破裂型椎间盘突出动物模型中自身免疫反应的实验研究

Experimental study on the abnormal autoimmunity in the model of ruptured lumbar disc herniation

中文关键词:  坐骨神经痛模型  自身免疫  抗原抗体复合物  细胞因子

英文关键词:Sciatica model without mechanical pressure  Abnormal autoimmunity  Antigen-antibody complexes  Cytokine

中文摘要:

  【摘要】 目的：从动物实验的角度探讨破裂型椎间盘突出动物模型中的自身免疫反应。方法：20只SD大鼠分为两组，实验组采用自体髓核移植于坐骨神经旁的方法建立破裂型椎间盘突出动物模型（坐骨神经痛模型）；对照组大鼠手术方法同实验组，但不放置髓核。术前及术后1、2、3周时采用爬坡实验及后肢机械缩爪阈值测定评估大鼠造模前后后肢运动能力及痛觉过敏的变化。术后3周时处死动物应用透射比浊法检测大鼠血清中免疫球蛋白IgG、IgM的含量，ELISA法测定血清中TNF-α、IL-6、IL-12的含量；采用免疫组化染色观察移植髓核中抗原抗体复合物沉积情况；应用BCA蛋白定量法观察坐骨神经中磷脂酶A2（PLA2）的活性。结果：所有大鼠模型建立前爬坡试验结果均为Ⅳ级，造模后对照组爬坡试验仍为Ⅳ级，实验组在造模后1、2及3周时爬坡试验均为Ⅲ级。实验组大鼠造模后1、2、3周时的后肢机械缩爪阈值分别为67.2±8.4、41.3±5.2及40.7±5.3mmHg，较术前（90.4±5.0mmHg）及对照组明显降低（P<0.01），出现较为明显的痛觉过敏。术后3周实验组大鼠血清中IgG含量4.98±0.96g/L及IgM含量1.45±0.37g/L较对照组（4.31±0.77g/L及0.79±0.35g/L）明显上升（P<0.05）；血清中TNF-α、IL-6、IL-12的含量（205.77±46.32pg/L，186.4±87.3pg/L，69.23±27.46pg/L）较对照组（11.01±2.53pg/L，85.0±13.2pg/L，21.65±11.93pg/L）均明显升高（P<0.05）。实验组大鼠移植髓核中出现抗原抗体复合物阳性沉积，阳性率为80%，明显高于对照组（P<0.01）；其坐骨神经中PLA2活性为0.0766±0.0039μmoL/（min·L），较对照组0.0006±0.0010μmoL/（min·L）明显升高（P<0.05）。结论：破裂型椎间盘突出动物模型中存在着由移植髓核引起的全身及局部异常的自身免疫反应，这可能是导致其疼痛的主要原因。

英文摘要:

  【Abstract】 Objectives: To investigate the abnormal autoimmunity in the model of ruptured lumbar disc herniation. Methods: 20 SD rats were divided into experimental group and control group, a lumbar disc herniation model without mechanical pressure(sciatica model) was established on Sprague Dawley rat by putting nucleus pulposus near the nerve root. The mechanical withdrawal thresholds of posterior limbs and slope-climbing test were measured at preoperation and 1, 2, 3 weeks after operation. 3 weeks later, the rats were sacrificed, and the immunoglobulin G(IgG) and immunoglobulin M(IgM) in serum were examined by turbidimetry. The presence and distribution of antigen-antibody complexes in nucleus pulposus were examined by immunohistochemical method. And the levels of cytokine TNF-α, IL-6, IL-12 were detected by enzyme linked immunosorbent assay, and the BCA method was used to analyze the activity of PLA2 in the sciatic nerve. Results: Before model establishment, all rats had IV grade slope-climbing, after surgery, the rats in experimental group had Ⅲ grade slope-cimbing, while the rats in control group maintained IV grade slope-climbing. After the model establishment, the mechanical withdrawal thresholds of posterior limbs significantly decreased (from 40.7±5.3mmHg to 90.4±5.0mmHg, P<0.01), the levels of IgG and IgM in serum elevated significantly(4.98±0.96g/L and 1.45±0.37g/L respectively, P<0.05), meanwhile the levels of cytokine TNF-α, IL-6, IL-12 elevated remarkably(205.77±46.32pg/L, 186.4±87.3pg/L and 69.23±27.46pg/L, P<0.05). There was positive presence of antigen-antibody complexes in the nucleus pulposus(P<0.01), and obvious inflammatory lesion in the sciatic nerve was observed after model establishment. Conclusions: There is abnormal autoimmunity in the sciatica model induced with no mechanical pressure, which may lead to pain.