Nucleolar changes in human phytohaemagglutinin-stimulated lymphocytes

Abstract

The nucleoli of lymphocytes from circulating peripheral blood and from phytohaemagglutinin (PHA)-stimulated cultures (from 2 h-96h) were studied using a silver method, RNA-specific fluorescent staining, and electron microscopy of ultrathin sections. In peripheral blood about 75% of the lymphocytes have one "ring-shaped" nucleolus composed of a distinct fibrillar centre surrounded by a dense pars fibrillaris and little granular material; the remaining lymphocytes showing two or more small "ring-shaped" nucleoli. With PHA stimulation, the number of cells with several nucleoli increases first (from 2 h--12 h). Next, cells containing one or, at most, two large nucleoli with nucleolonema devoid of fibrillar centers are seen (from 4 h on). 34 h after PHA, nucleoli of the "compact" type containing one or more fibrillar centres appear and comprise about 60% of the cells after 72 h. The appearance of more than one nucleolus per cell shortly after PHA administration suggests an activation of additional nucleolar organizer regions (NOR), which fuse to form one or two large nucleoli with nucleolonema. These are then transformed into "compact" nucleoli. The fibrillar centers stasin preferentially with silver. They contain nonchromosomal proteins and may serve as stores for nucleolar proteins. The fusion of activated NORs during the first cell cycle explains the relatively high frequency of satellite associations in first mitoses compared to later mitoses after stimulation.