The Role And Mechanism Of Complement C3in Rat Cancer Induced Bone Pain

Objective To investigate the role and mechanism of complement C3in rat cancerinduced bone pain (CIBP).Methods (1)39rats were divided into sham operated group(n=15) and CIBPgroup(n=24). Rats were measured mechanical pain threshold and killed on the7th，14th，21st day after operation，and the lumbar enlargement segment of spinal cord wasremoved for determination of C3or GFAP expression by real-time PCR orimmunohistochemistry assay．(2)6CIBP rats were randomly divided into2groups：vehicle control group and C3inhibitor compstatin intervention group at the eleventh day after successful molding.Sterile normal saline or compstatin15μ（l0.4mg/ml）was administrated twice daily forten days. Rats were killed on the21st day after molding, and the lumbar spina lenlargement segment was removed for determination of GFAP expression．(3) Astrocytes cultured in vitro were divided into control group and compstatin group,and C3inhibitor compstatin was added to the latter culture medium. The proliferationactivity of astrocyte was detected by EdU kit.Results (1) The pain threshold of CIBP rats was19.43±4.66on the14th day afteroperation, and it decreased significantly (P <0.05) compared with the preoperative(23.40±5.00). The pain threshold of CIBP rats was19.11±4.94on the21st day after operation, and it decreased significantly（P <0.05）compared with the preoperative(23.40±5.00) or sham group (27.88±5.33). Real-time PCR detecting the expressionlevel of spinal cord C3mRNA, the C3mRNA expression of CIBP rats was2.52±0.67on the7th day after operation，8.94±2.15on the14th day after operation and9.55±2.91on the21st day after operation；C3expression level on the14th or21st dayafter operation increased significantly（P <0.05）compared with the sham group.Immunofluorescence detecting the expression changes of spina l cord complement C3and GFAP， compared with sham group, C3expression of CIBP rat increasedobviously on the14th day after operation， and GFAP expression increasedsignificantly on the21st day after operation.(2) In the CIBP rats intrathecal injected compstatin, the expression of spinal GFAPdecreased significantly.(3) Astrocyte proliferation positive rate was34.5%±15.6%in compstatin group, andwas60.6%±6.6%in control group. The difference of astrocyte proliferation betweenthe two group was statistically significant (P <0.05).Conclusion The expression of spinal cord complement C3and the activity of werespinal astrocyte increased,and intrathecal complement inhibitor compstatin reducedspinal astrocyte activity in CIBP rats. Compstatin inhibited proliferation of astrocytescultured in vitro. Complement C3involved in the signal transduction of cancerinduced bone pain by mediating spinal cord astrocytes activity and proliferation.