Construction of a novel Shigella live-vector strain co-expressing CS3 and LTB/STm of enterotoxigenic E.coli

AIM: To construct and evaluate a polyvalent recombinant vaccine strain Shigella flexneri2a T32 against enterotoxigenic E.coli/(ETEC). METHODS: By using a host-plasmid balanced lethal system based on asd gene, a polyvalent recombinant strain was constructed to highly express CS3 and regularly express fusion enterotoxin of LIB subunit and mutant ST (LTB/STm) in a vaccine strain Shigella flexneri 2a T32 with specific deletion of asd gene. Fimbria CS3 was observed by immunofluorescence and electron microscopy assay. The security of LTB/STm was examined by ileal loop assay and suckling mouse assay. To evaluate this new candidate vaccine, it was compared with a previous vaccine strain in plasmid and protein level, growth assay and immunogenicity in Balb/c mice. RESULTS: The newly constructed vaccine expressed CS3 and grew better than the previously constructed vaccine except for the lower expression of LTB/STm. Serum IgG and mucosal IgA against CS3, LTB, ST, and host lipopolysaccharide (LPS) were produced after immunization of Balb/c mice by oral route with the new strain. The titers were not significantly different from the Balb/c mice with the previous strain. CONCLUSION: This novel candidate diarrheal vaccine can effectively induce serum and mucosal antibody responses against ETEC and Shigella.

Endostatin promotes the anabolic program of rabbit chondrocyte

Endostatin is a natural occurred angiogenesis inhibitor derived from collagenXVIII. So far its function during the angiogenesis process of bone formation and arthropathy has not been well studied yet. The present study addresses the function of endostatin in rabbit articular chondrocytes (RAC). We found that endostatin can promote RAC adhesion and spreading as well as its proliferation. In monolayer cultured RAC, CollagenII, TIMP1 and collagenXVIII transcription were up regulated by endostatin while collagenI and MMP9 were down regulated. Moreover collagenXVIII and endostatin antigens are present at synovial fluid. These findings indicate new function of endostatin as a homeostatic factor in cartilage metabolism.

Identification of alkA gene related to virulence of Shigella flexneri 2a by mutational analysis

AIM: In vivo induced genes are thought to play an important role during infection of host. ,AlkA was identified as an in vivo-induced gene by in vivo expression technology (IVET),but its virulence in Shigella flexneri was not reported. The purpose of this study was to identify the role of alkA gene in the pathogenesis of S. flexneriMETHODS: PCR was used to amplify alkA gene of S. flexneri 2a and fragment 028pKm. The fragment was then transformed into 2457T05 strain, a S flexneri 2a strain containing Red recombination system, which was constructed with a recombinant suicide plasmid pXLkd46. By in vivo homologous recombination, alkA mutants were obtained and verified by PCR and sequencing. Intracellular survival assay and virulence assay were used to test the intracellular survival ability in HeLa cell model and the virulence in mice lung infection model respectively.RESULTS: Deletion mutant of S. flexneri 2a alkA was successfully constructed by λ Red recombination system.The mutant exhibited significant survival defects and much significant virulence defects in mice infection assay.CONCLUSION: AlkA gene plays an important role in the infection of epithelial cells and is a virulent gene of Shigella spp.