| Porcine reproductive and respiratory syndrome (PRRS) is caused by porcine reproductive and respiratory syndrome virus(PRRSV) that is characterized by Reproductive failure in pregnant sows and respiratory problems of piglets and growing pigs.PRRSV is apt to vary and is a small, enveloped, positive-sense single stranded RNA virus which can cause persistent infection and immune suppression, it is provided with antigenic variability、macrophage tropism、antibody-dependent enhancement (Antibody-dependent enhancement, ADE) and so on. PRRSV invaded alveolar macrophages mainly, which is related to specific virus receptors on porcine alveolar macrophages. Receptor is the primary gateway to viruses, in the porcine alveolar macrophages there has been confirmed with three kinds of receptors present: sialoadhesin (Sialoadhesin, Sn), heparan sulfate (Heparin sulphate, HS)and the CD 163 molecule. HS assists PRRSV to adsorb onto PAM; the Sn-mediated endocytosis of PRRSV and adsorption;CD 163 molecule helps Sn annexation and makes PRRSV envelope and the cell membrane fusion so as to establish the channel to help PRRSV undressing shell.the viral genome RNA is released into the cytoplasm. Combination of PRRSV and Sn is necessary for endocytosis.This study first used online software to predict pigs sialoadhesin receptor Sn physicochemical properties, transmembrane signal peptide, peptide hydrophilic, antigenicity index, and the possibility of flexible surfaces, domains and motifs. Predicting the possibility of prokaryotic expression of Sn in vitro, in the case of considering swine sialoadhesin domain, pig Sn is expressed prokaryoticly by section, the corresponding polyclonal antibodies were prepared, using DEAE-cellulose ion-exchange chromatography to purify serum immunoglobulin IgG.To determine binding domain of the pig sialoadhesin receptor Sn and PRRSV, this study obtained porcine alveolar macrophages in vitro, and used murine anti-Sn IgG which was prepared and purified to close and dispose PAM cells respectively. PRRSV infected the cells 24h and 48h,then detect cell proliferation of the virus by established SYBR Green Ⅱ real-time quantitative PCR method.The results showed that, compared to PAM cells which were closed by the negative control group of rats IgG, the effect of the closure of each segment structual domains on Inhibition of virus proliferation varies after Inoculating with virus 24h and 48h, inhibition of the joint closure domains differ significantly, individual closed domains slightly decreased, Inhibition of the 9th,10th and 13th segment structual domains were obvious in 17 structual domains of Sn receptor predicted by software. It shows the integrity of the structure of sialic acid was crucial to viral adsorption, structual domains of the separate closed portion had also inhibiting effect. It suggested that this part of the domain may have its unique features, or may be associated with adherence of virus, but repeated trials and research is more needed.On this basis, we were prepared with different concentrations of infectious immune complexes, groping whether Sn receptors or CD 163 play a role in immune complexes infecting body, when the infectious immune complexes were four times diluted, Sn receptor N domain has little effect on the increasing value of immune complexes; mixed closure domains inhibited the increasing value of infectious immune complexes; when the infectious immune complexes were two times and four times diluted, the virus copies of the mixed closure domains group decreased compared to the control group and the difference was significant;when infected immune complexes were 8-fold diluted, closed Sn group was slightly higher compared with the control group, but the difference was not significant; compared with the control group, Inhibition role of virus proliferation compared with the inhibition role of separately enclose Sn or CD 163 was more apparent 48h later. The effect is not simply sum of Sn It indicating that the role of infectious immune complexes on body can be adjusted by Sn receptor to some extent, Sn receptor and CD 163 help the infectious immune complexes get into cells.Reported the possibility of different species about PRRSV into cells that is mediated by Sn receptor, the conclusion show that except pig Sn receptor, mouse and human Sn receptor also have the ability to bind PRRSV. PRRSV antigen-antibody double-negative pigs in the experiment is not easy to obtain, PRRSV may be lactate dehydrogenase increased disease virus (LDV) mutants obtained from hog through the intermediate host infected wild pigs, this study conducted a discussion on PRRSV infection in a mouse model:Part of mice immunized directly concentrated purified PRRSV, another part of mice immunized with vaccine firstly to make the body produce antibodies against PRRSV, then concentrate virus, collect serum, spleen, lungs of mice in different time periods post-infection.After PRRSV RNA extraction and nucleic acid detection. The results showed that PRRSV just absorb on rat alveolar macrophages but not infection. Indicating that when PRRSV invade the host cell, Sn receptors play a role in mediating endocytosis of the virus, but whether the viral infected host or not need other receptors participated in. |
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