.The patients with positive epidemic history had lower white blood cell,neutrophil and monocyte counts than those who did not had epidemic history (4.73 × 109/Lvs 7.28 × 109/L,2.73 × 109/L vs 4.79 × 109/L,and 0.36 × 109/L vs 0.64 × 109/L;F =9.486,10.130,9.785,P ＜ 0.05).And no significant difference of lymphocyte counts,hs-CRP and PCT between the two groups were observed (1.57 × 109/L vs 1.73 × 109/L,29.30 vs 35.76 mg/L,and 0.15 vs 0.09 μg/L;P ＞ 0.05).All the cases were infected by Brucella melitensis,and 33 of them were sensitive to general antibiotics in vitro.There were 40 cases discharged after treatment,and 34 cases still needed to increase antibiotic treatment courses.Most patients had good outcomes.Conclusions In Foshan,patients with irregular fever and rachialgia or arthralgia,and no significantly increased inflammation index,should be aware of brucellosis.We should strengthen the screening of brucellosis in Foshan.

Objective To investigate the prevalence and risk factors of restless leg syndrome (RLS) in hemodialysis patients after kidney transplantation failure.Methods Patients of hemodialysis after kidney transplantation failure were investigated by face-to-face interviews,from March to July,2015,at four dialysis units in Beijing.RLS was diagnosed according to the International RLS Study Group (IRLSSG) criteria.The severity of RLS was assessed using International RLS Rating scale.Besides,three validated sleep disorder questionnaires (Hamilton anxiety and depression scale,Epworth sleepiness scale and Pittsburgh sleep quality index) were completed by the patients at the same time.Results Ninety-four hemodialysis patients after kidney transplantation failure were enrolled;46 patients (48.94％) met the diagnosis of RLS,the average age was 53.44±l 1.89 years,and the median time of RLS onset after kidney transplantation failure was 46 months.The International RLS Rating scale scores of the patients were 17.26±7.81;76.0％ patients were above moderate.As compared with the non-RLS patients,patients with RLS used more erythropoietin (44/48 vs.46/46),less ferrila (30/48 vs.19/46),and few hypnotic medicine (10/48 vs.3/46),with significant differences (P＜0.05).The serum ion,serum ferritin and serum Vitamin B12 of patients with RLS were significantly lower as compared with non-RLS patients (P＜0.05);and poorer sleep quality and higher depression scale scores in the patients with RLS were noted as compared with those in the non-RLS patients (P＜0.05).Conclusion The prevalence of RLS in hemodialysis patients after kidney transplantation failure is high,low iron protein content,low serum iron content and low vitamin B12 levels may be risk factors for RLS.