Rivero J L, Talmadge R J, Edgerton V R
Department of Comparative Anatomy and Pathological Anatomy, Faculty of Veterinary Science, University of Cordoba, Spain.
Electrophoresis. 1997 Oct;18(11):1967-72. doi: 10.1002/elps.1150181115.
In adult horses, three myosin heavy chain (MyHC) isoforms can be identified by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and immunohistochemistry using specific anti-MyHC monoclonal antibodies. This report studies the suitability of a consistent SDS-PAGE technique for quantifying MyHC profiles in homogenized cryostate sections of equine gluteus medius muscle biopsies (n = 18). The method used (previously described by R. J. Talmadge and R. R. Roy; J. Appl. Physiol. 1993, 75, 2337-2340) resolved MyHCs in three bands: I, IIB or IIX, and IIA from the fastest to the slowest migration band. The success rate of the protocol for yielding three well-differentiated MyHC bands was 100% and a subsequent quantification by densitometry for each MyHC isoform was obtained in all 18 muscle biopsies. The results obtained with this electrophoretic method were compared with routine myofibrillar adenosine triphosphatase histochemistry and immunohistochemistry using specific anti-MyHC monoclonal antibodies. The percent composition of the three electrophoretically separated MyHC isoforms (I, IIA and IIB or IIX) showed strong positive correlation with percentages of the area occupied in the biopsies by the three major fiber types (I, IIA, and IIB) identified histochemically (r = 0.96, P < 0.001) and immunohistochemically (r = 0.94, P < 0.01). It can be concluded that the electrophoretic method used here for measuring MyHC content is a valid alternative for muscle fiber typing in horses. As it is less costly and time-consuming than both qualitative histochemistry and immunohistochemistry, the method offers new prospects for application in equine experimental studies and veterinary medicine.
在成年马中，通过十二烷基硫酸钠-聚丙烯酰胺凝胶电泳（SDS-PAGE）和使用特异性抗肌球蛋白重链（MyHC）单克隆抗体的免疫组织化学方法，可以鉴定出三种MyHC亚型。本报告研究了一种一致的SDS-PAGE技术在定量马臀中肌活检匀浆冷冻切片（n = 18）中MyHC谱方面的适用性。所使用的方法（先前由R. J. 塔尔梅奇和R. R. 罗伊描述；《应用生理学杂志》1993年，75卷，2337 - 2340页）将MyHCs分离为三条带：从迁移最快到最慢的带依次为I、IIB或IIX以及IIA。产生三条清晰区分的MyHC带的方案成功率为100%，并且在所有18份肌肉活检样本中均通过光密度法对每种MyHC亚型进行了后续定量。将这种电泳方法获得的结果与常规肌原纤维三磷酸腺苷酶组织化学以及使用特异性抗MyHC单克隆抗体的免疫组织化学结果进行了比较。通过电泳分离的三种MyHC亚型（I、IIA和IIB或IIX）的百分比组成与通过组织化学（r = 0.96，P < 0.001）和免疫组织化学（r = 0.94，P < 0.01）鉴定的活检样本中三种主要纤维类型（I、IIA和IIB）所占面积的百分比呈强正相关。可以得出结论，此处用于测量MyHC含量的电泳方法是马肌肉纤维分型的一种有效替代方法。由于该方法比定性组织化学和免疫组织化学成本更低且耗时更少，因此为马的实验研究和兽医学应用提供了新的前景。
