Effect of Toll-like receptor 2 and 4 of corneal fibroblasts on cytokine expression with co-cultured antigen presenting cells

Keratocytes are the first component to contact ocular pathogens when the epithelial barrier breaks down and the emerging evidences indicated keratocytes appeared to be one of the corneal cellular immune components. Little is known about the role of Toll-like receptors (TLRs) in keratocytes, although it has been well documented that keratocytes constitutively express various TLRs including TLR2 and TLR4. In this in vitro study, the authors focused on the role of keratocytes in corneal innate immune system and cross-talk of keratocytes with resident antigen presenting cells (APCs), especially through TLR2 and TLR4. Primary cultivated keratocytes (corneal fibroblasts) from C57BL/6 mice per se actively secreted pro-inflammatory cytokines, especially interleukin (IL)-6, with a dose-dependent manner in response to Pam3CSK4 or lipopolysaccharide (LPS) challenge. With co-culture of corneal fibroblasts with APCs per se, secretion of IL-6 and tumor necrosis factor (TNF)-α was markedly increased and it was counterbalanced by concurrent increase in IL-10 and tumor growth factor-β1. After Pam3CSK4 or LPS stimulation, this cytokine balance was completely broken down by overwhelming amplification of IL-6 and TNF-α secretion, especially in co-culture of corneal fibroblasts with macrophages, rather than with dendritic cells. Using corneal fibroblasts from TLR2 or TLR4 knockout mice, we could find the reversal of Pam3CSK4 or LPS-responsive dose-dependent increment in IL-6 and TNF-α. These results implied that corneal fibroblasts and their TLRs could be key components for the ocular homeostasis and pathogen-associated ocular innate immunity.

► Corneal fibroblasts secrete pro-inflammatory cytokines after TLR2/TLR4 stimulation. ► Corneal fibroblasts with APCs per se significantly increase IL-6/TNF-α secretion. ► TLR2/TLR4 stimulation amplifies this IL-6/TNF-α secretion with dose-dependent fashion. ► TLR2/TLR4 knockout reverses TLR-responsive IL-6/TNF-α secretion pattern. ► Corneal fibroblast is a innate immune cell, whose role is best played with APCs.