Primary porcine CD11R1+ antigen-presenting cells isolated from small intestinal mucosa mature but lose their T cell stimulatory function in response to cholera toxin treatment

Antigen-presenting cells (APCs) in the small intestinal mucosa perform dual functions of maintaining tissue homeostasis and of protecting against intestinal pathogens as key inducers of both innate and adaptive immune responses. Intestinal APCs are thus important regulators of intestinal immunity and also potential target cells for mucosal adjuvants such as cholera toxin (Ctx), which was used successfully in several oral vaccination studies in pigs. The aims of the present study were (1) to isolate porcine small intestinal APCs and evaluate the feasibility of using these cells for functional in vitro studies and (2) to determine the response of intestinal APCs to Ctx. Microscopic and flow cytometric analyses using antibodies to CD1, CD11R1, CD16, and SIRPα (SWC3) revealed the presence of multiple subsets of MHC-II++ APCs in porcine small intestinal mucosa. The α-integrin subunit CD11R1 was most frequently expressed and therefore chosen as a selection marker. CD11R1+ cells were enriched from total lamina propria cells to >90% purity by immunomagnetic separation. Within the CD11R1 cells, we identified two populations with distinct forward and side scatter characteristics: (1) APCs identified by their high expression of MHC-II and consisting of SIRPα+ and SIRPα− subsets, and (2) contaminating eosinophils. In culture, intestinal APCs spontaneously matured, as shown by significant (>5-fold) increase in CD80/CD86 expression. The SIRPα+ APCs quickly disappeared from the cultures, likely due to increased apoptotic cell death. However, the observed spontaneous changes in the isolated cell population did not mask the effects of stimulation with Ctx, which resulted in a 2.5-fold increase in the expression of maturation markers CD80/CD86, but significant loss of T cell stimulatory function, corroborating previous results obtained with MoDC.