Comparison between proliferative changes and nuclear events during ageing of human fibrobalsts in vitro

Abstract

Age-related alterations in nucleoproteins were examined in resting human embryonic fibroblasts at various population doubling levels (PDL) with three techniques for electron microscopy. Conventional glutaraldehyde fixation showed nuclear modifications consistent with previously published studies. Miller's technique permitted the detection of transcription complexes in young as well as in old cells. The technique involving the ultrathin sectioning of partially loosened nucleoproteins revealed at high PDL a marked rarefaction of the chromatin threads and the appearance of large parts of the lamina densa devoid of attached threads. In addition, the nucleolar filamentous masses were considerably less tufty than at earlier PDL, suggesting a decrease in the ribosomal transcriptional activity. These changes in nucleoprotein organization were rare before the 40th PDL, then increased to about 5% of the nuclei when the percentage of cells initiating DNA during the first 24 h remained at low levels. They were the rule for the last 6 to 7 PDL and coincided with the time when the maximal densities started to fall rapidly. They appeared after treatment with a loosening medium at neutral and at alkaline pH.