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目的:研究核转录因子NF-κB信号在一次大强度运动后泛素蛋白酶体途径降解骨骼肌蛋白质的作用。方法:42只雄性SD大鼠随机分为2组,对照组24只和注射组18只,进行1次大强度上坡跑运动(25m/min,5%坡度,1 h)。运动前1 h,注射组腹腔注射0.25 ml/100 g体重的8%PDTC(NF-κB信号阻断剂)溶液;对照组腹腔注射0.25 ml/100 g体重pH7.4PBS溶液。运动前从对照组中取材6只,做为安静时对照,运动后即刻和运动后1、6 h分别从每组取材6只。用EMSA测定腓肠肌细胞核NF-κB结合DNA活性,Western Blot测定细胞质IκBα蛋白含量,Real-Time PCR测定MuRF1、MAFbxmRNA含量,HPLC测定肌肉3-MH含量。结果:1)对照组IκBα蛋白含量在运动后即刻(P<0.01)、运动后1 h(P<0.01)较安静时非常显著降低,注射组IκBα蛋白含量在运动后即刻(P<0.05)运动后1 h(P<0.01)显著高于对照组。2)对照组运动后1 h细胞核NF-κB结合DNA能力非常显著高于安静时(P<0.01)和运动后即刻(P<0.01),注射组运动后即刻(P<0.05)、运动后1 h(P<0.01)显著低于同时间的对照组。3)对照组运动后1、6 h腓肠肌MAFbx、MuRF1 mRNA含量较安静时和运动后即刻非常显著升高(P<0.01),而注射组运动后1 h显著低于对照组(P<0.05)。4)对照组运动后即刻、运动后6 h3-MH含量非常显著高于安静时和运动后1 h(P<0.01)。注射组运动后6 h显著低于对照组(P<0.05)。结论:1次大强度运动后,骨骼肌泛素连接酶基因表达增加,激活泛素蛋白酶体途径活性,加强蛋白质降解。阻断NF-κB信号,可降低泛素连接酶基因表达,有助于减少运动后骨骼肌蛋白质降解。
Objective: To study the effect of NF-κB signaling on degradation of skeletal muscle protein by ubiquitin-proteasome pathway after a high-intensity exercise. Methods: Forty-two male Sprague-Dawley rats were randomly divided into two groups: control group (n = 24) and injection group (n = 18). One hour before exercise, the injection group was intraperitoneally injected with 0.25 ml / 100 g body weight of 8% PDTC (NF-κB signal blocker) solution; the control group was intraperitoneally injected with 0.25 ml / 100 g body weight of pH7.4 PBS solution. Six rabbits were selected from the control group before exercise, and were taken as control when quiet. Six rabbits were drawn from each group immediately after exercise and at 1,6 hours after exercise. The activity of NF-κB-binding DNA in gastrocnemius muscle cells was measured by EMSA. The content of IκBα in cytoplasm was determined by Western Blot. The contents of MuRF1 and MAFbx mRNA were determined by Real-Time PCR. The content of 3-MH was determined by HPLC. Results: 1) The content of IκBα in the control group was significantly decreased immediately after exercise (P <0.01) and 1 h after exercise (P <0.01), and the content of IκBα protein in the injected group was increased immediately after exercise (P <0.05) After 1 h (P <0.01) was significantly higher than the control group. 2) Compared with resting (P <0.01) and immediately after exercise (P <0.01), the NF-κB DNA binding capacity of the control group was significantly higher than that of the control group 1 h after exercise (P <0.05) h (P <0.01) was significantly lower than the control group at the same time. 3) Compared with the control group, the contents of MAFbx and MuRF1 mRNA in the gastrocnemius muscle increased significantly at 1 and 6 h after exercise (P <0.01), while those at 1 h after injection were significantly lower than those in the control group (P <0.05) . 4) Immediately after exercise, the level of 3-MH in 6 h after exercise was significantly higher than that at rest and 1 h after exercise (P <0.01). The injection group 6 h after exercise was significantly lower than the control group (P <0.05). Conclusion: After a period of intensive exercise, the ubiquitin ligase gene expression in skeletal muscle increased, activating ubiquitin proteasome pathway activity and enhancing protein degradation. Blocking NF-κB signaling reduces ubiquitin ligase gene expression and helps to reduce skeletal muscle protein degradation after exercise.