The failure of rotavirus (RV) vaccines in infants underscores a need for more effective vaccination strategies and an improved understanding of the development of mucosal immunity to human rotavirus (HRV) in infants. High levels of maternal antibodies to RV in the neonate's circulation and in breast milk may interfere with successful immunization against RV and hinder assessment of active immunity to RV in vaccine trials. Thus, to facilitate RV vaccine development we will investigate the development of mucosal immunity to RV in neonates and delineate the effect of maternal antibodies on immunity to RV. Because of the difficulty in analyzing mucosal immune responses in infants, we will rely on the use of gnotobiotic (Gn) pigs for our studies. The neonatal Gn pig is an ideal animal because it is highly susceptible to HRV infection and disease, it resembles humans in gastrointestinal physiology and it is born without maternal antibodies. We will investigate the development of systemic and local (intestinal) antibody and cellular immune responses to oral inoculation with virulent HRV (mimic natural exposure) and live attenuated HRV (mimic oral vaccines) and parenterally administered HRV recombinant proteins (mimic subunit vaccines) in three experimental groups of Gn pigs: pigs devoid of maternal RV antibodies (immunologically virgin); pigs with circulating maternal RV antibodies (represent formula-fed infants); and pigs with maternal circulating and milk RV antibodies (represent breast- fed infants). Finally selected immunoenhancers (mucosal adjuvants, cytokines) will be evaluated for their ability to enhance the development of active immunity to HRV and to counter suppressive effects of maternal antibodies on active immune responses to HRV. Antibody responses to HRV and RV proteins (VP4 and VP7) in the experimental Gn pigs will be detected and quantitated by neutralization, isotype and epitope-blocking ELISA and ELISPOT assays. Cellular immune responses will be assessed by lymphoproliferation assays and cytokine profiles. Cytokines will be examined by ELISPOT, RT-PCR and northern blot with an emphasis on delineating Th1- and Th2-like cells. To interpret the results of lymphoproliferation and cytokine analysis, we will phenotype lymphoid cells from the intestine using monoclonal antibodies and flow cytometry. Our data should elucidate the components and tissue sites of the immune system affected by maternal RV antibodies and identify future targets for immunoenhancement of the neonatal immune response to oral vaccines.
Showing the most recent 10 out of 24 publications
