Objective: To investigate the expression regulation of inflammation cytokines interleukin 4 (IL-4), IL-6, IL-13, and tumor necrosis factor α (TNF-α) in rats with sciatic nerve defect following olfactory ensheathing cell (OEC) transplantation.
Methods: The primary OEC for cell culture and identification was dissociated from the olfactory bulb of the green fluorescent protein-Sprague Dawley (GFP-SD) rat. One hundred SD rats were randomly divided into 2 groups, and the right sciatic nerve defect (10 mm in length) model was made, then repaired with poly (lactic acid-co-glycolic acid) (PLGA). The mixture of equivalent cultured GFP-OEC and extracellular matrix (ECM) was injected into both ends of PLGA nerve conduit in the experimental group (n=55), and the mixture of DMEM and ECM in the control group (n=45). The general situation of rats was observed after operation. At 6 hours, 1 day, 3 days, 1 week, 2 weeks, 3 weeks, 4 weeks, and 6 weeks, the inflammatory cytokines were detected by Western blot. At 2, 4, and 6 weeks, the survival of GFP-OEC was observed in the experimental group. At 9 weeks, HE staining was used to observe the morphology of nerve tissue, and the sensory and motor function and the electrophysiological index were detected.
Results: The cultured primary cells were GFP-OECs by immunofluorescence staining. Compared with the control group, the experimental group showed significantly increased expression level of IL-4 at 2-6 weeks (P<0.05), significantly decreased expression level of IL-6 and TNF-α at 3 days and 1 week (P<0.05) and significantly increased expression level of IL-13 at 1 day and 3-6 weeks (P<0.05) by Western blot detection. At 2, 4, and 6 weeks, the surviving GFP-OEC of regenerative nerve end was observed in the experimental group under the fluorescence microscope. At 9 weeks, regenerative nerve tissue was loose, and cell morphology was irregular in the experimental group, while the regenerative nerve tissue had vesicular voids and the cell number decreased significantly in the control group. At 9 weeks, the functional recovery of sciatic nerve in the experimental group was better than that of the control group, showing significant difference in the lateral foot retraction time, sciatic nerve function index, muscle action potential latency, and the amplitude of compound muscle action potential (P<0.05).
Conclusions: OEC can promote the anti-inflammation cytokines expression of IL-4 and IL-13 and inhibit the pro-inflammatory cytokines expression of IL-6 and TNF-α, which can improve the local inflammatory microenvironment of sciatic nerve and effectively promote the structure and function recovery of sciatic nerve.
目的: 探讨嗅鞘细胞（olfactory ensheathing cell，OEC）移植修复大鼠坐骨神经缺损后，其在修复过程中对坐骨神经局部微环境内炎性因子表达的调节作用。.
方法: 取绿色荧光蛋白（green fluorescent protein，GFP）转基因SD大鼠嗅球，原代培养GFP-OEC并鉴定。取100只SD大鼠随机分为两组，制备右侧坐骨神经缺损（10 mm长）模型后，采用聚乳酸-聚羟基乙酸共聚物[poly（lactic acid-co-glycolic acid），PLGA]神经导管修复缺损处，实验组（n=55）于导管两端注入GFP-OEC与细胞外基质（extracellular matrix，ECM）的混合液，对照组（n=45）注入等量DMEM与ECM的混合液。术后观察大鼠一般情况，于6 h、1 d、3 d以及1、2、3、4、6周Western blot检测IL-4、IL-6、IL-13和TNF-α的表达；术后2、4、6周实验组取材观察GFP-OEC存活情况；9周两组取材行HE染色观察神经组织形态，并行坐骨神经感觉及运动功能检测、神经电生理检测。.
结果: 经免疫荧光染色鉴定，原代培养细胞为GFP-OEC。Western blot检测显示，与对照组比较，2～6周实验组IL-4相对表达量明显升高，3 d、1周时IL-6及TNF-α相对表达量降低，1 d以及3～6周时IL-13相对表达量升高，比较差异有统计学意义（P＜0.05）。术后2、4、6周荧光显微镜下观察，实验组神经移植体内的神经再生端均有GFP-OEC存活；术后9周，HE染色示实验组再生神经组织较松散，细胞形态不规则；对照组再生神经组织成泡状空隙，细胞明显减少。术后9周，实验组大鼠坐骨神经功能恢复程度优于对照组，后足回缩时间、坐骨神经功能指数、肌肉动作电位潜伏期及复合肌肉动作电位波幅比较，差异均有统计学意义（P＜0.05）。.
结论: 坐骨神经缺损后植入OEC可促进抗炎因子IL-4和IL-13的表达，同时抑制促炎因子IL-6和TNF-α的表达，改善了坐骨神经局部的炎症微环境，进而有效地促进坐骨神经结构和功能的修复重建。.
Keywords: Anti-inflammation cytokine; Olfactory ensheathing cell; Peripheral nerve regeneration; Pro-inflammatory cytokine; Rat.