Keratinocytes differentiate in response to inhibitors of deoxyribonucleotide synthesis

We previously reported that methotrexate (MTX) caused an irreversible inhibition of growth and induced terminal differentiation of human keratinocytes. These effects of methotrexate were prevented by thymidine and thus, were attributed to depletion of thymine deoxyribonucleotides. The purpose of the research reported in this paper was to determine whether differentiation was induced by the general class of agents which are known to interfere with synthesis or utilization of deoxyribonucleotides. Agents examined included 5-fluorouracil, 5-bromodeoxyuridine, hydroxyurea, high-dose thymidine, aphidicolin, and AG#85, a newly reported thymidylate synthase inhibitor. All these agents increased the expression of involucrin and increased the amount of cornified envelope protein at doses that inhibited proliferation by > 75%. We demonstrated, however, that in our cell culture system not all conditions producing inhibition of proliferation induced differentiation; withdrawal of growth factors and supplemental amino acids inhibited proliferation but did not increase involucrin expression or production of cornified envelope protein.