药膳加贴敷对衰老大鼠骨骼肌抗氧化能力及细胞凋亡影响的实验研究
发布时间：2019-03-21 17:12
【摘要】：目的:本实验基于“脾主肌肉四肢”理论,选用D-半乳糖(D-a lactose,D-gal)致衰老大鼠为研究对象,采用补脾益气药膳灌胃加穴位贴敷的疗法对衰老模型大鼠进行干预,观察衰老大鼠骨骼肌形态学改变、抗氧化能力及cas pase-3和caspase-8的表达影响,了解骨骼肌质量的渐进性下降与细胞凋亡和抗氧化能力之间的内在联系和作用机制。为延缓肌肉衰减综合征(Sarcopenia)的发生提供理论依据,并最终为临床干预Sarcopenia提供有效的护理措施,以便提高老年人日常活动能力、自理能力,减少跌倒及卧床的机率,改善老年人生活质量,降低医疗保健成本及家庭和社会的经济负担。材料与方法:选取SPF(Specific pathogen Free,SPF)级3月龄健康雄性Spr ague-DaWley(SD)大鼠36只,随机分为空白组、模型空白组和药膳贴敷疗法组,每组12只。模型空白组和药膳贴敷组每日腹部皮下注射7.5%D-gal 125mg/kg,空白组每日腹部皮下注射0.9%生理盐水125mg/kg,造模持续6周。6周后,药膳贴敷组给与药膳灌胃(10ml·kg-1·d-1)和穴位贴敷干预,空白组和模型空白组给与等剂量的生理盐水和空白贴敷干预,干预持续4周。最后一次灌胃24小时后,进行腹腔麻醉,经腹主动脉取血,迅速剥离股直肌。通过HE染色镜下观察衰老大鼠股直肌细胞形态结构的变化,PCR检测股直肌含半胱氨酸的天冬氨酸蛋白水解酶3(Cysteinyl aspa rtate specifi c proteinase-3,Caspase-3)和Caspase-8mRNA的表达,ELISA检测血清中谷胱甘肽过氧化物酶(glutathione peroxidase,GSH-Px)、超氧化物歧化酶(superoxide dismutase,SOD)的活性和丙二醛(Mal ondialdehyde,MDA)的含量及股直肌GSH-Px、SOD、Cas pase-3 Caspase-8的活性和MDA的含量。采用SPSS17.0统计分析软件和Microsoft Office Excel 2007进行数据处理分析,每组数据用平均数±标准差(Mean±S)表示,组间差异采用单因素方差分析,差异显著性水平定义为P0.05。结果:1.模型空白组和药膳贴敷组大鼠都有衰老的体貌特征,但药膳贴敷组大鼠较模型空白组大鼠衰老特征有所减轻;三组大鼠体重均有增加,模型空白组大鼠略低于空白组,但组间比较无显著性差异(P0.05)。2.光镜下:空白组大鼠为正常骨骼肌组织学特征,模型空白组和药膳贴敷组大鼠都出现了不同程度的肌细胞萎缩或退行性改变,而药膳贴敷组大鼠肌细胞的萎缩程度比模型空白组较轻。3.血清学指标:模型空白组大鼠血清中GSH-Px、SOD的活性较空白组显著下降(P0.05),而MDA的含量较空白组显著增加(P0.05);与模型空白组相比,药膳贴敷组大鼠血清中GSH-Px、SOD的活性显著提高(P0.05),而MDA的含量显著降低(P0.05)。4.骨骼肌组织生化指标:与空白组相比,模型空白组大鼠骨骼肌中GSH-Px、SOD的活性显著下降(P0.05),MDA的含量及Caspase-3、Caspase-8的活性显著提高(P0.05);与模型空白组相比,药膳贴敷组大鼠骨骼肌中GSH-Px、SOD显著提高(P0.05),MDA的含量及Caspase-3、Caspase-8的活性显著下降(P0.05)。5.骨骼肌组织分子生物学指标:与空白组相比,模型空白组大鼠骨骼肌中Caspase-3、Caspase-8mRNA表达明显增加(P0.05);与模型空白组相比,药膳贴敷组大鼠模型组大鼠骨骼肌中Caspase-3、Caspase-8mRNA(P0.05)表达明显减少。结论:1.药膳加穴位贴敷疗法能够改善老年大鼠的体貌、状态及骨骼肌组织形态学特征,并且能够减缓肌肉衰减综合征的发生及发展。2.药膳加穴位贴敷疗法能够提高衰老大鼠血清及骨骼肌中GSH-Px、SOD的活性,降低MDA的含量,从而提高骨骼肌的抗氧化能力,减轻骨骼肌的氧化损伤,延缓骨骼肌的衰老。3.药膳加穴位贴敷疗法能够降低衰老大鼠骨骼肌细胞中Caspase-3和Caspase-8的活性,下调Caspase-3mRNA和Caspase-8mRN A的表达,进而减少细胞凋亡,延缓骨骼肌的衰老。
[Abstract]:Objective: To study the changes of skeletal muscle morphology in aging rats by using D-galactose (D-gal)-induced aging rats to study the aging model rats based on the "spleen main muscle and limbs" theory. The relationship between the progressive decline of skeletal muscle mass and the cell apoptosis and the anti-oxidation ability was studied by the effect of the anti-oxidation ability and the expression of caspase-8. The invention provides a theoretical basis for delaying the occurrence of the muscle attenuation syndrome (Sarcoenia), and finally provides effective nursing measures for the clinical intervention Sarcoenia, so as to improve the daily activity capability of the old people, self-adjustment, reduce the probability of falling and bedridden, improve the quality of life of the old people, And the economic burden of the health care cost and the family and the society is reduced. Materials and Methods:36 healthy male Sprague-DaWley (SD) rats were randomly divided into blank group, model blank group and medicated diet therapy group. The daily abdominal subcutaneous injection of the model blank group and the medicated diet group was 7.5% D-gal 125 mg/ kg. The daily abdomen of the blank group was injected with 0.9% normal saline of 125 mg/ kg and the model was established for 6 weeks. After 6 weeks, the medicated diet was administered to the medicated diet (10 ml 路 kg-1 路 d-1) and the acupoint application for intervention. The blank group and the model blank group were given equal dose of normal saline and blank for intervention, and the intervention lasted for 4 weeks. After the last administration for 24 hours, the abdominal cavity was anesthetized, the blood was taken from the abdominal aorta, and the rectus femoris was quickly separated. The changes of the morphological structure of the straight muscle cells of the aging rats were observed by HE staining, and the expression of cysteinyl aspa rate specific c protein-3 (Caspase-3) and Caspase-8 mRNA was detected by PCR, and the glutathione peroxidase (GSH-Px) in the serum was detected by ELISA. The activity of superoxide dismutase (SOD) and the content of MDA and the activity of GSH-Px, SOD, Cas ase-8 and the content of MDA were studied. The data was analyzed by SPSS17.0 statistical analysis software and Microsoft Office Excel 2007. The mean standard deviation (Mean-S) of each group of data indicated that the difference between the groups was defined as P0.05 with a single-factor analysis of variance. Results:1. In the model blank group and the medicated diet group, the aging characteristics of the rats in the model blank group and the medicated diet group were reduced; the weight of the three groups of rats was increased, and the rats in the blank group of the model group were slightly lower than that of the blank group, but no significant difference was found between the groups (P0.05). Under the light microscope, in the blank group, the histological characteristics of the normal skeletal muscle, the blank group of the model group and the rats of the medicated diet group had different degrees of myocyte atrophy or degeneration, and the extent of the atrophy of the myoblasts in the medicated diet group was less than that of the blank group of the model. The results showed that the activity of GSH-Px and SOD in the serum of the model blank group was significantly lower than that in the blank group (P0.05). Compared with the blank group of the model, the activity of GSH-Px and SOD in the serum of the medicated diet group was significantly increased (P0.05). The content of MDA was significantly lower (P0.05). Compared with the blank group, the activity of GSH-Px and SOD in the skeletal muscle of the model blank group decreased significantly (P0.05). Compared with the blank group of the model, the activity of GSH-Px in the skeletal muscle of the medicated diet group was higher than that of the blank group (P0.05). The content of MDA and the activity of Caspase-3 and Caspase-8 decreased significantly (P0.05). Compared with the blank group, the expression of Caspase-3 and Caspase-8 mRNA in the skeletal muscle of the model group was significantly lower than that of the blank group (P0.05). Conclusion:1. The medicated diet plus acupoint application can improve the physical appearance, state and skeletal muscle tissue morphology of the old rats, and can slow down the occurrence and development of the muscle attenuation syndrome. The medicated diet plus acupoint application can improve the activity of GSH-Px and SOD in the serum and skeletal muscle of the aging rats, and decrease the content of MDA, so as to improve the antioxidant capacity of the skeletal muscle, reduce the oxidative damage of the skeletal muscle and delay the aging of the skeletal muscle. It is possible to reduce the activity of Caspase-3 and Caspase-8 in the skeletal muscle cells of aging rats, and to reduce the expression of Caspase-3 mRNA and Caspase-8 mRN A, so as to reduce the cell apoptosis and delay the aging of the skeletal muscle.
【学位授予单位】：辽宁中医药大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R248
本文编号：2445141
[Abstract]:Objective: To study the changes of skeletal muscle morphology in aging rats by using D-galactose (D-gal)-induced aging rats to study the aging model rats based on the "spleen main muscle and limbs" theory. The relationship between the progressive decline of skeletal muscle mass and the cell apoptosis and the anti-oxidation ability was studied by the effect of the anti-oxidation ability and the expression of caspase-8. The invention provides a theoretical basis for delaying the occurrence of the muscle attenuation syndrome (Sarcoenia), and finally provides effective nursing measures for the clinical intervention Sarcoenia, so as to improve the daily activity capability of the old people, self-adjustment, reduce the probability of falling and bedridden, improve the quality of life of the old people, And the economic burden of the health care cost and the family and the society is reduced. Materials and Methods:36 healthy male Sprague-DaWley (SD) rats were randomly divided into blank group, model blank group and medicated diet therapy group. The daily abdominal subcutaneous injection of the model blank group and the medicated diet group was 7.5% D-gal 125 mg/ kg. The daily abdomen of the blank group was injected with 0.9% normal saline of 125 mg/ kg and the model was established for 6 weeks. After 6 weeks, the medicated diet was administered to the medicated diet (10 ml 路 kg-1 路 d-1) and the acupoint application for intervention. The blank group and the model blank group were given equal dose of normal saline and blank for intervention, and the intervention lasted for 4 weeks. After the last administration for 24 hours, the abdominal cavity was anesthetized, the blood was taken from the abdominal aorta, and the rectus femoris was quickly separated. The changes of the morphological structure of the straight muscle cells of the aging rats were observed by HE staining, and the expression of cysteinyl aspa rate specific c protein-3 (Caspase-3) and Caspase-8 mRNA was detected by PCR, and the glutathione peroxidase (GSH-Px) in the serum was detected by ELISA. The activity of superoxide dismutase (SOD) and the content of MDA and the activity of GSH-Px, SOD, Cas ase-8 and the content of MDA were studied. The data was analyzed by SPSS17.0 statistical analysis software and Microsoft Office Excel 2007. The mean standard deviation (Mean-S) of each group of data indicated that the difference between the groups was defined as P0.05 with a single-factor analysis of variance. Results:1. In the model blank group and the medicated diet group, the aging characteristics of the rats in the model blank group and the medicated diet group were reduced; the weight of the three groups of rats was increased, and the rats in the blank group of the model group were slightly lower than that of the blank group, but no significant difference was found between the groups (P0.05). Under the light microscope, in the blank group, the histological characteristics of the normal skeletal muscle, the blank group of the model group and the rats of the medicated diet group had different degrees of myocyte atrophy or degeneration, and the extent of the atrophy of the myoblasts in the medicated diet group was less than that of the blank group of the model. The results showed that the activity of GSH-Px and SOD in the serum of the model blank group was significantly lower than that in the blank group (P0.05). Compared with the blank group of the model, the activity of GSH-Px and SOD in the serum of the medicated diet group was significantly increased (P0.05). The content of MDA was significantly lower (P0.05). Compared with the blank group, the activity of GSH-Px and SOD in the skeletal muscle of the model blank group decreased significantly (P0.05). Compared with the blank group of the model, the activity of GSH-Px in the skeletal muscle of the medicated diet group was higher than that of the blank group (P0.05). The content of MDA and the activity of Caspase-3 and Caspase-8 decreased significantly (P0.05). Compared with the blank group, the expression of Caspase-3 and Caspase-8 mRNA in the skeletal muscle of the model group was significantly lower than that of the blank group (P0.05). Conclusion:1. The medicated diet plus acupoint application can improve the physical appearance, state and skeletal muscle tissue morphology of the old rats, and can slow down the occurrence and development of the muscle attenuation syndrome. The medicated diet plus acupoint application can improve the activity of GSH-Px and SOD in the serum and skeletal muscle of the aging rats, and decrease the content of MDA, so as to improve the antioxidant capacity of the skeletal muscle, reduce the oxidative damage of the skeletal muscle and delay the aging of the skeletal muscle. It is possible to reduce the activity of Caspase-3 and Caspase-8 in the skeletal muscle cells of aging rats, and to reduce the expression of Caspase-3 mRNA and Caspase-8 mRN A, so as to reduce the cell apoptosis and delay the aging of the skeletal muscle.
【学位授予单位】：辽宁中医药大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R248
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