| The gene vaccine compared with the norpmal protein vaccinesand others can induce body immunity more completely. Recentresearch shows that the gene vaccine can availably stimulate the bodyto produce stronger cellular immunity, particularly the effect ofcytotoxic T lymphocytes. Because the gene vaccine is in the bodythrough turn to record, the deluxe space structure of the glycoproteinreflection antigen that translation system synthesize, not only caninduce the host CD4+ the body fluid immunity ,but also can induce theCD8+ the cell immunity. This research examines the immunity inadopting the HSV-1 protection antigen gene gB to set up full longpcDNA3-gB and truncated pcDNA3-gB the small rat T lymphoidcell variety, the CTL activity of the second cluster and the levelthat express the cell factor IL-2, the IFN γmRNA.The cell immunity should take place to the HSK paroxysm toemphasize the function to the clearance of the HSV-1 within the body.The gB is a main target antigen that infects with the lymphoid cell ofthe host HSV-1 cell toxicity T, is also HSV-1 especially the differentCTL identifies to order. HSV original hair the infection induces thestrong CTL reaction of aim at the gB namely. The cell expression ofthe infection combines to pass to present the gB quickly, stiring up theparticularity CD8+ availably lymphoid cell. The cell that CTL kill theinfection directly, and secrete the replication of the cell factorinterference virus, induce to not infect with the cell into the virussafeappearance. Be in conjunction with the function through severalaspects, it can resist the virus infection effectively, limit the HSV-1 inthe nervous system of sow to spread, promote the more match of theinfection.In this research, the FCM examine CD4+ of the adoptionantibody marking CD8+ the variety of the second cluster of lymphoidcell, count the method examination HSV-gB gene vaccine to inducethe output particularity CTL of small rat to respond through a flowtype of marking cell, use to converse to record to come together theenzyme chain to respond( RT-PCR) the method, inquiry into the cellof spleen in the expression that IL-2, the IFN γmRNA is after genethe vaccine immunity. As a result showing that these two kinds ofgene vaccines can induce the small rat to produce stronger cellimmunity, performance the BALB/ c small rat CD4+ the lymphoid cellincrease, the CTL activity strengthen, the cell factor IL-2 and theexpression level of the IFN-r mRNAs increase highly. Objective: Having already set up full long(2673 bps) andtruncated(1482 bps) two kinds of herpes simplex virus I typeglycoprotein B DNA vaccine. After small rat of immunity, inquiryinto the influence of it upon the small rat immunity function, andcarry on the comparison of both vaccines, make fully variousimmunity index signs of more excellent gB HSV-1 gene vaccine, forprevent from and cure the herpes simplex virus infection laying thefoundation. Methods: After immunizing small rat, take the spleen cell to beused for the examination. First the FCM of the applied antibodymarking, second cluster of lymphoid cell of the examination.Thelymphoid cells have two mainly second cluster, then lymphoidcell( Th) CD4+ of the assistance and the cell poison/ repress thelymphoid cell( Ts) CD8+.CD4+ lymphoid cell has the activity ofregulate the immunity reaction, the assistance B cell creatingantibody, and secrete the cell factor, but CD8+ the lymphoid cell thenhas the immunity to repress and the cell toxicity function.Theprinciple: make use of various single gram antibody and lymphoid cellsurface of antigen combine, many color fluorescence dyestuffs ofmatch, with opening a distinction of various different function, thenget opposite comparison of each cluster. The next in order, we adopt the poisonous activity of the cell thatFCM of marking cell counts the examination of cell to lead. The gB isa main target antigen that infects with the lymphoid cell of the hostHSV-1 cell toxicity, is a HSV-1 especially the different CTL identifiesto order. The principle uses the first fluorescence dye stuff CFDA-SEmarking target cell first, distinguishing the effect cell and cell oftargets in order. Distinguish the cell of target that film break alreadywith the second DNA fluorescence dye stuff PI again, the PI carries onkilling the wound to the cell of target at the particularity CTL, the cellmembrane transparence contain small change, can immediately enterthe cell, thus certain death percentage of the cell. As a result havethe high intelligent, can immediately examine the death of the cell inearly time at killing to harm. End, to examine the IL-2 and the IFN γmRNA, adopt RT-PCRmethods that spleen cells of the small rat express. After the genevaccine immunity cell of B propagates the conversion, and promotethe T expression of the lymphoid cell IL-2, the IFN γmRNA. But IL-2,the IFN γagain is to promote lymphoid importance of cell propagateand divide of poisonous T of cell adjust control the factor. The aboveboth side is in conjunction with the function, stiring up from the non-particularity immunity and particularity immunity both side theimmunity of the small rat. In order to examine the expression level ofthe IL-2, the IFN γmRNA, the usage of reverse transcriptionpolymerase chain reaction inquiry into the IL-2, the IFN γmRNAafter gene the vaccine immunity of expression. The principle of theRT-PCR is: Withdraw the organization or total RNA within cells, bethe template with the mRNA in which, adopt the Oligo( dT) or leadthe thing exploitation to converse to record the reversal of enzyme torecord random cDNA. Take cDNA as again the template to carry onPCR expand, then acquire the purpose gene or examine the geneexpression.The result is full long pcDNA3-gB and truncated pcDNA3-gBhaving the function of the BALB/ c rat spleen CD4+ immunitylymphoid cell increment.The immunity a CD8+ of the gene vaccinethe T lymphoid cell and CD4+/ CD8+ although compare thephysiology brine and the empty immunity of quality matched controlto increase high, the covariance did not show the difference obviously.Consider the possible reason is that the existence of repressing T cell,or is because of experiment error margin, this still needs the furtherresearch.The particularity cell poison of two kinds of genes vaccineimmunity set responds to show the high in the matched control and theimmunity matched control of pcDNA3 of the physiology brine. Kill toharm the effect along with the increment of the effect targetcomparison but increase. Express two kinds of DNA vaccines has thestronger immunity activity. Full long pcDNA3-gB and truncated shortpcDNA3-gB to promote the rat spleen cell the cell factor IL-2 and theIFN –r mRNA express. Truncated pcDNA3-gB compares withwhole long pcDNA3-gB, the result is better in stir up immunity. Several results are comprehensive and we can calculate, full longpcDNA3-gB and truncated pcDNA3-gB two gene vaccines canstrengthen the immunity dint of the small rat availably.And from fulllong pcDNA3-gB and truncated the comparison of two genes of... |
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