Two highly conserved and with abundant quantity of lipoproteins in outer membrane of pathogenic species,but not in saprophytic species of leptospira ,LipL32 and LipL21,were selected to construct the fusion gene DNA vaccine pVAX1/LipL21-LipL32,and its ability to induce immune responses in BALB/c mice inoculated with this recombinant DNA vaccine was investigated in the present study.Expression of the fusion-protein LiPL21-LiPL32 was demonstrated in HEK293 cells following transfection with the fusion gene DNA vaccine and the immune responses induced after intramuscular inoculation with this DNA vaccine in BALB/c mice was then evaluated by microscopic agglutination test (MAT),meanwhile the ELISA assay was used to detect the cytokines induced.It was demonstrated that significant level of specific antibodies agglutinating antigens of Leptospira interrogans could be detected by MAT after DNA vaccine inoculation.The production of cytokines IL-10 and TNF-β in mice inoculated with DNA vaccine pVAX1/LipL21-LipL32 was significantly increased in comparison with that of the group inoculated with pVAX1 alone.These results indicate that the recombinant DNA vaccine pVAX1/LipL21-LipL32 may be of potential value to design and develop new generation of vaccines against leptospirosis.

Hemolysins of Leptospira interragans have been shown to be the virulence factor in the pathogenesis of leptospirosis and 10 potential hemolysin genes were charecterized by genomic annotation of L.interrogans serovar.Lai strain 56601. In the present study, the LA0202 gene supposed to encode one of the new potential hemolysin was cloned and the protein encoded was purified. The purified protein was shown to have highly hemolytic activity as demonstrated on the sheep blood agar plate. It was also confirmed that the LA0202 protein-mediated hemolysis on sheep erythrocytes was osmotically protected by PEG6000. Meanwhile, this protein could induce pore formation on sheep erythrocytes and cause damages on the membrane of human L-02 liver cells. In addition, it could induce apoptosis of human L-02 liver cells after treatment of cells with this protein for 24 hours. It is evident that LA0202 protein acting as a pore-formong hemolysin can induce cytotoxic damage on mammalian cells.

Objective:　To compare the dynamic changes of int erleukin-1 (IL-1), interleukin-6 (IL-6), and tumor necrosis factor (TNF) in intermingled skin graft with those in other types of skin grafts in rats. 　　Methods:　A 10%-15% third-degree burn was created in 180 Spreg ue-Dawley (SD) rats. After removing the scar, skin grafts were performed on the open wounds immediately with autoskin (aus, n=54), allosk in (als, n=54) and intermingled skin (n=36). That is to say, in the intermingled skin graft, a big piece of alloskin (mals) was grafted first, and 3 days later, small pieces of autoskin (maus) wer e embedded in the alloskin. The rest 36 rats were taken as the controls. And the biological activities of IL-1, IL-6 and TNF in graft sheets in each group wer e detected after skin graft. 　　Results:　The levels of IL-1, IL-6 and TNF in the aus group de creased steadily after their initial elevations, whereas in the als group they i ncreased significantly and kept on the peak level in the later phases. In the in termingled group, there appeared a lowest IL-1 level in the mals and a highest one in the maus simultaneously at 7 (4) days (The number out of parenthesis is t he days after transplanting with alloskin sheets, and the number in parenthesis is the days after embedding autoskin sheets in the intermingled skin graft. Simi larly hereinafter.) after skin graft (P<0.01), and the high level in the maus abruptly decreased at 14 (11) days after skin graft. At exactly the same phase on day 7 (4), a prominent peaked IL-6 in the mals occurr ed. In the later phases, the levels of TNF remained relatively low both in the m als and in the maus. From day 7 (4) on, each cytokine fluctuation in the mals sy nchronized with that in the maus. The longer the post transplantation period las ted, the more the positive cytokine correlated between the mals and the maus. 　　Conclusions:　The low levels of IL-1 and TNF may be important f actors to lighten the intensity of local rejection in the intermingled skin graf t. The temporarily peaked IL-6 is both an inducer which induces the production of local IL-1 receptor antagonists and soluble TNF receptors and a signal which indicates a local enhancement of Th2 cells. The mild rejection process and th e synchronized cytokine level during the later phases suggest a possible chimeri sm between the mals and the maus.

Abstract: Objective To express Mycobacterium tuberculosis Rv2626c protein in Escherichia coli (E. coli) and study the antigenicity of the purified recombinant Rv2626c protein. Methods The amino acid sequence of Rv2626c protein from Mycobacterium tuberculosis H37Rv strain (accession number: CCP45424.1) in GenBank was retrieved and converted into the corresponding DNA sequence according to the codon preference of E. coli. This DNA sequence was synthesized and cloned into pET24a(+) plasmid to construct pET24a(+)-Rv2626c recombinant plasmid. This plasmid was transformed into E. coli BL21(DE3) cells, and the expression of Rv2626c protein was induced under various conditions of isopropyl β-D-thiogalactopyranoside (IPTG) concentrations, temperature, and period. The recombinant Rv2626c protein was identified by SDS-PAGE and Western Blot. The recombinant Rv2626c protein was purified by nickel chelate affinity chromatography and used to immunize violet blue rabbits to prepare anti-Rv2626c anti-serum. The specificity and titer of the serum were respectively detected by Western Blot and enzyme-linked immunosorbent assay (ELISA). Results The recombinant plasmid pET24a(+)-Rv2626c was successfully constructed. SDS-PAGE analysis showed that recombinant Rv2626c was expressed in the recombinant plasmid transformed E. coli with IPTG induction, with a molecular weight of about 14 500, and the size was consistent with the expectation. The optimal expression condition for recombinant Rv2626c protein was at 31 ℃ with 1.0 mmol/L IPTG for 6 hours. The target protein was mainly present in a soluble form, which was consistent with the results of Western blot. The hyperimmunized serum with recombinant Rv2626c protein vaccination showed good specificity, with a titer of 1∶ 256 000 detected by ELISA. Conclusions Mycobacterium tuberculosis Rv2626c protein is successfully expressed in E. coli, and the purified protein has good purity and antigenic activity, laying the foundation for further reveals of its biological functions.

【Objective】 To investigate the correlation of monocytes and high-density lipoprotein cholesterol ratio(MHR) and albumin with the severity of coronary artery lesions in patients with unstable angina pectoris. 【Methods】 We enrolled 342 patients with unstable angina pectoris. According to the Gensini score of their coronary angiography results, they were divided into Gensini≤ 20 group, 2040 group. The differences in biochemical indicators between the groups were compared, and the correlation between the different indicators and the Gensini score was analyzed. According to the MHR quartile grouping, there were differences between the comparison groups. LDL-C was divided into subgroups and then subjected to multifactor Logistic regression analysis. 【Results】 MHR differed significantly among low, moderate and high grade lesions (P<0.05). Subgroup analysis showed that in low LDL-C group, Gensini score was positively correlated with MHR(P<0.05), while in high LDL-C group, Gensini score was negatively correlated with albumin(P<0.05). Multivariate Logistic regression analysis showed that the MHR level of patients with high Gensini score was 102.375 times higher than that of patients with low Gensini score(P<0.05). In the group with high LDL-C, the serum albumin level in the group with low Gensini score was 1.431 times that in the group with high Gensini score and 1.218 times that in the group with moderate Gensini score(all P<0.05). 【Conclusion】 In patients with unstable angina pectoris, especially when LDL-C levels are not high, both high MHR and low serum albumin are independent risk factors for the severity of coronary artery disease.