K562 human erythroleukemia cells demonstrate commitment

Abstract

Commitment, ie, the decision to express a differentiated phenotype and to terminate proliferation irreversibly in the absence of inducer, was investigated in K562 human erythroleukemia cells. Cells were cultured for 0, 1, 2, 3, or 4 days with inducer and then plated in medium containing methylcellulose without inducer. Daily after plating, hemoglobin content was scored by benzidine staining, and growth was assessed by estimating the cell number per colony. With all inducers used, three types of colonies were found, those containing only benzidine-positive cells, those containing only benzidine-negative cells, and those containing both cell types (mixed colonies). Thymidine produced a progressive increase in the percentage of positive and mixed colonies and a progressive fall in the percentage of negative colonies. Whereas negative colonies grew at an exponential rate with a generation time of about 20 hours, positive colonies reached an average maximum size of 16 cells, representing a total of four divisions. Butyrate had a similar effect, except that the rise was greater for mixed colonies than for positive colonies, and the plateau in positive colony size was less evident. In contrast, CO2 depletion or hemin treatment induced an increase in the fraction of cells staining benzidine positive that was lost rapidly upon removal of the inducing condition. Thus, of the four conditions, thymidine and butyrate caused commitment, whereas hemin and CO2 depletion did not. Thus K562 cells, like Friend cells, demonstrate commitment, but, unlike Friend cells, demonstrate a significant rate of commitment in the absence of inducer and hence form a significant percentage of mixed colonies with or without inducer.

Control of ribosomal protein synthesis by Microprocessor complex

“…The firefly luciferase activity was normalized by the renilla luciferase activity to normalize transfection efficiency. For hemin treatment, K562 cells were treated with 25 μM hemin (Sigma-Aldrich, T8768) for 24 h, followed by benzidine ([1,1'-Biphenyl]-4,4'-diamine) staining as previously described (Rowley et al, 1985;Tomoda et al, 1991).…”

Modulation of homeobox gene expression alters the phenotype of human hematopoietic cell lines.

“…All cytochemical stains were performed using commercial staining kits following manufacturer's suggestions (Sigma, St Louis, MO). Heme content in K562 cells was assessed by direct benzidine staining of individual cells in liquid suspension or of individual colonies in semi-solid medium (Rowley et al, 1985). Since the endogenous heme content of HEL cells is too low for visual scoring, heme content in HEL cells was measured spectrophotometrically following reaction of cell lysates with benzidine; all samples were run in triplicate and the experiment was repeated twice (Clarke et al, 1982).…”

Effect of second‐messenger modulators in K‐562 cell differentiation: Dual action of calcium/phospholipid‐dependent protein kinase in the process of differentiation

“…It has been shown that Ara-C and thymidine belong t o the former group and hemin to the latter group (Rowley et al, 1985;Ohlsson-Whilhelm et al, 1987;Dean et al, 1981). Commitment refers to an irreversible process that leads to expression of the differentiation-related phenotype, i.e., benzidine(B)-positivity, in the absence of an inducer (Gusella et al, 1976;Housman et al 1978; Levenson and Housman, 1979a-c).…”