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Semi-automated Analysis of Mouse Skeletal Muscle Morphology and Fiber-type Composition.
Tyagi, Sidharth; Beqollari, Donald; Lee, Chang Seok; Walker, Lori A; Bannister, Roger A.
Afiliação
  • Tyagi S; Department of Medicine-Cardiology Division, University of Colorado School of Medicine.
  • Beqollari D; Department of Medicine-Cardiology Division, University of Colorado School of Medicine.
  • Lee CS; Department of Molecular Physiology and Biophysics, Baylor College of Medicine.
  • Walker LA; Department of Medicine-Cardiology Division, University of Colorado School of Medicine.
  • Bannister RA; Department of Medicine-Cardiology Division, University of Colorado School of Medicine; Roger.Bannister@ucdenver.edu.
J Vis Exp ; (126)2017 08 31.
Article em En | MEDLINE | ID: mdl-28892032
For years, distinctions between skeletal muscle fiber types were best visualized by myosin-ATPase staining. More recently, immunohistochemical staining of myosin heavy chain (MyHC) isoforms has emerged as a finer discriminator of fiber-type. Type I, type IIA, type IIX and type IIB fibers can now be identified with precision based on their MyHC profile; however, manual analysis of these data can be slow and down-right tedious. In this regard, rapid, accurate assessment of fiber-type composition and morphology is a very desirable tool. Here, we present a protocol for state-of-the-art immunohistochemical staining of MyHCs in frozen sections obtained from mouse hindlimb muscle in concert with a novel semi-automated algorithm that accelerates analysis of fiber-type and fiber morphology. As expected, the soleus muscle displayed staining for type I and type IIA fibers, but not for type IIX or type IIB fibers. On the other hand, the tibialis anterior muscle was composed predominantly of type IIX and type IIB fibers, a small fraction of type IIA fibers and little or no type I fibers. Several image transformations were used to generate probability maps for the purpose of measuring different aspects of fiber morphology (i.e., cross-sectional area (CSA), maximal and minimal Feret diameter). The values obtained for these parameters were then compared with manually-obtained values. No significant differences were observed between either mode of analysis with regards to CSA, maximal or minimal Feret diameter (all p > 0.05), indicating the accuracy of our method. Thus, our immunostaining analysis protocol may be applied to the investigation of effects on muscle composition in many models of aging and myopathy.
Assuntos

Texto completo: 1 Coleções: 01-internacional Base de dados: MEDLINE Assunto principal: Fibras Musculares Esqueléticas / Cadeias Pesadas de Miosina Limite: Animals / Humans / Male Idioma: En Revista: J Vis Exp Ano de publicação: 2017 Tipo de documento: Article

Texto completo: 1 Coleções: 01-internacional Base de dados: MEDLINE Assunto principal: Fibras Musculares Esqueléticas / Cadeias Pesadas de Miosina Limite: Animals / Humans / Male Idioma: En Revista: J Vis Exp Ano de publicação: 2017 Tipo de documento: Article