从阴阳自和角度探讨灰树花多糖对CIF模型线粒体自噬蛋白PINK1和Parkin的调节效应

Effects of Grifola Frondosa Polysaccharides on Mitophagy Proteins PINK1 and Parkin in CIF Model from the Perspective of Yin and Yang

目的:探讨灰树花多糖调节化疗相关性疲劳(CIF)模型骨骼肌线粒体异常的机制。方法:将60只6~8周龄SPF级C57BL/6小鼠随机分为空白组、模型组、灰树花多糖低剂量组(5 mg·kg-1)和灰树花多糖高剂量组(10 mg·kg-1),每组15只。将除空白组以外的小鼠腹腔注射5-氟尿嘧啶(5-Fu,40 mg·kg-1),每天给药1次,连续5 d,同时灰树花多糖低、高剂量组分别给予不同剂量灰树花多糖灌胃,空白组和模型组给予等量生理盐水灌胃,期间测量体质量及抓力。第13天处死动物取腓肠肌,测SOD和MDA含量,透射电镜观察腓肠肌线粒体形态,Western Blot法检测线粒体自噬相关蛋白PINK1和Parkin的表达情况。结果:与空白组相比,模型组抓力显著下降(P<0.05),SOD显著降低(P<0.01),MDA显著升高(P<0.01),灰树花多糖组抓力显著高于模型组(P<0.05),灰树花多糖高剂量组SOD显著高于模型组(P<0.05),灰树花多糖组MDA显著低于模型组(均P<0.01)。电镜结果显示模型组骨骼肌线粒体肿胀变形,且有大量空泡变性,线粒体内嵴排列紊乱或不清晰,灰树花多糖组骨骼肌肌节仍有部分排列紊乱或断裂现象,但可见较多自噬的线粒体。Western Blot结果显示模型组线粒体自噬相关蛋白PINK1和Parkin的表达显著下降(P<0.01),灰树花多糖可显著增加PINK1和Parkin的表达(P<0.01)。结论:灰树花多糖可以改善CIF骨骼肌SOD与MDA失衡,增强线粒体自噬相关蛋白PINK1和Parkin的表达,促进线粒体自噬。

Objective:To investigate the regulating mechanism of grifola frondosa polysaccharide on mitochondrial abnormalities in skeletal muscle of chemotherapy induced fatigue(CIF) model. Method:Sixty 6~8 week SPF C57 BL/6 mice were randomly divided into 4 groups, namely normal group, model group, low dose grifola frondosa polysaccharide group(5 mg·kg-1) and high dose grifola frondosa polysaccharide group(10 mg·kg-1). The mice were injected 5-fluorouracil(40 mg·kg-1) daily for five days except normal group. At the same time,low dose grifola frondosa polysaccharide group and high dose grifola frondosa polysaccharide group were given different doses of grifola frondosa polysaccharide. Body mass and grip strength were measured during the experimental period. On the 13 rd day, the mice were sacrificed to inspect the contents of SOD and MDA in gastrocnemius muscle of CIF mice. The morphology of gastrocnemius muscle mitochondria was observed by transmission electron microscope. The expressions of mitophagy related proteins PINK1 and Parkin were detected by Western Blot. Result:The grip strength of the model group decreased significantly(P<0.05),SOD decreased significantly(P<0.01) and MDA increased significantly(P<0.01) compared with those of the normal group.The grip strength of grifola frondosa polysaccharide group was increased significantly(P<0.05), SOD increased significantly(P<0.05) and MDA decreased significantly(P<0.01) compared with those of the model group(P<0.01). Electron microscopy results showed that there were mitochondria swelling, vacuolar degeneration and mitochondrial crista disorder(or not clearly) in skeletal muscle of model group, but there was more mitochondrial autophagy in grifola frondosa polysaccharide groups though skeletal muscle sarcomere disorder or fracture still existed. Western blot results showed that the PINK1 and Parkin proteins expressions in model group decreased significantly(P<0.01), but grifola frondosa polysaccharides significantly increased the expressions of PINK1 and Parkin protein(P<0.01). Conclusion:Grifola polysaccharides can improve the imbalance of SOD and MDA in CIF skeletal muscles and promote the expressions of mitophagy related protein PINK1 and Parkin.