Intrathecal Injection Of Liproxstatin-1 Alleviates Bone Cancer Pain In Rats

Objective:To investigate whether intramural injection of Liproxstatin-1 can relieve bone cnacer pain(BCP)in rats.Methods:Healthy adult female SD rats with body weigh of 160 ～ 180 g were selected to construct the rat model of bone cancer pain by injecting breast cancer cell line Walker256 into the tibia marrow cavity.The experiment was divided into two parts.The first part: 40 female SD rats were randomly divided into 5 groups,namely Normal group,Sham group and BCP 3subgroups : BCP 9 d group,BCP 12 d group,BCP 15 d group(group according to the corresponding days after molding)with 8 rats in each group.Ambulatory Score(AS)and Mechanical Paw Withdrawal Threshold(MPWT)were measured before modeling(0 d)and at1,3,6,9,12,15,18 d post-modeling,respectively.The bone destruction was observed by X-ray radiography of the tibia.Real-time fluorescence quantitative PCR(qRT-PCR)and Western blot were used to determine the expression of GPX4 and ACSL4 in the spinal dorsal horn of rats in each group.The second part: 32 female SD rats were randomly divided into four groups,namely Sham group,BCP group,BCP+DMSO group,and BCP+Lip-1 group.The BCP+DMSO group and BCP+Lip-1 group were intraminally injected with DMSO or Liproxstatin-1(ferroptosis inhibitor)at 6,7 and 8 days after modeling.The changes of AS score and MPWT were determined at 0,1,3,6,9,12,15 and 18 days.The expressions of GPX4 and ACSL4 in the spinal dorsal horn of the four groups at 15 days were determined by Western blot.Results : Compared with the Normal group,the AS score of the rats in the BCP group gradually increased(P < 0.01),and the MPWT gradually decreased(P < 0.01)from 6 to 18 days after modeling.X-ray films showed that the bone mineral density of rats in the BCP group began to decrease gradually after 6 days of modeling,and the bone ring was broken,which continued to deteriorate with time.qRT-PCR and Western blot results showed that:Compared with the Normal group,the expression levels of GPX4 mRNA and protein in the spinal dorsal horn of rats in the BCP group were significantly decreased at 9,12,and 15 days(P < 0.05 or P < 0.01).The expression levels of ACSL4 mRNA and protein were significantly increased(P < 0.05 or P < 0.01).Compared with the BCP group,the AS score of the BCP+Lip-1 group was significantly decreased(P < 0.05),and the MPWT was significantly increased(P < 0.05)within 9-18 days.In BCP+Lip-1 group,GPX4 protein expression level was significantly increased(P < 0.05),while ACSL4 protein expression level was significantly decreased(P < 0.05).Conclusion:Intrathecal injection of Liproxstatin-1 can relieve bone cancer pain in rats,and ferroptosis may be involved in the process of bone cancer pain in rats.