[Cell kinetic analysis with flow cytometry as a disdifferentiation mechanism in human neuroblastoma cells in vitro]

The established cell line NB6 (human neuroblastoma) was perturbed in vitro, in order to evaluate the disdifferentiation mechanism of malignant neoplastic cells during cultivation. The methods for perturbation were as follows: 1) 50%-cell removal from dishes; 2) 0.1 mg/ml of vincristine (VCR) contact; and 3) 10Gy of beta-radiation. Specimens were harvested every 24 hours until 72 hours, simultaneously stained for DNA and RNA with acridine orange (AO), and cell kinetically analyzed for DNA/RNA/cell-distribution with the aid of flow cytometry (FCM). Thus disdifferentiated cells and/or newly cell cycle traversing cells, were evaluated. The new subpopulations were considered to be recruited from G1A and/or Gq (Darzynkieowicz et al.) into the cell cycle after perturbations. Cells of the new subpopulations had smaller volume compared with the control cells. Furthermore, in the current study, newly cell cycle traversing cells of NB6 appeared to have a disturbed cell cycle traverse in G2B phase from the clonological changing patterns of FCM cytograms. These new subpopulations were revealed to be able to passage up to five or six times. Though it was impossible to make them appear, these new cycling subpopulations might be capable of remaining as permanent clones in vitro.