Biallelic NF2 gene inactivation is frequently found in human malignant mesothelioma. In order to assess whether NF2 hemizygosity may enhance susceptibility to asbestos fibres, we investigated the Nf2 status in mesothelioma developed in mice presenting a heterozygous mutation of the Nf2 gene (Nf2 KO3/ þ ), after intraperitoneal inoculation of crocidolite fibres. Asbestos-exposed Nf2 KO3/ þ mice developed tumoural ascites and mesothelioma at a higher frequency than their wild-type (WT) counterparts (Po0.05). Six out of seven mesothelioma cell lines established from neoplastic ascitic fluids of Nf2 KO3/ þ mice exhibited loss of the WT Nf2 allele and no neurofibromatosis type 2 protein expression was found in these cells. The results show the importance of the NF2 gene in mesothelial oncogenesis, the potential association of asbestos exposure and tumour suppressor gene inactivation, and suggest that NF2 gene mutation may be a susceptibility factor to asbestos.

The control of DNA integrity in mammalian cells is important to maintain the cell homeostasis and prevent neoplastic transformation. Control of cell division and cell death permits repair or elimination of damaged cells. Since asbestos fibers can produce DNA damage, chromosome alterations and apoptosis in several sorts of cells, including mesothelial cells, it was interesting to investigate cell cycle disturbances in rat pleural mesothelial cells (RPMC) treated with asbestos fibers. Cell cycle analyses were performed in RPMC exposed to crocidolite (10 and 20 microg/cm2) and chrysotile (5 and 10 microg/cm2) for different times (4 to 48 h). Both fiber types entailed a G2/M accumulation in agreement with a delay in the mitosis course. Chrysotile fibers produced a G0/G1 accumulation associated with a time-dependent p53 and p21 expression. Crocidolite exposure resulted in a delay in the G1/S transition paralleling a low rate of p53 expression. These results are in agreement with a DNA damaging potential of asbestos fibers since similar results were found following RPMC exposure to gamma rays. In asbestos-treated RPMC, a low rate of apoptosis was found suggesting that RPMC may follow a DNA repair pathway that could contribute to the formation of DNA lesions. In addition, the cell cycle disturbances at the G2/M checkpoint suggest that genetically altered cells have progressed through the cycle and support the already published findings on the ability of asbestos fibers to impair cell division.

The role of reactive oxygen metabolites in the toxic effects of asbestos on pleural mesothelial cells is not well defined. We exposed rat pleural mesothelial cells (RPMC) to chrysotile and crocidolite fibers (0-40 micrograms/cm2) in the presence or absence of catalase and superoxide dismutase (SOD). Cell injury was measured using the colorimetric 3-4 (5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and DNA damage was evaluated in terms of unscheduled DNA synthesis (UDS). Catalase (100 U/ml) and SOD (250 U/ml) protected RPMC against asbestos-induced cytotoxicity and DNA damage. However, the inactivated enzymes and bovine serum albumin also showed some protection, suggesting that the effect of antioxidant enzymes may be partly related to their protein nature. These results suggest that oxygen derivatives are partly involved in the toxic effects of asbestos on cultures of RPMC. The presence of extracellular proteins may also decrease asbestos-produced toxicity by reducing the degree of RPMC-fiber interaction.