Fas与Fasl在慢性阻塞性肺疾病模型大鼠骨骼肌细胞的表达
摘要
目的:研究慢性阻塞性肺疾病(chronic obstructive pulmonarydisease,COPD)模型大鼠营养不良骨骼肌细胞凋亡和Fas、Fas1的表达,探讨Fas与Fas1是否参与调节COPD模型大鼠骨骼肌萎缩。
方法:将100只健康Wistar大鼠随机分为模型组80只,对照组20只。模型组采用1-29天,31-60天熏香烟,第30天气管内注入猪胰蛋白酶(PEE)和熏香烟的方法建立COPD大鼠模型。对照组不熏香烟,在模型组气管内注入PEE的同一天采用同一术式在气管内注入等量注射用生理盐水。再饲养至第90天,以低至对照组大鼠平均体重的90%作为判断营养不良的标准。然后将实验大鼠分为:对照组、COPD体重正常组和COPD营养不良组。处死全部大鼠,进行如下测定:
1、肺组织标本采集
2、膈肌和左侧趾长伸肌标本采集
3、测量大鼠体重与趾长伸肌重量。
4、采用TUNEL技术检测膈肌、趾长伸肌细胞凋亡。
5、采用免疫组化法测定骨骼肌Fas、Fas1蛋白质表达。
结果:
1.肺组织石蜡切片经HE染色,光学显微镜下可见COPD模型组大鼠有明显的COPD的特征性病理改变。
2.COPD模型组大鼠趾长伸肌肌纤维明显萎缩。
3.COPD体重正常组体重及趾长伸肌明显低于对照组,COPD营养不良组体重及趾长伸肌明显低于对照组和COPD体重正常组。
4.COPD模型营养不良组大鼠骨骼肌凋亡率明显高于COPD体重正常组,且两组COPD模型大鼠的骨骼肌凋亡率明显高于对照组。
5.Fas与Fas1在COPD营养不良组表达最强,COPD体重正常组次之,对照组最弱。
结论:
COPD模型大鼠骨骼肌存在明显的细胞凋亡率增加,在COPD模型大鼠发生营养不良时更为明显。凋亡相关基因Fas与Fas1的表达增加在COPD模型大鼠骨骼肌细胞凋亡的发生机制起着重要的作用。
Objective: To investigate apoptosis of skeletal muscles in rats with chronic obstructive pulmonary diseases, and then to measure expression of Fas and Fasl. To research the impact of Fas、Fasl on skeletal amyotrophy with the COPD mode rats.
Method: one hundred healthy male Wistar rats were randomly divided into two groups: normal control group (n=20)and model group (n=80). The COPD rat model were established by intratracheal instillation of PEE (20u/100g.BW) and being exposed to cigarette smoke for 8 weeks, 30 minutes per day. The control group was not exposed to smoke but was in intratracheal instilled of the same amount of normal saline on the same day. Malnutrition was defined when the weight of the rats in the model group was lower than 10% of the mean body weight of the control group. All rats were re-divided into 3 groups after 90 days: the control group, the non-malnutrition group, the malnutrition group. All of rats put to death. We get diaphragmatic muscle and long extensor muscle digits from rats, and made paraffin section.
1、we sampled from pneumono-tissue
2、we sampled from diaphragmatic muscle and long extensor muscle of toes.
3、We measured the weight of rats and the weight of long extensor muscle digits.
4、The rates of apoptotic in skeletal muscle were examined by Terminal deoxy nucleotidyl Transferase-mediated dUTP nick end labeling (TUNEL) method.
5、The expression of apoptosis related genes Fas and Fasl were measured by immunohistochemical technique.
Result:
1、The paraffin section of pneumono-tissue displayed the pathological character coincided with pathological character of COPD.
2、The muscle fiber of the COPD model rat was analosis.
3、The weight of long extensor muscle digits of the malnutrition group were significantly decreased compared with that of the control group, the non- malnutrition group. The weight of long extensor muscle digits of the non-malnutrition group were significantly decreased compared with that of the control group. The weight of the malnutrition group were significantly decreased compared with that of the control group, the non- malnutrition group. The weight of the non-malnutrition group were significantly decreased compared with that of the control group.
4、Apoptotic nucleus was not observed in normal skeletal muscle. The rate of apoptosis of the malnutrition group were significantly higher than those of the control group, the non- malnutrition group. The rate of apoptosis of the non-malnutrition group were significantly higher than those of the control group.
5、The optical density of Fas of the malnutrition group were significantly higher than those of the control group, the non- malnutrition group. The optical density of Fas of the non-malnutrition group were significantly higher than those of the control group. The optical density of Fasl of the malnutrition group were significantly higher than those of the control group, the non- malnutrition group. The optical density of Fasl of the non-malnutrition group were significantly higher than those of the control group.
Conclusion: Apoptosis is closely related to muscular atrophy, The rate of apoptosis was obviously found in the COPD model rats,especially in the malnutritional rats. The increase of Fas、Fasl gene expression play important roles in apoptosis of skeletal muscle cell.
方法:将100只健康Wistar大鼠随机分为模型组80只,对照组20只。模型组采用1-29天,31-60天熏香烟,第30天气管内注入猪胰蛋白酶(PEE)和熏香烟的方法建立COPD大鼠模型。对照组不熏香烟,在模型组气管内注入PEE的同一天采用同一术式在气管内注入等量注射用生理盐水。再饲养至第90天,以低至对照组大鼠平均体重的90%作为判断营养不良的标准。然后将实验大鼠分为:对照组、COPD体重正常组和COPD营养不良组。处死全部大鼠,进行如下测定:
1、肺组织标本采集
2、膈肌和左侧趾长伸肌标本采集
3、测量大鼠体重与趾长伸肌重量。
4、采用TUNEL技术检测膈肌、趾长伸肌细胞凋亡。
5、采用免疫组化法测定骨骼肌Fas、Fas1蛋白质表达。
结果:
1.肺组织石蜡切片经HE染色,光学显微镜下可见COPD模型组大鼠有明显的COPD的特征性病理改变。
2.COPD模型组大鼠趾长伸肌肌纤维明显萎缩。
3.COPD体重正常组体重及趾长伸肌明显低于对照组,COPD营养不良组体重及趾长伸肌明显低于对照组和COPD体重正常组。
4.COPD模型营养不良组大鼠骨骼肌凋亡率明显高于COPD体重正常组,且两组COPD模型大鼠的骨骼肌凋亡率明显高于对照组。
5.Fas与Fas1在COPD营养不良组表达最强,COPD体重正常组次之,对照组最弱。
结论:
COPD模型大鼠骨骼肌存在明显的细胞凋亡率增加,在COPD模型大鼠发生营养不良时更为明显。凋亡相关基因Fas与Fas1的表达增加在COPD模型大鼠骨骼肌细胞凋亡的发生机制起着重要的作用。
Objective: To investigate apoptosis of skeletal muscles in rats with chronic obstructive pulmonary diseases, and then to measure expression of Fas and Fasl. To research the impact of Fas、Fasl on skeletal amyotrophy with the COPD mode rats.
Method: one hundred healthy male Wistar rats were randomly divided into two groups: normal control group (n=20)and model group (n=80). The COPD rat model were established by intratracheal instillation of PEE (20u/100g.BW) and being exposed to cigarette smoke for 8 weeks, 30 minutes per day. The control group was not exposed to smoke but was in intratracheal instilled of the same amount of normal saline on the same day. Malnutrition was defined when the weight of the rats in the model group was lower than 10% of the mean body weight of the control group. All rats were re-divided into 3 groups after 90 days: the control group, the non-malnutrition group, the malnutrition group. All of rats put to death. We get diaphragmatic muscle and long extensor muscle digits from rats, and made paraffin section.
1、we sampled from pneumono-tissue
2、we sampled from diaphragmatic muscle and long extensor muscle of toes.
3、We measured the weight of rats and the weight of long extensor muscle digits.
4、The rates of apoptotic in skeletal muscle were examined by Terminal deoxy nucleotidyl Transferase-mediated dUTP nick end labeling (TUNEL) method.
5、The expression of apoptosis related genes Fas and Fasl were measured by immunohistochemical technique.
Result:
1、The paraffin section of pneumono-tissue displayed the pathological character coincided with pathological character of COPD.
2、The muscle fiber of the COPD model rat was analosis.
3、The weight of long extensor muscle digits of the malnutrition group were significantly decreased compared with that of the control group, the non- malnutrition group. The weight of long extensor muscle digits of the non-malnutrition group were significantly decreased compared with that of the control group. The weight of the malnutrition group were significantly decreased compared with that of the control group, the non- malnutrition group. The weight of the non-malnutrition group were significantly decreased compared with that of the control group.
4、Apoptotic nucleus was not observed in normal skeletal muscle. The rate of apoptosis of the malnutrition group were significantly higher than those of the control group, the non- malnutrition group. The rate of apoptosis of the non-malnutrition group were significantly higher than those of the control group.
5、The optical density of Fas of the malnutrition group were significantly higher than those of the control group, the non- malnutrition group. The optical density of Fas of the non-malnutrition group were significantly higher than those of the control group. The optical density of Fasl of the malnutrition group were significantly higher than those of the control group, the non- malnutrition group. The optical density of Fasl of the non-malnutrition group were significantly higher than those of the control group.
Conclusion: Apoptosis is closely related to muscular atrophy, The rate of apoptosis was obviously found in the COPD model rats,especially in the malnutritional rats. The increase of Fas、Fasl gene expression play important roles in apoptosis of skeletal muscle cell.
引文
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