脊髓水平神经型一氧化氮合酶在骨癌痛发生中的作用

The role of neuronal nitric oxide synthase in the development of bone cancer pain

目的探讨脊髓水平神经型一氧化氮合酶（neuronal nitric oxide synthase，nNOS）在小鼠骨癌痛发生中的作用。方法72只C3H／Hej小鼠随机分为肿瘤组（n=40只）和假手术组（n=32只）。肿瘤组再分为5组（n=8）：17组（肿瘤细胞接种后7d），T10组（接种后10d）和T14组（接种后14d），L组（接种后14d＋L-NMMA）和C组（接种后14d＋溶剂），假手术组相应分为S7组（假手术7d）、S10组（假手术10d）、S14组（假手术14d）和S组（假手术14d＋溶剂）。将含10^5个纤维肉瘤NCTC2472细胞的最小必需培养基（α-MEM）注射到小鼠右侧股骨远端骨髓腔内，制作骨癌痛模型。假手术组注入不含肿瘤细胞的α-MEM。按照分组在相应时间点处死小鼠，用RT-PCR的方法检测脊髓腰膨大nNOS mRNA水平。术后14d，L组小鼠鞘内注射NOS抑制剂L-NMMA 50μg，给药前及给药后2h、12h、24h检测小鼠痛行为学的变化：机械痛缩足阈值（PWMT）和热痛缩足潜伏期（PWTL）。结果肿瘤细胞接种后10d，脊髓水平nNOS mRNA表达较假手术组增高（P〈0．05），而接种后14d nNOS mRNA水平与假手术鼠无差别；鞘内注射L-NMMA 50μg明显改善早期骨癌痛小鼠的痛行为。结论脊髓水平nNOS可能在小鼠骨癌痛发生的早期起作用，而癌痛的维持依赖于其他类型一氧化氮合酶（nitric oxide synthase，NOS）。

Objective To explore the role of neuronal nitric oxide synthase （nNOS） in the development of bone pain cancer of mice. Methods 72 C3 H/Hej mice were divided randomly into tumor group （ n = 40） and sham group （ n = 32）. Tumor group was divided randomly into T7, T10, T14, L （L-NMMA） and C group （vehicle）. The Sham group mice were further divided randomly into S7 group, S10 group, S14 group and S group （vehicle）. Osteosarcoma NCTC 2472 cells were implanted into the intramedullary space of the fight femurs of mice to induce ongoing bone cancerrelated pain behaviors. The sham group was inoculated by α-MEM without any cells. Mice were killed according to the corresponding time. Reverse transcriptive polymerase chain reaction （RT-PCR） was applied to analysis the expression of nNOS mRNA. The mice were observed the changes of pain behaviors in L Group, such as paw withdrawal mechanical threshold （PWMT） and paw withdrawal thermal latency （PWTL） , at 2 h, 12 h and 24 h after intrathecal administration of 50 μg L-NMMA. Results The levels of nNos mRna in spinal cord of tumor mice were increased （ P 〈 0.05 ） at 10 d after incucation and decreased at 14 d. The pain behaviors were attenuated by intrathecal administration of 50 μg L-NMMA. Conclusion nNOS in spinal cord may participate in the development of bone cancer pain in the early time, and the maintenance of bone cancer pain may depend on the other type of NOS.