Methotrexate ameliorates T cell dependent autoimmune arthritis and encephalomyelitis but not antibody induced or fibroblast induced arthritis

Abstract

Objective: To investigate the mode of action of methotrexate (MTX) in different types of models for rheumatoid arthritis (RA) and multiple sclerosis (MS).

Methods: Models for RA and MS were selected known to have different pathogenesis--that is, fibroblast induced arthritis in SCID mice, collagen induced arthritis (CIA), anticollagen II antibody induced arthritis (CAIA), and experimental autoimmune encephalomyelitis (EAE) in (Balb/c x B10.Q)F1 and B10.Q mice, and Pristane induced arthritis in DA rats (PIA). The MTX treatment was started 1 day after the onset of disease and continued for 14 days to compare effects on the different models.

Results: All models known to be critically dependent on T cell activation (CIA, PIA, and EAE) were effectively down regulated by titrated doses of MTX. In contrast, no effects were seen on fibroblast induced arthritis or CAIA. No effects were seen on the levels of anticollagen II antibodies in the CIA experiment.

Conclusion: The data show that MTX has strong ameliorative effect on both classical models of RA, like CIA and PIA, but also on a model for MS, EAE. It also suggests that MTX operates only in diseases which are preceded by, and dependent on, T cell activation. A comparison of CAIA and CIA suggested that MTX operates independently of arthritogenic antibodies. These results demonstrate that different animal models reflect the complexity of the corresponding human diseases and suggest that several models should be used for effective screening of new therapeutic agents.

Figure 1

(A) MTX treatment of CIA. Day 0 indicates the onset of arthritis for each mouse and MTX is given at different doses starting 1 day after the onset. The mice were treated with 0.1 mg MTX/kg (n = 5), 2.5 mg MTX/kg (n = 3), 5 mg MTX/kg (n = 3) and a control group was treated with PBS (n = 5). The mean score of all mice for each day is given with the SEM. *Indicates significant differences in severity scores, p<0.05, for the groups treated with 2.5 mg/kg and 5 mg/kg in comparison with the control group. (B) Anti-CII antibody levels in the CIA experiment. The mice were bled at day 14 after the onset of arthritis. Mean values and SEM are indicated. No significant differences according to MTX treatment were seen.

Figure 2

MTX treatment of CAIA. One group of mice were treated with MTX (2.5 mg/kg; n = 13) and a control group were treated with PBS (n = 13) starting 1 day after the onset of arthritis. Control mice were treated with PBS only. The mean score of all mice for each day is given with the SEM. No significant effect of MTX was seen.

Figure 3

MTX treatment of PIA. Day 0 indicates onset of arthritis of each rat, and MTX was given at different doses starting 1 day after the onset. The rats were treated with 0.05 mg MTX/kg (n = 6), 0.075 mg MTX/kg (n = 6), 0.1 mg MTX/kg (n = 13), and a control group were treated with PBS (n = 6). The mean score of all rats for each day is given with the SEM. *Indicates significant differences in severity scores, p<0.05, for all MTX treated groups in comparison with the control group. In the group with highest MTX dose (0.1 mg MTX/kg) eight rats died at different times during the experiment.

Figure 4

MTX treatment of fibroblast induced arthritis in SCID mice. MTX was given at different doses (5 mg/kg, 2.5 mg/kg, 0.1 mg/kg) and PBS was given in the control group (n = 9 in each group) starting 1 day after the injection of the fibroblasts, when all mice had already developed arthritis. Scoring started at day 1 when the arthritis began and treatment was given from this day and then daily until day 14. The mean diameter of the injected left knee of all mice in each group for each day is given with the SEM. No significant effect of MTX was seen.

Figure 5

Histological aspect of a left SCID mouse knee joint 14 days after injection of 5x10⁵ LS48 cells into the articular space. Original magnification x16 (A) and x40 (B). LS48 cells form a dense tissue attaching to and invading the articular structures. Stained with haematoxylin and eosin.

Figure 6

MTX treatment of EAE. Day 0 indicates onset of encephalomyelitis of each mouse and MTX was given at different doses starting 1 day after the onset. The mice were treated with 0.1 mg MTX/kg (n = 11), 2.5 mg MTX/kg (n = 10), 5 mg MTX/kg (n = 10) and a control group were treated with PBS (n = 12). The mean score of all mice each day is given with the SEM. *Indicates significant differences in encephalomyelitis severity scores, p<0.05 in comparison with the control group.