The Study Of Tat Protein Enhance HIV-1 Vaccine Immunity Level

AIDS (Acquired Immune Deficiency Syndrome) is caused by the HIV(Human Immunodeficiency Virus). The virus destroys the body'sability to fight infections and certain cancers. As the AIDS epidemiccontinues to spread across the world, it became the focus that thetreatment strategies of its infection and the prevention of its spreading.Chemotherapeutic has been limited to cost and the strict nature of theadministration regimens and high mutation rate in(maybe of) virus.These restrictions support that there is a need for the development ofeffective vaccine, and only by using vaccine can control HIV-1 spread.More and more evidences suggest that cytotoxic T lymphocyte(CTL) responses play an important role in the protection against HIVviruses. Several groups have observed that viral clearance wasassociated with specific CTL activity in acute cases of HIV-1 infection.HIV-1 specific CTLs were also found in a number of chronicallyexposed prostitutes in Gambia who continue to resist infection withHIV-1. Long-term non-progressor HIV-infected individuals, chroniclow virus load and is related with high frequency CTL. Cytotoxic Tlymphocytes (CTL) not only recognize processed viral fragmentspresented on the infected cell surface and, as a result, destroyvirus-infected cells, but also target all viral gene products that areexpressed during viral replication. Targeting viral proteins through thedevelopment of specific CTL responses could aid in lowering (help tolower) the viral load by interfering with viral assembly.DNA vaccine or nucleic acid immunization is an importantvaccination strategy that delivers DNA constructs encoding for aspecific immunogen into the host. Intramuscular injection of DNAvaccine leads to uptake of the plasmid by host cells and expression ofthe protein Antigen. The protein enters the Ag-processing pathways,resulting in strong and persistent humoral and cellular immuneresponse. This vaccination technique is being explored as animmunization strategy against a variety of infectious diseases,including HIV-1 infection. The immune responses elicited by DNA vaccination haveafforded a degree of protection in nonhuman primate againstnonpathogenic AIDS virus. However, these immune responses havenot been of a magnitude sufficient to protect against pathogenic viralchallenges. We therefore were interested in exploring strategies foraugmenting DNA vaccine-elicited immune response. Several studieshave shown that exogenous Tat protein was able to translocate throughthe plasma membrane and to reach the nucleus to transactivate theviral genome . A region of the Tat protein centered on a cluster ofbasic amino acids has been assigned to this translocation activity.Recent data have demonstrated that chemical coupling of aTat-derived peptide (extending from residues 49 to 57)-T9(RKKRRQRRR) to several proteins allowed their functionalinternalization into several cell lines or tissues. According to those reports, we design strategys that tat and T9fused to gag/pol of HIV-1 separately, then tat protein or T9 proteinhelp fusion proteins delivering into cells. The fusion protein enters theAg-processing pathways, resulting in strong and persistent humoraland cellular immune response. We successful constructed DNA vaccine, such as â‘ VR1012-gag/pol â‘¡VR1012-tat-gag/pol â‘¢VR1012-T9-gag/pol â‘£VR1012-tat and transfected eukaryote such as 293,COS7. Theexpression of target protein was examined and demonstrates thecorrectness of DNA vaccine construction. The protein expression inthe pure VR1012-tat group is lower than that in the VR1012-tat +VR1012-gag/pol, even we add the same weight VR1012-tat plasmid.The reason might be that the contransfection of VR1012-gag/pol andVR1012-tat affect the transfection Efficiency. Intramuscular injectionof DNA vaccine is performed on 6to 8 week old female Balb/c miceevery two weeks. We test the sera at various times followingvaccination. The humoral immune response was steadily enhanced astime went on. At the 10th week, the humoral immune response isstrong enough for the test, and the mice were given the recombinantMVA as boost. Mice were killed after boost 5 days, and the spleen...