尾部悬吊对达乌尔黄鼠比目鱼肌形态及 mATP 酶活性的影响

采用尾部悬吊法建立后肢骨骼肌废用的动物模型,以肌球蛋白ATP酶(mATPase)法测定比目鱼肌的mATPase活性,依据mATPase染色结果进行肌纤维分型,并测量肌纤维横截面积(Cross-section area,CSA),首次观察了尾部悬吊对达乌尔黄鼠比目鱼肌湿重、CSA和梭外肌、梭内肌纤维mATPase活性的影响,并与尾部悬吊大鼠进行了比较。旨在探讨冬眠动物骨骼肌在废用状态下的变化。结果显示,尾部悬吊14d可使大鼠比目鱼肌湿重体重比下降35.52%(P<0.001),Ⅰ型肌纤维CSA和Ⅱ型肌纤维CSA分别下降18.91%和20.68%(P<0.05);肌纤维平均CSA减少20.45%(P<0.01)。比目鱼肌中Ⅰ型肌纤维的构成比由对照组的80.61%降低为66.83%,Ⅱ型肌纤维的构成比由19.39%增加到33.17%(P<0.001);梭内肌纤维mATPase活性增强,核袋1纤维的mATPase染色由阴性(-)转变为强阳性( ),核袋2纤维和核链纤维由阳性( )转变为强阳性( )。而达乌尔黄鼠在尾部悬吊14d后,比目鱼肌湿重仅比对照组下降0.05%,Ⅰ、Ⅱ型肌纤维CSA与平均CSA分别比对照组减少0.84%、0.63%和0.37%,均无明显差异(P>0.05);与对照组相比,比目鱼肌中Ⅰ型肌纤维的构成比从82.55%减少到77.30%,Ⅱ型肌纤维的构成比由17.45%增加到22.70%(P<0.05);梭内肌纤维mATPase活性亦明显升高,核袋1纤维的mATPase染色由对照组的阴性(-)转化为强阳性( ),核袋2纤维及核链纤维则由对照组的阳性( )转化为核袋2纤维呈阳性( ),核链纤维则呈弱阳性( )。结果表明:尾部悬吊可致大鼠比目鱼肌明显萎缩;达乌尔黄鼠比目鱼肌则无明显萎缩;两者比目鱼肌梭内、外肌mATPase活性均明显升高。     

模拟失重条件下大鼠比目鱼肌肌梭神经营养因子3表达降低

本研究旨在探讨模拟失重对大鼠比目鱼肌肌梭神经营养因子3(neurotrophin-3,NT-3)表达的影响。采用大鼠尾部悬吊法建立模拟失重动物模型,按体重配对原则随机将大鼠分为5组,即尾悬吊3d组、7d组、14d组、21d组和正常同步对照组。采用免疫组织化学ABC染色法及酶联免疫吸附实验法(ELISA)检测大鼠比目鱼肌肌梭NT-3的表达。结果显示,大鼠比目鱼肌梭外肌中未见到NT-3表达;正常对照组大鼠比目鱼肌肌梭中,核袋1和核袋2纤维NT-3呈现强阳性表达;模拟失重后,梭内肌纤维的NT-3免疫染色反应进行性降低;NT-3的ELISA定量检测结果显示,正常组、尾悬吊3d组、7d组、14d组和21d组大鼠比目鱼肌NT-3的含量分别为(14.23±1.65)、(14.11±1.53)、(13.09±1.47)、(12.45±1.51)和(9.85±1.52)pg/mg。统计比较显示,尾悬吊14d后,大鼠比目鱼肌NT-3的含量较正常对照组明显减少(P<0.05);而尾悬吊21d后,大鼠比目鱼肌NT-3的表达进一步减少(P<0.01)。以上结果表明,模拟失重可致大鼠比目鱼肌肌梭NT-3的表达明显减少,并且随着模拟失重时...     

改良的大鼠模拟失重模型制备方法*

目的: 介绍一种改良的尾部悬吊使后肢去负荷的制备大鼠模拟失重模型的方法。方法: 90只成年雄性SD大鼠随机分为对照组,经典尾吊组和改良尾吊组(每组30只)。经典尾吊组利用医用胶带和纱布制作大鼠尾套后悬吊大鼠尾部。改良尾吊组在上述操作基础之上,在尾套内增加聚乙烯发泡棉隔层,以缓冲纱布对尾部的挤压,保证远端血液循环。对照组尾部不做特殊处理。尾吊4周后观察尾部损伤和尾套脱落等并发症,测量大鼠体质量及右侧比目鱼肌湿重。结果: 与对照组相比,经典尾吊组的比目鱼肌湿重/体质量比值显著减少,但体质量无显著差异,大鼠尾部远端出现缺血坏死损伤的发生率为40.0%,尾套脱落的发生率为26.7%,模型成功率为33.3%;与经典尾吊组相比,改良尾吊组的尾部损伤程度明显降低,远端缺血坏死率为13.3%, 尾套脱落率为3.3%,模型成功率为83.3% (P均＜0.05)。结论: 采用改良尾吊法建立大鼠模拟失重模型能够显著减少鼠尾坏死和尾套脱落发生率,简单易行,提高了模型制备的成功率。     

大鼠和家兔生后发育各阶段比目鱼肌纤维的比较

为研究大鼠与家兔骨骼肌各类型肌纤维的数量和二维分布以及生后发育对其影响,取生后2d和2、4、6、8、10周龄(体重10g和32、95、190、280、320g)大鼠及生后2d和2、4、8、12、16、20、24周龄(体重100g和220、400、750、1200、1600、2100、2500g)家兔的比目鱼肌做琥珀酸脱氢酶染色。实验结果表明,大鼠和家兔比目鱼肌纤维被分成Ⅰ型(SO),ⅡX型(FO)和ⅡA型(FOG)3型。使用图像分析系统分析每型肌纤维在生后发育各阶段的相关变化,大鼠和家兔比目鱼肌中：Ⅰ型纤维分布于整块肌肉,其数量随着生后发育而增加。幼体ⅡX型纤维分布在整块肌肉中,其数量随生后发育而减少;ⅡA型分布在肌肉中深层,数量几乎无变化;至成体时只有少量的ⅡX和ⅡA分布在肌表层。整个发育期间未见ⅡB型纤维。ⅡA型纤维直径最大,Ⅰ型中等,而ⅡX型最小。家兔3型肌纤维的平均横切面积比大鼠的大。这些结果表明大鼠和家兔后肢肌各种类型肌纤维的数量比例和分布随生长过程发生改变。     

肌卫星细胞在失重肌萎缩中的可塑性变化及机制

肌卫星细胞在骨骼肌生长发育和出生后骨骼肌损伤修复中起着重要的作用,但是有关肌萎缩中肌卫星细胞的可塑性变化、作用及其机制尚不清楚.本研究采用小鼠尾悬吊模拟失重效应诱导失重肌萎缩,动态分析了失重肌萎缩发生过程中不同类型肌纤维的肌卫星细胞数量和增殖、分化潜能可塑性的改变,发现在失重肌萎缩过程中,处于安静状态的肌卫星细胞显著增多、激活增殖的肌卫星细胞显著减少,而具有成肌分化潜能的肌卫星细胞有持续减少趋势.此外,在失重肌萎缩比目鱼肌单根肌纤维移出的体外培养中,证明了失重肌萎缩肌纤维肌卫星细胞可塑性降低的特征性变化.进一步,通过对比分析Smad3基因敲除及其同窝野生型小鼠,在失重肌萎缩中肌卫星细胞可塑性的差异性变化,揭示了Smad3在调控失重肌萎缩肌卫星细胞可塑性变化中的关键作用.     

家兔与大鼠腓肠肌的生后发育比较

研究分析了家兔、大鼠腓肠肌在生后各年龄阶段的内侧头、外侧头的内侧亚体、中间亚体或外侧浅亚体、外侧亚体或外侧深亚体内快慢肌纤维的发育情况，应用大体解剖结合组织化学方法确定了其肌亚体，并进行琥珀酸脱氢酶染色、图像分析两型肌纤维的直径特征，以及肌构筑学与肌诱发电位的测量。结果表明：家兔在生后1个月时，内侧亚体从其深面凸现于内侧头与外侧亚体之间，中间亚体居于内侧亚体远端；大鼠内侧头未能区分肌亚体，其外侧头分为内侧亚体、外侧浅亚体，而外侧深亚体居于外侧浅亚体的深面呈重叠状：生后2、3天均未能分出Ⅰ、Ⅱ型肌纤维，也未见有原始肌束；Ⅰ、Ⅱ型肌纤维比例随年龄增长而变化，内侧亚体的Ⅱ型肌纤维比例在家兔与大鼠的生后发育中始终占优，而在其它各肌亚体内，Ⅱ型肌纤维的比例在发育中不恒定，直至成年后Ⅱ型肌纤维才趋于稳定并占优。在生长发育过程中，各肌亚体内Ⅱ型肌纤维的直径在各年龄段均大于Ⅰ型肌纤维。生后6个月家兔外侧头内侧亚体(FL／CSA)比值越大，倾向于速度型构筑；内侧头、中间亚体和外侧亚体(CSA／MW)比值越大，倾向于力量型构筑。大鼠腓肠肌外侧头内侧亚体乙酰胆碱酯酶染色显示葡萄状运动神经末梢支配慢肌纤维，斑点状运动终板位于快肌纤维。     

大鼠与家兔生后各年龄阶段跖肌纤维的比较

应用建立在肌球蛋白重链异构体基础上的标准肌动球蛋白ATP酶和琥珀酸脱氢酶组织化学方法,分析大鼠和家兔出生后发育各年龄阶段跖肌纤维型分布。在生后2周至24周龄的大鼠和家兔Ⅰ、ⅡX型肌纤维百分比例减少,而ⅡA、ⅡB型纤维则增加。进行大量单肌纤维的组织化学特征的比较和相关性探讨。结果显示动物平均体重与跖肌的平均湿重随生后发育逐渐增加,Ⅰ、ⅡX、ⅡA及ⅡB型纤维均在生后各年龄组的全部肌肉内被发现,但出生后2日龄组是个例外。在生后发育期间,雄性大鼠和家兔ⅡB型纤维的平均肌纤维型构成要大于雌性大鼠和家兔,而雄性大鼠和家兔Ⅰ、ⅡX、ⅡA型三种氧化组织化学分类的肌纤维型构成均小于雌性大鼠和家兔。大鼠Ⅰ、ⅡX、ⅡA和ⅡB型纤维的平均横切面积显然要比家兔的同类型肌纤维要小。在大鼠和家兔可见明显的性别差异。大鼠和家兔的ⅡX型纤维横切面积是最小的,Ⅰ、ⅡA型纤维呈中等大小,ⅡB型纤维最大。该重要的测试有助于我们深入研究啮齿类动物快肌纤维生理特征的适应。     

尾悬吊大鼠比目鱼肌‘肌肉生长抑制素’基因的表达水平

目的确定尾悬吊大鼠比目鱼肌组织中‘肌肉生长抑制素’(myostatin,MSTN)基因的表达水平。方法在测定尾悬吊14、30 d大鼠后肢比目鱼肌相对湿重的基础上,采用real-time RT-PCR方法,在基因水平对尾悬吊14、30 d大鼠比目鱼肌中MSTN mRNA水平进行检测。结果尾悬吊14、30 d后比目鱼肌的相对湿重分别下降了11%和19%。real-time RT-PCR结果表明,大鼠尾悬吊14、30 d后,比目鱼肌中MSTN mRNA表达水平均有所提高,分别是对照大鼠的2.2倍和3.5倍。结论尾悬吊大鼠后肢去负荷后可诱导抗重力肌-比目鱼肌组织中MSTN mRNA表达水平的上调。     

幼龄、亚成年、成年达乌尔黄鼠比目鱼肌Ⅰ和Ⅱ型肌纤维的比例与脏器指数的比较

选取达乌尔黄鼠(Spermophilusdauricus)24只,按年龄分为幼龄、亚成年和成年3组。采用Ca2 -ATPase法测定比目鱼肌的mATPase活性,计算Ⅰ型和Ⅱ型肌纤维的比例,并称量达乌尔黄鼠脏器重量,计算脏器指数。结果显示(1)成年组和亚成年组达乌尔黄鼠比目鱼肌中Ⅱ型肌纤维比例均极显著地低于幼龄组;成年组Ⅱ型肌纤维比例也有明显低于亚成年组的趋势;(2)幼龄组达乌尔黄鼠的胸腺指数显著高于亚成年组和成年组,脾脏指数显著高于亚成年组;肝脏指数则均极其显著低于亚成年组和成年组;成年组与亚成年组相比,肝脏指数显著增高,其他指数无显著性差异。以上结果提示,在生长发育过程中,达乌尔黄鼠比目鱼肌的mATPase活性逐渐降低,因而其Ⅱ型肌纤维的比例逐渐减小;胸腺指数、脾脏指数均逐渐降低,肝脏指数则逐渐升高。     

血管紧张素Ⅱ对废用性骨骼肌萎缩肌纤维类型的影响

目的:观察血管紧张素Ⅱ在废用性肌萎缩过程中对骨骼肌肌纤维类型转化的影响。方法:以雌性SD大鼠为研究对象,按随机配对原则分成4组(n=8):即假手术组(C)、AngⅡ组(CA)、悬吊组(T)和悬吊+AngⅡ(TA)。ATPase染色法分析评价各组肌纤维类型的转化情况。结果:T组与C组相比,Ⅱ型肌纤维比例显著升高(P<0.05),而TA组与T组相比Ⅱ型肌纤维比例下降(P<0.05)。结论:在骨骼肌萎缩过程中,肌纤维类型出现Ⅰ型肌纤维向Ⅱ型肌纤维类型的转化;而血管紧张素Ⅱ在骨骼肌萎缩过程中抑制了肌纤维类型由慢向快的转化。     

Intrafusal muscle fibers of duck muscle spindles

Serial transverse paraffin sections of intrafusal muscle fibers of spindles from the extensor pollicis and the extensor digitorum communis of ducks show that only one type of intrafusal muscle fiber exists, based on the mid-equatorial nucleation pattern, diameter, and length. Although the overall range in fiber diameter at the mid-equatorial region is between 4.2-20.0 microns, the average caliber is 10.4 +/- 3.18 microns (S.D.) for spindles of the extensor pollicis and 9.3 +/- 2.11 microns (S.D.) for spindles of the extensor digitorum communis muscles. The range in spindle length for the extensor pollicis is 290-2,090 microns, average 1,120 +/- 569 microns (S.D.), and for the extensor digitorum communis 1,160-2,500 microns, average 1,745 +/- 367 microns (S.D.). The range in number of fibers per spindle for the extensor pollicis muscle is 5-12, average 8.2, and for the extensor digitorum muscle it is 1-11. In the extensor digitorum communis, there appear to be two groups, based on fiber number. Spindles of one group have a range of 5-11 fibers per spindle with an average of 7.2, whereas the second group has a range of 1-4 with an average of 2.7 fibers per spindle. The second group of spindles constitutes 52.5% of the 40 spindles studied, and of these 7.5% were monofibril spindles, 15.0% difibril, 17.5% trifibril, 12.5% quadrifibril spindles.     

Morphology of muscle fibres in amphibian submandibular muscle

The microscopic organization and ultrastructure of the submandibular muscle of 10 species of Amphibia were compared. Among other fibre features the diameter of fibres, their content of mitochondria and fat, organization of sarcomeres: morphology of Z-line, M-band and sarcoplasmic reticulum were taken into consideration and 4 main types of muscle fibres were distinguished. They correspond to tonic (slow) and phasic (red, white and intermediate) ones. Slight variety of fibre morphology and of fibre elements among the examined species was found. Special attention to the variety of fibre morphology among the established types has been paid and the existence of continuous "spectrum" of fibres was suggested. The correlation of frequency of fibres of particular types with the body size, gular oscillation frequency, and some other characteristics of the submandibular muscle in the examined species was discussed. Also the zonal arrangement of muscle according to the fibre types, as well as possible dynamic nature of muscle fibres were emphasised.     

Novel clearance of muscle proteins by muscle cells

Blood levels of cardiac troponins (cTn) and myoglobin are analysed when myocardial infarction (MI) is suspected. Here we describe a novel clearance mechanism for muscle proteins by muscle cells. The complete plasma clearance profile of cTn and myoglobin was followed in rats after intravenous or intermuscular injections and analysed by PET and fluorescence microscopy of muscle biopsies and muscle cells. Compared with intravenous injections, only 5 % of cTnT, 0.6 % of cTnI and 8 % of myoglobin were recovered in the circulation following intramuscular injection. In contrast, 47 % of the renal filtration marker FITC-sinistrin and 81 % of cTn fragments from MI-patients were recovered after intramuscular injection. In addition, PET and biopsy analysis revealed that cTn was taken up by the quadriceps muscle and both cTn and myoglobin were endocytosed by cultured muscle cells. This local clearance mechanism could possibly be the dominant clearance mechanism for cTn, myoglobin and other muscle damage biomarkers released by muscle cells.     

Functional dependence of single muscle receptors on muscle activity

In experiments on chloraloso-urethane anesthetized cats changes in spontaneous and induced spike activity of single muscle spindles and Golgi's receptors following a direct and/or indirect electrical stimulation of the muscle were studied. It was found that contractile activity of the plantar (phase) and heel (tonic) muscles decreased the spike activity of both muscle spindle and Golgi's receptors, the decrease in the phase muscle spindle activity being more considerable than in the tonic one.     

Formation of muscle spindles in regenerated avian muscle grafts

Summary Pigeon muscles lacking muscle spindles were grafted into sites which normally have a muscle containing spindles. The reciprocal transplantations were also made. After two to eight months, the graft of the donor muscle without spindles had regenerated into a muscle containing muscle spindles. The reciprocal grafts, muscles containing spindles transplanted to a site lacking spindle innervation, had neither muscle spindles nor remnants of the spindles. These experiments demonstrate that 1) the innervation is required for formation of the spindle; 2) the original spindles do not survive transplantation; and 3) parts of the original spindle are not required for spindle regeneration.This work was supported in part by NSF grants PCM 77-15960 and PCM 79-16540