The effects of low-dose methotrexate on thymidylate synthetase activity in human peripheral blood mononuclear cells.
OBJECTIVE
Methotrexate (MTX) in low doses is widely used in the treatment of rheumatoid arthritis (RA) and it is not known whether its effects are due to immunosuppressive and/or anti-inflammatory actions. High concentrations of MTX inhibit the activity of thymidylate synthetase (TS) and dihydrofolate reductase essential for DNA synthesis. This study investigated the effects of low-dose MTX on TS activity and proliferation in human peripheral blood mononuclear cells (PBMC).


METHODS
The MTX concentrations in our experiments were chosen according to the plasma concentrations measured in 8 RA patients treated with MTX. The effect of MTX on TS activity and DNA synthesis were measured in stimulated normal PBMC and in PBMC obtained from 6 RA patients treated with oral MTX before and 2 hours after intake of their weekly MTX dose. The effect of MTX on the TS mRNA concentration was also investigated in order to elucidate its effect on TS production.


RESULTS
Low-dose MTX significantly inhibited TS activity and the proliferation of stimulated PBMC independent of the mode of activation. Interestingly, the concentration of TS mRNA in normal PBMC was upregulated by the presence of MTX. Finally, there was no difference between TS activity measured before and after MTX intake in 6 RA patients on long-term MTX treatment.


CONCLUSIONS
We show that low concentrations of MTX inhibit TS activity in vitro. An in vivo effect cannot, however, be proven given our study design. The role of these in vitro findings is discussed, particularly in relation to the in vivo effects of MTX.
Hornung
N
N
Department of Clinical Biochemistry, Aarhus University Hospital, Aarhus, Denmark. Bankhornung@post.tele.dk
Stengaard-Pedersen
K
K
Ehrnrooth
E
E
Ellingsen
T
T
Poulsen
J H
JH
D000328:Adult; D000368:Aged; D002455:Cell Division / Q000187:drug effects; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D005260:Female; D006801:Humans; D008297:Male; D008727:Methotrexate / Q000008:administration & dosage* / Q000097:blood / Q000494:pharmacology; D008875:Middle Aged; D009000:Monocytes / Q000166:cytology / Q000187:drug effects / Q000201:enzymology*; D012333:RNA, Messenger / Q000097:blood; D012016:Reference Values; D013940:Thymidylate Synthase / Q000097:blood* / Q000235:genetics
D012333:RNA, Messenger; D013940:Thymidylate Synthase; D008727:Methotrexate