The Immune Function Of Exosomes Secreted By DF-1Cells Induced By ALV-J

Avain leukemia is a tumor infectious disease caused by avain leukosis virus (ALV) topoultry certain cellular component hyperplasa in hematopoetic tissue. Avain leukosis virus(ALV) is a group of enveloped retrvirus, can also cause body severe immunosuppressionresulting in a deline in productivity and frequent mixed infection, secondary infection, besidesdirect harm to the flocks. For a long time,due to the various of avain leukosis virus subgroups,the virsu itself speciality and the complexity of the disease, though both native and foreignscientists have done a lot of research, yet the immunosupressive mechanism is still not veryclear.The founding of Exosomes brings a good surprise to the research of avain leukosis virusimmunosupression, bridging the gap between avain leukosis virus and effector cells. Thefamous Trojan hypothesis said: retrovirus utilize host cell inherent Exosomes formation andrelease pathway to produce infectious viral particles, using Exosomes reuptake way forminganother infectious mode without depending on the envelope protein and non-receptormediating to infect other cells. In recent years, numerous studies have confirmed that somevirus really can infect normal cells with extracellular volume.To confirm ALV infected DF-1cells secrete Exosomes (Exo-J) containing some viralingredients, we analysis the extracted Exosomes protein by SDS-PAGE electrophoresis andWesternblot experiment after making it denaturation. We found that there is a differencebetween the Exosomes secreted by normal DF-1cells and ALV-J infected DF-1cells in about26KD protein. Cut off the strip from the plastic and disgest with enzyme, analysising byliquid phase and mass spectrometry, the result show that both Exo and Exo-J contain thecharacteristic proteins, such as HSP70, pyruvate kinase,14-3-3, actin and so on. HSP70protein family often used as a iconic protein to identify Exosomes. Exo-J contain the env,gag and pol protein of the ALV-J. Avain leukosis virus gag geneang pol gene have a high degree of homology among the subgroups. The gag gene encodesprotein27KDa,19KDa,15KDa,12KDa,27KDa protein is the mutual gs subgroup-specificantigen among all subgroups. The pol genes encode virus reverse transcriptase(RT) andintegrase(IN). The reverse transcriptase helps to complete the procession of the virus RNAwhich used as cell gene expression reverse transcripte into proviral DNA. The integrase isuesd for proviral DNA integrated into the cell chromosome. The envelope protein, as the envgene encodes the viral envelope protein glycosylation-protein, including membrane surfaceglycoprotein subunit(SU) and transmembrane glycoprotein subunits(TM). The SU containsvirus-receptor determinant, decide the subgroup speciality and host range. The TM mediatevirus and cell fusion process. The presence of of virus protein illustrates that ALV-J take someof Exosomes as its own membrane.In this study, in order to clarify the Exosomes and virus effects on lymphocytes, wedesign the experiment that Exosomes immune function on the spleen lymphocytes outside.Separating42days SPF chicken spleen lymphocytes, adding a dose of Exosomes and fosterappropriate time, detecting the antibody level of CD4+and CD8+. The result show that Exohas no immunosupression on lymphocytes and maintain at normal levels. Exo-J has apromotion on the CD4+and CD8+. But as the dose increase, the amount of CD4+and CD8+degrees, and is similar to the actual case, the more virus infecting body or cells, the moreExosomes secreted and the immunosupression effect is more obvious. For further study of theExo-J and ALV-J are immunosuppressive synergy design Exo-J and ALV-J on proliferation ofspleen lymphocytes in vitro.Added Exo-J and ALV-J to the isolated white blood cells of thespleen, according to the results separately to join the Exo-J (20μg), ALV-J group, CD4+,CD8+significantly below the control group, ALV-J group of inhibitory effects was stronger,but when both effects, their inhibition did not increase, but weaker than a single (ALV-J orExo-J) group, contrary to the expected result, This phenomenon may be due to competitiveExo-J and ALV-J in the same system with the same cell receptor-related, resulting inweakened immune suppression, this speculation further experiments are needed for authentication.Research has shown that Env protein specially integrate with cell surface receptor is thekey contion of the virus entering the cell, replicating and inducing tumor. Env protein play abig role in infecting host cells, also the virus specific receptor in the infected cells can beclosed by the virus producing gp85protein, which can limit reinfection. This anti-viral actionaccelerate the variation of ALV, and broaden the host range. In this immune selectingmechanism much related to the Env protein,the pathogenicity of the ALV will be stronger andstronger. Previous studies also that Env protein is closely related with the oncogenic type ofALV. The mass spectromety show that Exo-J contain the Env protein of ALV-J, and show theinterference effect on immune cell among the Exosomes effecting experiment. This may bethe action of Env protein carried by Exo-J. Therefore,it is the research direction of the futureto determine the immunosuppressive gene region.In summary, the study on the extraction of Exosomes have been improved, Exosomesstudies of the physical properties and biological activity, find Exo-J carrying gag and envproteins of ALV-J, immune cells has an obvious immune suppression, immunosuppressionmay play a decisive role in the process of ALV-J, Interaction between Exo-J and ALV-J in thehigh-dose and found immune rejection when you weaken the immune cells, this may beimpossible to bring the Exo-J to the in vitro system remote function results.