The biochemical basis of changes in normal and mutant human skin fibroblasts during ageing in culture : an investigation into the free radical theory of ageing

From the pattern of inheritance of normal ageing, this process inevitably results from either abnormal aggregation of subunits of enzymes, abnormal feedback inhibition of enzymes, receptor mutations, membrane defects, or deposition of abnormal fibrillar proteins (Vogel and Motulsky, 1986). The process of ageing should, due to the principle of universality, occur at a site common to all cells : general metabolism, chromatin structure and function ( eg. DNA replication and gene expression) or the physiology of the membrane. The cell type to be studied should be fully functional as to these three sites, and the theories potentially able to explain the ageing process should all encompass changes at any of these aspects . of cellular function. The system to be used throughout this thesis is the cultured human skin fibroblast, which still expresses genes, replicates DNA, has a general metabolism and has functional membranes. This cell type alos undergoes a loss of proliferative potential both in vivo and in culture (Martin et al.,1970). Furthermore, any kind of a phenotype expressed in fibroblast-like cells can theoretically be used for prenatal diagnosis of human genotypes (eg. Werner's syndrome and Spielmeyer-Vogt syndrome). The following postulates can be derived now. First, free radicals should cause the accumulation of lipid peroxides or decay products, which should further yield to the accumulation of autofluorescence. Second, free radicals, or the reactive products resulting from them should impair cellular proliferation to the extent as to cause an irreversible growth arrest of initially dividing cells. Third, crosslinking of proteins by reactive breakdown products of lipid peroxides should occur. Fourth, the capacity of the systems implied in the detoxification of free radicals, or the reactive products thereof, should decrease in capacity during ageing of cultured cells.