Acute and chronic suppression of leukotriene B4 synthesis ex vivo in neutrophils from patients with rheumatoid arthritis beginning treatment with methotrexate.
OBJECTIVE
To compare the cumulative effects of oral methotrexate (MTX) therapy (after 6-8 weeks) with the acute effects (24 hours after a dose) on arachidonic acid metabolism by the 5-lipoxygenase (5-LO) pathway in neutrophils from patients with active rheumatoid arthritis (RA) who were beginning therapy with MTX.


METHODS
Neutrophils and monocytes were isolated from whole blood from 7 patients with RA, immediately before and 24 hours after their first weekly dose of 7.5 mg of MTX, and again after their dose at 6-8 weeks.


RESULTS
Total immunoreactive leukotriene B4 (LTB4) formation in neutrophils activated ex vivo with calcium ionophore A23187 was significantly suppressed (by 33%) before the 6-8-week dose, compared with the level before the first dose (mean +/- SEM 8.29 +/- 1.24 ng/10(6) cells at predose 6-8 weeks versus 12.29 +/- 2.13 ng/10(6) cells at predose 1; P = 0.03). Reductions were also observed after the first dose (27%; P = 0.07) and after the 6-8-week dose (43%; P = 0.05) compared with the respective predose levels. MTX treatment produced significant reductions in the total generation of 5-LO pathway products (5-hydroxyeicosatetraenoic acid + 6-trans-LTB4 + LTB4 + omega-oxidation products of LTB4) by calcium ionophore-activated neutrophils, as quantitated by integrated optical density after resolution on reverse-phase high-performance liquid chromatography. Decreases were observed after the first dose (26%; P = 0.025), immediately before the 6-8-week dose (23%; P = 0.05), and after the 6-8-week dose (47%; P = 0.0033) compared with levels before the first dose, and after the 6-8-week dose compared with the level before it (32%; P = 0.04). The generation of LTB4 by calcium ionophore-activated monocytes was not significantly affected by MTX therapy.


CONCLUSIONS
The significant decreases in the formation of omega-oxidation products of LTB4 and in the total generation of neutrophil 5-LO pathway products in the absence of a significant change in the release of 3H-arachidonic acid or the generation of platelet-activating factor suggest that the activity of the 5-LO enzyme in neutrophils is inhibited. We conclude that weekly oral MTX therapy in patients with active RA inhibits neutrophil 5-LO pathway product generation in a pattern consistent with inhibition of the activity of the 5-LO enzyme; an effect is observed after the first dose. The inhibition of 5-LO is cell-selective and cumulative, with a superimposed incremental inhibition observed after the weekly MTX dose.
Sperling
R I
RI
Department of Medicine, Harvard Medical School, Boston, MA 02115.
Benincaso
A I
AI
Anderson
R J
RJ
Coblyn
J S
JS
Austen
K F
KF
Weinblatt
M E
ME
D000328:Adult; D000368:Aged; D001094:Arachidonate 5-Lipoxygenase / Q000378:metabolism; D001172:Arthritis, Rheumatoid / Q000188:drug therapy* / Q000378:metabolism / Q000473:pathology; D002634:Chemotaxis, Leukocyte / Q000187:drug effects; D006801:Humans; D007975:Leukotriene B4 / Q000037:antagonists & inhibitors* / Q000096:biosynthesis; D008727:Methotrexate / Q000097:blood / Q000627:therapeutic use*; D008875:Middle Aged; D009000:Monocytes / Q000378:metabolism; D009504:Neutrophils / Q000187:drug effects / Q000378:metabolism*; D013997:Time Factors
D007975:Leukotriene B4; D001094:Arachidonate 5-Lipoxygenase; D008727:Methotrexate