Objective:To explore the selection of strategies for early reperfusion therapy and its impact on prognosis in patients with acute ST segment elevation myocardial infarction (STEMI).Methods:The treatment data and 3-year follow-up results of acute myocardial infarction (AMI) patients in 49 hospitals in Hebei Province from January to December 2016 were collected. Patients with STEMI who received either intravenous thrombolytic therapy (ITT) or primary percutaneous coronary intervention (PPCI) within 12 hours of onset were enrolled. Baseline data, the time from the first diagnosis to the start of reperfusion (FMC2N for ITT patients and FMC2B for PPCI patients), vascular recanalization rate, in-hospital mortality, 1-year mortality, and 3-year mortality were compared between ITT and PPCI groups. The efficacy and prognosis of ITT and PPCI at different starting time of reperfusion (FMC2N≤30 minutes, FMC2N > 30 minutes, FMC2B≤120 minutes, FMC2B > 120 minutes) were analyzed.Results:A total of 1 371 STEMI patients treated with ITT or PPCI were selected, including 300 patients in the ITT group and 1 071 patients in the PPCI group. 1 055 patients were actually followed up (205 patients in the ITT group and 850 patients in the PPCI group), with a rate of 79.4%. There were no significant differences in age, gender, and previous history between the two groups. The time from the first diagnosis to the start of reperfusion in the ITT group was shorter than that in the PPCI group [minutes: 63 (38, 95) vs. 95 (60, 150), U = -9.286, P = 0.000], but was significantly longer than the guideline standard. Compared with the ITT group, the vascular recanalization rate in the PPCI group was higher [95.5% (1 023/1 071) vs. 88.3% (265/300), P < 0.01], and in-hospital mortality was lower [2.1% (22/1 071) vs. 6.7% (20/300), P < 0.01], but there were no significant differences in the 1-year mortality and 3-year mortality [5.3% (45/850) vs. 4.4% (9/205), 9.5% (81/850) vs. 9.3% (19/205), both P > 0.05]. Between ITT group and PPCI group with different reperfusion starting time, the FMC2N > 30 minutes group had the lowest vascular recanalization rate and the highest in-hospital mortality. Pairwise comparison showed that the vascular recanalization rate of the FMC2B ≤ 120 minutes group and the FMC2B > 120 minutes group were significantly higher than those of the FMC2N > 30 minutes group [95.5% (654/685), 95.6% (369/386) vs. 88.0% (220/250), both P < 0.008], the in-hospital mortality was significantly lower than that of the FMC2N > 30 minutes group [2.0% (14/685), 2.1% (8/386) vs. 7.6% (19/250), both P < 0.008]. There was no significant difference in 1-year mortality (χ 2 = 2.507, P = 0.443) and 3-year mortality (χ 2 = 2.204, P = 0.522) among the four groups. Conclusions:For STEMI patients within 12 hours of onset, reperfusion therapy should be performed as soon as possible. PPCI showed higher infarct related artery opening rate and lower in-hospital mortality compared with ITT, and had no effect on 1-year and 3-year mortality.

Objective To explore the action mechanism of tauroursodeoxycholic acid(TUDCA)promoting intracellular clearance of Burkholderia pseudomallei(B.pseudomallei)in RAW264.7 macrophages.Methods After TUDCA of different concentrations were used to treat RAW264.7 cells pre-infected with B.pseudomallei for 8 h or not,flow cytometry was applied to detect the apoptosis of the infected and control cells.In addition,another endoplasmic reticulum stress(ERS)inhibitor 4-PBA was used to detect the apoptosis and proliferation of host cells after B.pseudomallei infection with Annexin-V/PI double staining and MTT cell proliferation assay.Furthermore,after transfected with CHOP siRNA,Western blotting and flow cytometry were employed to detect the effect of TUDCA on the expression levels of Caspase-3 and Caspase-12 and the changes in apoptotic rate after B.pseudomallei infection,respectively.Finally,the effect of TUDCA on intracellular multiplication of infected RAW264.7 cells were observed to estimate the CFU value in the presence and absence of CHOP siRNA.Results Under different concentrations of TUDCA,100 or 200 μmol/L TUDCA significantly reduced B.pseudomallei-induced apoptosis in RAW264.7 cells(P＜0.05).Meanwhile,both TUDCA and 4-PBA treatment could decrease the apoptosis induced by B.pseudomallei infection by ERS(P＜0.05).Further,the expression levels of Caspase-3 and Caspase-12 were obviously increased after B.pseudomallei infection compared with uninfected groups,but their expression levels in the siCHOP group was significantly lower than that in the siC group.Besides,flow cytometry also showed that TUDCA could reduce apoptosis induced by B.pseudomallei infection(P＜0.05),but no significant effect of TUDCA on apoptosis was observed under CHOP knockdown.Finally,intracellular CFU assay indicated that TUDCA treatment promoted the host cell clearance of B.pseudomallei(P＜0.05),but no such effect was observed in siCHOP group.Conclusion In B.pseudomallei infected RAW264.7 cells,TUDCA promotes the intracellular clearance of the bacteria by inhibiting ERS-induced apoptosis.

Objective To express recombinant Burkholderia pseudomallei(B.pseudomallei)type Ⅲ secretion system BipD protein,prepare its polyclonal antibodies and verify their immunological traits.Methods The recombinant pET-28a-BipD plasmid was generated,and the pET-28a-BipD-carried E.coli BL21(DE3)bacteria were induced with isopropyl-β-d-thiogalactoside(IPTG)to express recombinant BipD(rBipD)protein.The rBipD was obtained by affinity chromatography using His Trap column,then mixed with Fredrick's adjuvant to immunize BALB/c mice by intraperitoneal injection in order to obtain anti-rBipD polyclonal antibodies.The immunoreactivity of rBipD was detected by Western blot assay using rabbit anti-melioidosis serum and the serum from melioidosis patients.The immunogenicity of rBipD was evaluated using Western blotting and immunofluorescence staining.Finally,rBipD was used to establish an indirect ELISA to detect serum antibodies of clinical melioidosis patients.Results The recombinant plasmid pET-28a-BipD was successfully constructed and transformed into E.coli BL21(DE3)to induce rBipD expression with IPTG treatment.The obtained rBipD had a relative molecular weight of 36×103 and a purity of 95.4％,and had good immunogenicity and immunoreactivity.It could induce the production of specific antibodies after immunizing mice,and mouse polyclonal antibodies against rBipD were prepared with the titer of 1∶512 000.rBipD of 5.0 μg/mL produced specific immune response with the serum of melioidosis patients,but had no specific reaction with the serum of tuberculosis patients,with statistical difference(P＜0.01).Conclusion rBipD with immunological activity is successfully prepared and purified,and its polyclonal antibodies are also developed,which provide a good tool for clinical immunological diagnosis and study of immune mechanism of B.pseudomallei infection.

Objective To analyze the mechanism that Hcp1 protein in type Ⅵ secretion system of Burkholderia pseudomallei(B.pseudomallei)mediates the formation of multinucleated giant cells(MNGCs)when host cells are infected by the bacterium.Methods The mutant strain(BPC006 Δhcp1)and complementation strain(BPC006 Δhcp1::hcp1)were constructed by homologous recombination and plasmid complement technology,respectively.After RAW264.7 cells were infected with B.pseudomallei,the localization of Hcp1 in host cells was analyzed by immunofluorescence staining.The localization was further verified by cytoplasmic-membrane isolation in 293T cells after transfecting pCDNA4.1-Hcp1.The biological significance and effect of Hcp1 were explored by the anti-Hcp1 polyclonal antibody blocking and the formation of MNGC was detected by Giemsa staining.Results Western blotting showed that BPC006 Δhcp1 could not express Hcp1,while BPC006 Δhcp1::hcp1 restored Hcp1 expression.The above results proved that the mutant and complement strains were successfully constructed.Both cellular immunofluorescence co-localization and cytoplasmic-membrane isolation experiments showed that Hcp1 localized to host cell membranes.Last but not least,compared with the control group,anti-Hcp1 polyclonal antibodies inhibited the formation of MNGC(P＜0.01).Conclusion Hcp1 protein in type Ⅵ secretion system of B.pseudomallei is able to translocate to the RAW264.7 cell membranes and plays an important role in the formation of MNGCs.