PRETREATMENT WITH OLEIC-ACID ACCELERATES THE ENTRANCE INTO THE MITOTIC-CYCLE OF EGF-STIMULATED FIBROBLASTS

We have previously demonstrated that pretreatment of several cell line s with cis-unsaturated fatty acids, like oleic acid, blocks epidermal growth factor (EGF)induced early ionic signals, and in particular the [Ca2+](i) rise. In the present work we show that this blockade does no t alter EGF-stimulated cellular proliferation evaluated by direct cell counting, but induces a powerful enhancement in the pulsed thymidine incorporation assay. The lack of effect of oleic acid on EGF-stimulate d cellular proliferation was confirmed by repeated cell counts, cumula tive thymidine incorporation, and protein synthesis, but a clear syner gistic effect between oleic acid and EGF was again obtained by means o f time course experiments with pulsed thymidine. Combined flow cytomet ry analysis and cell counts at earlier times in EGF-stimulated cells s howed that oleic acid accelerates the entrance of cells into the repli cative cycle leading to an earlier cell division. Afterward, these ole ic acid-pretreated cells became delayed by an unknown compensatory mec hanism in such a way that at 48 h post-EGF, the cell count in control and oleic acid-pretreated cells was equal. In conclusion (a) oleic aci d accelerates or enhances the EGF mitogenic action and (b) in the long term cells compensate the initial perturbation with respect to untrea ted cells. As a side observation, the widely employed pulsed thymidine incorporation method as a measure of cell division could be extremely misleading unless experimental conditions are well controlled. (C) 19 95 Academic Press, Inc.