Chemokines produced by mesothelial cells: huGRO-alpha, IP-10, MCP-1 and RANTES

Abstract

Recently we showed the in vivo relevance of chemokines in cases of bacterial peritonitis in continuous ambulatory peritoneal dialysis (CAPD) patients. Mesothelial cells, the most numerous cells in the peritoneal cavity, are hypothesized to function as a main source of chemokine production. We investigated the time- and dose-dependent expression patterns of four chemokines by mesothelial cells at the mRNA and protein level in response to stimulation with physiological doses of proinflammatory mediators that are present at the site of bacterial inflammation. Besides the chemokines huGRO-alpha (attractant for neutrophils), MCP-1 and RANTES (monocyte attractants), the expression and production of IP-10 was analysed. Mesothelial cells were cultured and stimulated with either IL-1beta, tumour necrosis factor-alpha (TNF-alpha) or IFN-gamma or combinations of these. The time- and dose-dependent mRNA expression of the chemokines was determined by Northern blot analysis and the protein production by ELISA. It was concluded that mesothelial cells could indeed be triggered by the mentioned stimuli to induce mRNA and protein production (huGRO-alpha and IP-10) or to augment constitutive protein production (MCP-1). However, RANTES mRNA and protein production could only be induced in some cases and only in small amounts. The chemokine response of mesothelial cells was regulated differentially, depending on the stimulus and the chemokine measured. In distinct cases, combination of the stimuli led to synergy in mRNA expression and protein production. The presented in vitro data support our hypothesis that mesothelial cells in vivo are the main source of relevant chemokines in response to proinflammatory mediators, suggesting an important role for mesothelial cells in host defence.

Fig. 1

Northern blot analysis and quantification of mRNA expression of huGRO (a), IP-10 (b) and MCP-1 (c) by human peritoneal mesothelial cells after stimulation with increasing doses of IL-1β, tumour necrosis factor-alpha (TNF-α), and IFN-γ. The graphs show the chemokine mRNA/β-actin mRNA expression ratio in densitometric units to correct for RNA loading differences, and indicate the quantity of mRNA expression.

Fig. 2

Northern blot analysis and quantification of mRNA expression of huGRO (a), IP-10 (b) and MCP-1 (c) by human peritoneal mesothelial cells after stimulation for various different periods (2, 4, 8, 12 and 24 h) with IL-1β, tumour necrosis factor-alpha (TNF-α), IFN-γ or combinations of these cytokines. *Combination of IL-1β and IFN-γ; †combination of TNF-α and IFN-γ. Control gives data of unstimulated cultures. The graphs show the chemokine mRNA/β-actin mRNA expression ratio in densitometric units to correct for RNA loading differences, and indicate the quantity of mRNA expression.