Charakterisierung der Ratten-Makrophagenzellinien R2 und NR8383 und Untersuchung des Einflusses ausgewählter mineralischer Fasern und Partikel auf die Freisetzung immunologisch wirksamer Substanzen

Summary For several years mineral fibers have been used as a replacement for asbestos because of its potential to cause asbestosis and neoplasm. The main goal of this study was to find out how the substitutes titanium dioxide, glaswool, rockwoll and ceramic fibers interact with macrophages and to find out how they react to this materials in order to get evidence of the pathogenicity of the newly used materials. We studied the reaction of the cells to the materials in terms of vitality and their production of inflammatory cytokines like TNF-alpha, IL-1beta, MCP-1 and the release of nitroxy radicals (NO). In my paper the pleura macrophage celline R2 and the alveolar celline NR8383 of the rat was used. In order to determine whether these cells are suitable for the following assays we first had to characterize them. We used LPS and IFN-gamma as well known stimuli for macrophages in various concentrations. In both cell lines there was a dose dependend release of NO, TNF-alpha, MCP-1 and IL-1beta by LPS. IFN-gamma did not lead to a significant change of the expression of these mediators. When both stimulants were combined the release of NO was raised significantly. When low concentrations of IFN-gamma (1 U/ml) and LPS were added there was an increased release of TNF-alpha, MCP-1 and IL-1beta. In higher doses of IFN-gamma (10, 100 U/ml) there was a decreased release of the LPS induced production. When literature is taken into account our results indicate that the alveolar macrophage celline NR8383 and the pleura celline R2 react like primary cells and are suitable for our experiments. After 24 hour stimulation of the macrophage cellines with the used materials in the concentrations 2, 10 and 50 microgramm/cm2 the production of proinflammatory cytokines was measured. Christoballite and Chrysotile were used as a positive control. The results indicate a high biological reactivity for titanium dioxide. When the materials interacting with the cellines are examined under the microscope it became clear that the dimension like length, width and mass and their interaction with macrophages vary to a high degree. In order to get comparable results phagocytable units instead of mass/cm2 was used in the following experiments. Using equal phagocytable units of the materials in combination with IFN-gamma as an additional stimulus all materials lead to an increased production of NO by the macrophages. All materials induced an increased release of TNF-alpha by the cellines. All materials but glaswoll and titanium dioxide stimulated the macrophages to the production of MCP-1. An important problem interpreting in vitro results with cellines is the transfer of the results to primary cells. In order to show differences in species primary human monocytes were used in an identical manner for further experiments. After the application of titanium dioxide, christoballite and asbestos in combination with IFN-gamma human monocytes reacted with the release of MCP-1. Since there was a difference of the reaction of macrophage cellines to those of human monocytes differences in species and in tissue specific cells are here discussed. To sum up the results one can say that all used mineral fibers are able to induce a proinflammatory reaction in macrophages and monocytes. In principle all used mineral fibes are suspected of inducing a pneumoconiosis. In order to make a prediction of the pathogenic potential of mineral fibers further informations in addition of the biologic reacticity are necessary. The biopersistence is of a great importance. It is determined by the clearance of the mineral fibers in the lung, their translocation to the interstitium and their solubility. In order to determine the biopersistence in vivo experiments are inevitable. The developed in vitro test can help to describe the biologic reactivity and in addition to the biopersitence of these materials a prediction of the pathogenic potential of mineral fibers may be possible.