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Immunolabelling, histochemistry and in situ hybridisation in human skeletal muscle fibres to detect myosin heavy chain expression at the protein and mRNA level

Published online by Cambridge University Press:  24 August 2001

A. L. SERRANO
Affiliation:
Department of Comparative Anatomy and Pathological Anatomy (Muscle Biology Laboratory), University of Cordoba, Spain. Department of Biomedical Sciences and CNR Centre of Muscle Biology and Physiopathology, University of Padova, Italy;.
MARGARITA PÉREZ
Affiliation:
Departamento de Ciencias Morfológicas y Fisiología, Universidad Europea de Madrid, Spain
A. LUCÍA
Affiliation:
Departamento de Ciencias Morfológicas y Fisiología, Universidad Europea de Madrid, Spain
J. L. CHICHARRO
Affiliation:
Departamento de Enfermería, Universidad Complutense de Madrid, Spain
E. QUIROZ-ROTHE
Affiliation:
Department of Comparative Anatomy and Pathological Anatomy (Muscle Biology Laboratory), University of Cordoba, Spain.
J. L. L. RIVERO
Affiliation:
Department of Comparative Anatomy and Pathological Anatomy (Muscle Biology Laboratory), University of Cordoba, Spain.
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Abstract

The distribution of muscle fibres classified on the basis of their content of different myosin heavy chain (MHC) isoforms was analysed in vastus lateralis muscle biopsies of 15 young men (with an average age of 22 y) by correlating immunohistochemistry with specific anti-MHC monoclonal antibodies, myofibrillar ATPase (mATPase) histochemistry and in situ hybridisation with probes specific for MHC β-slow, MHC-IIA and MHC-IIX. The characterisation of a large number of individual fibres was compared and correlated on a fibre-to-fibre basis. The panel of monoclonal antibodies used in the study allowed classification of human skeletal muscle fibres into 5 categories according to the MHC isoform they express at the protein level, types I, I+IIA, IIA, IIAX and IIX. Hybrid fibres coexpressing two isoforms represented a considerable proportion of the fibre composition (about 14%) and were clearly underestimated by mATPase histochemistry. For a very high percentage of fibres there was a precise correspondence between the MHC protein isoforms and mRNA transcripts. The integrated methods used demonstrate a high degree of precision of the immunohistochemical procedure used for the identification and quantification of human skeletal muscle fibre types. The monoclonal antibody S5-8H2 is particularly useful for identifying hybrid IIAX fibres. This protocol offers new prospects for muscle fibre classification in human experimental studies.

Type
Paper
Copyright
© Anatomical Society of Great Britain and Ireland 2001

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