Effects of Astragalus Polysaccharide on Immune Responses of Porcine PBMC Stimulated with PRRSV or CSFV

BACKGROUND: Astragalus polysaccharide (APS) has been used as an immunomodulator that can enhance immune responses, whereas the immunomodulatory effects of APS on porcine peripheral blood mononuclear cells (PBMCs) exposed to porcine reproductive and respiratory syndrome virus (PRRSV) and classical swine fever virus (CSFV) have not been investigated. METHODOLOGY/PRINCIPAL FINDINGS: Porcine PBMCs were cultured in complete RPMI media in the presence of the R98-strain of PRRSV (5×10(4) TCID(50)/ml) or C-strain of CSFV (10(3) TCID(50)/ml) with or without APS. The expression of mRNA for CD28, cytotoxic T-lymphocyte antigen 4 (CTLA-4), transforming growth factor-β (TGF-β), interleukin 2 (IL-2) and IL-10 was assayed by TaqMan real-time RT-PCR. The expression of mRNA for CD28 and CTLA-4 increased at 24 h after stimulation of PBMCs with CSFV and the increased production of CTLA-4 was confirmed by western blot analysis, whereas the increases were inhibited by the addition of APS. In addition, APS alone upregulated IL-2 and TGF-β mRNA expression in PBMCs and the addition of APS had the capacity to prevent a further increase in IL-2 mRNA expression in PBMCs during CSFV or PRRSV infection, but had no effect on TGF-β mRNA expression. The production of tumor necrosis factor-alpha (TNF-α) increased at 12 h after stimulation with PRRSV or CSFV, but not with PRRSV plus APS or CSFV plus APS, whereas the addition of APS to PBMCs infected with PRRSV or CSFV promoted IL-10 mRNA expression. CONCLUSIONS: We suggested that APS had immunomodulatory effects on cells exposed to PRRSV or CSFV. It might be that APS via different mechanisms affects the activities of immune cells during either PRRSV or CSFV infection. This possibility warrants further studies to evaluate whether APS would be an effective adjuvant in vaccines against PRRSV or CSFV

Rapid Evolution of the CD8(+) TCR Repertoire in Neonatal Mice

Currently, there is little consensus regarding the most appropriate animal model to study acute infection and the virus-specific CD8(+) T cell (CTL) responses in neonates. TCRb high-throughput sequencing in naive CTL of differently aged neonatal mice was performed, which demonstrated differential V beta family gene usage. Using an acute influenza infection model, we examined the TCR repertoire of the CTL response in neonatal and adult mice infected with influenza type Avirus. Three-day-old mice mounted a greatly reduced primary NP(366-374)-specific CTL response when compared with 7-d-old and adult mice, whereas secondary CTL responses were normal. Analysis of NP(366-374)-specific CTL TCR repertoire revealed different V beta gene usage and greatly reduced public clonotypes in 3-d-old neonates. This could underlie the impaired CTL response in these neonates. To directly test this, we examined whether controlling the TCR would restore neonatal CTL responses. We performed adoptive transfers of both nontransgenic and TCR-transgenic OVA((257-264)-specific) (OT-I) CD8(+) T cells into influenza-infected hosts, which revealed that naive neonatal and adult OT-I cells expand equally well in neonatal and adult hosts. In contrast, nontransgenic neonatal CD8(+) T cells when transferred into adults failed to expand. We further demonstrate that differences in TCR avidity may contribute to decreased expansion of the endogenous neonatal CTL. These studies highlight the rapid evolution of the neonatal TCR repertoire during the first week of life and show that impaired neonatal CTL immunity results from an immature TCR repertoire, rather than intrinsic signaling defects or a suppressive environment

Validation of Abnormal Self-Antigens as Targets for Tumor Rejection by CAR T Cells

Aberrant posttranslational modifications of cellular proteins represent a broad repertoire of putative tumor-specific targets. In general, vaccines targeting these aberrant self-antigens have only generated modest immune responses. In contrast, genetically modified T cells that express chimeric antigen receptors (CARs) demonstrate robust responses against associated targets, and have been clinically effective in treating hematologic cancers. However, in solid tumors the full potential of CAR T cell therapy is limited by the availability of cell surface antigens with sufficient cancer-associated expression to which single-chain variable fragments (scFvs) can be designed. Thus, the majority of CAR targets to date have been normal self-antigens on dispensable hematopoietic tissues or overexpressed shared antigens. Here, we validate abnormal self-antigens as targets for tumor rejection through the use of a novel CAR. We targeted the cancer-associated Tn glycoform of MUC1 in a variety of cancers. The anti-Tn-MUC1 CAR demonstrated target-specific cytotoxicity and successfully controlled tumor growth in xenograft models of T cell leukemia and pancreatic cancer. These finding demonstrate the therapeutic efficacy of CAR T cells directed against abnormal self-antigens and the potential for targeting tumor-specific glycoproteins in future cancer immunotherapies