Epidemiologic Survey Of Bovine Viral Diarrhea In Beijing Region, Study Of IFN-α/β Response In Persistently Infected Catttle And Immunomodulatory Effects Of Forsythoside A

To determine the epidemiological characterization of bovine viral diarrhea virus (BVDV) and identify cattle persistently infected (PI) with the virus, a total of 4,327 samples of Holstein dairy cows were screened over a three-year period in Beijing region of China. All samples were tested for BVDV antigen and genome using antigen capture enzyme-linked immunosorbent assay (ELISA) and reverse transcription-PCR (RT-PCR). Eighteen BVD viruses were isolated and 12 isolates were from PI cattle. In addition,1,000 serum samples were randomly selected and tested for BVDV antibodies and the prevalence of BVDV antibody positive animals was 93.4%. Genetic analysis of these isolates in 5’-UTR regions revealed that 15 isolates were assigned to the BVDV-lm (6 isolates), BVDV-la (5 isolates), BVDV-1d (3 isolates), and BVDV-lb (1 isolate). And the other 3 isolates were clustered into putative ’BVDV-lq’.Moreover, to investigate the innate immune responses in PBMCs of PI cattle,6 clinically normal PI cattle and 6 healthy cattle were selected. The mRNA expression of interferon-a (IFN-a), IFN-β, myxovirus (influenza virus) resistance 1 (Mxl), Toll-like receptors (TLRs), RIG-I-like receptors (RLRs), and protein expression of interferon regulatory factor (IRF) 3 and IRF-7 in PBMCs were determined. The mRNA expression of IFN-a, IFN-β,and TLR-7 decreased in PBMCs from PI cattle. Furthermore, decreased IFN-α/β mRNA expression correlated with the decreased expression of IRF-3 and IRF-7. Taken together, these results reveal that PI cattle lead to the widespread infection of BVDV in Beijing region and ncpBVDV can inhibit the expression of IFN-α/β in PBMCs in PI cattle which are associated with down-regulation of TLR-7 and IRF3/7. Our results suggest a role for TLR-7 and IRF-7 in ncpBVDV-mediated evasion of type I IFN response.Here we reported that Forsythoside A (FTA), a constituent of the Forsythia suspense Vahl, had the potential role in enhancing 4-1BB and OX40 signals in BVDV-infected bovine peripheral blood mononuclear cells (PBMCs). The PBMCs mRNA expressions of CD28, cytotoxic T-lymphocyte antigen 4 (CTLA-4),4-1BB,4-1BBL, and OX40 were investigated and we found FTA could enhance the 4-1BB,4-1BBL and OX40 mRNA expression following virus infections. Moreover, the 4-1BB expression was determined by western blot. Sequentially, their adaptors TRAF-1 (tumor necrosis factor receptor related factor 1) and TRAF-2 and down-stream apoptotic factors Bcl-xL, Bim were also monitored. It is seems that FTA could affect the four factors’ genes transcription levels except TRAF-1. Moreover, the production of IFN-y and IgG2a in supernatant of cell cultures was detected by ELISA and the results indicated that FTA could ameliorate the immune homeostasis for bovine PBMCs infected with BVDV. Therefore, these data point to an unrecognized role of FTA in modulation of costimulatory members of 4-1BB and OX40 in BVDV-infected bovine PBMCs.