Effect of intrathecal methotrexate on activation of spinal astrocytes in a rat model of bone cancer pain

Abstract

Objective To evaluate the effect of intrathecal methotrexate on activation of spinal astrocytes in a rat model of bone cancer pain (BCP). Methods Sixty female unmated Sprague-Dawley rats, aged 5-7 weeks, weighing 150-180 g, were randomly divided into 3 groups (n=20 each)using a random number table: sham operation group (group S), BCP group (group P), and BCP + methotrexate group (group PM). BCP was induced by injecting Walker-256 cancer cells into the medullary cavity of tibia.On day 7 after BCP, methotrexate 100 μg (diluted to 15 μl in artificial cerebrospinal fluid)was injected intrathecally over 10 s in group PM, and artificial cerebrospinal fluid 15 μl was injected intrathecally over 10 s in S and P groups.The mechanical paw withdrawal threshold (MWT)was measured at 1 day before BCP and 3, 7, 14 and 21 days after BCP.Five rats were sacrificed after measurement of the pain threshold at 7, 14 and 21 days after BCP, and the lumbar enlargement segments of the spinal cord were harvested for detection of the expression of glial fibrillary acidic protein (GFAP)by immuno-histochemistry.Five rats were sacrificed after measurement of the pain threshold at 14 days after BCP, and the expression of GFAP in the spinal cord was detected by Western blot. Results Compared with group S, the MWT was significantly decreased at 3, 7, 14 and 21 days after BCP in C and CM groups, the expression of GFAP was significantly up-regulated at each time point after BCP in group C, and the expression of GFAP was significantly up-regulated at 7 and 14 days after BCP in group CM (P<0.05). Compared with group C, the MWT was significantly increased, and the expression of GFAP was significantly down-regulated at 14 and 21 days after BCP in group CM (P<0.05). Conclusion The mechanism by which intrathecal methotrexate reduces BCP may be related to inhibition of spinal astrocyte activation in the rats. Key words: Methotrexate; Injections, spinal; Pain; Bone neoplasms; Astrocytes