目的 探讨华蟾素对骨癌痛（cancer-induced bone pain，CIBP）大鼠镇痛作用机制。方法 筛选痛阈值满足条件的雌性SD大鼠构建CIBP模型，华蟾素各组大鼠在造模第7天开始ip低、中、高浓度的华蟾素注射液，假手术组与模型组ip等量的生理盐水，连续给药7 d，各组大鼠分别在造模前和造模后及单次注射华蟾素后的0.5、1、2、4、6、8、24 h检测大鼠机械痛阈值与热痛阈值，Western blotting检测丝裂原激活的蛋白激酶（mitogen-activated protein kinases，MAPKs）相关蛋白的表达，ELISA法检测脊髓细胞因子白细胞介素-1β（Interleukin-1β，IL-1β）、肿瘤坏死因子-α（TNF tumor necrosis factor-alpha，TNF-α）、单核细胞趋化蛋白（monocyte chemoat tractant protein，MCP-1）的含量。结果 大鼠造模后，模型组大鼠的热痛阈值、机械痛阈值明显降低（P<0.01）；予以华蟾素干预后，大鼠的热痛阈值、机械痛阈值升高，且浓度越高，痛阈值越高（P<0.05、0.01）；单次给药后，华蟾素在ip后6 h作用最强，而后作用缓慢减弱；但华蟾素对正常大鼠的痛阈值无影响（P>0.05）。Western blotting的结果显示，模型组大鼠脊髓MAPKs相关蛋白的活化水平增加（P<0.05）；华蟾素能下调大鼠脊髓中活化的氨基末端激酶（c-Jun N-terminal kinases，JNK）、p38蛋白的表达量，而p-ERK蛋白的表达未见明显差异（P>0.05）。ELISA结果显示，模型组大鼠脊髓中TNF-α、IL-1β、MCP-1的含量明显增多（P<0.05），华蟾素能显著抑制其释放（P<0.05）。结论 华蟾素可能是通过抑制脊髓MAPKs信号通路相关蛋白（JNK、p38）的活化，减少细胞因子释放，从而发挥缓解CIBP的作用。

Objective To investigate the analgesic mechanism of cinobufagin in rats with bone cancer pain. Methods Female SD rats meeting the conditions of pain threshold were selected to construct cancer-induced bone pain (CIBP) model. On the 7th day after modeling, the sham group and the model group were administrated by saline, while the treatment groups were administrated with the low, medium and high concentrations of cinobufagin for consecutive 7 d. The pain behavior (mechanical withdrawal threshold and thermal pain threshold) was tested before modeling and after modeling, and single injection of cinobufagin after 0.5, 1, 2, 4, 6, 8 and 24 h at the first day. The expression of MAPKs protein was detected by Western Blotting, and the content of spinal cytokines (IL-1β, TNF-α, MCP-1) was detected by ELISA. Results The mechanical pain threshold and thermal pain threshold were significantly decreased in the model group, compared with the sham group (P<0.01). After injected with cinobufagin, the pain threshold increased with a certain concentration dependence (P<0.05, 0.01). After a single dose, the effect of cinobufagin was the strongest at 6 h after intraperitoneal injection, and then gradually decreased. However, cinobufagin had no effect on the pain threshold of normal rats (P>0.05). Protein levels of MAPKs were increased in the model group, while the levels of JNK and p38 were decreased in the cinobufagin group (P<0.05), while had no effect on the levels of p-ERK (P>0.05). ELISA results showed that cinobufagin significantly decreased the content of cytokines in the spinal cord, when compared with the model group (P<0.05). Conclusion Cinobufagin can inhibit the expression of MAPKs proteins in the spinal cord of the rat model with bone cancer pain, ultimately decrease the content of IL-1β, TNF-α, and MCP-1 to alleviate the pain during the process of cancer pain.

R285.5

国家自然科学基金资助项目（81503381）