Radiation response of proliferating and quiescent subpopulations isolated from multicellular spheroids

Two subpopulations enriched in cells with a G1-like content of DNA were isolated from EMT6/Ro spheroids using centrifugal elutriation. The techniques of two-step acridine orange staining followed by flow cytometry, and continuous [3H]-thymidine labelling agreed qualitatively that one of these subpopulations predominantly consisted of proliferating G1 cells, while the other contained about four times more quiescent G0/G1 cells. These two subpopulations had similar median cell volumes and DNA contents, but the cell volume distributions were different. The clonogenicity was greater in the 'proliferating' subpopulation than the 'quiescent' subpopulation. When cell number seeded was corrected for viability, regrowth studies showed that there was a longer time (25 h) for the 'quiescent' subpopulation than the 'proliferating' subpopulation (10 h) before any increase in cell number was observed. In addition, relative to the 'proliferating' cells, the 'quiescent' cells were more sensitive when exposed to 137Cs gamma-ray radiation. The D0's were similar between the two subpopulations (D0 = 1.6 Gy and 1.8 Gy for the 'proliferating' G1 and 'quiescent' G0/G1 subpopulation, respectively), but the width of the shoulder of the radiation survival curve was reduced in the 'quiescent' subpopulation (Dq = 2.3 Gy vs. 5.3 Gy).