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The case-based teaching was inserted in the lecture-based teaching in clinical microbiol-ogy. The teachers conducted the case-based teaching through proper selection of cases, leading students to discuss and analyze cases. The students explored and reported from the cases independently. Finally, the teachers summarized and evaluated from the case-based teaching. The combination of case-based teaching and lecture-based study stimulated the students' interest in learning and motivation, consolidated their theoretical knowledge, cultivated their abilities of self-learning and clinical idea, and made the correlation between the theory and the clinical practice closely. These methods improved the teaching quality of clinical microbiology.
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Regulatory T cells (Treg),a group of negative regulatory cells,have four subsets:CD4+Treg,CD8 + Treg,NKT Treg and DN Treg cells.They play an essential role in the inhibitive immune-regulation and might be the key factors of neoplasms immune escape.These mechanisms include inhibiting the effector cell function by inhibitory cytokines,killing effector cells by granzyme and perforin,competition and inhibiting IL-2,and affecting Treg differentiation and proliferation by regulating the function of CTLA-4,etc.Tumor immunotherapies targeting Treg and related immunosuppressive factors,such as remove Treg or controling the numbers and functions,enhances the immune response against tumors,which might offer a new method of tumor immunotherapy.
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Glucose metabolism of cancer cells presents Warburg effect.In resent years,more and more experiments demonstrate that the therapeutics based on aerobic glycolysis pathway in cancer cells restrict energy and inhibit tumor proliferation through the inhibition of a variety of moleculars,genes and signal pathways.These can provide an opportunity for targeted cancer therapies and possess enormous potential advantages and broad prospects in clinical application.
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Cancer stem cells are a subclass of stem-cell-like tumor cells.They have an apparently higher tumorigenicity than other cell populations within the same tumor.They play a Very important role in genesis,development and maintenance of cancer.The data from research in leukemia and solid timors suggest that in each cancer tissue,only a small fraction of the cells have the ability to initiate tumor,which are called cancer stem cells.In recent years,with in-depth research on stem cells and cancer stem cells,accumulating evidence suggests that cancer stem cells are the cause of malignant tumor metastasis and recurrence and are resistant current callcer drugs.Drugs targeting cancer stem cells are urgently needed. Flow cytometry and magnetic cell sorting using the identified tumor stem cell surface markers are mainly used to separate cancer stem cells or stem cell-like cells. In this article,the concept of cancer stem cells and the commonly used approaches to isolate and identify cancer stem cells,especially for gastric cancer stem cells will be reviewed.
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Objective To investigate the adjuvant effect of dimo-thylidioctyl ammonium bromide (DDA) and/or DDA-BCG polysaccharide nucleic acid( BCG-PSN), which was combined with a Mycobacterium tuberculosis fusion protein AMM ( Ag 8 5 B - MPT64190-198 - Mtb8.4 ) to boost BCG primed immunization. Methods DDA with or without BCG PSN was mixed with the fusion protein AMM to construct the boosting vaccine. Mice were immunized with BCG and then boosted twice with AMM formulated with the adjuvant DDA with or without BCG-PSN. PBS or BCG vaccination without boosting was used as control. The humoral and cell-mediated immune responses were analyzed by ELISA and ELISPOT. Moreover, the protective efficacy of BCG prime-AMM subunit vaccine boosting against Mycobacterium tuberculosis infection was analyzed. Results With in vitro stimulation of Ag85B and PPD( purified protein derivative) antigen, the number of IFN-γ secreting cells from the mice boosted twice by AMM/DDA/BCG-PSN and AMM/DDA were higher than BCG and PBS group (P <0.05). The CFU in lungs of mice boosted with AMM/DDA/BCG-PSN was less than that of PBS group(P <0.05), while the CFU of AMM/DDA-boosted mice was less than that of BCG and PBS group(P < 0.05).However, fewer lesions were seen in lungs of mice immunized with BCG alone or BCG-prime-AMM/DDA/BCG-PSN boosting than the other groups. Conclusion DDA is an idea adjuvant for tuberculosis subunit vaccine;BCG-PSN might play a role in alleviating the immunity-mediated pathology.
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Objective To construct protective immunity to Mycobncterium tuberculosis latent infection, a novel fusion protein consisting of HspX, the 190 to 198 peptide of Mpt64 and Ag85B, which were confirmed to be the effective protective antigens mainly expressed in the dormancy and exponential phase of growth, was constructed and its immunogenicity was investigated. Methods Ag85B and Mpt64190-198-HspX sequences were amplified by PCR and cloned into plasmids pET-28a. The fusion protein, Ag85BMpt64190-198-HspX (AMH) was expressed in E. coli BL21 and purified with Ni-NTA resins. C57BL/6 mice were immunized three times at 2-week intervals subcutaneously with AMH formulated with the adjuvant composed of dimethyl-dioctyldecyl ammonium bromide (DDA) and BCG polysaccharide nucleic acid (BCGPSN). Humoral and cell-mediated immunity responses were analyzed at five weeks after the last injection. Results AMH was expressed stably in E. coli and could be purified well by Ni-NTA affinity chromatography. C57BL/6 mice immunized with AMH subunit vaccine generated specific cellular and humoral immunologic response to the stimulation of Ag85B, Mpt64190-198 and HspX. Conclusion It suggested that AMH was a promising candidate antigen of tuberculosis subunit vaccine.
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Objective To investigate the boosting efficiency of a subunit vaccine consisting of the fusion protein Ag85B-Mpt64190-198-Mth8.4 (AMM) , dimethyl-dioctyldecyl ammonium bromide (DDA) and BCG polysaceharide nucleic acid (BCG-PSN) on the primed inoculation with BCG. Methods The AMM subunit vaccine was composed of fusion protein AMM, adjuvant DDA and BCG-PSN. The first mouse experi-mental group was immunized with BCG first, then boosted with the AMM subanit vaccine in the 10th week. The second experimental group was boosted with the AMM subunit vaccine in the 8th week and the 10th week respectively with a two weeks interval after the primed with BCG. Two control groups were treated re-spectively with physiological saline alone and BCG alone. After the primed inoculation, ELISPOT and ELISA were used for the detection of the cell-mediated and humoral immune response in week 14 and week 22 re-spectively. Furthermore, the immunized mice were challenged with live BCG to mimic tuberculosis infection in the 22nd week after the primed inoculation. Subsequently the T cell typing and humoral response were de-tected by flow cytometry and ELISA, respectively. Results ( 1 ) The level of secreting IFN-γ: 14 weeks af-ter the primed inoculation,with the stimulation of the specific antigen-Ag85B, the number of cells secreting IFN-γ in the second experimental group (135±14) was more than BCG alone immunized group (19±16), t = 10. 98, P < 0.01. In the 22nd week, the number of cells secreting IFN-γ in the second experimental group (208±11) was still more than BCG alone group (57±18), t =6.43, P <0.01. (2) The level of humoral immune response: the IgG1 antibody titer in the second experimental group was obviously higher than that in the first experimental group. However, the ratio of IgG2a to IgG1, as the index reflecting the Thl-type immune response, in the experimental group 2 was lower than that in the experimental group 1. (3) The contents of CD4+ CD25+ T cells after challenged with live BCG strain: the first and the second ex-perimental groups were both higher than the BCG alone group (t1 = 3.08, t2 = 3.16, P < 0.05 ). Conclu-sion Boosting the BCG-pfimed mice with tuberculosis AMM subunit vaccine twice can induce higher level of cell-mediated and humoral immune response than BCG alone, which could activate the regulative immune response at the same time.
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Objective To construct the recombinant secretion type BCG-Eg95 vaccine of Echinococcus granulosus (rsBCG-Eg95). Methods BCG-Ag85B signal sequence with 117 bp and Eg95 gene with 471 bp were amplified from the genome of BCG and pGEX-4T-Eg95 by PCR,respectively. BCG-Ag85B signal coding gene and Eg95 gene were cloned into E. coli-BCG shuttle-vector pMV261 to get the recombinant plasmid pSMEg95,which was confirmed by restriction endonuclease digestion,PCR amplification and gene sequencing. These recombinant plasmids were introduced into BCG by electroporation for the construction of rsBCG-Eg95 vaccine. The rsBCG-Eg95 positive clones were screened by Kan+ and identified by PCR amplification. Results BCG-Ag85B signal sequence coding gene and Eg95 coding gene were successfully cloned into pMV261,which was confirmed by restriction endonuclease digestion,PCR amplification and sequencing of the plasmid pSMEg95. The plasmids were introduced into BCG and confirmed as the recombinant secreting BCG-Eg95 vaccine of E. granulosus (rsBCG-Eg95). Conclusion The recombinant secretion type BCG-Eg95 vaccine (rsBCG-Eg95) of E. granulosus with BCG-Ag85B signal sequence and Eg95 gene has been constructed.