Evaluating the innate immune response of the porcine uterus to vaccine adjuvants for the development of an intrauterine vaccine

Abstract

Mucosal vaccination is a widely sought after method of vaccination for both its ease of delivery and because it can stimulate a strong local mucosal immune response. As the vast majority of pathogens enter the body through mucosal surfaces, a strong mucosal immune response has the potential to eliminate pathogens before them crossing the epithelial barrier, and can, in the case of the uterus, protect the developing fetuses from pathogens. For a mucosal vaccine composed of inactivated antigens to be effective, it must be formulated with potent adjuvants that will initiate a strong immune response in the uterus. This thesis aimed to evaluate the capacity of uterine epithelial cells to respond to vaccine adjuvants and to subsequently initiate and modulate the local uterine immune response of an intrauterine vaccine in sows. Because uterine epithelial cells (UECs) are the first point of contact for an intrauterine vaccine, we established an in vitro model using primary uterine epithelial cells to enable us to evaluate what receptors are present on these cells that could be targeted by the adjuvants in an intrauterine vaccine. Previous research showed that a triple adjuvant combination including polyI:C, a host defense peptide (HDP) and polyphosphazene (PCEP) administered as part of a mucosal vaccine induced strong humoral immunity. We tested these adjuvants as well as muramyl dipeptide and LPS alone and in combination with each other in the in vitro system. In vitro stimulations resulted with polyI:C alone and in combination with other potential adjuvants induced significant pro-inflammatory cytokines and chemokines, which was not further improved by the inclusion of HDP and PCEP. To determine if the observed response in vitro resulted in similar responses in vivo, the triple adjuvant combination was administered in vivo into the uterus of sows in combination with a standard semen dose. Twenty four hours post-breeding, epithelial cells had significantly increased CCL2 expression in response to the adjuvants combined with semen compared to the response generated to semen alone. Additionally animals bred with adjuvants included in the semen dose had a significant decrease in the proportion of γδ T cells and monocytes in the blood, however, there was no significant increase of these cells in the lumen indicating the γδ T cells were not being recruited into the uterine lumen. Following evaluation of infiltrated monocytes and macrophages in the tissue, no significant difference was observed in these cells levels compared to animals bred with semen alone. Lastly, returning to the in vitro UEC model, I took steps to quantify whether secretions from stimulated uterine epithelial cells impacted monocyte to DC differentiation or moDC activation. Secretions from triple adjuvant-stimulated uterine epithelial cells were not able to induce recruitment of monocytes nor did the secretions impact on the moDCs capacity to endocytose antigen, mature in response to stimulation or induce proliferation in T cells. The established primary UEC culture system established and characterized was observed to closely resemble the behaviour of UECs in vivo for studying the responses of adjuvants and could be further utilized for continuing the develop UECs role in the uterine immune response to stimulants and pathogens. Although uterine epithelial cells respond to vaccine adjuvants, the magnitude of this response is unable to significantly impact the immune response to vaccine adjuvants delivered by the intrauterine route in a semen dose.