Staphylococcus (S). aureus is a leading cause of infectious keratitis associated with extended wear of contact lenses, accounting for approximately one-quarter of confirmed cases, with a gradual increase recently in the number of S. aureus keratitis in both the US and China. Under normal condition, the cornea is highly resistant to infection despite its constant exposure to a wide array of microorganisms. To cause infection, the opportunistic bacteria such as S.aureus are likely to first confront the epithelium with compromised barrier such as that caused by contact lens wearing. The epithelial cells that form the first line of defense against microbial pathogens, in turn must be able to recognize and respond appropriately to the invading pathogen. It is now well-established fact that recognition of invading microorganisms by epithelial as well as other mammalian cells includes the action of the Toll-like receptor (TLR) family. It plays a central role in early innate immunity by sensing the presence of microbial pathogens and initiating a response to eliminate them. These receptors recognize the highly conserved structural motifs called pathogen-associated molecular patterns (PAMPs), Bacterial lipoproteins (LP) are one of the PAMPs, they are a family of cell wall components found in a wide variety of bacteria.
In this study, we used Triton X-114 extraction method prepared LP isolated from S. aureus. S. aureus LP(saLP), then used it to stimulate HUCL cell, a telomerase -immortalized human corneal epithelial cell (HCEC) line. Our result showed that S. aureus LP(saLP) prepared by Triton X-114 extraction stimulated the activation of NF-κB, JNK, and P38 signaling pathways in HUCL cells. In HUCL cells after lipoprotein lipase treatment, or in human embryonic kidney (HEK) cell lines expressing functional TLR9, or in Hela cells expressing functional TLR4, the extracts failed to stimulate NF-κB activation, indicating lipoprotein nature of the extracts. saLP induced the up-regulation of a variety of inflammatory cytokines and chemokines (IL-6, IL-8, ICAM-1) and antimicrobial molecules (hBD-2, LL-37, and iNOS), and homeostasis genes (Mn-SOD) at both the mRNA level and protein level. Similar inflammatory response to saLP was also observed in primarily cultured HCECs using the production of IL-6 as readout. Moreover, TLR2 neutralizing antibody blocked the saLP-induced secretion of IL-6, IL-8 and hBD2 in HUCL cells. Our findings suggest that saLP activates TLR2 and contributes to innate immune response in the cornea to S. aureus infection via production of proinflammatory cytokines and defense molecules.