Table 2.

a,b Methotrexate (MTX)-induced effects on cytokine production by purified CD19⁺ B lymphocytes (a) and CD14⁺ monocytes (b) from the peripheral blood of healthy donors

a. B cells

b. Monocytes

c. Monocytes

d. Monocytes

B cells and monocytes were isolated using immunomagnetic beads and stimulated with lipopolysaccharide (LPS) plus Staphylococcus aureus Cowan's antigen (SAC) in the presence of indicated amounts of MTX. Supernatants were collected after 48 h and assayed for cytokine concentration by specific ELISA. Data are expressed as percentage of cytokine concentration in the absence of MTX (100%) and represent mean values ± s.d. of three independent experiments. Baseline cytokine levels (pg/ml) ± s.d. were as follows: IL-6, 2536 ± 1655 (CD14⁺) and 1632 ± 2173 (CD19⁺); IL-10, 15 379 ± 23 197 (CD14⁺) and 517 ± 422 (CD19⁺); TNF, 9005 ± 5857 (CD14⁺)

Dose-dependent LPS effects on TNF production by CD14⁺ monocytes (absolute values ± s.d.). Maximal TNF production was observed at 1 μg/ml LPS. There was no significant MTX-induced reduction in TNF production at any LPS concentration tested.

Effects of MTX on cytokine production by purified monocytes after 7 days. CD14⁺ monocytes were isolated using immunomagnetic beads and stimulated with LPS, SAC plus granulocyte-macrophage colony-stimulating factor (GM-CSF) in the presence of indicated amounts of MTX for 7 days. Supernatants were taken and analysed for cytokine concentration by ELISA. Data are expressed as percentage of cytokine concentration in the absence of MTX (100%) and represent mean values ± s.d. of three independent experiments.