Examine the Correlation between Heat Shock Protein IbpA and Heat Tolerance in Cronobacter sakazakii / 生物医学与环境科学（英文）

We used a proteomic approach to identify IbpA in Cronobacter sakazakii (C. sakazaki), which is related to heat tolerance in this strain. The abundance of IbpA in C. sakazakii strains strongly increased after heat shock. C. sakazakii CMCC 45402 ibpA deletion mutants were successfully constructed. The C. sakazakii CMCC 45402 ΔibpA and wild-type strains could not be distinguished based on colony morphology on LB agar plates or biochemical assays. The growth of the C. sakazakii CMCC 45402 ΔibpA mutant in heat shock conditions was indistinguishable from that of the isogenic wild-type, but showed greater heat resistance than E. coli O157:H7 strain CMCC 44828. This study suggests that the absence of a single ibpA gene has no obvious effect on the phenotype or heat resistance of the strain C. sakazakii CMCC 45402.

Akt-centered amplification loop plays a critical role in vascular endothelial growth factor/stromal cell-derived factor 1-α cross-talk and cardioprotection / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Vascular endothelial growth factor (VEGF) is one of major mediators of angiogenesis and survival factor in some tissue, however, its direct effects on cardiomyocytes remain poorly understood.</p><p><b>METHODS</b>Rat neonatal ventricular myocytes were cultured in vitro. Akt phosphorylation was measured by Western blotting; the expression of stromal cell-derived factor α (SDF-1α)/CXCR4 axis was evaluated by real-time PCR and Western blotting. LY294002 and AMD3100 were used to interfere with the signaling of VEGF and SDF-1α/CXCR4 axis. Cardiac myocytes viability and injury were evaluated by trypan blue staining and lactate dehydrogenase (LDH) release.</p><p><b>RESULTS</b>Treatment of neonatal rat ventricular myocytes with VEGF induced phosphorylation of Akt in a dose and Flk-1 dependent manner. VEGF attenuated H2O2 induced cardiac myocyte death. The phosphoinositol-3-kinase (PI3K) inhibitor, LY294002 and Flk-1 antibody abolished the beneficial effects of VEGF on H2O2 induced cell death. In the mean time SDF-1α-CXCR4 axis was up-regulated by VEGF through PI3K-Akt signaling and contributed to the protective effects of VEGF on H2O2 induced cell death. Interestingly, SDF-1α also promoted production of VEGF in cultured cardiac myocytes and LY294002 reversed the up-regulation of VEGF induced by SDF-1α.</p><p><b>CONCLUSION</b>VEGF has direct protective effects on cardiomyocytes; a crosstalk between VEGF and SDF-1α through PI3K-Akt serves a survival role in cardiomyocytes in vitro.</p>

Nitric oxide induces heat shock protein 72 production and delayed protection against myocardial ischemia in rabbits via activating protein kinase C / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Nitric oxide (NO) is a biologically active molecule which has been reported to protect the heart against ischemia and reperfusion injury in different species. This study aimed to test the hypothesis that nitric oxide may induce the expression of heat shock protein 72 (HSP72) which may protect the heart against ischemia.</p><p><b>METHODS</b>Rabbits were given intravenous saline or S-nitroso-N-acetylpenicillamine (SNAP), a nitric oxide donor, or Zaprinast, an inhibitor of cyclic guanosine monophosphate (GMP)-phosphodiesterase, which may increase myocardial cyclic GMP content. Twenty-four hours later, the rabbits were either sampled to measure HSP72, or induced with a 30-minute coronary occlusion followed by a 120-minute reperfusion, and then the infarct size was measured. Meanwhile, chelerythrine (CHE, an inhibitor of protein kinase C) was given intravenously 5 minutes before SNAP injection and the effect on HSP72 expression and infarct size was determined.</p><p><b>RESULTS</b>Twenty-four hours after pretreatment, immunoblotting showed HSP72 expression increased in the SNAP group compared with control groups, and this was blocked by CHE. Myocardial infarct size in the SNAP group was smaller than that of the control group ((32.4 +/- 5.8)% vs (51.1 +/- 4.7)%, P < 0.05). Pretreated with CHE abolished the infarct size-limiting effect of SNAP ((46.0 +/- 5.1)%). Pretreatment with Zaprinast neither induced HSP72 expression nor reduced infarct size ((55.4 +/- 5.4)%).</p><p><b>CONCLUSION</b>NO induced HSP72 expression and a delayed protection to the heart via the activities of protein kinase C by a cyclic GMP-independent pathway.</p>

Preparation of monoclonal antibody and development of enzyme-linked immunosorbent assay specific for Escherichia coli O157 in foods / 生物医学与环境科学（英文）

<p><b>OBJECTIVE</b>To prepare monoclonal antibodies (MAb) and antisera specific for Escherichia coli (E. coli) O157, and to develop a sandwich enzyme-linked immunosorbent assay (ELISA) to detect E. coli O157 in foods.</p><p><b>METHODS</b>Spleen cells from BALB/c mice immunized with the somatic antigen of E. coli O157:H7 were fused with murine Sp2/0 myeloma cells. The hybridoma cell line specific for E. coli O157 was established after having been subcloned. Antisera specific for E. coli O157 was prepared by intravenous injection into New Zealand rabbits with a stain of E. coli O157:H7. The sandwich ELISA was developed with the polyclonal antibody as the capture antibody and the MAb 3A5 as the detection antibody. The inoculated ground poultry meat and pasteurized milk were tested to confirm efficiency of the method.</p><p><b>RESULTS</b>MAb 3A5 specific for E. coli O157 and O113:H21 belonged to subtype IgM. The ascetic titers of the antibody was 1:1x10(6). No cross-reactivity of the MAb was observed with strains of Salmonella spp, Yersinia enterocolitica, Shigella dysenteriae, etc. The purified polyclonal antibody had a titer of 1:1x10(5) with E. coli O157. The detection limit of this sandwich ELISA was 10(3)-10(4) cfu E. coli O157/mL in pure culture with a high specificity, which was characterized by every non-O157 strain with negative response. With 10h enrichment procedure, E. coli O157:H7 recovered well from inoculated ground poultry meat and pasteurized milk at levels of 0.1 cfu/g and 0.1 cfu/mL.</p><p><b>CONCLUSION</b>MAb 3A5 specific for E. coli O157 and O113:H21 can be produced by immunizing BALB/c mice with a strain of E. coli O157:H7. Then a sandwich ELISA can be developed with the polyclonal antibody as the capture antibody and the MAb 3A5 as the detection antibody. The method is proved to be a sensitive and specific technique to detect low number of E. coli O157 in food.</p>

Application of green fluorescence protein in the detection of E. coli O157:H7 in food / 中华预防医学杂志

<p><b>OBJECTIVE</b>To introduce green fluorescence protein (GFP) into E. coli O157:H7 and improve the detection methods for this bacteria, and to study the kinetics of E. coli O157:H7.</p><p><b>METHODS</b>The plasmid pGFP was transferred into E. coli O157:H7. The characteristic of the new built O157:H7-pGFP strain was evaluated. Some food samples were inoculated with the recombinant strain under certain temperature to imitate different storage circumstances. The contaminated E. coli O157:H7 were counted after certain time.</p><p><b>RESULTS</b>The pGFP was stable in E. coli O157:H7. The E. coli O157:H7-pGFP inoculated in ground poultry meat and pasteurized milk were enriched to 35 000 approximately 200 000 times in 12 h under higher storage temperature (37 degrees C), whereas the quantity decreased slowly under lower temperature (4 degrees C).</p><p><b>CONCLUSION</b>The recombinant strain with the characters of ampicillin resistance and green fluorescence under UV 365 nm was a useful tool in detection methods improvement and bacteria survival studies.</p>