Serum creatine kinase activity and its changes after a muscle biopsy

Summary. The significance of the absolute elevations of serum creatine kinase (CK) levels after intense exercise and injuries was studied by measuring CK activities from seven healthy active males during a 2-week period, with a muscle biopsy taken between the first and second week. Most of the subjects (three lifters and two runners) carried on their normal exercise activities, while two lifters stopped training during the 2 weeks. The weight of the biopsy, number of fibres, percentage of fibre types, and cross-sectional areas of the muscle fibres were measured. The CK levels of the nonactive subjects and runners remained consistently low during the control week, whereas those of the lifters were usually 500% greater than those of the other two groups, and fluctuated with the intensity of their workouts. A muscle biopsy, having a mean weight of 71.3 mg and containing 1800 fibres, increased the CK values by approximately 100 units litre^-1 (U1^-1) in most of the subjects. One runner injured his right hamstring muscles 2 days prior to the biopsy, and his CK values rose from 50 to 4400 U I^-1. The increases in CK after the biopsy were not related to fibre type, activity, weight of the biopsy, or number or size of fibres removed. These results indicate that:

• 1 CK values are consistently lower in normal subjects and runners than in lifters.
• 2 Weight training results in chronic elevations of CK.
• 3 Compared to a muscle biopsy, muscular injury dramatically increases CK levels.
• 4 Elevation of serum CK is observed as early as 1 h after an intense weight-lifting session.
• 5 The elevation of serum CK by 100 U 1^-1 is associated with damage to approximately 2000 fibres.

     

Isoforms of creatine kinase: MM in the study of skeletal muscle damage

Isoforms of creatine kinase (CK) MM have been analysed in plasma from normal subjects and patients with muscular dystrophy using isoelectric focusing techniques. Most plasma samples analysed contained three isoforms of CK-MM of isoelectric points 7.26 (MMI), 6.85 (MMII) and 6.45 (MMIII) although in some plasma samples two additional isoforms of isoelectric points 7.12 and 6.65 were seen. Patients with muscular dystrophy were found to have a generally higher proportion of CK-MMI in their plasma than normal subjects and this was relatively unaffected by large variations in the total creatine kinase activity. By comparison eccentric exercise in normal subjects was found to result in a large increase in total plasma CK activity which then declined to normal over a period of approximately 6 days. CK-MMI was found to increase initially followed by CK-MMII and CK-MMIII. Analysis of the isoforms in biopsy samples of human muscle revealed the presence of two of the bands found in plasma (CK-MMI and MMII) and a third muscle specific isoform, while incubation of muscle homogenates in plasma induced the formation of CK-MMIII and the two isoforms of pI 7.12 and 6.65. It is concluded that analyses of CK-MM isoforms in human plasma can provide useful information on the extent and relative time course following an episode of muscle damage but that in patients with muscular dystrophy the large variations in plasma CK activity are not reflected in the proportion of CK found in each isoform.(ABSTRACT TRUNCATED AT 250 WORDS)     

重组人肌酸激酶MM同工酶的酶动力学研究

目的:对纯化后重组肌酸激酶MM同工酶(CK-MM)的米氏常数(Km)进行研究,为其作为参考品或校准品应用于临床奠定基础。方法:将重组酶置于自制类人血清基质中,按IFCC参考方法配制试剂,采用HITACHI全自动生化分析仪测定重组酶与天然酶[底物分别是磷酸肌酸(PCr)和腺苷二磷酸(ADP)]的Km常数。结果:重组酶KmPCr值高于天然酶1.4倍,Km ADP重组酶与天然酶近似。结论:重组CK-MM的酶动力学常数基本符合参考方法的测定要求,初步具备作为基因工程人源酶校准品的基本特点。     

重组人肌酸激酶MM同工酶在不同保存基质中的稳定性和互通性观察

目的观察重组人肌酸激酶MM同工酶(CK-MM)在不同基质中的稳定性和互通性,筛选出适合的基质制备校准品用于CK测定的标准化。方法将重组酶分别置于自制类人血清基质、抗冻基质、仿有证参考物配方基质及人血清基质中,用常规测定方法观察重组酶的稳定性;分别采用参考方法、常规方法(罗氏试剂和中生试剂)测定不同基质中的重组酶活性和不同浓度的人血浆总CK活性,观察重组酶在不同方法间的互通性。结果重组酶在自制类人血清基质和仿有证参考物配方基质中-20℃冻存可稳定25d和23d,在抗冻基质中至少可稳定240d。自制类人血清基质和仿有证参考物配方基质中重组酶在参考方法与常规方法间有互通性,人血清基质和抗冻基质中的低值重组酶样品在两方法间具有互通性。结论自制基质和有证参考物配方基质中的重组酶具有较好的稳定性和互通性,初步具备了校准品的基本特点。本研究为进一步研制CK测定的参考品,包括校准品和定值控制品提供了思路。     

Development and performance of an enzyme immunoassay to detect creatine kinase isoenzyme MB activity using anti-mitochondrial creatine kinase monoclonal antibodies

Abstract

Objective: The MB fraction of creatine kinase (CK-MB) has long been used as a cardiac marker. It is known that the CK-MB immunoinhibition method lacks selectivity and accuracy, because the appearance of macro CK type 2, corresponding to mitochondrial creatine kinase (MtCK) in some patient serum may render CK-MB activity measured by conventional method abnormally high. Thus, to improve the specificity and accuracy of the CK-MB assay, we developed two types of monoclonal anti-MtCK antibodies against sarcomeric MtCK and ubiquitous MtCK, and present herein the performance of a new method using these antibodies. Material and methods: The performance of our test for detecting CK-MB activity was compared with other methods, and the range of CK-MB activities in normal human serum was investigated. Results: The two types of monoclonal antibodies developed by us were isoenzyme-specific to sMtCK or uMtCK. The correlation coefficients of our method and conventional method to electrophoresis were 0.973 and 0.873, respectively. The mean CK-MB activity in normal human serum by our method and the conventional method was 2.4 and 11.7 U/L, respectively. Thus, our data indicated that about 80% of CK-MB activity, determined using the conventional method, seems to correspond to the MtCK activity. Conclusion: Our method is novel in offering higher accuracy of measuring true CK-MB contents in human serum as compared to the conventional method. The possibility of accurately estimating CK-MB activity by our method which can inhibit MtCKs in healthy person and patient serum is likely to bring a break-through in clinical diagnostics.     

碱性成纤维细胞生长因子对培养的骨骼肌细胞生长作用的研究

目的观察碱性成纤维细胞生长因子(bFGF)对体外培养的新生Wistar大鼠骨骼肌细胞存活、分化及促增殖分裂影响。方法应用无血清培养技术,采用四唑盐(MTT)微量比色法,乳酸脱氢酶(LDH)活性、肌酸磷酸激酶(CK)活性测定及蛋白定量检测。结果bFGF干预组MTT微量比色的ΔOD值、LDH活性、CK活性及蛋白质合成均高于对照组。结论外源性bFGF明显促进细胞的存活、生长及增殖分化。     

运动干预对胰岛素抵抗大鼠骨骼肌蛋白激酶B活性和蛋白与基因表达的影响

目的：探讨一次性游泳运动对胰岛素抵抗大鼠骨骼肌蛋白激酶B（protein kinaes, PKB）mRMA表达、蛋白总量（t- PKB）及磷酸化PKB（p-PKB）的影响。方法：SD大鼠随机分为正常饮食组、高脂饮食组、高脂饮食+运动组。分别对高脂饮食组与高脂饮食+运动组饲以高脂饲料,4周后对高脂饮食+运动组大鼠运动干预2h。测血液葡萄糖和胰岛素浓度;Western blot法检测骨骼肌t-PKB蛋白表达、p-PKB磷酸化水平;RT-PCR法分析PKB mRNA表达。结果：高脂饮食组血糖和胰岛素浓度明显高于对照组;高脂饮食+运动组血糖和胰岛素浓度低于高脂饮食组;高脂饮食组PKB mRNA表达下降;高脂饮食+运动组 PKB磷酸化水平和蛋白总量高于高脂饮食组。结论：运动干预改善高脂饮食诱发胰岛素抵抗大鼠对胰岛素的敏感性,增加PKB磷酸化水平和蛋白与基因的表达。     

电针疗法对运动后肌肉损伤相关因子活性的影响

目的:通过观察电针疗法对单纯性中度抗阻训练后肌肉损伤相关因子活性的影响,探究其对运动肌肉损伤可能的保护作用。方法:30例男性志愿者(年龄20—25周岁)按随机数表法分为治疗组15例和对照组15例。两组受试者接受为期5d,每天3组,每组20次的腿举训练。训练前接受评估,使腿举重量为受试者最大腿举重量的70%。每次训练结束后治疗组给予30min的电针治疗,选择足三里-条口、承山-委中两组对穴;对照组不做特殊处理,嘱充分休息。训练前及每次治疗结束30min后抽取受试者血液样本以检测肌酸激酶(creatine kinase,CK)和乳酸脱氢酶(lactic dehydrogenase,LDH)活性。训练前后采用等速肌力测试系统检测膝关节屈伸肌峰力矩值(peak torque,PT)和平均功率(average power,AP)。结果:首次训练后两组受试者血液样本中CK和LDH活性均显著提升(P0.01),5d内对照组两项指标均高于治疗组(P0.01),治疗组CK在第3天后开始下降;5d训练治疗后治疗组膝关节屈伸肌峰力矩值和平均功率高于对照组,较训练前也有所提高(P0.05),对照组训练前后无明显改变(P0.05)。结论:中度的抗阻训练可以对肌肉组织产生一定的损伤,电针治疗能够降低相关因子的活性,促进损伤肌肉和运动能力的恢复,能够有效地运用到运动过后肌肉损伤的康复治疗中。     

Possible involvement of dopamine D-1 and D-2 receptors in diazepam-induced hyperphagia in rats

Possible involvement of dopamine receptors in diazepam-induced (1 mg/kg, subcutaneous (sc] hyperphagia was studied in nondeprived rats. Pretreatment with the selective D-1 antagonist, SCH23390 (0.03 mg/kg, sc) inhibited diazepam-induced hyperphagia. In addition, pretreatment with the preferential D-2 antagonists, haloperidol (0.1 to 0.3 mg/kg, sc) and clebopride (0.1 to 0.3 mg/kg, sc) inhibited diazepam-induced hyperphagia in a dose-dependent manner. Pretreatment with co-administration of SCH23390 (0.1 mg/kg, sc) and clebopride (0.03 mg/kg, sc) completely inhibited this hyperphagia. The selective D-2 antagonist, sulpiride (40 mg/kg, sc) and the peripheral D-2 antagonist, domperidone (10 mg/kg, sc) did not affect diazepam-induced hyperphagia. However, sulpiride (10 micrograms, icv) or domperidone (2 micrograms, icv) administered centrally inhibited this hyperphagia. The highest dose of haloperidol (0.3 mg/kg, sc) or clebopride (0.3 mg/kg, sc) and higher doses of SCH23390 (0.01 and 0.03 mg/kg, sc) or SCH23390/clebopride (0.01/0.03 and 0.01/0.1 mg/kg, sc) tended to decrease spontaneous feeding in non-deprived rats. In addition, the highest dose of haloperidol, clebopride or SCH23390/clebopride inhibited spontaneous feeding in deprived rats. Interestingly, diazepam-induced hyperphagia was inhibited significantly by doses of haloperidol (0.1 mg/kg, sc), clebopride (0.1 mg/kg, sc) and SCH23390/clebopride (0.003/0.03 and 0.003/0.1 mg/kg, sc) which did not affect spontaneous feeding in non-deprived or deprived rats. Pretreatment with alpha-methyl-p-tyrosine (40 mg/kg, IP x 2, 6 and 2 h prior to diazepam administration) failed to inhibit this hyperphagia. Furthermore, pretreatment with a large dose of haloperidol (5 mg/kg, sc, 4 days before diazepam administration) augmented the sub-hyperphagic effect to diazepam (0.5 mg/kg, sc). Thus, these findings suggest that hyperphagia to diazepam is mediated in part by both dopamine D-1 and D-2 receptors in non-deprived rats.     

毒鼠强对鼠心肌、骨骼肌的作用及与血清酶升高的关系

目的 研究毒鼠强对鼠心肌、骨骼肌的影响及与血清酶升高的关系。方法 用同种大鼠30只,将其分成3组：对照组、半致死量组（0．02mg／kg）及致死量组（0．04mg／kg）;每组雌雄各半。按大鼠的体重将毒鼠强粉溶于生理盐水后灌入胃内,对照组则灌入等量的生理盐水。于死亡立即或1h后取血清测心肌肌钙蛋白TnⅠ及肌酸激酶（CK）、天门冬酸氨基转移酶（AST）、乳酸脱氢酶（LDH）、α-羟丁酸脱氢酶（α-HBDH）,并取死亡或处死大鼠的心肌和大腿骨骼肌做病理检查。结果大鼠中毒后10～60min出现四肢或全身抽搐;致死量组6只大鼠于中毒后20-60分钟死亡。血清心肌肌钙蛋白TnⅠ中毒组较对照组增高（P〈0．01）,致死量组较半致死量组增高（P〈0．01）;CK、AST、LDH、α-HBDH中毒组高于对照组（P〈0．01）,致死量组高于半致死量组（P〈0．01）;中毒大鼠心肌、骨骼肌组织均有变性、坏死;中毒剂量越大,损伤越明显;骨骼肌的损害比心肌损害更严重的。结论毒鼠强急性中毒后,血清酶学异常升高;其心肌和骨骼肌组织均有显著损伤。异常升高的血清酶是毒鼠强对心肌和骨骼肌双重损伤的结果。     

营养与运动负荷

本文针对运动负荷引起的肌肉适应现象,着力介绍了适应过程中肌肉的相关营养生化改变以及不同类型和强度的运动对机体能量代谢的影响。     

不同运动方式对自发性高血压大鼠骨骼肌纤维类型与代谢的影响

目的:探讨不同运动方式对自发性高血压大鼠(spontaneously hypertensive rats,SHR)骨骼肌纤维类型及代谢的影响,为高血压患者最佳运动处方制定提供科学依据和有效方法。方法:30只3月龄雄性SHR随机分为安静组(SHR sedentary,SHR-S)、中等强度持续有氧运动组(SHR moderate continuous aerobic training,SHR-MCAT)和高强度间歇运动组(SHR high-intensity interval training,SHR-HIIT),同时以10只雄性Wistar-Kyoto大鼠作为正常血压对照组(WKY)。WKY和SHR-S组安静饲养,SHR-MCT、SHR-HIT组分别进行8周持续和间歇跑台运动。实验后利用递增负荷跑台实验测定力竭时间(exhaust time,ET),分离胫骨前肌(快肌,Ⅱ型肌纤维为主)和比目鱼肌(慢肌,Ⅰ型肌纤维为主),利用比色法测定柠檬酸合酶(citrate synthase,CS)(有氧代谢标志物)和乳酸脱氢酶(lactate dehydrogenase,LDH)(无氧代谢标志物)...     

α-硫辛酸抑制慢性低氧大鼠骨骼肌线粒体的氧化应激**

背景：α-硫辛酸被誉为“全能抗氧化剂”和“线粒体营养剂”,但其是否可用于慢性低氧骨骼肌的防护,及其相关机制尚不清楚。目的：观察α-硫辛酸对慢性低氧大鼠骨骼肌线粒体活性氧生成及抗氧化酶系的作用,并探讨α-硫辛酸作用的相关信号通路。方法：将36只 SD 大鼠随机分为常氧对照组、单纯低氧组和单纯低氧+α-硫辛酸组。低氧干预为常压低氧帐篷,氧体积分数设定为11.3%;α-硫辛酸干预为标准饲料中添加α-硫辛酸(0.25%)。各种干预均持续4周。结果与结论：α-硫辛酸显著上调Sirtuin-3表达,提高线粒体ATP合成活力和膜电位,上调线粒体态3呼吸速率、呼吸控制比和磷氧比,下调态4呼吸速率,促进并上调锰超氧化物歧化酶、谷胱甘肽过氧化物酶、过氧化氢酶等线粒体抗氧化酶系活性,从而抑制线粒体H2O2产生速率,降低线粒体丙二醛含量。表明α-硫辛酸可提高慢性低氧骨骼肌线粒体能量代谢效率,抑制活性氧生成。并通过提高线粒体抗氧化酶系活性,抑制低氧诱导的氧化应激。α-硫辛酸对低氧骨骼肌线粒体的保护效应可能与其上调态3呼吸速率有关。     

白细胞介素6对2型糖尿病大鼠骨骼肌糖代谢的影响

目的：通过研究白细胞介素6（IL－6）对2型糖尿病（T2DM）大鼠骨骼肌葡萄糖转运体4（GLUT4）和糖原含量的影响,探讨其影响T2DM糖代谢的机制。方法：健康雄性SD大鼠随机分为4组：正常对照组（N组）10只,糖尿病IL-6抗原干预组（A组）10只,糖尿病IL－6抗体干预组（B组）8只,糖尿病对照组（C组）8只。测定血糖、血清胰岛素、血脂等指标;取股四头肌比色法测定肌糖原含量,免疫组化法测定骨骼肌GLUT4蛋白表达。结果：T2DM大鼠骨骼肌糖原含量和GLUT4蛋白表达较正常大鼠明显降低,IL-6抗体干预可以升高T2DM大鼠骨骼肌糖原含量和GLUT4蛋白表达。结论：T2DM大鼠骨骼肌GLUT4蛋白表达减少;IL－6抗体阻滞通过增加GLUT4蛋白表达促进骨骼肌对糖的利用。     

83例窒息新生儿血清肌钙蛋白Ⅰ及肌酸激酶同工酶检测分析

目的 探讨窒息新生儿血清肌钙蛋白Ⅰ(cTnⅠ)、肌酸激酶同工酶(CK-MB)的变化及临床意义.方法 对83例窒息新生儿(其中轻度窒息48例、重度窒息35例)与30例健康新生儿(对照组),采用微粒子化学发光法和干化学分析法,进行血清cTnⅠ和CK-MB水平检测.结果 对照组cTnⅠ和CK-MB平均水平分别为(0.03±0.02)ng/mL、(37.0±12.3)U/L;轻度窒息组cTnⅠ和CK-MB平均水平分别为(0.12±0.05)ng/mL、(71.0±19.3)U/L;重度窒息组cTnⅠ和CK-MB平均水平分别为(0.19±0.14)ng/mL、(92.0±25.4)U/L.结论 新生儿窒息后血清cTnⅠ和CK-MB水平可明显升高,通过检测血清cTnⅠ和CK-MB水平不仅可用来判断窒息对患儿心肌损害的程度,亦可间接反映患儿窒息程度,有助于早期诊断及治疗.     

耐力运动对大鼠骨骼肌PKB磷酸化与表达的影响*

目的：探讨耐力运动对大鼠骨骼肌蛋白激酶B（PKB）mRNA表达、蛋白总量（t-PKB）及磷酸化PKB（p-PKB)的影响。方法：SD大鼠随机分为对照组和运动组。运动组分为1 h/d、1.5 h/d组,运动7周,最后一次运动后24h和48h取材,分为1 h/d运动24h取材组、1 h/d运动48h取材组、1.5 h/d运动24h取材组、1.5 h/d运动48 h取材组。测定血清葡萄糖和胰岛素浓度;采用Western blot法检测t-PKB 和p-PKB水平;用RT-PCR法分析PKB mRNA的表达。结果     

运动和胰岛素对高血糖大鼠骨骼肌JNK和p38信号传导系统的调节作用

目的：观察运动和胰岛素对高血糖大鼠骨骼肌细胞应激激活蛋白激酶-JNK和p38信号通道活性的作用，为糖尿病的运动疗法提供分子生物学依据。方法：12周龄、体重200-230g的SD大鼠18只，腹腔内注射链脲佐菌素，建立持续高血糖模型。然后随机分为运动组、胰岛素组和对照组。运动组采用活动平板，坡度10％，速度20m／min，运动20min后取标本；胰岛素组给予皮下注射胰岛素，30min后取标本；对照组不参与运动也不给予胰岛素注射。运用化学发光法测定骨骼肌JNK和p38活性。结果：骨骼肌JNK活性在运动组分别高于对照组和胰岛素组，胰岛素组JNK活性与对照组无显著差异。骨骼肌p38活性在运动组和胰岛素组较对照组显著增高。结论：在高血糖状态下，运动使得骨骼肌JNK和p38激酶的活性增高，提示通过对JNK和p38信号转导通道的调节作用，参与了骨骼肌结构和代谢功能的适应性改变；胰岛素可以激活骨骼肌p38通道但对JNK的活性无影响，提示通过对p38的激活，改善了葡萄糖转载体的内在活性。     

Comparison of serum cardiac specific troponin-I with creatine kinase,creatine kinase-MB isoenzyme,tropomyosin, myoglobin and C-reactive protein release in marathon runners: cardiac or skeletal muscle trauma?

Problems arise in distinguishing skeletal from cardiac muscle trauma on the basis of serum enzyme tests following severe muscle exercise. The contributions of cardiac and skeletal sources have been assessed in eleven marathon runners by measuring pre- and post-race serum levels of cardiac-specific myofibrillar troponin-I together with total creatine kinase, creatine kinase-MB isoenzyme, myoglobin, myofibrillar tropomyosin and C-reactive protein. Total creatine kinase, creatine kinase-MB isoenzyme, tropomyosin and myoglobin were significantly elevated above pre-race levels in all runners between 1 h and 128 h post-race. Neither mean cardiac troponin-I nor C-reactive protein was elevated post-race. Nine out of sixty-three samples fulfilled conventional positive criteria for cardiac muscle damage on the basis of combined creatine kinase and creatine kinase-MB isoenzyme levels. Six runners had one or more positive samples. No samples had levels above twice the upper normal limit for either cardiac troponin-I or C-reactive protein. Correlation analysis of levels in each sample indicated skeletal and not cardiac muscle as the source of raised serum protein.     

运动对2型糖尿病大鼠骨骼肌细胞腺苷酸活化蛋白激酶表达和活性的影响

目的观察运动对正常大鼠和2型糖尿病大鼠骨骼肌中腺苷酸活化蛋白激酶（AMPK）的蛋白表达和活性的影响。方法采用OLETF大鼠作为2型糖尿病组，LETO大鼠作为正常对照组，并分别随机分为2组，即运动组和对照组，每组5只。运动组进行急性游泳运动3h后，采用Western blot法对大鼠比目鱼肌和趾长伸肌中AMPK蛋白进行检测。结果OLETF大鼠和LETO大鼠的比目鱼肌和趾长伸肌中AMPKα蛋白表达和磷酸化均无明显差异（P〉0．05）；OLETF大鼠和LETO大鼠在急性运动3h后，趾长伸肌中AMPKαThr^172。磷酸化明显高于对照组（P〈0．05），比目鱼肌中AMPKα蛋白表达和Thr^172。磷酸化则无明显变化（P〉0．05）。结论AMPKα的表达和激活在2型糖尿病大鼠比目鱼肌和趾长伸肌中无缺陷，AMPK对运动的反应在不同肌纤维类型中存在差异，AMPK可能参与了快肌（趾长伸肌）中运动所引发的信号传导机制。