摘要
目的:观察腹腔注射雷公藤内酯醇(Triptolide,T10)对完全弗氏佐剂(complete Freund’s adjuvant,CFA)诱导的大鼠慢性炎性痛行为的改善作用并探讨其机制。方法:SD大鼠随机分为四组,即Saline+Veh组、Saline+T10组、CFA+Veh组和CFA+T10组。后两组大鼠于右侧足底注射CFA制备大鼠慢性炎性痛模型,前两组则在相同部位注射生理盐水。第二组和第四组大鼠分别于造模前1 h给予T10腹腔注射,随后每12 h给予腹腔注射药物1次,持续用药7 d;第一组和第三组则以相同方式给予100 ml/kg的生理盐水作为对照。采用辐射热法和机械刺激法连续观察给药后大鼠的痛行为;应用免疫组织化学染色和Western Blot方法观察连续给予T10 3d和7 d后大鼠腰膨大平面脊髓背角内小胶质细胞和星形胶质细胞的活化程度;应用实时定量PCR(RT-PCR)方法观察连续给药7 d后大鼠腰5背根神经节内炎性因子,如白细胞介素-6(interleukin-6,IL-6)、白细胞介素-1β(interleukin-1 beta,IL-1β)和肿瘤坏死因子-α(tumour necrosis factor-α,TNF-α)的表达变化。结果:(1)与CFA+Veh组相比,CFA+T10组大鼠机械缩足阈值及辐射热痛潜伏期显著下降(P<0.05),并持续到造模成功后7 d,表明腹腔注射T10可以明显改善CFA诱导的大鼠机械痛敏和辐射热痛敏。(2)免疫组织化学染色和Western Blot结果显示:造模后3 d和7 d后CFA+Veh组大鼠脊髓背角出现大量CFA诱导活化的小胶质细胞和星形胶质细胞;小胶质细胞标记物IBa-1和星形胶质细胞标记物GFAP的表达量分别在CFA造模后3 d和7 d显著增高(P<0.05),而给予T10 3 d和7 d后炎性痛大鼠腰膨大平面IBa-1和GFAP的活化程度均显著减轻(P<0.05)。(3)RT-PCR结果显示:与Saline+Veh组相比,CFA造模7 d后腰5背根神经节内炎性因子IL-1β、IL-6 mRNA和TNF-αmRNA的表达显著增高(P<0.05),腹腔注射T10 7 d后上述炎性因子的表达则显著降低(P<0.05)。结论:腹腔注射T10可以显著改善CFA诱导的大鼠慢性炎性痛,其机制可能与抑制脊髓背角小胶质细胞和星形胶质细胞的活化以及炎性因子IL-1β、IL-6和TNF-α的合成有关。
Objective：To observe the improvement and potential mechanism of intraperitoneal injection of tripterygium wilfordii hook F（ T10） on complete Freund＇s adjuvant（ CFA）-induced chronic inflammatory pain behavior in rats.Methods：SD rats were divided into 4 groups randomly,Saline ＋ Veh group,Saline ＋ T10 group,CFA ＋ Veh group and CFA ＋ T10 group,and 6 rats in each group.For the last two groups,CFA was injected into the right hindpaw to establish the inflammatory pain model;while the normal saline was injected into the same regions in first two groups.For the second and forth group rats,T10 was injected intraperitoneally 1 h before the injection of CFA and each 12 h after the iniection of CFA for 7 d.At the same time,a same dose of saline instead of T10 was used in the first and third group rats as control groups.Radiant heat and mechanical stimulation methods were applied to investigate the of pain behavior continually.Immunohistochemical staining and Western Blot were used to qualitatively and semi-quantitatively analyze the microglia and astrocyte activation in spinal cord dorsal horn after intraperitoneal injection of T10 on 3 d and 7 d.Real-time PCR（ RT-PCR） was applied to qualitatively analyze the inflammation factors,such as interleukin-6（ IL-6）,interleukin-1 beta（ IL-1β）,tumor necrosis factor-alpha（ TNF-α） in lumbar 5 ganglia after a 7 d consecutive intraperitoneal injection of T10.Results：（ 1） Compared with CFA ＋ Veh group,mechanical withdraw threshold and the thermal withdraw latency showed significant decrease in CFA ＋ T10 groups（ P〈0.05）,which also lasted until to the ends of the experiment on 7d,indicating intraperitoneal injection of T10 can significantly improve CFA induced mechanical and thermal hyperalgesia in rats.（ 2） Immunohistochemical staining and Western Blot results showed that on 3 d and 7 d after CFA injection,large number of CFA induced activation of microglia and astrocyte were observed respectively at spinal dorsal horn in CFA ＋Veh group rats.The expression of microglia marker IBa-1 and astrocyte marker GFAP both increased significantly on day3 and 7 respectively after CFA injection（ P〈0.05）.However,after administration of T10 for 3 or 7 d the IBa-1 and GFAP activation on lumbar enlargement in inflammatory pain rats were obviously diminished（ P〈0.05）.（ 3） RT-PCR results showed that the mRNA expression of inflammatory factors such as IL-1β,IL-6 and TNF-α in L5 spinal nerve ganglion were significantly increased on 7 d after injection of CFA（ P〈0.05）,and the 7 days＇ intraperitoneal administration of T10 could distinctly depress the increase tendency（ P〈0.05）.Conclusion：Intraperitoneal injection of T10 can significantly improve the CFA-induced chronic inflammatory in rats.The underlying mechanism of the potential anti-allodynia effect of T10 may be related to the suppression of spinal microglial cells and astrocyte activition.
出处
《神经解剖学杂志》
CAS
CSCD
北大核心
2014年第6期621-628,共8页
Chinese Journal of Neuroanatomy
基金
国家自然科学基金(81371238)
