龟鹿益神配方颗粒对慢性疲劳大鼠行为和骨骼肌线粒体的影响

发布时间：2018-08-14 18:01

【摘要】：目的：建立慢性疲劳大鼠模型，观察龟鹿益神配方颗粒对慢性疲劳大鼠一般情况和行为学指标（力竭游泳时间、鼠尾悬吊静止时间）的影响，电镜下观察慢性疲劳大鼠骨骼肌线粒体微观结构的变化，检测血清琥珀酸脱氢酶活性的变化，探讨慢性疲劳综合征的发病机理；观察龟鹿益神配方颗粒对慢性疲劳大鼠骨骼肌线粒体微观结构和血清SDH的影响，探讨其作用机制，为龟鹿益神配方颗粒治疗CFS的临床应用提供实验依据和理论支持。方法：雄性SPF级SD大鼠40只，体重200±20克，适应性饲养1周后，随机抽取16只不进行造模，其余24只进行造模。采用强制力竭游泳，束缚和夹尾复合因素刺激，构建慢性疲劳大鼠模型。造模成功后，未造模者随机分为两组，分别为正常组和正常对照组；造模者随机分为3组，分别为模型组、苁蓉益肾组、龟鹿益神组。从造模结束次日起，在正常饲养的基础上，正常组、模型组给予生理盐水灌胃；正常对照组、龟鹿益神组给予龟鹿益神配方颗粒混悬液灌胃；苁蓉益肾组给予苁蓉益肾颗粒混悬液灌胃。各组大鼠均给药1次/d，连续14天。造模前、造模后及末次给药后均测一次大鼠体重、力竭游泳时间及鼠尾悬吊静止时间。上述实验结束后，大鼠经腹腔麻醉后，取各组大鼠右后肢骨骼肌组织一块，电镜下观察线粒体结构。经腹主动脉采血分离上清液，采用ELISA法测定各组大鼠血清SDH活性。采用IBM SPSS19.0统计软件对数据进行处理。结果：（1）造模结束后，各组造模大鼠体重增长缓慢甚至减轻，饮食量、饮水量减少，眯眼懒动，便溏，毛发暗淡失去光泽。与正常组比较，正常对照组体重、力竭游泳时间、鼠尾悬吊静止时间无显著性差异(P0.05)；模型组、龟鹿益神组、苁蓉益肾组大鼠体重减轻、力竭游泳时间缩短、鼠尾悬吊静止时间延长均有显著性差异(P 0.05)。给药结束后，与正常组相比，正常对照组力竭游泳时间延长有显著性差异，龟鹿益神组无显著性差异，两组大鼠体重、鼠尾悬吊静止时间均无显著性差异(P0.05)；苁蓉益肾组体重减轻有显著性差异（P0.05），力竭游泳时间和鼠尾悬吊静止时间均缩短无显著性差异(P0.05)；模型组大鼠体重减轻、力竭游泳时间变短、鼠尾悬吊静止时间延长均有显著性差异（P0.05）。与模型组相比，龟鹿益神组、苁蓉益肾组大鼠体重明显增加、力竭游泳时间明显延长、鼠尾悬吊静止时间缩短均有显著性差异（P0.05）。与苁蓉益肾组相比，龟鹿益神组大鼠体重增加、鼠尾悬吊静止时间缩短均无显著性差异（P0.05），力竭游泳时间延长有显著性差异（P0.05）。（2）模型组大鼠骨骼肌肌原纤维排列紊乱，Z线、M线模糊且排列紊乱，线粒体形状不规则，体积较正常组明显偏小，多数线粒体出现空泡，膜溶解并相互连接。与模型组相比，龟鹿益神组和苁蓉益肾组大鼠骨骼肌肌原纤维排列较整齐，Z线、M线清晰且整齐，线粒体体积大，空泡样变少。与正常组对比，正常对照组大鼠骨骼肌排列更整齐，Z线、M线清晰且排列更有规则，线粒体结构更完整，且体积大。（3）与正常组相比，正常对照组血清SDH活性明显降低，有显著差异（P0.05）；龟鹿益神组大鼠血清SDH活性轻度升高，无显著性差异（P0.05）；苁蓉益肾组、模型组大鼠血清SDH活性升高，有显著性差异（P0.05）。与模型组比较，苁蓉益肾组、龟鹿益神组血清SDH活性降低，有显著性差异（P0.05）。与苁蓉益肾组相比，龟鹿益神组血清SDH活性降低，有显著性差异（P0.05）。结论：（1）本实验采用非病毒感染的慢性应激刺激方法，即强制力竭游泳，束缚和夹尾激怒的复合因素刺激，构建慢性疲劳大鼠模型。此方法操作性强，能够模拟导致人类疲劳的内外环境，可复制出人体的慢性疲劳状态。（2）龟鹿益神配方颗粒可以延长慢性疲劳大鼠力竭游泳时间，缩短鼠尾悬吊静止时间，保护线立体结构的完整性，降低血清SDH的活性。（3）本实验表明，慢性疲劳大鼠骨骼肌及其线粒体结构易损伤，且不易恢复，血清SDH活性明显升高。龟鹿益神配方颗粒可增强骨骼肌及其线粒体膜的稳定性，促进其损伤的恢复，，增强线粒体功能，降低血清SDH活性。提示龟鹿益神配方颗粒可通过保护线粒体结构、增强其功能从而达到抗疲劳的作用，进一步阐释了慢性疲劳综合征的发病机制及龟鹿益神配方颗粒治疗该病的作用机理，为临床应用龟鹿益神配方颗粒治疗慢性疲劳综合征提供了实验依据。（4）龟鹿益神配方颗粒可作为CFS的预防性用药。
[Abstract]:Objective: To establish a chronic fatigue rat model and observe the effects of Guilu Yishen Formula Granules on the general condition and behavioral indexes (exhaustive swimming time, resting time of rat tail suspension) of chronic fatigue rats. To explore the pathogenesis of chronic fatigue syndrome, observe the effect of Guilu Yishen Formula Granule on the microstructure of skeletal muscle mitochondria and serum SDH in rats with chronic fatigue, and explore its mechanism of action, so as to provide experimental basis and theoretical support for the clinical application of Guilu Yishen Formula Granule in treating CFS.
METHODS: Forty male SPF SD rats weighing 200 65 The control group was randomly divided into three groups: model group, Cistanche deserticola Yishen group and Guilu Yishen group. On the basis of normal feeding, normal group was given normal saline by gastric lavage; normal control group, Guilu Yishen formula suspension was given by gastric lavage; Cistanche deserticola Yishen group was given Cistanche deserticola Yishen formula suspension. The rats in each group were given the suspension once a day for 14 consecutive days. The body weight, exhaustive swimming time and tail suspension rest time were measured before modeling, after modeling and after the last administration. The supernatant of abdominal aorta was collected and the SDH activity was determined by ELISA. The data were processed by IBM SPSS 19.0 statistical software.
Results: (1) After modeling, the body weight of rats in each group increased slowly or even decreased, diet, water intake decreased, squint lazy, stool, hair dull loss of luster. Compared with the normal group, there was no significant difference in body weight, exhaustive swimming time, tail suspension rest time (P 0.05); model group, Guilu Yishen group, cirong Yishen group There were significant differences in weight loss, exhaustive swimming time and tail suspension rest time between the two groups (P 0.05). There were significant differences in weight loss (P 0.05), exhaustive swimming time and tail suspension rest time (P 0.05), body weight loss, exhaustive swimming time shortened and tail suspension rest time prolonged in model group (P 0.05). The weight of rats in Yishen group was significantly increased, the time of exhaustive swimming was significantly prolonged, and the rest time of tail suspension was significantly shortened (P 0.05).
(2) The myofibrils of skeletal muscle in model group were arranged disorderly, Z-line, M-line were blurred and arranged disorderly, the shape of mitochondria was irregular, and the volume of mitochondria was smaller than that of normal group. Most of mitochondria were vacuole, membrane dissolved and connected with each other. Compared with the normal group, the skeletal muscles of the normal control group were arranged more orderly, Z-line and M-line were clear and arranged more regularly, and the mitochondrial structure was more complete and larger.
(3) Compared with the normal group, the serum SDH activity of the normal control group decreased significantly (P 0.05); the serum SDH activity of the Guilu Yishen group increased slightly (P 0.05); the serum SDH activity of the Cistanche Yishen group increased significantly (P 0.05). Compared with the model group, the serum SDH activity of the Cistanche Yishen group and the Guilu Yishen group increased slightly (P 0.05). The activity of serum SDH in Guilu Yishen group was significantly lower than that in Cistanche deserticola Yishen group (P 0.05).
CONCLUSIONS: (1) A rat model of chronic fatigue was established by means of non-viral chronic stress stimulation, i.e. forced exhaustive swimming, restraint and tail-clip irritation. This method can simulate the internal and external environment of human fatigue and replicate the chronic fatigue state of human body.
(2) Guilu Yishen Formula Granule can prolong the exhaustive swimming time of rats with chronic fatigue, shorten the resting time of rat tail suspension, protect the integrity of the three-dimensional structure of the line, and reduce the activity of serum SDH.
(3) The results showed that the structure of skeletal muscle and its mitochondria was easy to be damaged, and it was not easy to recover, and the activity of serum SDH was obviously increased. Guilu Yishen formula granule could enhance the stability of skeletal muscle and its mitochondrial membrane, promote the recovery of injury, enhance the function of mitochondria and reduce the activity of serum SDH. The mechanism of chronic fatigue syndrome and the mechanism of Guilu Yishen Formula Granule in treating chronic fatigue syndrome were further elucidated, which provided experimental basis for clinical application of Guilu Yishen Formula Granule in treating chronic fatigue syndrome.
(4) Gui Lu Yi Shen formula granule can be used as a preventive drug for CFS.
【学位授予单位】：河南中医学院
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R285.5

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