Studies On The Responses Of IL-10，IL-12and TNF-α After CSFV Immunized Pigs Were Inoculated With PRRSV Nsp2Δ1882-2241Attenuated Vaccine

Porcine reproductive and respiratory syndrome (PRRS) is a fulminatinginfectious diseases now has become one of the major epidemics of the large-scale pigfarms, which characterized by reproductive disorder in sows and respiratory systemdiseases in piglets.Usually, the immunization against swine fever virus is inactive dueto the suppression of interfere with the immune system of pigs by PRRSV andresultantly, co-infection of PRRSV and classical swine fever virus often occurs, if outof controll in time, it may cause a large number of pig deaths and huge economiclosses.In order to study the identity of cellular immune response to HP-PRRSV(TJM-F92) attenuated vaccine in pigs with different CSFV immune responsiveness,in this study, double immune models were established with classical swine fever andporcine reproductive and respiratory syndrome vaccine using specific pathogen-freeexperimental animals，the dynamic changes of the three cytokines in serum andPBMC culture supernatant at different times after the two vaccination were assayed.Designing the following test, Fifty-two blood samples of the3-week-old weanedclinical health pigs from one PRRSV negtive pig farm were collected to detect theantibodies against CSFV, PRRSV, pseudorabies virus (PRV), and porcine circovirus2(PCV2) using ELISA kits, PRRSV,CSFV, and PCV2were screened byRT-PCR. PRV gpI antibody were detected using ELISA.15healthy piglets wererandomly selected from22ultimately finalized free of PRRSV, CSFV, PRV andPCV2and were individually housed in sterile animal facilities and animal feeds andtools used have been approved by IACUC, then inoculated with swine fever livevaccine at the25th day after birth.All the pigs inoculated with1mL (106TCID50/mL)HP-PRRSV (TJM-F92) attenuated vaccine at the14thday after CSFV inoculation.The serum and PBMC were separated from pigs anterior vena cava blood respectivelyat the14th and28th day after PRRSV inoculation. CSFV, PRRSV antibody and the levels of IL-10, IL-12p40, TNF-α in serum were measured by ELISA. Supernatantswere collected after PBMC stimulated with PHA in vitro for24h, using competitiveinhibition ELISA determination of the concentration of IL-10, IL-12p40, and solidphase sandwich ELISA determination of the concentration of TNF-α. The test resultsof CSFV antibody-positive group called the CSFV high antibody group, CSFVantibody-negative as the CSFV low antibody group, then the test animals weregrouped according to the PRRSV antibody detection result. The CSFV high antibodygroup was divided into PRRSV high antibody group and PRRSV low antibody group,CSFV low antibody group is also divided into the group of PRRSV high-antibody andthe low-PRRSV antibody.Using ANOVA (One-Way ANOVA) analysis and the minimum significantdifference (LSD) to compare concentration differences of the cytokine in serum andPBMC culture supernatant between different antibody groups.The results show thatPRRSV antibody positive rate from6.67%at the14thday rising to80%at the28thday;differences of IL-10, IL-12p40, TNF-α in each group were not significant at the14thday and the28thday after vaccination(p>0.05); the concentration of IL-10in serumand PBMC culture supernatant at the28thday was significantly lower than at the14thday (p <0.01) in CSFV high antibody group after vaccination, however, the levels ofIL-12p40was significantly higher than the levels of14thday(p<0.01);theconcentration of TNF-α in serum and PBMC culture supernatant at28thday washigher than the levels of14thday (p <0.05).The results show that the cellular immune response in most of CSFV immunepigs after inoculated with PRRS nsp2Δ1882-2241attenuated vaccine is normal andgood immune status of CSFV can help the host to produce a good cellular immuneresponse to PRRSV attenuated vaccine; there has the response of cytokines in the casethat humoral immunity of PRRS does not reach the level of protection at the14thday,indicating that the PRRS cellular immunity generated earlier than humoral immunity;PRRSV antibody levels is high and positive rate is80%at the28thday, so it can beused as a monitoring time of the PRRSV (TJM-F92) vaccine antibody levels andPRRSV antibody levels at the28thday are more appropriate than the14d in evaluation of the immune effect of vaccine; both the level of IL-12p40significantlyincreased and IL-10down-regulated were an important mechanisms of PRRSV genedeletion vaccine. In addition, TNF-α also play a role in vaccine immune response.In this study, we first established experimental animal models inoculated withHP-PRRSV (TJM-F92) attenuated vaccine in non-specific status (immune swinefever vaccine). Analyzing the response characteristics of cytokines to PRRSV-specificin different CSFV immune status to find out the correlation of these cytokines inPRRSV immune process and provide a scientific basis for the further study of PRRSpathogenic law, immune regulation and cellular immune response mechanism.