Multiple intestinal 'loops' provide an in vivo model to analyse multiple mucosal immune responses

Mucosal immunity plays an important role in preventing disease but the induction of protective mucosal immune responses remains a significant challenge. We describe a novel in vivo model to analyze the induction of multiple mucosal immune responses in the small intestine. A sterile segment of intestine ('intestinal-segment'; 2-3 m long) was surgically prepared in the jejunum of 4-6-month-old lambs. This 'intestinal-segment' was then subdivided into consecutive segments, designated as 'loops' (15-20 cm long), that included a Peyer's patch (PP), or 'interspaces' (15-70 cm long), that lacked a visible PP. All 'loops' were sterile when collected 1-4 weeks post-surgery and there was no macroscopic or histological evidence of altered lymph or blood flow. Flow cytometric analysis of cells isolated from PP, mucosal epithelium (IEL) and the lamina propria (LPL) revealed no significant alterations in the cell populations present in 'loop' tissues. The functional integrity of M-cell antigen uptake in sterile intestinal 'loops' was evaluated by comparing the immune response induced by varying doses of soluble versus particulate porcine serum albumin (PSA formulated in alginate microspheres). A dose-dependent, PSA-specific antibody-secreting cell response was restricted to PP present in 'loops' injected with particulate PSA. These observations suggested that PP present in sterile 'loops' were functional and this conclusion was confirmed by detecting cholera toxin-specific antibody-secreting cells and secreted antibody in PP and intestinal contents, respectively, of immunized 'loops.' Thus, each 'loop' provided an independent site to analyze antigen-uptake and the induction of mucosal immune responses by a variety of antigen or vaccine formulations.