MESANGIAL REGULATION OF GLOMERULAR LEUKOCYTE TRAFFIC

该项目的长期目标是开发一个清晰的

The long term objective of this project is to develop a clear understanding of the process of glomerular monocyte infiltration, which is believed to be an important cause of acute glomerular dysfunction and chronic glomerular sclerosis. This laboratory has shown that the production of monocyte chemoattractant protein-1 (MCP-1), a monocyte specific chemotactic peptide, may be induced in glomerular mesangial cells. It was thus postulated that the glomerular mesangial cell regulates monocyte traffic via the production of MCP-1. This hypothesis will initially be studied using an in vitro model of glomerular injury, created by exposing cultured human mesangial cells to complement components, immune complexes, and lipoproteins, stimuli which have been shown to initiate glomerular injury in vivo. Mesangial MCP-1 expression will be assessed by Northern analysis of mesangial mRNA. MCP-1 secretion into the culture supernatants will be confirmed by immunoadsorption with a specific anti-human MCP-1beta antibody, and functional activity will be measured using a monocyte chemotaxis assay. To understand how such diverse stimuli may activate mesangial MCP-1 synthesis, a common mechanism of action will be sought. Possibilities which will be investigated include: 1) stimulus induction of another mesangial cytokine (e.g. IL-1, PDGF) which ultimately regulates MCP-1 gene expression, and 2) signal transduction through a common second messenger system (e.g. protein kinase C). this model will also be used to investigate mesangial elaboration of chemotaxis inhibitors, providing further evidence for a regulatory role of the mesangial cell in glomerular inflammation. The effect of anti-inflammatory agents in this system will be examined in order to test the susceptibility of MCP-1 expression to pharmacologic interventions. Further studies will be conducted using tissue obtained from human renal biopsy specimens and rodent kidneys. Evidence for the presence of MCP-1 in human glomerular disease will be sought using immunohistochemical analysis of the renal biopsy material; in situ hybridization for MCP-1 mRNA will also be performed to identify the glomerular source of MCP-1 as the mesangial cell. The time course of glomerular MCP-1 mRNA expression following induction of glomerulonephritis will be assessed (by Northern analysis and in situ hybridization) in a rodent model of monocyte-dependent glomerular injury, and correlated to the evolving leukocyte infiltration. Finally, neutralizing antibodies to MCP-1 will be given to rodents during the induction of glomerulonephritis in an attempt to abrogate the monocyte infiltration and the subsequent development of proteinuria. These studies will thus define a central role for the mesangial cell in the regulation of the initial phase of glomerular injury, and document the importance of the novel pro-inflammatory cytokine MCP-1 in recruiting monocytes to the glomerulus. Understanding the mechanisms of mesangial expression of MCP-1 could provide a basis for therapeutically modulating the release of this cytokine, thus preventing or ameliorating the acute glomerular damage and progressive renal insufficiency caused by infiltrating monocytes.