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Immunology is a kind of important basic subject for medical education, involving theories and technologies covering many aspects of disease occurrence, diagnosis, prevention and treatment, and is also the source of new means of medical treatment in the future. Therefore, successful teaching of medical immunology is associated with high-level medical undergraduate education. An appropriate textbook is critical for curriculum construction of immunology. However, little analysis of immunology textbooks has been published. We selected several famous and popular English immunology textbooks and compared their teaching objectives, methods, and contents, concluding the features and targeting readers of each textbooks. Our research results could provide some advices for students to learn immunology theory and researchers to utilize immunology methods, and also give a glimpse of the development trend and direction of immunology.
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In China,few research findings in basic medicine could be translated to clinical practice,which is known as the disconnection between basic research and clinical application.Such phenomenon is mainly due to lack of ability in translational medicine,particularly,lack of training in clinical research and translational medicine.Now the emergence of translational medicine has put forward new requirements for graduate education in medical school,and demanding that the patient-centered concept be strengthened.As the major force of scientific and technological innovation,graduate students should receive long-term systematic trainings and conduct original researches.In addition,advanced courses in clinical research and translational medicine should be offered to graduate students ;furthermore,translation research platforms should be built up for them so as to improve the capacity of scientific research and innovation.
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Objective To construct and express a prokaryotic expression vector carrying the gene of FimH 1-156 that comprises human lysosome membrane protein 2 P41-49 gene ,and to express and purify the fusion protein .Methods FimH1-156 gene was cloned from plasmid pPKL241 by PCR ,and inserted into vector pET-28a(+ ) to obtain prokaryotic expression plasmid pET-28a-FimH . After transforming Escherichia coli BL21(DE3) with pET-28a-FimH ,fusion protein FimH1-156 was expressed under induction .The target fusion protein was purified ,and its antigenicity was detected through Western blot .Results The expressed recombinant pro-tein was purified ,the expression of protein was the highest when IPTG was 1 mmol/L and 4h after induction ,it was expressed as include body form ,and the expressed protein was identified to react with monoclonal antibodies 6 × His by Western blotting .Conclu-sion We cloned FimH1-156 fusion protein expressed genes successfully ,constructed prokaryotic expression vector ,and won the in-clusion body purification of FimH1-156 fusion protein .
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Ph.D candidate education is the highest level of higher education. Training model of Ph.D candidate in Medical College of Georgia and University of Manitoba)has vivid characters compared with that in China,which is reflected by the training objective,qualification of students and tutors,culti-vating procedures and admission requirements for graduation. This kind of cultivating model performs stringent selection and can gradually pick out persons who are really fit for the scientific research. Ph.D candidate quality in the two universities is guaranteed by systemic and deep courses learning,immediate update of knowledge and strict evaluation system. The goal of this article is to provide experience and ref-erence for improving the education quality of medical Ph.D candidates in China.
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Medical Immunology is a multidisciplinary basic discipline,but its' abstract and complex leading most beginners daunting.Teachers should emphasize on ‘ transposition thinking' and stick the teaching concept of ‘ students-centered and teacher-guided learning' to the whole teaching process.Teacher should always make transposition thinking on how to stimulate students learning interest,how to guide students to learn autonomously,and how to make problem-based learning.This is important to build self-learning platform for the students and to achieve‘teaching' and ‘learning'win-win.
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The curriculum system of Proteomics was analyzed based on the teaching practice,the characteristics of ability training and gradation teaching were summarized and the prospect of curriculum optimization was proposed.These measures were conceived to enrich the course content and teaching methods for Proteomics course.
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Objective To investigate the immunogenicity of immunodominant cytotoxic T lymphocyte epitope E749-57 of human papilloma virus (HPV) 16 oncoprotein E7 chaperoned by heat shock protein (HSP)110. Methods Mouse HSP110 gene was cloned into prokaryotic expression vector pQE-80L for the expression of HSP110 protein, which was purified using Ni-NTA column. SDS-PAGE and Western-blot were conducted to confirm the purified mHSP110 protein, which was subsequently incubated with E749-57 peptide under heat shock condition, and high-performance liquid chromatography (HPLC) was used to evaluate the binding efficiency of the recombinant protein and E749-57 peptide. Twenty mice were divided into 4 groups to be immunized with mHSP110 protein, E749-57 peptide, mHSP110-E749-57 complex and phosphate buffered saline (PBS),respectively. Two weeks after the last immunization, spleen cells were collected from the immunized mice and divided into 2 parts: one were stimulated by E749-57 peptide followed by the detection of CD8+ INF-γ+ T cells with flow cytometry; the other one were subjected to MTT analysis for the estimation of cell proliferation. The mHSP110-E749-57 complex was also used to immunize TC-1 tumor bearing mice to observe its anti-tumor effect.Results The full-length 2577 bp-sized mHSP110 gene was amplified from mouse liver cDNA and cloned into pQE-80L vector. Direct sequencing confirmed the correctness of the cloning. SDS-PAGE and Western-blot demonstrated the successful purification of mHSP110. HPLC assay showed that the purified mHSP110 protein could bind with E749-57 to form a relatively stable protein complex. The percentage of IFN-γ+ CD8+ T cells in and proliferation index of spleen cells from the complex-immunized mice were statistically higher than those from the other 3 groups of mice. Moreover, the complex could obviously inhibit the growth of TC-1 tumor in mice. Conclusion The mHSP110-E749-57 complex could enhance the generation of specific cytotoxic T lymphocytes and exert anti-tumor effects in mice.
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Objective:To predict and analyze the secondary structure and B cell epitopes of Izumo protein.Methods: The secondary structure and flexible regions of Izumo protein were predicted by the methods of Chou-Fasman,Gamier-Robson and Karplus-Schulz.Moreover,hydrophilicity plot,surface probability plot and antigenic index of Izumo protein were predicted by the methods of Kyte-Doolitde,Emini and Jameson-Wolf,respectively.Results: Izumo protein contained moreαhelix regions.There were several centers ofαhelix in the regions of 6-17,30-40,88-99,103-120,153-160,173-188,249-260,283-297,334-338 and 339-346 of Izumo protein,and several centers of βsheet in the regions of 21-25,198-200,245-248 and 320-323.Moreover,many distinct B cell epitopes in Izumo protein possibly localized in the regions of 3642,62-66,94-99,118-122,129-132,151-154,161-164,173-177,205-208,212-216,256-265,271-276,283-288,314-318 and 336-350.Conclusion:These results are helpful for identification of the dominant B cell epitopes and the functional domains of Izumo protein.
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Objective To investigate the differential expressions of microRNA after HBV integration. Methods A human microRNA microarray was used to analyze the microRNA expression profile in HepG2.2.15 cell line, a novel model of HBV infection. HepG2 cells served as a mock control. Then the microRNA with differential expression were proved using real-time PCR. Results HBV infection induced 6 microRNA down-regulated (hsa-miR-30a-5p, hsa-miR-24, hsa-let-7a, hsa-let-7c, hsa-let-7f and hsa-miR-23b) and 3 microRNA up-regulated (hsa-miR-194, hsa-miR-200a and hsa-miR-345). Conclusion HBV integration can induce the changes of microRNA expression profile in hepatocytes.
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Objective: To identify the mutation points of Cu/Zn superoxide dismutase(SOD1) gene in an amyotrophic lateral sclerosis(ALS) family with a unique phenotype,and to compare the value of single strand conformation polymorphism(SSCP) and denaturing high performance liquid chromatography(DHPLC). Methods: Five exons of SOD1 gene were amplified by PCR. The difference of these products were analyzed by PCR-SSCP and DHPLC.DNA sequencing was used to examine the mutation. Results: ①Mutations were found in exons 2 and 5 in several family members.DNA sequencing revealed that a base pair insertion occurred in the codon area of exon 2 and in the non-codon area of exon 5.②The results of DHPLC tests proved double peaks in one member with ALS symptoms(Ⅲ1),which indicated the possibility of mutation in SOD1 exon 4.DNA sequencing revealed that there was a heterozygote,with a mutation of GAA to GGA in exon 4 in the member with double peak. Conclusion: ①The mutations in exons 2,4,5 were proved.Insertion of exon 2 may be responsible for the disease of the ALS family in Chongqing.②Compared with PCR-SSCP,DHPLC technique has been proven to be a rapid and reliable method for screening mutation site in large samples.
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Objective To identify the point mutation of Cu/Zn superoxide dismutase(SOD1) gene in an amyotrophic lateral sclerosis(ALS) family and observe the value of denaturing high performance liquid chromatography(DHPLC). Methods DHPLC and DNA sequencing were used to examine SOD1 gene of the ALS family which had not been found mutation by PCR-SSCP. Results DHPLC tests proved double peaks in one member(Ⅲ_1), Which indicated the possibility of mutation in SOD1 exon 4. DNA sequencing revealed that there was a heterozygote,with mutation of GAA to GGA in exon 4, and with a substitution of glutacid by glycine. Conclusion As compared with PCR-SSCP, DHPLC technique has proved to be a rapid and reliable method for screening mutation site in large samples.
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Objective To obtain high-yield and easy-purification severe acute respiratory syndrome coronavirus(SARS-CoV) S1 protein with biological activity and to study the activity of S1 protein and its antibody further.Methods SARS-CoV S1 gene was inserted into yeast expression vector pMET?A by ligation reaction.The recombinant plasmid was verified by enzyme digestion and sequencing,followed by being transformed into yeast host strain PMAD11 with electroporation.After induced with methanol,the S1 gene expression was verified with overlay assay and Western blotting.Results The positive clones of S1 gene into pMET?A were approved by restriction enzyme digestion and sequencing.The expression of S1 protein was confirmed subsequently by overlay assay and Western blotting.Conclusion SARS-CoV S1 gene has been cloned and expressed in yeast p.methanolica,which can provide experimental data for next study on the activity of this protein and its antibody during SARS-CoV infection.
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Objective To study the immunity suppressive effect of the staphylococcal enterotoxin as a super-antigen and investigate its mechanism.Methods BALB/c mice aged 8-12 weeks were randomly assigned to receive 0.2 ml injection of 50 ?g/ml staphylococcal enterotoxin B(SEB)(n=20) or 0.2 ml physiological saline(n=20).One day later,all mice were sacrifice to collect the splenocytes which were employed to detect the expression of TGF-?1 and to countthe cells expressing CD4 and CD25 by flow cytometry(FCM).Results FMC showed that a remarkable increase of cells that expressed CD4 and CD25 in the SEB-primed splenocytes as compared with the saline primed splenocytes.Conclusion SEB,which is used as a superantigen in vivo,can induce the regulatory cells bearing suppressive activity.This may be partial mechanism of SEB-induced hyporesponsiveness.
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Objective To explore a new simpler method for the preparation of recombinant alpha virus as a novel vaccine at the DNA level. Methods Plasmids expressing ? gal protein and helper plasmids were transfected into BHK cells. Virus in culture supernatant of the transfected BHK cells were collected and purified and used to infect BHK cells in vitro to identify the expression of target gene and the titre of the recombinant virus. Results Recombinant virus with high titre, prepared by this method, could be expressed well in mammalian cells in vitro . Conclusion High titre recombinant alpha virus can be produced at the DNA level and this method can be applied for vaccine preparation and gene therapy.
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Objective To establish animal tumor model expressing human oncogene HER2/neu and to evaluate the model by immunological method. Methods The recombinant eukaryotic plasmid expressing HER2/neu gene was transfected into P815 tumor cells. The tumor cell line P815 expressing HER2/neu gene stably was screened by limiting dilution method. The expression of target gene in host cells was determined by Western blotting. Following the establishment of animal tumor model expressing HER2 antigen in DBA/2 mice, tumor specific response was induced in the spleen cells of DBA/2 mice which were inoculated with recombinant plasmids. Results P815 cell line expressing HER2/neu oncogene stably was obtained. HER2/neu specific CTLs could be induced from immunized DBA/2 mice. Conclusion HER2/neu expressing tumor animal model is established successfully and the in vivo immune response can be induced after immunization with HER2/neu genetic vaccine.
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Objective To obtain Chinese hamsterovary (CHO) cell line expressing human decay accelerating activity (hDAF) stably and to observe the protective effect of hDAF on heterologous cells under the circumstance of complement activation. Methods The eukaryotic expression vector DAF pcDNA3.1 was constructed and then transfected into CHO cells by lipofection. Monoclones of cells expressing hDAF stably were screened by the method of limiting dilution. hDAF expression was detected by flow cytometry. The decay accelerating activity of hDAF was determined by assay of C3 deposition and 51Cr release. Results The expression vector DAF pcDNA3.1 was successfully constructed, and monoclones of cells expressing hDAF were obtained. CHO cells expressing hDAF could decrease C3 deposition and attenuate the killing effect of activation of the complement system. Conclusion We have obtained CHO cell clones expressing hDAF stably, which is helpful for the further studies of the relationship of the structure with the functions of hDAF.
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Objective To investigate the effect of thymosin ?1 (T?1) on cellular immune function in gastric cancer patients through observing its treatment on the differentiation of T-lymphocyte subsets from screened peripheral blood mononuclear cells (PBMCs). Methods PBMCs were obtained by centrifugation of blood samples from 18 healthy subjects and 32 patients with gastric cancer,and then cultured in the presence of culture medium with addition of T?1 at 50,10 and 1 ?g/ml for 2 d. T lymphocyte subsets (such as CD4+,CD8+ and CD4+ CD25+ Foxp3+ T cells) and Th1/Th2 multiplex cytokines were detected by flow cytometry (FCM). Results After PBMCs isolated from healthy people and patients were incubated with or without T?1,there was no significant change in percentage of CD4+,CD8+ peripheral lymphocyte subsets and ratio of CD4+/CD8+. There was no obvious change in the percentage of CD4+ CD25+ Foxp3+ T lymphocyte subsets in the normal control,but a significant increase was observed in the cells from patients with gastric cancer after treatment (P
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Objective To explore how to trigger an HLA Ⅰ restricted CD8 + T cell response to exogenously synthesized peptides in vitro . Methods A new panel of therapeutic peptides based on the immunodominant B and CTL epitopes of HBV PreS 2 region and HBcAg and the tetanus toxoid common T helper epitopes were synthesized by Merrifield solid phase peptide synthesis, and HLA A2 + human PBMCs were used to investigate the immunological properties of the mimetic peptides. Results The results demonstrated that the peptides could trigger vigorous CD8 + HBV specific CTL responses in vitro specifically and effectively. Conclusion The results reveal that T helper plus B epitopes designing with the introduction of short and flexible linker can remarkably improve the immunogenicity of short peptides and hence produce effective CTL responses in vitro .
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Objective To predict the HLA A *0201 restricted CTL epitopes (nonamers) in SARS coronavirus (SARS CoV) E protein. Methods The CTL epitopes of E protein were predicted by using supermotif method combined with quantitative matrix method. Results Thirteen HLA A *0201 restricted CTL epitope candidates were predicted in SARS CoV E protein. Conclusion The prediction of the CTL epitopes in SARS CoV E protein will benefit the identification of CTL epitopes by experiment. Those results are of importance for immune recognition and vaccine design for SARS CoV.
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Objective:To identify HLA-A2-restricted CTL epitope of breast cancer differentiation antigen NY-BR-1.Methods:The HLA-A2-restricted CTL epitope of breast cancer differentiation antigen NY-BR-1 is predicted by combination quantitative motif method and the molecular dynamics.The three epitope were assayed their affinity to HLA-A2.Results:The affinity to HLA-A2 of NY-BR-1_ 1043-1051 is a best.Conclusion:NY-BR-1_ 1043-1051 is a HLA-A2 restricted epitope.