Proinflammatory cytokine responses of cultured equine keratinocytes to bacterial pathogen-associated molecular pattern motifs.

The research paper explores equine keratinocyte responses to different bacterial molecular patterns, with its findings indicating that these keratinocytes respond more to Gram-negative bacterial patterns compared to Gram-positive ones.

Objective

The objective of the study was to investigate how equine keratinocytes react to variations of pathogen-associated molecular pattern molecules (PAMPs) from both Gram-negative and positive bacteria. Furthermore, the study analyses cytokine response to these PAMPs to better understand horse skin diseases.

Methodology

• Keratinocytes, a type of skin cell, were isolated from two horses and cultivated in a controlled environment.
• These cells were then subjected to PAMPs associated with specific types of bacteria, namely lipopolysaccharide (LPS), lipoteichoic acid (LTA), peptidoglycan (PGN), bacterial DNA (CpG), and flagellin.
• After exposing the cells to these PAMPs for a duration of 4 or 24 hours, cells were collected for RNA extraction.
• The concentration of three important inflammatory cytokines – interleukin-1beta (IL-1beta), interleukin-6 (IL-6), and CXCL8 – was assessed using real time-quantitative PCR.

Results

• The study reveals that LPS exposure leads to increased concentrations of IL-1beta, IL-6, and CXCL8 mRNA, while flagellin exposure results in elevated IL-6 and CXCL8 mRNA levels compared to the control group.
• The responses were overall more significant after 4 hours of exposure.
• No increases in cytokine expression were observed after exposure to LTA, PGN, or CpG, suggesting an absence of functional TLR2, TLR9, or NOD receptor signaling.

Conclusions

• The research indicates that equine keratinocytes respond more acutely to PAMPs associated with Gram-negative bacteria, compared to those tied to Gram-positive bacteria.
• The findings suggest that this could potentially explain the higher incidence of injury to epidermal structures in cases of Gram-negative sepsis in horses, attributable to a lack of TLR signaling response to Gram-positive PAMPs in the horse’s skin cells.