Pam3Cys-SKKKK (Pam3Cys) is a synthetic lipopeptide, which acts as a Toll-like receptor (TLR) 2 agonist on cells of the innate immune system. Recent studies showed that TLRs are also expressed on cells of the adaptive immune system. The aim of this study was to investigate the effect of Pam3Cys on porcine lymphocytes. For this purpose, porcine peripheral blood mononuclear cells (PBMC) were stimulated with this lipopeptide in vitro and their proliferation was analyzed. Already after two days of in vitro culture strong proliferation of B cells was observed by measurement with Ki-67- and CFSE- fluorescence by flow cytometry. These B cells produced high amounts of IgM and IgG molecules as measured by ELISA. Analysis of TLR2 expression in porcine B cells demonstrated, that TLR2 molecules could be found neither extra- nor intracellular. In further experiments, monocytes and dendritic cells from PBMC were depleted by magnetic cell separation or pure B cells were obtained by FACS sorting. In vitro cultures from these purified B cells showed only a reduced proliferation and production of antibodies after stimulation with Pam3Cys. Therefore it seems that Pam3Cys demonstrates both, a direct as well as an indirect effect on porcine B cells.
In addition, it was investigated whether the porcine IgG antibodies were produced from memory B cells and whether we could induce a polyclonal pathogen-specific B cell memory response against pathogens which each pig has contacted before. However, pathogen-specific antibodies in cell culture supernatants of PBMC stimulated with Pam3Cys could neither be detected in fattened pigs from the slaughterhouse nor in pigs experimentally infected with PRRSV. In contrast porcine T cells did not show specific responses after stimulation with Pam3Cys, although this is described for human memory T cells in literature. TLR2 expression at protein level could not be detected in porcine T cells as well.
B cell / T cell / Toll-like receptor / Pam3Cys / porcine / lymphocytes / lipopeptide / immune system