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Title:   Author  item
                                            Tuo, Wenbinitem
                                            DAVIS, W - WASHINGTON STATE PULLMANitem
                                            Fetterer, Raymonditem
                                            Jenkins, Markitem
                                            Boyd, Patriciaitem
                                            GASBARRE, L - USDA ARS BELTSVILLE MDitem
                                            Dubey, Jitender
  Submitted to: Parasite Immunology
                                    Publication Type: Peer Reviewed Journal
                                    Publication Acceptance Date: 2/3/2004
                                    Publication  Date: 3/8/2003
                                    Citation: Tuo, W., Davis, W., Fetterer, R.H., Jenkins, M.C., Boyd, P., Gasbarre, L.C., Dubey, J.P. 2003. Establishment of short-term cd4+ T cell lines recognizing Neospora caninum antigen. Parasite Immunology. 26:243-246.Interpretive Summary: Neosporosis causes pregnancy loss in cattle.  The presence of N. caninum-specific antibodies in circulation may not necessarily provide protection, suggesting neosporosis requires both cell-mediated and humoral immune response for effective protection. The objective of this study was to establish short-term CD4+ T cell lines from peripheral blood mononuclear cells of N. caninum-infected cows. These CD4+ T cells will be used to identify immunodominant Neospora antigens for the development of neosporosis vaccine. Crude N. caninum tachyzoite antigen was prepared from in vitro derived Neospora tachyzoites. Eight of 13 Neospora antigen immunized and N. caninum challenged cows exhibited high levels of proliferation and interferon-gamma production by PBMC in response to antigen stimulation and therefore, some of these cows were used as blood donors to establish CD4+ T cell lines. Multiple Neospora antigen-specific CD4+ T cell lines were established and maintained continuously for 11 weeks in culture by weekly re-stimulation with Neospora tachyzoite antigen and antigen-presenting cells. All cell lines responded highly to antigen stimulation between weeks 3 and 11. Flow cytometric analysis showed that these T cells were composed of primarily CD4+ T cells between week 2 and 8 with a gradually expansion of gamma/delta+ T cells. The results indicate that Neospora antigen-specific CD4+ T cell lines can be established and maintained without the presence of T cell growth factors and may be used to identify immunodominant Neospora antigens.  This research will enhance our understanding of bovine cell-mediated immunity to neosporosis and may facilitate development of a proven neosporosis vaccine.
                                Technical Abstract: Neosporosis causes pregnancy loss in cattle.  The presence of N. caninum-specific antibodies in circulation may not necessarily provide protection, suggesting neosporosis requires both cell-mediated and humoral immune response for effective protection. The objective of this study was to establish short-term CD4+ T cell lines from peripheral blood mononuclear cells of N. caninum-infected cows. These CD4+ T cells will be used to identify immunodominant Neospora antigens for the development of neosporosis vaccine. Crude N. caninum tachyzoite antigen was prepared from in vitro derived Neospora tachyzoites. Eight of 13 Neospora antigen immunized and N. caninum challenged cows exhibited high levels of proliferation and interferon-gamma production by PBMC in response to antigen stimulation and therefore, some of these cows were used as blood donors to establish CD4+ T cell lines. Multiple Neospora antigen-specific CD4+ T cell lines were established and maintained continuously for 11 weeks in culture by weekly re-stimulation with Neospora tachyzoite antigen and antigen-presenting cells. All cell lines responded highly to antigen stimulation between weeks 3 and 11. Flow cytometric analysis showed that these T cells were composed of primarily CD4+ T cells between week 2 and 8 with a gradually expansion of gamma/delta+ T cells. The results indicate that Neospora antigen-specific CD4+ T cell lines can be established and maintained without the presence of T cell growth factors and may be used to identify immunodominant Neospora antigens.  This research will enhance our understanding of bovine cell-mediated immunity to neosporosis and may facilitate development of a proven neosporosis vaccine.


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