牛磺酸对骨骼肌缺血/再灌注损伤大鼠肌浆网功能的影响
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摘要
目的探讨牛磺酸对骨骼肌缺血/再灌注(I/R)损伤的防治作用及其机制。方法选择Wistar大鼠18只,随机分为对照组、模型组、牛磺酸组,每组6只。模型组、牛磺酸组制备骨骼肌I/R损伤模型。牛磺酸组制模前30 min经左颈外静脉注射0. 6 mol/L的牛磺酸1. 5 m L/kg,模型组经左颈外静脉注射生理盐水1. 5 m L/kg。对照组仅左经颈外静脉注射生理盐水1. 5 m L/kg,不制备骨骼肌I/R损伤模型。模型组与牛磺酸组再灌注2 h,对照组同时间,麻醉后完整分离左侧股内侧肌群。取一半称湿重,彻底干燥后称干重,计算湿重/干重;取另一半制备组织匀浆,检测脂质过氧化产物丙二醛(MDA)含量及髓过氧化物酶(MPO)活性。制备三组骨骼肌肌浆网,采用定磷法检测肌浆网Ca~(2+)-ATPase活性。采用液体闪烁计数仪检测3H-Ryandine配体-受体最大结合量(Bmax)及解离常数(Kd);采用液体闪烁计数仪检测45Ca~(2+)的放射活性,以此反映肌浆网Ca~(2+)摄入量和释放量。结果模型组与牛磺酸组骨骼肌湿重/干重及MPO活性和MDA含量均明显高于对照组,但牛磺酸组上述指标明显低于模型组(P均<0. 01)。模型组与牛磺酸组肌浆网Ca~(2+)-ATPase活性、3H-Ryandine配体-受体Bmax、Ca~(2+)摄入量均明显低于对照组,3H-Ryandine配体-受体Kd均明显高于对照组(P均<0. 01);但牛磺酸组肌浆网Ca~(2+)-ATPase活性、3HRyandine配体-受体Bmax、Ca~(2+)摄入量均明显高于模型组,3H-Ryandine配体-受体Kd明显低于模型组(P均<0. 01)。各组加入反应液1 min时Ca~(2+)释放速度比较无统计学差异(P均> 0. 05),加入反应液3、5 min时模型组与牛磺酸组Ca~(2+)释放量均明显低于对照组(P均<0. 01),但牛磺酸组Ca~(2+)释放量明显高于模型组(P <0. 01)。结论牛磺酸对骨骼肌I/R损伤具有防治作用,其机制可能与抑制氧化应激反应及调节肌浆网Ca~(2+)稳态有关。
Objective To explore the preventive and therapeutic effects of taurine on ischemia/reperfusion( I/R) injury in skeletal muscle and its mechanism. Methods Eighteen Wistar rats were selected and randomly divided into 3 groups: the control group,model group,and taurine group,with 6 rats in each group. The I/R injury model of skeletal muscle was prepared in the model group and taurine group. In the taurine group,0. 6 mol/L taurine was injected via the left external jugular vein 30 min before model making. The model of skeletal muscle I/R injury was not prepared in the control group. After the rats in the model group and taurine group were reperfused for 2 h,as well as in the control group,the rats were anesthetized and the left femoral medial muscles were separated. Half of the tissue was weighed as wet weight,after thorough drying,the dry weight was weighed,and then we calculated the wet weight/dry weight,on the other hand,half tissue was taken to prepare homogenate to measure the content of malondialdehyde( MDA) and the activity of myeloperoxidase( MPO). The skeletal muscle sarcoplasmic reticulum was prepared in the three groups. The maximum binding capacity( Bmax) of Ryandine with3 H mark and the dissociation constant( Kd) of receptor and ligand were measured by liquid scintillation counter. The radiative activity of45 Ca~(2+)was measured by liquid scintillation counter to reflect the intake and release of sarcoplasmic reticulum Ca~(2+). Results The ratios of wet/dry weight,MPO activity and MDA level of the skeletal muscles in the model group and the taurine group were significantly higher than those in the control group,but the above indicators in the taurine group were significantly lower than those in the model group( all P < 0. 01). The Ca~(2+)-ATPase activity of sarcoplasmic reticulum,3 H-Ryandine ligand-receptor Bmax and Ca~(2+)intake in the model group and the taurine group were significantly lower than those in the control group,and3 H-Ryandine ligand-receptor Kd were significantly higher than those in the control group. However,the Ca~(2+)-ATPase activity of sarcoplasmic reticulum,3 H-Ryandine ligand-receptor Bmax and Ca~(2+)intake were significantly higher in the taurine group than in the model group,and the Kd of3 HRyandine receptor and ligand were significantly lower in the model group( all P < 0. 01). There was no statistically significant difference in the release rate of Ca~(2+)at 1 min of addition of the reaction solution between groups( all P > 0. 05). At 3 and 5 min of addition of the reaction solution,the release rate of Ca~(2+)in the model group and the taurine group was significantly lower than that in the control group( all P < 0. 01),but the release rate of Ca~(2+)in the taurine group was significantly higher than that in the model group( P < 0. 01). Conclusion Taurine has a preventive effect on skeletal muscle I/R injury,and its mechanism may be related to the inhibition of oxidative stress response and regulation of the Ca~(2+)homeostasis of sarcoplasmic reticulum.
Objective To explore the preventive and therapeutic effects of taurine on ischemia/reperfusion( I/R) injury in skeletal muscle and its mechanism. Methods Eighteen Wistar rats were selected and randomly divided into 3 groups: the control group,model group,and taurine group,with 6 rats in each group. The I/R injury model of skeletal muscle was prepared in the model group and taurine group. In the taurine group,0. 6 mol/L taurine was injected via the left external jugular vein 30 min before model making. The model of skeletal muscle I/R injury was not prepared in the control group. After the rats in the model group and taurine group were reperfused for 2 h,as well as in the control group,the rats were anesthetized and the left femoral medial muscles were separated. Half of the tissue was weighed as wet weight,after thorough drying,the dry weight was weighed,and then we calculated the wet weight/dry weight,on the other hand,half tissue was taken to prepare homogenate to measure the content of malondialdehyde( MDA) and the activity of myeloperoxidase( MPO). The skeletal muscle sarcoplasmic reticulum was prepared in the three groups. The maximum binding capacity( Bmax) of Ryandine with3 H mark and the dissociation constant( Kd) of receptor and ligand were measured by liquid scintillation counter. The radiative activity of45 Ca~(2+)was measured by liquid scintillation counter to reflect the intake and release of sarcoplasmic reticulum Ca~(2+). Results The ratios of wet/dry weight,MPO activity and MDA level of the skeletal muscles in the model group and the taurine group were significantly higher than those in the control group,but the above indicators in the taurine group were significantly lower than those in the model group( all P < 0. 01). The Ca~(2+)-ATPase activity of sarcoplasmic reticulum,3 H-Ryandine ligand-receptor Bmax and Ca~(2+)intake in the model group and the taurine group were significantly lower than those in the control group,and3 H-Ryandine ligand-receptor Kd were significantly higher than those in the control group. However,the Ca~(2+)-ATPase activity of sarcoplasmic reticulum,3 H-Ryandine ligand-receptor Bmax and Ca~(2+)intake were significantly higher in the taurine group than in the model group,and the Kd of3 HRyandine receptor and ligand were significantly lower in the model group( all P < 0. 01). There was no statistically significant difference in the release rate of Ca~(2+)at 1 min of addition of the reaction solution between groups( all P > 0. 05). At 3 and 5 min of addition of the reaction solution,the release rate of Ca~(2+)in the model group and the taurine group was significantly lower than that in the control group( all P < 0. 01),but the release rate of Ca~(2+)in the taurine group was significantly higher than that in the model group( P < 0. 01). Conclusion Taurine has a preventive effect on skeletal muscle I/R injury,and its mechanism may be related to the inhibition of oxidative stress response and regulation of the Ca~(2+)homeostasis of sarcoplasmic reticulum.
引文
[1]Schaffer SW,Jong CJ,Ito T,et al. Effect of taurine onischemiareperfusion injury[J]. Amino Acids,2014,46(1):21-30.
[2]Kulthinee S,Rakmanee S,Michael Wyss J,et al. Taurine supplementation ameliorates the adverse effects of perinatal taurine depletion and high sugar intake on cardiac ischemia/reperfusion injury of adult female rats[J]. Adv Exp Med Biol,2017(975):741-755.
[3]王丽娟,徐超,赵玉梅.对大鼠局灶性脑缺血-再灌注损伤的全脑保护作用[J].脑与神经疾病杂志,2017,25(10):617-622.
[4]侯阳阳,赵利军,郑艳妮,等.后处理对大鼠肢体缺血再灌注后骨骼肌的保护作用[J].中国现代医学杂志,2014,24(3):11-15.
[5]Periasamy M,Maurya SK,Sahoo SK,et al. Role of SERCA pump in muscle thermogenesis and metabolism[J]. Compr Physiol,2017,7(3):879-890.
[6]Lamboley CR,Murphy RM,Mc Kenna MJ,et al. Sarcoplasmic reticulum Ca2+uptake and leak properties,and SERCA isoform expression,in typeⅠand typeⅡfibres of human skeletal muscle[J]. J Physiol,2014,592(6):1381-1395.
[7]Mc Grew SG,Wolleben C,Siegl P,et al. Positive cooperativity of ryanodine binding to the calcium release channel of sarcoplasmic reticulum from heart and skeletal muscle[J]. Biochemistry,1989,28(4):1686-1691.
[8]Fauconnier J,Roberge S,Saint N,et al. Type 2 ryanodine receptor:a novel therapeutic target in myocardial ischemia/reperfusion[J]. Pharmacol Ther,2013,138(3):323-332.
[9]Zhu S,Xu T,Luo Y,et al. Luteolin enhances sarcoplasmic reticulum Ca2+-ATPase activity through p38 MAPK signaling thus improving rat cardiac function after ischemia/reperfusion[J]. Cell Physiol Biochem,2017,41(3):999-1010
[10] Li C,Hu M,Wang Y,et al. Hydrogen sulfide preconditioning protects against myocardial ischemia/reperfusion injury in rats through inhibition of endo/sarcoplasmic reticulum stress[J]. Int J Clin Exp Pathol,2015,8(7):7740-7751.
[11]Sun B,Wei J,Zhong X,et al. The cardiac ryanodine receptor,but not sarcoplasmic reticulum Ca2+-ATPase,is a major determinant of Ca2+alternans in intact mouse hearts[J]. J Biol Chem,2018. Pii:jbc. RA118. 003760.
[12]Sukhotnik I,Aranovich I,Ben Shahar Y,et al. Effect of taurine on intestinal recovery following intestinal ischemia-reperfusion injury in a rat[J]. Pediatr Surg Int,2016,32(2):161-168.
[13]Yamori Y,Liu L,Ikeda K,et al. Distribution of twenty-four hour urinary taurine excretion and association with ischemic heart disease mortality in 24 populations of 16 countries:results from the WHO-CARDIAC study[J]. Hypertens Res,2001,24(4):453-457.
[14]Imura H,Caputo M,Parry A,et al. Age-dependent and hypoxiarelated differences in myocardial protection during pediatric open heart surgery[J]. Circulation,2001,103(11):1551-1556.
[15]Oriyanhan W,Yamazaki K,Miwa S,et al. Taurine prevents myocardial ischemia/reperfusion-induced oxidative stress and apoptosis in prolonged hypothermic rat heart preservation[J]. Heart Vessels,2005,20(6):278-285.
[16]Geyer N,Diszházi G,Csernoch L,et al. Bile acids activate ryanodine receptors in pancreatic acinar cells via a direct allosteric mechanism[J]. Cell Calcium,2015,58(2):160-170.
[2]Kulthinee S,Rakmanee S,Michael Wyss J,et al. Taurine supplementation ameliorates the adverse effects of perinatal taurine depletion and high sugar intake on cardiac ischemia/reperfusion injury of adult female rats[J]. Adv Exp Med Biol,2017(975):741-755.
[3]王丽娟,徐超,赵玉梅.对大鼠局灶性脑缺血-再灌注损伤的全脑保护作用[J].脑与神经疾病杂志,2017,25(10):617-622.
[4]侯阳阳,赵利军,郑艳妮,等.后处理对大鼠肢体缺血再灌注后骨骼肌的保护作用[J].中国现代医学杂志,2014,24(3):11-15.
[5]Periasamy M,Maurya SK,Sahoo SK,et al. Role of SERCA pump in muscle thermogenesis and metabolism[J]. Compr Physiol,2017,7(3):879-890.
[6]Lamboley CR,Murphy RM,Mc Kenna MJ,et al. Sarcoplasmic reticulum Ca2+uptake and leak properties,and SERCA isoform expression,in typeⅠand typeⅡfibres of human skeletal muscle[J]. J Physiol,2014,592(6):1381-1395.
[7]Mc Grew SG,Wolleben C,Siegl P,et al. Positive cooperativity of ryanodine binding to the calcium release channel of sarcoplasmic reticulum from heart and skeletal muscle[J]. Biochemistry,1989,28(4):1686-1691.
[8]Fauconnier J,Roberge S,Saint N,et al. Type 2 ryanodine receptor:a novel therapeutic target in myocardial ischemia/reperfusion[J]. Pharmacol Ther,2013,138(3):323-332.
[9]Zhu S,Xu T,Luo Y,et al. Luteolin enhances sarcoplasmic reticulum Ca2+-ATPase activity through p38 MAPK signaling thus improving rat cardiac function after ischemia/reperfusion[J]. Cell Physiol Biochem,2017,41(3):999-1010
[10] Li C,Hu M,Wang Y,et al. Hydrogen sulfide preconditioning protects against myocardial ischemia/reperfusion injury in rats through inhibition of endo/sarcoplasmic reticulum stress[J]. Int J Clin Exp Pathol,2015,8(7):7740-7751.
[11]Sun B,Wei J,Zhong X,et al. The cardiac ryanodine receptor,but not sarcoplasmic reticulum Ca2+-ATPase,is a major determinant of Ca2+alternans in intact mouse hearts[J]. J Biol Chem,2018. Pii:jbc. RA118. 003760.
[12]Sukhotnik I,Aranovich I,Ben Shahar Y,et al. Effect of taurine on intestinal recovery following intestinal ischemia-reperfusion injury in a rat[J]. Pediatr Surg Int,2016,32(2):161-168.
[13]Yamori Y,Liu L,Ikeda K,et al. Distribution of twenty-four hour urinary taurine excretion and association with ischemic heart disease mortality in 24 populations of 16 countries:results from the WHO-CARDIAC study[J]. Hypertens Res,2001,24(4):453-457.
[14]Imura H,Caputo M,Parry A,et al. Age-dependent and hypoxiarelated differences in myocardial protection during pediatric open heart surgery[J]. Circulation,2001,103(11):1551-1556.
[15]Oriyanhan W,Yamazaki K,Miwa S,et al. Taurine prevents myocardial ischemia/reperfusion-induced oxidative stress and apoptosis in prolonged hypothermic rat heart preservation[J]. Heart Vessels,2005,20(6):278-285.
[16]Geyer N,Diszházi G,Csernoch L,et al. Bile acids activate ryanodine receptors in pancreatic acinar cells via a direct allosteric mechanism[J]. Cell Calcium,2015,58(2):160-170.