Expression of pattern recognition receptors in porcine uterine epithelial cells in vivo and in culture

Preservation of a pathogen free uterine environment is critical for maintaining healthy swine herds with high reproductive performance. Considering that uterine epithelial cells are the most numerous and thus likely point of cellular contact for pathogens in the uterus, we hypothesize that these cells may be critical for activating the immune system to clear uterine infections. Although uterine epithelial cells have not been well characterized in pigs, studies in several other species have shown that these cells express several pattern recognition receptors (PRR) and thus may act as sentinels for the uterine immune response. To characterize PRR expression in the porcine uterine epithelia, we used laser-capture microdissection to isolate epithelial cells lining the porcine uterus to quantify in vivo mRNA expression levels for select PRRs. As well, primary uterine epithelial cells (UECs) were isolated, cultured, polarized and PRR expression was quantified. Immunohistofluorescence and immunofluorescence were used to determine subcellular localization of TLR3, TLR4 and TLR9 in both uterine tissue and in polarized primary UECs. Finally, polarized primary UECs were stimulated with ligands for TLR3, TLR4, TLR9 and NOD2 to determine their functional innate immune response. Uterine epithelial cells (in vivo and in vitro) were shown to express TLR1-7, TLR9, NOD1, NOD2, NLRP3, NLRP6, NLRX1, RIG1, MDA5 and LGP2. Subcellular localization of in vivo and polarized primary UECs exhibited TLR3 and TLR9 localized to the apical cell surface whereas TLR4 was localized to the intracellular space. Polarized primary UECs stimulated with TLR3, TLR4 and TLR9 ligands showed induced secretion of IL-6, IL-13 and IL-10, respectively indicating that these receptors were functional. These results indicate that pig uterine epithelial cells are functional innate immune cells that may act as sentinels to protect against uterine infection.