Secretion of IL-6, Monocyte Chemoattractant Protein-1, Macrophage Inflammatory Protein-1α, and TNFα by Cultured Intact Human Peritoneum

Abstract

The peritoneum is an important site of host defence. The mesothelial cells, lining the peritoneum, and the fibroblasts found in the layers below are potent sources of a variety of mediators. Furthermore, granulocytes, mast cells, and macrophages, either resident or attracted by inflammatory processes, are interspersed within the tissue. We investigated the production of mediators by samples of fresh human peritoneum. The method described here has the advantage that the cellular composition of the human peritoneum remains intact. Samples of peritoneum were excised at the beginning of elective abdominal operations in infection-free patients. The tissue was placed across the wells of a microtitre plate, fixed in place by the plate cover and incubated with culture medium with or without lipopolysaccharide (LPS) for up to 5 h. The accumulation of IL-6, monocyte chemoattractant protein-1 (MCP-1), macrophage inflammatory protein-1α (MIP-1α) and TNFα in culture supernatants was measured by ELISA. Production of MCP-1 and IL-6 occurred spontaneously during incubation and was enhanced by as much as 4-fold in the presence of different concentrations of LPS (0.5–500 ng/ml) in a dose-dependent manner. MIP-1α and TNFα were detected in culture supernatants of LPS-stimulated samples with concentrations about 8 times as high as those of samples cultured with no such stimulus. The addition of IL-1β resulted in an increase in the release of IL-6 and MCP-1, similar to that observed with LPS stimulation, but failed to increase the production of TNFα. MIP-1α production was only marginally enhanced by IL-1β. In conclusion, our experimental system is suitable for the investigation of chemokine and cytokine production by the human peritoneum, with the aim of assessing aspects of local immunocompetence.