Chang-hong Shi

Fourth Military Medical University, Xi’an, Liaoning, China

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Publications (24)14.83 Total impact

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    ABSTRACT: To investisate the inhibition of Hsp-16.3 on the autophagosomes formation of macrophages. Mouse RAW264.7 macrophages were induced by rapamycin (50 ng/μL) following infection with M.tuberculosis H37Rv strains, thereafter, co-incubated with Hsp16.3 protein (25 μg/mL). The effects of Hsp16.3 protein on the autophagosomes formation was observed with transmission electron microscope. The expression of autophagy-related genes (atg8) for macrophages was detected by Western blotting. It was found that rapamycin-induced autophagy of macrophages infected with M.tuberculosis H37Rv enhanced localization of mycobacteria with autophagosomes. Hsp16.3 protein inhibits autophagosome formation and affects M.tuberculosis survival inside infected macrophages. Furthermore, Hsp16.3 protein significantly increased M.tuberculosis colony forming units (CFU), and decreased the expression of microtubule-associated protein light chain-3 (LC3) expression level (P<0.05). The results showed that Hsp16.3 protein inhibits the formation of autophagosomes by regulating the expression of LC3 protein.
    No preview · Article · Dec 2011 · Xi bao yu fen zi mian yi xue za zhi = Chinese journal of cellular and molecular immunology
  • Xue Gao · Li-Mei Wang · Yin-Lan Bai · Hong Jiang · Yuan Li · Chang-Hong Shi · Hai Zhang · Ying Xue
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    ABSTRACT: Ferric uptake regulator A of Mycobacterium tuberculosis (MTB), which belongs to the Fur superfamily, is situated immediately upstream of katG encoding catalase-peroxidase, a major virulence factor that also activates the pro-drug isoniazid. The feature and role of FurA in oxidative stress contribute to research on the pathogenesis of mycobacteria. In this study, four novel mouse monoclonal antibodies were generated using the prokaryotically expressed FurA protein as immunogen. The furA gene of M. tuberculosis H37Rv was inserted into a bacterial expression vector of pRSET-A and effectively expressed in Escherichia coli BL21(DE3). The expressed fusion protein existed as soluble form in cell lysates and was purified via Ni-NTA purification system. Using the fusion protein to immunize BALB/c mice, four monoclonal antibodies (H9H6, H9E12, H10H6, and H10H8) were produced. As shown by Western blot analysis and cell fluorescence microscopy assay, the four antibodies could recognize the FurA protein, respectively. Then we assessed the effect of iron on the expression of FurA in MTB H37Rv and we concluded that iron does not affect FurA expression. These results suggest that the antibodies against FurA may provide a powerful tool for elucidating FurA biofunctions and regulation mechanism in the pathogenesis of tuberculosis.
    No preview · Article · Aug 2011 · Hybridoma (2005)
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    ABSTRACT: The blood-testis barrier (BTB) plays an important role in male reproductive system. Lots of environmental stimulations can increase the permeability of BTB and then result in antisperm antibody (AsAb) generation, which is a key step in male immune infertility. Here we reported the results of male mice exposed to electromagnetic pulse (EMP) by measuring the expression of tight-junction-associated proteins (ZO-1 and Occludin), vimentin microfilaments, and transforming growth factor-beta (TGF-beta3) as well as AsAb level in serum. Male BALB/c mice were sham exposed or exposed to EMP at two different intensities (200kV/m and 400kV/m) for 200 pulses. The testes were collected at different time points after EMP exposure. Immunofluorescence histocytochemistry, western blotting, laser confocal microscopy and RT-PCR were used in this study. Compared with sham group, the expression of ZO-1 and TGF-beta3 significantly decreased accompanied with unevenly stained vimentin microfilaments and increased serum AsAb levels in EMP-exposed mice. These results suggest a potential BTB injury and immune infertility in male mice exposed to a certain intensity of EMP.
    Full-text · Article · Sep 2010 · Toxicology
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    ABSTRACT: To evaluate the immune responses and resistance against Mycobacterium tuberculosis (MTB) infection in the mice induced by HSP16.3 of MTB and its synthetic peptide. BALB/c mice were immunized subcutaneously 3 times at 2 week interval at the base of tail. The doses of HSP16.3 protein and synthetic peptide were both 50 microg each time. A single dose of BCG (5 x 10(6) CFU/mouse) was used to immunize the mice. The concentrations of specific antibodies in serum obtained at 0, 2, 4, 6, 8 weeks after the first immunization and the titer of serum obtained at 8th week, were analyzed by enzyme linked immunosorbent assay (ELISA). Four weeks after the final immunization, 8 mice from each group were sacrificed and single-cell suspensions of splenocytes were prepared, some of which were used for lymphocyte proliferation by MTT colorimetry with HSP16.3 stimulation, and the remaining cells were used for IFN-gamma level assay by sandwich ELISA. The remaining mice in each group were challenged intravenously with 10(5) colony forming units (CFU) of MTB H(37)Rv and were sacrificed 4 weeks after infection, and the number of bacteria in the spleens and lungs were determined by plating serial dilutions of homogenized tissue on Middlebrook 7H10 agar. The statistical significance of differences among means was assessed by an LSD-t test. The level of specific antibody to HSP16.3 protein and the peptide increased rapidly in the former 4 weeks and moderately in the later weeks. The average antibody-specific titers of 3 experiment groups (HSP16.3 protein + DDA + MPL, synthetic peptide + DDA + MPL and synthetic peptide + IFA) were higher than the BCG group. The indexes of spleen lymphocyte proliferation (SI) of the 3 experiment groups (3.13 +/- 0.18, 3.21 +/- 0.21 and 2.40 +/- 0.15) were significantly higher than the BCG group (1.67 +/- 0.12) and the saline group (1.04 +/- 0.09) respectively. The SI of HSP16.3 protein + DDA + MPL group (3.13 +/- 0.18) and synthetic peptide + DDA + MPL group (3.21 +/- 0.21) were higher than the synthetic peptide + IFA group (2.40 +/- 0.15). The IFN-gamma levels induced among the 3 experiment groups [(182 +/- 6), (194 +/- 9) and (179 +/- 8) mg/L] were lower than the BCG group [(275 +/- 10) mg/L], but higher than the saline group [(71 +/- 3) mg/L]. The IFN-gamma level induced among the 3 experiment groups did not show any marked difference. Although the protection induced by HSP16.3 protein + DDA + MPL, synthetic peptide + DDA + MPL and synthetic peptide + IFA all showed resistance against MTB H(37)Rv infection in the spleens or lungs (the bacterial logarithmic loads of spleen: 6.74 +/- 0.14, 6.60 +/- 0.13 and 6.81 +/- 0.28; the bacterial logarithmic loads of lung: 5.81 +/- 0.21, 5.74 +/- 0.27 and 6.65 +/- 0.32), none of them was better than the conventional BCG (the bacterial logarithmic loads of spleen and lung: 5.95 +/- 0.17 and 5.62 +/- 0.23). Both HSP16.3 and its synthetic peptide can be considered as TB vaccine candidates or effective components in TB vaccines.
    No preview · Article · Aug 2009 · Zhonghua jie he he hu xi za zhi = Zhonghua jiehe he huxi zazhi = Chinese journal of tuberculosis and respiratory diseases
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    ABSTRACT: To study the effect of autophagy in MTB's infection and the expression of its related gene. The formation of autophagy was induced by Rapamycin and observed by the transmission electron microscope. The cleaning role of autophagy to the MTB H37Rv virulent strain after its formation was detected by clone forming unit (CFU). Realtime PCR was used to detect the mRNA of the autophagy related gene was expressed. The RAW264.7 cell could form autophagosome under the induction of the Rapamycin, and it had the determinate cleaning role to the H37Rv strain in the cell after which formed. The mRNA of atg5, atg8 and atg12 which participated the formation of autophagy were expressed more, but the expression of atg7 had no change. Autophagy participated the process of immune response of anti-MTB. Atg5, atg8 and atg12 were the important molecule which control the formation of autophagy when MTB infected.
    No preview · Article · Mar 2009 · Xi bao yu fen zi mian yi xue za zhi = Chinese journal of cellular and molecular immunology
  • Li-Mei Wang · Yin-Lan Bai · Chang-Hong Shi · Hui Gao · Ying Xue · Hong Jiang · Zhi-Kai Xu
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    ABSTRACT: Developing a new generation of vaccines is important for preventing tuberculosis (TB). DNA vaccine is one promising candidate. In this study we evaluated the immunogenicity and protective efficacy of the DNA vaccine encoding the fusion protein of Mycobacterium tuberculosis heat shock protein 65 (Hsp65) with human interleukin-2 (hIL-2) in BALB/c mice. We showed that the DNA vaccine pcDNA-Hsp65-hIL-2 could induce high levels of antigen-specific antibody, IFN-gamma, CD4(+) and CD8(+) T cell production. When the immunized mice were infected with M. tuberculosis H37Rv, the organ bacterial loads in the DNA immunized group were significantly reduced compared to those of the saline control group, but the ability to reduce bacteria was not better than for BCG. The histopathology in lungs of the DNA vaccine immunized mice was similar to that of BCG immunized mice, which was obviously ameliorated compared to that of the saline control group. Overall, the DNA vaccine could afford protection against M. tuberculosis infection, though the protection efficacy was not as great as that of conventional BCG.
    No preview · Article · Jan 2009 · Apmis
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    ABSTRACT: To express Micrococcus luteus resuscitation promoting factor (Rpf) domain and its mutants in prokaryotic cells, and to investigate their bioactivity. The gene of Rpf domain and its mutants (E54K, E54A) were amplified by polymerase chain reaction (PCR) from the genome of Micrococcus luteus and cloned into pMD18-T vector. After sequenced, the Rpf domain and its mutant gene were subcloned into expression vector PGEX-4T-1, and transfected into E. coli DH5alpha. The expressed product was purified by affinity chromatography using GST Fusion Protein Purification bead. The aim proteins were identified by SDS-PAGE analysis and by Western blot with monoclonal antibodies against Rpf domain (mAb). The bioactivity of the proteins was analyzed by stimulating the resuscitation of Mycobacterium smegmatis. The sequences of the PCR products were identical to those of the Rpf domain and its mutant gene in GenBank. The relative molecular mass identified by SDS-PAGE analysis was consistent with that had been reported, which was also confirmed by Western blot analysis that there were specific bindings at 32 000 with Rpf domain mAb. The purified GST-Rpf domain could stimulate resuscitation of Mycobacterium smegmatis. Replacements E54A and especially E54K resulted in inhibition of Rpf resuscitation activity. Rpf domain and two kinds of its mutant protein were obtained, and its effects on the resuscitation of dormant Mycobacterium smegmatis were clarified.
    No preview · Article · Nov 2008 · Zhonghua jie he he hu xi za zhi = Zhonghua jiehe he huxi zazhi = Chinese journal of tuberculosis and respiratory diseases
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    ABSTRACT: To investigate the immunobiology of Rpf domain from Micrococcus luteus. BALB/c mice were immunized with Rpf domain three times at 2-week interval. ELISA was used to detect the title of the anti-Rpf domain antibody titer in the immunized mice sera. The spleen lymphocytes of the immunized mice were separated and the stimulation index (SI) was measured by MTT colorimetry. The levels of secreted IFN-gamma, IL-10 and IL-12 upon specific antigen stimulation was detected by ELISA. The Rpf domain immunized BALB/c mice were intravenously infected with 10(5) CFU MTB H37Rv. The number of CFU in the spleens was determined four weeks after final injection. The titer of the specific antibody in sera of the immunized BALB/c mice was 1:128 000. The SI of Rpf domain immunized group (2.10+/-0.12) was significantly higher than that of saline immunized group (0.90+/-0.21). The lever of IFN-gamma, IL-10 and IL-12 levels in culture supernatant of spleen lymphocytes from the fusion protein immunized mice was (1 126+/-36) ng/L, (368+/-13) ng/L and (289+/-14) ng/L, respectively, which was markedly higher than that of saline immunized group (P<0.01). Compared with normal saline immunized mice (6.64+/-0.13) four weeks after final injection, dramatic reduction in MTB replication was observed in the spleen (5.03+/-0.11) from the BALB/c mice immunized with fusion proteins. Rpf domain can be used as a candidate for a new TB vaccine.
    No preview · Article · Jul 2008 · Xi bao yu fen zi mian yi xue za zhi = Chinese journal of cellular and molecular immunology
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    ABSTRACT: To study the effect of electromagnetic pulse (EMP) exposure on the permeability of blood-testicle barrier (BTB) in mice. Adult male BALB/c mice were exposed to EMP at 200 kV/m for 200 pulses with 2 seconds interval. The mice were injected with 2% Evans Blue solution through caudal vein at different time points after exposure, and the permeability of BTB was monitored using a fluorescence microscope. The testis sample for the transmission electron microscopy was prepared at 2 h after EMP exposure. The permeability of BTB in mice was observed by using Evans Blue tracer and lanthanum nitrate tracer. After exposure, cloudy Evans Blue was found in the testicle convoluted seminiferous tubule of mice. Lanthanum nitrate was observed not only between testicle spermatogonia near seminiferous tubule wall and sertoli cells, but also between sertoli cells and primary spermatocyte or secondary spermatocyte. In contrast, lanthanum nitrate in control group was only found in the testicle sertoli cells between seminiferous tubule and near seminiferous tubule wall. EMP exposure could increase the permeability of BTB in the mice.
    Full-text · Article · Jul 2008 · Biomedical and Environmental Sciences
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    ABSTRACT: To express Micrococcus luteus Rpf domain in prokaryotic cells and prepare monoclonal antibodies against Rpf domain. The gene encoding Micrococcus luteus Rpf domain was amplified from genome of Micrococcus luteus by polymerase chain reaction(PCR), and inserted into cloning vector pUC-19. After sequenced, Micrococcus luteus Rpf domain gene was subcloned into the expression vector pPro-EXHT and transfected into E.coli DH5alpha. After induced by IPTG, the bacteria controlled by T7 promoter expressed the fused Micrococcus luteus Rpf domain protein with a hexahistidine tail at its N-terminal and the target protein was purified under denaturing conditions. Using this protein as antigen to immunize the BALB/c mice and prepare monoclonal antibodies against Micrococcus luteus Rpf domain. Then specifities and relative affinities of mAbs were identified by ELISA. The fusion protein was purified by metal chelate affinity chromatography under denaturing condition. Three cloned mAbs were prepared from the mice immunized by Rpf domain. All of them could recognize Rpf domain. specifically. The prepared mAbs against Rpf domain have strong specificity with high titers, which provides useful tools for further study of the function of Rpf domain in TB prevention.
    No preview · Article · Jun 2008 · Xi bao yu fen zi mian yi xue za zhi = Chinese journal of cellular and molecular immunology

  • No preview · Article · Jan 2008
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    ABSTRACT: The aim of this study was to explore the mechanism underlying the dual effect of androgen on prostate cancer cells and further explore its correlation with dopa decarboxylase (DDC), an androgen receptor (AR) coactivator and a traditional neuroendocrine differentiation (NED) marker. Cell proliferation and cycling after treatment with synthetic nonmetabolizable androgen R1881 was determined by the MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide) method and flow cytometry. Differential gene expression was analyzed by cDNA microarrays. DDC expression during the dual effect of R1881 was further explored with microarray, quantitative reverse transcriptase-polymerase chain reaction (RT-PCR), Western blot, and enzyme activity assays. Proliferation of LNCaP cells was inhibited by 1 nM R1881 but stimulated by 0.1 nM R1881. Compared with the untreated cells, 320 (2.26%; 170 up-regulated, 150 down-regulated) and 4608 (32.65%; 2046 up-regulated, 2562 down-regulated) genes were found to be expressed differentially in the 1 nM and 0.1 nM R1881-treated cells, respectively. The results were partially confirmed by RT-PCR and Western blot. The DDC gene was down-regulated in the 1 nM R1881-treated cells and up-regulated in 0.1 nM R1881- and 30 nM hydroxyflutamide-treated cells. The enzymatic activity of DDC in the latter 2 groups was also strengthened. Meanwhile, the NED markers CgA and synaptophysin were not affected by these AR activators. R1881 had a dose-dependent biphasic effect on LNCaP cell proliferation. AR coactivator DDC was respectively down- and up-regulated in high and low concentrations of R1881. DDC up-regulation by exogenous AR activators is not accompanied by up-regulation of definitive NED markers.
    Full-text · Article · Jul 2007 · Journal of Andrology
  • Li-Mei Wang · Chang-Hong Shi · Xiong-Lin Fan · Ying Xue · Yin-Lai Bai · Zhi-Kai Xu
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    ABSTRACT: Tuberculosis remains the leading cause of human death. Currently, Bacillus Calmette-Guérin (BCG) is the only available vaccine against tuberculosis but its efficacy is highly variable. Thus, developing new tuberculosis vaccines becomes an urgent task. In this study, we evaluated in BALB/c mice the humoral and cellular immune responses of recombinant BCG expressing the antigen ESAT-6 from Mycobacterium tuberculosis. Escherichia coli-BCG shuttle plasmid named pDE22-esat-6 was constructed by inserting the BamHI/EcoRI digested esat-6 gene PCR product into the similarly digested parental plasmid pDE22. BCG cells were transformed with pDE22-esat-6, which was named recombinant BCG (rBCG). BALB/c mice were immunized subcutaneously on the back with 100 microl normal saline containing 10(6) CFU of BCG or rBCG. They were sacrificed after 4 weeks to detect their humoral and cellular responses. There was no any significant differences in the growth characteristics between the conventional BCG and rBCG. In immunized mice, the IgG antibody titres of rBCG group were as high as 1:8000, which was significantly higher than that in BCG group (1:1400, P < 0.05). The elicited IFN-gamma level of rBCG group was (1993 +/- 106) pg/ml, which was also significantly higher than that in BCG group ((1463 +/- 105) pg/ml, P < 0.05). The splenocyte proliferation index of rBCG group reached 4.34 +/- 0.31, which was higher than that of BCG group (3.79 +/- 0.24, P < 0.05). rBCG secreted expressing antigen ESAT-6 stimulated stronger humoral and cellular immune responses than BCG did, and, therefore may be the better vaccine against mycobacterium tuberculosis.
    No preview · Article · Jul 2007 · Chinese medical journal
  • Chang-hong Shi · Zhi-kai Xu

    No preview · Article · Jan 2007 · Zhonghua jie he he hu xi za zhi = Zhonghua jiehe he huxi zazhi = Chinese journal of tuberculosis and respiratory diseases
  • Hai Zhang · Chang-hong Shi · Ying Xue · Yin-lan Bai · Li-mei Wang · Zhi-kai Xu
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    ABSTRACT: To study murine humoral and cellular immune response induced by fusion protein ESAT6-CFP10 and to examine its protective efficacy against M. tuberculosis (MTB) in mice. BALB/c mice were immunized subcutaneously on the back with fusion protein ESAT6-CFP10 that was transferred to nitro cellulose (NC) membrane beforehand. Stimulation index (SI) of the spleen lymphocytes of the immunized mice was measured by MTT colorimetry. The level of IFN-gamma and IL-2 and CTL upon antigen-specific stimulation were detected. The vaccinated BALB/c mice were intravenously infected with MTB H37Rv (10(5) CFU/mouse). Four weeks later the number of CFU in spleens was determined. The titer of serum specific antibody in BALB/c mice immunized with fusion protein ESAT6-CFP10 was 1:6,400. The SI of fusion protein immunized group (1.90+/-0.15) was significantly higher than that of saline-immunized group (0.9+/-0.15). The level of IFN-gamma and IL-2 induced by the fusion protein was 1.792+/-19 ng/L and 0.211+/-11 ng/L respectively, which was significantly higher than that of saline-immunized group and lower than that of BCG-immunized group. The specific killing activity of splenocytes was 36%. Compared with the saline-immunized mice (bacterial load was 6.51+/-0.13), MTB number (bacterial load was 5.24+/-0.15) was reduced dramatically in the spleens of BALB/c mice immunized with the fusion protein, but the protective efficacy of the mice immunized with BCG was higher than that of ESAT6-CFP10 vaccinated group. Fusion protein ESAT6-CFP10 can be used as a candidate for novel vaccines.
    No preview · Article · Aug 2006 · Xi bao yu fen zi mian yi xue za zhi = Chinese journal of cellular and molecular immunology
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    ABSTRACT: To explore the effect of androgen receptor (AR) on the expression of the cell cycle-related genes, such as CDKN1A and BTG1, in prostate cancer cell line LNCaP. After AR antagonist flutamide treatment and confirmation of its effect by phase contrast microscope and flow cytometry, the differential expression of the cell cycle-related genes was analyzed by a cDNA microarray. The flutamide treated cells were set as the experimental group and the LNCaP cells as the control. We labeled cDNA probes of the experimental group and control group with Cy5 and Cy3 dyes, respectively, through reverse transcription. Then we hybridized the cDNA probes with cDNA microarrays, which contained 8 126 unique human cDNA sequences and the chip was scanned to get the fluorescent values of Cy5 and Cy3 on each spot. After primary analysis, reverse transcription polymerase chain reaction (RT-PCR) tests were carried out to confirm the results of the chips. After AR antagonist flutamide treatment, three hundred and twenty-six genes (3.93%) expressed differentially, 97 down-regulated and 219 up-regulated. Among them, eight up-regulated genes might be cell cycle-related, namely CDC10, NRAS, BTG1, Wee1, CLK3, DKFZP564A122, CDKN1A and BTG2. The CDKN1A and BTG1 gene mRNA expression was confirmed to be higher in the experimental group by RT-PCR, while p53 mRNA expression had no significant changes. Flutamide treatment might up-regulate CDKN1A and BTG1 expression in prostate cancer cells. The protein expressions of CDKN1A and BTG1 play an important role in inhibiting the proliferation of cancer cells. CDKN1A has a great impact on the cell cycle of prostate cancer cells and may play a role in the cancer cells in a p53-independent pathway. The prostate cancer cells might affect the cell cycle-related genes by activating AR and thus break the cell cycle control.
    Full-text · Article · Jan 2006 · Asian Journal of Andrology
  • Chang-hong Shi · Xiao-wu Wang · De-sheng Zhu · Zhi-kai Xu · Yin-lan Bai · Ying Xue
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    ABSTRACT: To evaluate the humoral and cell-mediated immune responses induced by a genetic vaccine expressing the Ag85B-ESAT6 fusion protein, and to investigate its protective effect against Mycobacterium tuberculosis (MTB) challenge. Fifty BALB/c mice were randomized into 5 groups and subjected to the following treatments respectively: immunization with normal saline, BCG, pcDNA3, A(Z)-pcDNA3-E(F) and E(Z)-pcDNA3-A(F) for 3 times at 2-week intervals. The stimulation index (SI) of the splenic lymphocytes from the immunized mice was measured by the methyl thiazolyl tetrazolium (MTT) method, and the level of secreted IFN-gamma upon antigen-specific stimulation was detected by ELISA. The immunized mice were intravenously infected with 10(5) colony forming unit (CFU) of MTB H(37)Rv. The numbers of MTB CFU in spleens were determined 4 weeks later. The specific antibody titers in the sera of mice immunized with plasmid A(Z)-pcDNA3-E(F) and E(Z)-pcDNA3-A(F) were 1:1,000 and 1:1,500 respectively, and the SI was 2.2 and 2.4 respectively, while the SI of the normal saline group and the plasmid pcDNA3 immunized group was only 0.9 and 1.1 respectively. The IFN-gamma concentrations in cultured supernatant of splenic lymphocytes from mice immunized with plasmid A(Z)-pcDNA3-E(F) [(5.48 +/- 0.38) ng/ml] and E(Z)-pcDNA3-A(F) [(5.76 +/- 0.51) ng/ml] were significantly higher than those of the normal saline group [(0.50 +/- 0.25) ng/ml] and the plasmid pcDNA3 immunized group [(1.20 +/- 0.33) ng/ml, P < 0.05], but were not significantly different with that of the BCG immunized group [(5.55 +/- 0.31) ng/ml]. Compared with plasmid pcDNA3 immunized group, the bacterial load (lg, CFU/g) in spleen was 6.08 +/- 0.25 which dramatically reduced in mice immunized with recombinant plasmids, but the protective efficacy of mice immunized with plasmid A(Z)-pcDNA3-E(F) (4.63 +/- 0.11) or E(Z)-pcDNA3-A(F) (4.50 +/- 0.32) was lower than that of the BCG vaccination group (4.09 +/- 0.27). The cell-mediated immune response induced by genetic vaccine expressing the Ag85B-ESAT6 fusion protein was similar to that induced by BCG immunization.
    No preview · Article · Nov 2005 · Zhonghua jie he he hu xi za zhi = Zhonghua jiehe he huxi zazhi = Chinese journal of tuberculosis and respiratory diseases

  • No preview · Article · Jun 2005 · Chinese medical journal
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    ABSTRACT: To investigate the effects of rBCG vaccination containing foreign antigen Der p2 in the form of lipoprotein on murine immune response. 6 to 8 weeks old and newborn BALB/c mice were vaccined intraperitoneally with 10(6) CFU rBCG or BCG. At the same time, the control group was injected with saline. Six weeks later, all animals were injected with Der p2 (20 microg). After two weeks later, the concentrations of IL-4 and IFN-gamma in the serum and splenocyte culture supernatant (STLCS) were determined by ELISA, and Th subgroups were determined by double fluorescent staining and flow cytometry. After vaccination, the serum and STLCS from both rBCG-immunized and BCG-immunized group of adult and newborn BALB/c mice had significantly higher level of IFN-gamma and lower level of IL-4 than those from control groups. Besides, there was the larger percentage of CD4 (+) IFN-gamma (+) cells in spleen from rBCG-vaccined and BCG-vaccined mice than that from control group. However, the percentage of CD4 (+) IL-4 (+) cells in spleen cells from rBCG-vaccined and BCG-vaccined group was lower than that from control group. Moreover, the level of IFN-gamma in STLCS from rBCG-immunized was significantly higher, compared with that from BCG-immunized mice. At the same time, the percentage of CD4 (+) IFN-gamma (+) cells in spleen from rBCG-vaccined mice was larger than that from BCG-vaccined group. Both rBCG and BCG could stimulate Th1 predominant immune response, when injected intraperitoneally into adult or newborn BALB/c mice, The Der p2 expressed on the cell wall of BCG can work as the component of BCG and be recognized by the immune system of mice, therefore stimulates Der p2-specific Th1 predominant immune response. These data indicate that recombinant BCG-expressing antigens can be used as the antigen-specific vaccines against allergic diseases by regulating the balance of Th1/Th2.
    No preview · Article · Jun 2005 · Xi bao yu fen zi mian yi xue za zhi = Chinese journal of cellular and molecular immunology
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    Jie-Ran Shi · Chang-Hong Shi · Chang-Gui Wu · Jin-Shan Zhang
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    ABSTRACT: AIM: To investigate the influence of oral cavity and gastrointestinal tract environment on the biological behavior of recombinant Bacillus Calmette-Guerin (rBCG) that expresses with allergen Der p2 after oral inoculation. METHODS: Four groups of 6 to 8 week old Balb/c mice were vaccinated orally with 100 μL of 109 CFU with natural BCG or three kinds of rBCGs. The BCG CFU was counted in several tissue (oral pharyngeal lymphoid tissues (OLT), gut associated lymphoid tissues (GALT), spleen, lung and liver) and feces from the inoculated Balb/ c mice at different time after inoculation. The DNA and mRNA of Der p2 gene in various tissue were identified by PCR and RT-PCR, respectively. RESULTS: All three kinds of rBCGs were observed in mucosal-asscciated lymphoid tissues, indicating that they could penetrate the mucosa of oral cavity and gastrointestinal tract. The Der p2 gene was expressed by rBCG in above tissues. CONCLUSION: It is suggested that the oral cavity and gastrointestinal tract environment does not interrupt rBCG expressed Der p2 from inducing immune response after oral inoculation.
    Preview · Article · May 2005 · World Chinese Journal of Digestology