Modification of membrane permeability measured by Texas-Red during cell cycle progression and differentiation

Membrane permeability is a critical parameter which must be assessed in the course of new potential anticancer drug studies. Flow cytometry allows us to monitor the permeability status of living cells through the measurement of fluorochrome retention; conventionally this is done using fluorescein diacetate or Hoechst33342. In the present study we show that Texas-Red (TR) can be advantageously used for the same purpose because: (i) it possesses fluorescence spectra compatible with the simultaneous use of various fluorochromes; (ii) staining is rapid; (iii) it is a viability marker. TR uptake is realized through an active, energy dependent transport across the membrane. This new technique was applied to exponentially growing HL60 and their differentiated counterparts. We show that membrane permeability varies throughout the cell cycle progression and during differentiation. We also show that TR uptake is different between sensitive and cytotoxic drug resistant cells. This technique could be used, in anticancer pharmacology, to correlate phase specificity of cytotoxic drugs with membrane permeability, to understand better the mechanism of action of differentiating agents, and to document the apparition of resistant clones within populations of cells.