Exploration of gastrocnemius injury and its mechanism in rats with syndrome of spleen deficiency

FAN Fangxin; ZHU Haoru; QIAN Meng; SHI Tuo; ZHU Sitai; XIE Ming

doi:10.3969/j.issn.1006-2157.2023.01.013

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Exploration of gastrocnemius injury and its mechanism in rats with syndrome of spleen deficiency

Experimental Studies | 更新时间：2023-02-03

- Exploration of gastrocnemius injury and its mechanism in rats with syndrome of spleen deficiency
- Journal of Beijing University of Traditional Chinese Medicine Vol. 46, Issue 1, Pages: 77-86(2023)
- 作者机构：
  
  北京中医药大学中医学院　北京　100029
- 作者简介：
  
  Prof. XIE Ming, Ph. D. , Doctoral Supervisor. School of Traditional Chinese Medicine, Beijing University of Chinese Medicine, No.11, Beisanhuan Donglu Road, Chaoyang District, Beijing, 100029. E-mail: xieming603@263.net
- 基金信息：
  
  National Natural Science Foundation of China(81173193)
- DOI：10.3969/j.issn.1006-2157.2023.01.013
  CLC： R285.5
- Received：28 February 2022，
  
  Published Online：02 November 2022，
  
  Published：30 January 2023
- 稿件说明：
移动端阅览
FAN Fangxin, ZHU Haoru, QIAN Meng, et al. Exploration of gastrocnemius injury and its mechanism in rats with syndrome of spleen deficiency[J]. Journal of beijing university of traditional chinese medicine, 2023, 46(1): 77-86.
DOI：

FAN Fangxin, ZHU Haoru, QIAN Meng, et al. Exploration of gastrocnemius injury and its mechanism in rats with syndrome of spleen deficiency[J]. Journal of beijing university of traditional chinese medicine, 2023, 46(1): 77-86. DOI： 10.3969/j.issn.1006-2157.2023.01.013.

摘要

目的

观察脾虚证模型大鼠腓肠肌损伤情况并对其机制进行研究。

方法

将雄性Wistar大鼠(SPF/VAF级)按体质量随机分为对照组和模型组，模型组采用饮食失节联合游泳劳倦法复制脾虚证模型，对照组予以正常饲养。定期观察并记录2组大鼠的外观表征、体质量、饮食量和粪便含水率。实验第2、3、4周末，测定各组大鼠的游泳力竭时间、抓握力、尿D－木糖排泄率、脾脏指数、胸腺指数及腓肠肌湿质量，HE染色观察腓肠肌病理结构，酶联免疫吸附测定法检测血清中胃动素、胃泌素、白细胞介素－1β(IL-1β)、白细胞介素－6(IL-6)、肿瘤坏死因子-α(TNF-α)和腓肠肌组织中柠檬酸合酶、Ca

2＋

－ATP酶、IL-1β、IL-6、TNF-α含量。实验第4周末，用免疫组织化学法检测腓肠肌IL-1β、IL-6、TNF-α阳性表达水平，免疫荧光法检测腓肠肌切割型天冬氨酸蛋白水解酶3(cleaved-caspase 3)阳性细胞。

结果

与对照组比较，第2～4周模型组大鼠体质量、饮食量、游泳力竭时间、抓握力、腓肠肌湿质量和尿D－木糖排泄率下降(

0.05，

0.01)，粪便含水率增加(

0.01)；第3～4周模型组大鼠血清胃泌素下降(

0.05，

0.01)；第2周模型组大鼠腓肠肌湿质量/体质量和血清IL-6增加(

0.05，

0.01)；第4周模型组大鼠腓肠肌湿质量/体质量、脾脏指数、胸腺指数、腓肠肌柠檬酸合酶和Ca

2＋

－ATP酶水平、血清胃泌素、IL-1β和TNF-α含量下降(

0.01)，腓肠肌cleaved-caspase 3、IL-1β、IL-6、TNF-α含量增加(

0.01)。

结论

饮食失节联合游泳劳倦法复制的脾虚证模型大鼠具有中医脾虚证的外观表征和实验室相关指标的变化特点，腓肠肌出现肌力减退和肌萎缩，并随造模时间延长而加重，伴有全身免疫功能减退与血清、腓肠肌炎性因子含量增加，提示脾虚证骨骼肌受损涉及线粒体损伤和细胞凋亡，并与炎性机制有关。

Abstract

Objective

To investigate skeletal muscle injury and its mechanism in a rat model of spleen deficiency syndrome.

Methods

Male Wistar rats(SPF/VAF) were randomly divided into control group and model group according to body weight. The "improper diet combined with exhaustive swimming" method was used to duplicate the rat model with spleen deficiency syndrome. The control group was fed normally. We regularly recorded the appearance

body weight

dietary intake

and excrement water content of the two groups of rats. At the end of Week 2

and 4

the swimming exhaustion time

grip strength

urine D-xylose excretion rate

spleen index

thymus index

and gastrocnemius wet weight were measured

and the gastrocnemius was subjected to pathological examination by HE staining. Besides

at the end of Week 2

and 4

we used enzyme-linked immunoassay to determine the levels of motilin

gastrin

interleukin-1β (IL-1β)

interleukin-6 (IL-6)

and tumor necrosis factor-α (TNF-α) in serum

and the levels of citrate synthase

-ATPase

IL-1β

IL-6

and TNF-α in the gastrocnemius. At the end of Week 4

immunohistochemistry method was used to measure the positive expression of IL-1β

IL-6

and TNF-α

and immunofluorescence was used to detect the cells that were positive for cysteine aspartate proteolytic enzyme 3 (cleaved-caspase 3) in the gastrocnemius.

Results

Compared to the control group

the body weight

dietary intake

swimming exhaustion time

grip strength

gastrocnemius wet weight

and urine D-xylose excretion rate in the model group were significantly decreased from Week 2 to 4 (

0.05

0.01)

while excrement water content increased (

0.01). Serum gastrin in the model group was also significantly decreased at Week 3 and 4 (

0.05

0.01). Besides

the gastrocnemius wet weight to body weight ratio and serum IL-6 of the rats in the model group were increased at Week 2 (

0.05

0.01). At the end of Week 4

the gastrocnemius wet weight to body weight ratio

spleen index

thymus index

citrate synthase and Ca

-ATPase in the gastrocnemius

the levels of gastrin

IL-1β

and TNF-α in the serum of the model group decreased significantly (

0.01); and the levels of cleaved-caspase 3

IL-1β

IL-6

and TNF-α in the gastrocnemius increased significantly (

0.01).

Conclusion

The spleen deficiency syndrome rat model induced by "improper diet combined with exhaustive swimming" method in accordance with the appearance of traditional Chinese medicine syndrome of spleen deficiency and related laboratory indexes. The muscle strength and atrophy of the gastrocnemius was decreased with the modeling time. The systemic immune function of the model rats decreased

while the protein expression of systemic and gastrocnemius inflammatory factors increased. We presume that the skeletal muscle damage involves mitochondrial injury and apoptosis in the spleen deficiency syndrome rat model but is related to inflammatory mechanism.

关键词

Keywords

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