脊髓PI3K/Akt信号通路在大鼠骨癌痛维持中的作用：与小胶质细胞活化的关系

Role of spinal PI3K/Akt signaling pathway in maintenance of bone cancer pain in rats： the relationship with microglial activation

目的 评价脊髓磷脂酰肌醇-3-激酶/丝氨酸-苏氨酸蛋白激酶（PDK/Akt）信号通路在大鼠骨癌痛维持中的作用及其与小胶质细胞活化的关系.方法 健康雌性SD大鼠40只,体重180 ～200 g,采用随机数字表法将其分为5组（n=8）：假手术组（S组）、PI3K抑制剂LY294002组（L组）、骨癌痛组（BCP组）、骨癌痛＋二甲基亚砜组（BCP＋D组）、骨癌痛＋LY294002组（BCP＋L组）.采用左侧胫骨干骺端骨髓腔内接种Walker256乳腺癌细胞的方法制备大鼠骨癌痛模型.于接种后7-9 d,L组和BCP＋L组鞘内注射LY294002 2.5 μg/10μL,S组和BCP组鞘内注射生理盐水10μl,BCP＋D组鞘内注射5％ DMSO 10μL,1次/d.于接种前1d及接种后1、3、5、7、8、9d给药后测定机械痛阈.于接种后9d行为学测试结束后处死大鼠取L4-6脊髓组织,采用免疫荧光染色法检测脊髓背角小胶质细胞的活化水平.结果 与S组比较,BCP组、BCP＋D组和BCP＋L组机械痛阈降低,脊髓背角小胶质细胞活化水平升高（P＜0.01）.与BCP组和BCP＋D组比较,BCP＋L组机械痛阈升高,脊髓背角小胶质细胞活化水平降低（P＜0.05）.结论 脊髓PI3K/Akt信号通路可能通过活化背角小胶质细胞参与大鼠骨癌痛的维持.

Objective To evaluate the role of spinal phosphatidyl-inositol 3-kinase/Akt （PI3k/Akt） signaling pathway in the maintenance of bone cancer pain （BCP） in rats and its relationship with microglial activation.Methods Forty healthy female Sprague-Dawley rats,weighing 180-200 g,were randomly divided into 5 groups （n =8 each）：sham operation group （group S） ; PI3K inhibitor LY294002 group （group L） ; group BCP; BCP ＋ dimethyl sulfoxide （DMSO） group （group BCP ＋ D） ; BCP ＋ LY294002 group （group BCP ＋ L）.BCP was induced by inoculating Walker 256 mammary gland carcinoma cells into the medullary cavity of the left tibia.At 7-9 days after inoculation,LY294002 2.5 μg/10 μl was injected intrathecally in L and BCP ＋ L groups,normal saline 10 μl was injected intrathecally in S and BCP groups,and 5% DMSO 10 μl was injected intrathecally in BCP＋ D group once a day.Mechanical paw withdrawal threshold （MWT） was measured at 1 day before inoculation and 1,3,5,7,8 and 9 days after inoculation.The rats were sacrificed after MWT was measured on day 9 after inoculation and the L4-6 segments of the spinal cord were removed to determinate the activation of spinal microglia using immunofluorescence.Results Compared with group S,MWT was significantly decreased,and the activation of spinal microglia was increased in BCP,BCP ＋ D and BCP＋ L groups.Compared with BCP and BCP ＋ D groups,MWT was significantly increased,and the activation of spinal microglia was decreased in BCP ＋ D group.Conclusion Spinal PI3K/Akt signaling pathway is involved in the maintenance of BCP possibly through activating microglia in spinal dorsal horns of rats.