接种 Walker 256 乳腺癌细胞诱导骨癌痛大鼠背根神经节中 Nav1.8 的双侧下调。

BACKGROUND: Rapid and effective treatment of cancer-induced bone pain remains a clinical challenge and patients with bone metastasis are more likely to experience severe pain. The voltage-gated sodium channel Nav1.8 plays a critical role in many aspects of nociceptor function. Therefore, we characterized a rat model of cancer pain and investigated the potential role of Nav1.8. METHODS: Adult female Wistar rats were used for the study. Cancer pain was induced by inoculation of Walker 256 breast carcinosarcoma cells into the tibia. After surgery, mechanical and thermal hyperalgesia and ambulation scores were evaluated to identify pain-related behavior. We used real-time RT-PCR to determine Nav1.8 mRNA expression in bilateral L4/L5 dorsal root ganglia (DRG) at 16-19 days after surgery. Western blotting and immunofluorescence were used to compare the expression and distribution of Nav1.8 in L4/L5 DRG between tumor-bearing and sham rats. Antisense oligodeoxynucleotides (ODNs) against Nav1.8 were administered intrathecally at 14-16 days after surgery to knock down Nav1.8 protein expression and changes in pain-related behavior were observed. RESULTS: Tumor-bearing rats exhibited mechanical hyperalgesia and ambulatory-evoked pain from day 7 after inoculation of Walker 256 cells. In the advanced stage of cancer pain (days 16-19 after surgery), normalized Nav1.8 mRNA levels assessed by real-time RT-PCR were significantly lower in ipsilateral L4/L5 DRG of tumor-bearing rats compared with the sham group. Western-blot showed that the total expression of Nav1.8 protein significantly decreased bilaterally in DRG of tumor-bearing rats. Furthermore, as revealed by immunofluorescence, only the expression of Nav1.8 protein in small neurons down regulated significantly in bilateral DRG of cancer pain rats. After administration of antisense ODNs against Nav1.8, Nav1.8 protein expression decreased significantly and tumor-bearing rats showed alleviated mechanical hyperalgesia and ambulatory-evoked pain. CONCLUSIONS: These findings suggest that Nav1.8 plays a role in the development and maintenance of bone cancer pain.

背景：快速有效地治疗癌症引起的骨痛仍然是一个临床挑战，而患有骨转移的患者更有可能经历严重的疼痛。电压门控钠离子通道 Nav1.8 在伤害感受器功能的许多方面都起着关键作用。因此，我们构建了一种癌症痛大鼠模型，并研究了 Nav1.8 的潜在作用。

方法：本研究使用成年雌性 Wistar 大鼠。将 Walker 256 乳腺癌肉瘤细胞接种到胫骨中来诱导癌症痛。手术后，评估机械性和热痛觉过敏以及活动评分，以确定与疼痛相关的行为。我们使用实时 RT-PCR 来确定手术后 16-19 天双侧 L4/L5 背根神经节（DRG）中 Nav1.8mRNA 的表达。使用 Western 印迹和免疫荧光比较荷瘤和假手术大鼠 L4/L5 DRG 中 Nav1.8 的表达和分布。在手术后 14-16 天鞘内给予 Nav1.8 反义寡核苷酸（ODN）以敲低 Nav1.8 蛋白表达，并观察疼痛相关行为的变化。

结果：荷瘤大鼠在接种 Walker 256 细胞后第 7 天表现出机械性痛觉过敏和活动诱发的疼痛。在癌症痛的晚期（手术后第 16-19 天），通过实时 RT-PCR 评估的同侧 L4/L5 DRG 中 Nav1.8 mRNA 的归一化水平明显低于假手术组。Western 印迹显示荷瘤大鼠双侧 DRG 中 Nav1.8 蛋白的总表达显著降低。此外，免疫荧光显示，只有双侧 DRG 中小神经元中的 Nav1.8 蛋白表达明显下调。给予 Nav1.8 反义 ODN 后，Nav1.8 蛋白表达明显降低，荷瘤大鼠表现出机械性痛觉过敏和活动诱发的疼痛减轻。

结论：这些发现表明 Nav1.8 在骨癌痛的发生和维持中起作用。