Cofactors in in vitro induction of apoptosis in HL60 cells by all-trans retinoic acid (ATRA).
The aim of this study was to determine the culture conditions that could modulate the induction of apoptosis by all-trans retinoic acid (ATRA). Cell viability was evaluated by trypan blue test, differentiation by nitro blue tetrazolium test, and apoptosis by morphological analysis. ATRA induced apoptosis in HL60 cells only when more than 100,000 cells/mL were seeded, while differentiation was induced regardless of the seeded cell concentration. Reduction in the concentration of foetal calf serum or glutamine in the medium led to a weak increase in apoptosis. In contrast, a dramatic enhancement of apoptosis occurred when the culture medium was supplemented with glucose or when the culture pH was decreased. These characteristics were independent of the mechanism of action of ATRA, but the action of glucose could be of significance in diabetic patients. An exchange of supernatants after 3 days of culture showed that supernatants from control cultures seeded at high cell density were better apoptosis inducers than supernatants from cultures treated with ATRA, but seeded at low cell density. Factor(s) in this supernatant which induced apoptosis was (were) removed by ultrafiltration. In conclusion, our results showed that ATRA alone cannot induce apoptosis, but can do so in conjunction with cofactors. The depletion of some components of the medium and the appearance of secreted macromolecule(s) could be cofactor(s) in the induction of apoptosis.
Carpentier
Y
Y
Institut J Godinot, Reims, France.
Mayer
P
P
Bobichon
H
H
Desoize
B
B
D017209:Apoptosis / Q000187:drug effects* / Q000502:physiology; D002454:Cell Differentiation / Q000187:drug effects; D002466:Cell Nucleolus / Q000187:drug effects / Q000648:ultrastructure; D002470:Cell Survival / Q000187:drug effects; D002843:Chromatin / Q000187:drug effects / Q000648:ultrastructure; D003470:Culture Media; D005947:Glucose / Q000494:pharmacology*; D005973:Glutamine / Q000494:pharmacology*; D018922:HL-60 Cells; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D009685:Nuclear Envelope / Q000187:drug effects / Q000648:ultrastructure; D014212:Tretinoin / Q000494:pharmacology*
D002843:Chromatin; D003470:Culture Media; D005973:Glutamine; D014212:Tretinoin; D005947:Glucose