Previously, we demonstrated that Lactobacillus jensenii TL2937 attenuates the inflammatory response triggered by activation of Toll-like receptor 4 (TLR-4) in porcine intestinal epithelial cells. In view of the critical importance of antigen-presenting cell (APC) polarization in immunoregulation, the objective of the present study was to examine the effect of strain TL2937 on the activation patterns of APCs from swine Peyer's patches (PPs). We demonstrated that direct exposure of porcine APCs to L. jensenii in the absence of inflammatory signals increased expression of interleukin-10 (IL-10) and transforming growth factor β in CD172a+ APCs and caused them to display tolerogenic properties. In addition, pretreatment of CD172a+ APCs with L. jensenii resulted in differential modulation of the production of pro- and anti-inflammatory cytokines in response to TLR4 activation. The immunomodulatory effect of strain TL2937 was not related to a downregulation of TLR4 but was related to an upregulation of the expression of three negative regulators of TLRs: single immunoglobulin IL-1-related receptor (SIGIRR), A20, and interleukin-1 receptor-associated kinase M (IRAK-M). Our results also indicated that TLR2 has an important role in the anti-inflammatory activity of L. jensenii TL2937, since anti-TLR2 antibodies blocked the upregulation of SIGIRR and IRAK-M in CD172a+ APCs and the production of IL-10 in response to TLR4 activation. We performed, for the first time, a precise functional characterization of porcine APCs from PPs, and we demonstrated that CD172a+ cells were tolerogenic. Our findings demonstrate that adherent cells and isolated CD172a+ cells harvested from swine PPs were useful for in vitro study of the inflammatory responses in the porcine gut and the immunomodulatory effects of immunobiotic microorganisms.