他莫昔芬对大鼠脊髓损伤后炎性反应及细胞凋亡的影响

目的检测他莫昔芬对脊髓损伤大鼠核因子kappa B抑制蛋白激酶复合体/核因子kappa B（IKK/NF-κB）通路及细胞凋亡的影响,研究其神经保护机制。方法将51只Sprague-Dawley（SD）大鼠随机分为3组（每组17只）：假手术组（仅行椎板切除术）、对照组（脊髓损伤后硬膜下注射2%二甲基亚砜5 m L/kg）和他莫昔芬组[脊髓损伤后硬膜下注射他莫昔芬5 mg/kg（溶于2%二甲基亚砜1 mg/m L）]。改良Allen法（25 g·cm）制作T12节段脊髓损伤模型。术后24 h采用免疫印迹法（Western blot）检测损伤节段脊髓核因子kappa B p65（NF-κB p65）、磷酸化核因子κB抑制因子α（p I-κBα）和活性半胱天冬酶-3（caspase-3）表达水平,应用Gel Pro Analyser 4.0定量灰度值（OD）;比色法检测脊髓组织中髓过氧化物酶（MPO）活性。统计方法采用单因素方差分析和Bonferrni检验。结果与假手术组相比,对照组和他莫昔芬组术后24 h炎症相关指标NF-κB p65[（0.31±0.03）比（3.17±0.12）比（1.55±0.07）,P〈0.05]、p I-κBα[（0.12±0.01）比（0.98±0.05）比（0.53±0.03）,P〈0.05]的表达水平及MPO活性较高[（0.06±0.01）U/mg比（0.64±0.03）U/mg比（0.35±0.02）U/mg,P〈0.05],但他莫昔芬组低于对照组,组间差异有统计学意义（P〈0.05）;此外,凋亡相关的活性caspase-3在假手术组表达水平极低,另外两组则明显升高,其中对照组升高更为显著[（0.08±0.01）比（1.03±0.05）比（0.36±0.02）,P〈0.05]。结论他莫昔芬能够调控脊髓损伤部位的细胞凋亡,并减轻炎性反应,从而发挥神经保护作用。

Effect of Tamoxifen on inflammation and apoptosis after spinal cord injury in rats

Objective To investigate neuroprotective effect of Tamoxifen after spinal cord injury（SCI） by examining factors influencing IKK/NF-κB pathway and apoptosis in rats. Methods 3 groups of SD rats（n=17 each） were randomly assigned to sham group（laminectomy was performed only without SCI）, control group（SCI＋subdural injection of 2%dimethylsulfoxide 5 m L/kg） and Tamoxifen group（SCI ＋subdural injection of same volume of 5 mg/kg Tamoxifen dissolved in 2% dimethylsulfoxide）. SCI was induced by using the modified weight-drop method（25 g·cm） at T12 level.24 hours after surgery, the expression of NF-κB p65, phosphorylated I-κB α and active caspase-3 from a 10 mm spinal cord segment containing the injury epicenter were detected by Western blot; optical density（OD） value of each protein band was scanned and digitized by Gel Pro Analyser. The activity of myeloperoxidase（MPO） was tested by densitometry.One-way analysis of variance was used to analyze the results followed by a Bonferroni post-hoc test for multiple comparisons. Results A comparison of these 3 groups showed that compared with the sham group, the expression of NF-κB p65 [（0.31±0.03） vs（3.17±0.12） vs（1.55±0.07）, P〈0.05] and p I-κB α[（0.12±0.01） vs（0.98±0.05） vs（0.53±0.03）, P〈0.05] increased, the activity of MPO [（0.06±0.01） U/mg vs（0.64±0.03） U/mg vs（0.35±0.02） U/mg, P〈0.05]were higher in the control group and Tamoxifen group. And the administration of Tamoxifen significantly reduced the expression of NF-κB p65 and phosphorylated I-κBα compared to control, and the lower the activity of MPO confirmed less infiltration of leukocytes to the injury site in Tamoxifen group. It also attenuated the expression of active caspase-3[（0.08±0.01） vs（1.03±0.05） vs（0.36±0.02）, P〈0.05] resulting in the reduction of apoptosis. Conclusion The potential neuroprotective effects of Tamoxifen may relate to the attenuation of inflammation and apoptosis after spinal cord injur