Enhancement of antinociception by coadministration of minocycline and a non-steroidal anti-inflammatory drug indomethacin in naïve mice and murine models of LPS-induced thermal hyperalgesia and monoarthritis

Abstract

Background: Minocycline and a non-steroidal anti-inflammatory drug (NSAID) indomethacin, have anti-inflammatory activities and are both used in the management of rheumatoid arthritis. However, there are no reports on whether coadministration of these drugs could potentiate each other's activities in alleviating pain and weight bearing deficits during arthritis.

Methods: LPS was injected to BALB/c mice intraperitoneally (i.p.) to induce thermal hyperalgesia. The hot plate test was used to study thermal nociception in naïve BALB/c and C57BL/6 mice and BALB/c mice with LPS-induced thermal hyperalgesia and to evaluate antinociceptive effects of drugs administered i.p. Monoarthritis was induced by injection of LPS intra-articularly into the right hind (RH) limb ankle joint of C57BL/6 mice. Weight bearing changes and the effect of i.p. drug administration were analyzed in freely moving mice using the video-based CatWalk gait analysis system.

Results: In naïve mice indomethacin (5 to 50 mg/kg) had no significant activity, minocycline (25 to 100 mg/kg) produced hyperalgesia to thermal nociception, however, coadministration of minocycline 50 mg/kg with indomethacin 5 or 10 mg/kg produced significant antinociceptive effects in the hot plate test. A selective inhibitor of COX-1, FR122047 (10 mg/kg) and a selective COX-2 inhibitor, CAY10404 (10 mg/kg) had no significant antinociceptive activities to thermal nociception in naïve mice, however, coadministration of minocycline, with CAY10404 but not FR122047 produced significant antinociceptive effects. In mice with LPS-induced hyperalgesia vehicle, indomethacin (10 mg/kg) or minocycline (50 mg/kg) did not produce significant changes, however, coadministration of minocycline plus indomethacin resulted in antinociceptive activity. LPS-induced RH limb monoarthritis resulted in weight bearing (RH/left hind (LH) limb paw pressure ratios) and RH/LH print area ratios deficits. Treatment with indomethacin (1 mg/kg) or minocycline (50 mg/kg) had no effects on the weight bearing and print area ratios deficits of monoarthritic mice. However, combination of minocycline plus indomethacin restored weight bearing and paw print area ratios of monoarthritic mice similar to that observed in non-arthritic control mice.

Conclusions: Coadministration of indomethacin or a selective COX-2 inhibitor, CAY10404 with minocycline potentiates their effects and results in antinociception against thermal nociception, reduction of thermal hyperalgesia and alleviation of weight bearing deficits in monoarthritic mice at doses where either drug alone has no significant activity. Thus, the coadministration of lower doses of a NSAID or a selective COX-2 inhibitor plus minocycline could be useful in the management of inflammatory pain and arthritis.

Figure 1

Coadministration of minocycline with indomethacin produces antinociceptive effects in the hot plate test. Effect of intraperitoneal treatment with indomethacin, minocycline or a combination of minocycline plus indomethacin in a hot plate test in naïve BALB/c mice. Time course of the reaction latency times for minocycline (50 mg/kg, n = 15), indomethacin (10 mg/kg, n = 16), minocycline + indomethacin (n = 16) or their vehicles (n = 16). Each point represents the mean ± S.E.M of the values obtained from 15 to 16 animals. Statistically significant differences in comparison with drug vehicle: * p < 0.05 and ** p < 0.01; and between minocycline or indomethacin and the combination of minocycline + indomethacin treatments: ^# p < 0.05 and ^{# #} p < 0.01 (two-way ANOVA followed by Bonferroni test).

Figure 2

Lipopolysaccharide (LPS)-induces thermal hyperalgesia in BALB/c mice. Time course of the reaction latency time to the hot plate test after administration of LPS (1 mg/kg, n = 5, i.p.) or its vehicle (n = 8). Each point represents the mean ± S.E.M of the values obtained from 5-8 animals. Statistically significant differences in comparison with drug vehicle at the same time point post treatment: * p < 0.05 and ** p < 0.01 (two-way ANOVA followed by Bonferroni test).

Figure 3

Coadministration of minocycline with indomethacin enhances their effects and alleviates LPS-induced hyperalgesia. Effect of treatment with indomethacin, minocycline or a combination of minocycline plus indomethacin on LPS-induced hyperalgesia at day 7 post lipopolysaccharide (LPS, 1 mg/kg, i.p.) inoculation in a hot plate test in BALB/c mice. A: Hypernociceptive effects of LPS, 1 mg/kg, i.p. in the hot plate model of inflammatory thermal hyperalgesia in BALB/c mice at 7 days post inoculation. Each point represents the mean ± S.E.M of the values obtained from 6 vehicle-treated and 29 LPS-treated animals. Statistically significant differences in comparison with vehicle inoculated mice at the same time point post treatment: ** p < 0.01 (two-way ANOVA followed by Bonferroni test). B: Percentage change in reaction latency times from baseline values (taken at 7 days post LPS) at 1 hour after treatment with minocycline (50 mg/kg, n = 15), indomethacin (10 mg/kg, n = 14), minocycline + indomethacin (n = 16) or their vehicles (n = 12) in hot plate test. Each bar represents the mean ± S.E.M of the values obtained from 12 to 16 animals. Statistically significant differences in comparison with drug vehicle: * p < 0.05 (two-way ANOVA followed by Bonferroni test).

Figure 4

Coadministration of minocycline with indomethacin alleviates weight bearing deficits in mice with LPS-induced monoarthritis. Effects of indomethacin 1 mg/kg, minocycline 50 mg/kg or a combination of minocycline 50 mg/kg plus indomethacin 1 mg/kg on A: weight bearing (measured as ratio of light intensity between right hind (RH) and left hind (LH) limbs) and B: ratio of RH/LH print area of mice with LPS-induced arthritis. The drugs or their vehicles were administered at 2 days post-LPS administration and their effects measured at 1 hour after drug treatment. Each point represents the mean ± S.E.M of the values obtained from 6 to 9 animals. Non-arthritic control (n = 9) and arthritic mice (LPS-inoculated) treated with vehicle (n = 7), indomethacin 1 mg/kg (n = 7), minocycline 50 mg/kg (n = 8) or a combination of minocycline 50 mg/kg plus indomethacin 1 mg/kg (n = 6). Statistically significant differences in comparison with non-arthritic control (vehicle only injected) group: * p < 0.05 and ** p < 0.01 (two-way ANOVA followed by Bonferroni test). C and D: Percentage change in RH/LH light intensity and print area ratios from LPS-induced monoarthritic mice at 1 hour after drug treatment. Each bar represents the mean ± S.E.M of the values obtained from 6 to 8 animals. Arthritic mice (LPS-inoculated) treated with vehicle (n = 7), indomethacin 1 mg/kg (n = 7), minocycline 50 mg/kg (n = 8) or a combination of minocycline 50 mg/kg plus indomethacin 1 mg/kg (n = 6). Statistically significant differences in comparison with drug vehicle treated group: * p < 0.05 and ** p < 0.01; and between indomethacin or minocycline alone versus minocycline plus indomethacin combination treated mice: ^# p < 0.05 and ^{# #} p < 0.01 (one-way ANOVA followed by Newman-Keuls multiple comparison test).