Upregulation of TNF-α Production by IFN-γ and LPS in Cultured Canine Keratinocytes: Application to Monosaccharides Effects

Abstract

Activated keratinocytes play a key role in the cutaneous immune system by their interactions with other cell types through the production of cytokines with both autocrine and paracrine activity. But there is little knowledge about epidermal cytokines in the dog. In this study, cultured canine keratinocytes were stimulated by human recombinant interferon gamma (IFN-gamma) and lipopolysaccharide (LPS) and cell supernatants were tested for tumour necrosis factor alpha (TNF-alpha) concentration using a cell viability assay on a murine cell line. We show that IFN-gamma in combination with LPS significantly increases TNF-alpha secretion by canine keratinocytes. The best stimulations were obtained using confluent cultures and the association of IFN-gamma (400 ng/ml) and LPS (40 microg/ml). The experimental protocol we describe represents a new method for studying keratinocyte activation and its modulation in the dog. We provide an example of application of our method: the study of the effects of different monosaccharides on canine keratinocyte activation.

Adjuvanting Allergen Extracts for Sublingual Immunotherapy: Calcitriol Downregulates CXCL8 Production in Primary Sublingual Epithelial Cells

“…In the past, TLR2 ligands were shown to be promising adjuvants for allergy treatment in some studies (16)(17)(18), whereas other studies with mouse asthma models showed that TLR2 ligands could enhance proallergenic responses (40,41). In canine epidermal keratinocytes, LPS and Pam3CSK4 also promoted the secretion of proinflammatory and proallergenic mediators (42)(43)(44). Additionally, the TLR3 ligand, poly I:C, had a significant lethal effect on the sublingual epithelial cells, as was observed for bronchial epithelial cells as well as in epidermal keratinocytes (45,46).…”

Current understanding of the pathophysiologic mechanisms of canine atopic dermatitis

“…Canine keratinocytes have already been shown to produce pro-inflammatory agents such as granulocyte macrophage colony-stimulating factor, IL-8, and tumor necrosis factor-α in response to allergen and bacterial components such as Der f1 and lipopolysaccharide, respectively. 97,98 An additional keratinocyte cytokine of great interest that has been implicated in AD in humans includes thymic stromal lymphopoietin. Thymic stromal lymphopoietin is an IL-7-like cytokine that stimulates dendritic cells to induce naïve T cells to differentiate into Th2-like cells.…”

A review of the roles of keratinocyte‐derived cytokines and chemokines in the pathogenesis of atopic dermatitis in humans and dogs

“…Microarray analysis demonstrated that treatment of human keratinocytes with TNF‐alpha induced transcription of genes associated with not only immune and inflammatory responses, but also with tissue remodelling, cell motility, cell cycle regulation and apoptosis, suggesting that TNF‐alpha has a multifunctional effect on keratinocytes . In dogs, production of TNF‐alpha was observed in keratinocytes stimulated with IFN‐gamma and LPS . One study has reported that activation via PAR‐2 increased the transcription of tnf‐alpha in CPEK .…”

“…57 In dogs, production of TNF-alpha was observed in keratinocytes stimulated with IFN-gamma and LPS. 58 One study has reported that activation via PAR-2 increased the transcription of tnf-alpha in CPEK. 40 TNF-alpha was also shown to induce transcription of CC chemokine ligand (CCL) 17 and CCL28 in CPEK, suggesting that canine keratinocytes likely express TNF receptor, similar to human keratinocytes.…”