GPR160受体参与骨癌痛大鼠脊髓中枢敏化的作用

Contribution of GPR160 receptor to the development of central sensitization of the spinal cord in rats with Bone cancer pain

摘要:

目的：检测骨癌痛（Bone cancer pain，BCP）大鼠脊髓（Spinal cord, SC）中GPR160受体的表达，探讨GPR160受体在骨癌痛发生发展过程中的作用。方法：雌性SD大鼠91只，随机分为五组：正常组（Naive组，n=11）、假手术组（Sham组，n=28）、骨癌痛组（BCP组，n=36）、BCP+GPR160-siRNA组（BCP+siGpr160组，n=8）、BCP+ control siRNA组（BCP+ sieGfp组，n=8）。采用Von Frey测定机械缩足反射阈值（Paw withdrawal threshold，PWT），检测大鼠机械痛阈的变化；CATWALK步态分析系统分析大鼠步态数据；Western Blot检测脊髓GPR160受体蛋白表达；Real-time PCR检测mRNA表达；ELISA和Western Blot检测TNF-α、IL-1β及IL-6蛋白表达。结果：①BCP大鼠造模后第6d， 左足PWT出现明显降低，并持续至术后18d（P <0.001）。在BCP造模后第18d，出现了明显的癌细胞浸润和骨质破坏。②与Sham组相比，BCP组大鼠脊髓中GPR160的蛋白和mRNA均明显的增加。③与Sham组相比，BCP 大鼠PWT明显下降，而鞘内注射GPR160-siRNA后，能明显逆转这一效应；与 BCP+sieGfp组相比，BCP+ siGpr160组大鼠脊髓GPR160蛋白表达明显下降（P <0.01）。④BCP组大鼠脊髓中促炎因子TNF-α、IL-1β、IL-6的表达明显升高，而鞘内注射GPR160-siRNA可以逆转这一作用。结论：GPR160受体参与骨癌痛大鼠脊髓中枢敏化，其机制可能与增加促炎因子表达有关。

Abstract:

Objective: To detect the expression of GPR160 receptors in the spinal cord of rats with bone cancer pain, and to investigate the role of GPR160 receptors in the development of bone cancer pain. Methods: Ninety-one female SD rats were randomly divided into five groups: Naive group (n=11), Sham group (n=28), BCP group (n=36), BCP + GPR160-siRNA group (BCP+siGpr160, n=8), BCP+ control siRNA group (BCP+ sieGfp, n=8). Paw withdrawal threshold (PWT) was assessed with von Frey. Rats gaits were evaluated with Catwalk gait analysis. Western blotting was used to detect the expression of GPR160 protein in spinal cord. Real-time PCR was used to detect the mRNA of GPR160 in spinal cord. The protein of TNF-α, IL-1β and IL-6 was detected with ELISA and Western Blot. Results: PWT was significantly decreased 6 days after BCP and lasted until 18 days (P <0.001). There was obvious infiltration of cancer cells and bone destruction 18 days after BCP. Compared with control group, the expression of GPR160 and pro-inflammatory cytokines TNF-α, IL-1β and IL-6 in BCP group was significantly increased. However, intrathecal injection of GPR160-siRNA remarkably attenuated the expression of it compared with BCP+sieGfp group. Conclusion: These results suggest that GPR160 receptors could be involved in central sensitization, and the mechanism may be related to the increased expression of pro-inflammatory cytokines.