Cell size and RNA content correlate with cell differentiation and proliferative capacity of rat keratinocytes

Abstract

Keratinocytes from rat skin were separated according to their size in a specially designed unit-gravity sedimentation chamber. The fractions obtained with this technique showed clear morphological differences, and analysis of size distribution confirmed that size was the criterion for separation. Simultaneous DNA and RNA staining of the fractions with acridine orange and subsequent flow cytometric analysis enabled one to classify cells into resting, proliferating, and differentiating stages. Cell size was not directly correlated with proliferation in situ as determined with acridine orange flow cytometry, nor with proliferative capacity in culture as assayed by BrdU/Hoechst flow cytometry. The smallest cells, exhibiting low DNA and RNA content, which do not proliferate in vivo, required a prolonged period of serum stimulation in vitro to initiate RNA and DNA synthesis. Cells of intermediate size exhibited early RNA synthesis and maximal proliferative capacity, whereas the largest cell population displayed no RNA synthesis in culture and the least proliferative capacity. In conclusion, these results suggest that RNA synthesis early after serum stimulation, in addition to a specific, optimal cell size, correlates with the proliferative capacity of keratinocytes in cell culture.

Cell kinetic disturbances induced by treatment of human diploid fibroblasts with 5-azacytidine indicate a major role for DNA methylation in the regulation of the chromosome cycle

“…A prolongation of the S and G2 phase at the expense of the duration of the G1 compartment, together with a striking arrest of cells in the G2 phase of the cell cycle as observed in 5AC-treated cultures of AFFL and HDFL cells, has previously been encountered in peripheral blood lymphocytes from patients with Fanconi's anemia (Kubbies et al 1985). Our results highlight the variability in the duration of the GI phase of the cell cycle in response to prolongation of the S or G2 compartment (Stancel et al 1981;Johnston and Singer 1983) and support the view that cell growth during the G1 phase is only remotely related to, transit through the chromosome cycle (Shields et al 1978;Baserga 1984;Poot et al 1990). It is tempting to suggest that the prolongation of the G2 phase may shift the putative decision point for initiation of the next round of DNA replication (believed to be localized in the G1 compartment of the cell cycle) before mitosis.…”

Mucosal keratinocyte isolation: a short comparative study on thermolysin and dispase

“…2,20 In the present study, an increase in size was seen as a result of culturing the cells as cells differentiate, corresponding to earlier reports on cell diameter changes of keratinocytes in culture. 13,17 The data in the present study show similar cell-size distributions between both tested protocols suggesting that no differences occur upon culture in vitro.…”

Changes in the ultrastructure of cytoskeleton and nuclear matrix during HaCaT keratinocyte differentiation

“…It is likely that the overall transcriptional activity in differentiated cells is low, which is in accordance with the finding of the reduced number of RNP granules in the matrix. This conclusion is supported by flow cytometric studies demonstrating low RNA content in differentiated keratinocytes (43, 44).…”