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Axial distribution of myosin binding protein-C is unaffected by mutations in human cardiac and skeletal muscle.

Vydyanath A, Gurnett CA, Marston S, Luther PK - J. Muscle Res. Cell. Motil. (2012)

Bottom Line: Myosin binding protein-C (MyBP-C), a major thick filament associated sarcomeric protein, plays an important functional and structural role in regulating sarcomere assembly and crossbridge formation.Due to the difficulty of obtaining normal human tissue, we compared the distribution to the A-band structure in normal frog skeletal, rat cardiac muscle and in cardiac muscle of MyBP-C-deficient mice.Very similar overall profile averages were obtained from the C-zones in cardiac HCM samples and skeletal DA-1 samples with MyBP-C gene mutations, suggesting that mutations in MyBP-C do not alter its mean axial distribution along the thick filament.

View Article: PubMed Central - PubMed

Affiliation: Faculty of Medicine, National Heart and Lung Institute, Imperial College London, London, UK. a.vydyanath@imperial.ac.uk

ABSTRACT
Myosin binding protein-C (MyBP-C), a major thick filament associated sarcomeric protein, plays an important functional and structural role in regulating sarcomere assembly and crossbridge formation. Missing or aberrant MyBP-C proteins (both cardiac and skeletal) have been shown to cause both cardiac and skeletal myopathies, thereby emphasising its importance for the normal functioning of the sarcomere. Mutations in cardiac MyBP-C are a major cause of hypertrophic cardiomyopathy (HCM), while mutations in skeletal MyBP-C have been implicated in a disease of skeletal muscle-distal arthrogryposis type 1 (DA-1). Here we report the first detailed electron microscopy studies on human cardiac and skeletal tissues carrying MyBP-C gene mutations, using samples obtained from HCM and DA-1 patients. We have used established image averaging methods to identify and study the axial distribution of MyBP-C on the thick filament by averaging profile plots of the A-band of the sarcomere from electron micrographs of human cardiac and skeletal myopathy specimens. Due to the difficulty of obtaining normal human tissue, we compared the distribution to the A-band structure in normal frog skeletal, rat cardiac muscle and in cardiac muscle of MyBP-C-deficient mice. Very similar overall profile averages were obtained from the C-zones in cardiac HCM samples and skeletal DA-1 samples with MyBP-C gene mutations, suggesting that mutations in MyBP-C do not alter its mean axial distribution along the thick filament.

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Light microscopy analysis of muscle biopsy and cardiac myectomy samples taken at ×20 (LHS panels) and ×100 (RHS panels). Paragon staining of semi thin sections of a ss-W236R, b ss-Y856H, c MH1, d M4 and e M9 is shown; the stain highlights cellular components in dark blue and connective tissue pink. Sarcomeric striations are observed in all sections at ×100. Bar = 10 μm. (Color figure online)
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Fig2: Light microscopy analysis of muscle biopsy and cardiac myectomy samples taken at ×20 (LHS panels) and ×100 (RHS panels). Paragon staining of semi thin sections of a ss-W236R, b ss-Y856H, c MH1, d M4 and e M9 is shown; the stain highlights cellular components in dark blue and connective tissue pink. Sarcomeric striations are observed in all sections at ×100. Bar = 10 μm. (Color figure online)

Mentions: Histological examinations performed on 0.5 μm thick plastic sections of the MyBP-C DA-1 patient samples are shown in Fig. 2. The morphology of the samples was highlighted by Paragon stain, which renders cellular components dark blue and connective tissue such as collagen pink. At lower magnification the staining of the muscle fibres is seen in blue, and at 100×, the striation pattern due to the sarcomeres in the longitudinal orientation can be clearly recognized in all the samples (both cardiac and skeletal). Light micrographs recorded with 20× objective and 100× oil immersion objective are shown in Fig. 2. The skeletal mutants displayed variable levels of staining for connective tissue (Fig. 2a, b). The amount of connective tissue appears to be more abundant in ss-Y856H (Fig. 2b) compared to ss-W236R (Fig. 2a). The ss-W236R tissue sections also show signs of severe bending of the myofibrils in some regions (indicated by "*" symbol in Fig. 2a). This is not an effect of sample mishandling because the surrounding fibres are normal (marked with a + symbol). This type of misalignment was not obvious for ss-Y856H under the light microscope since most of the fibres in the sample were in the transverse orientation, but was more clearly observed while imaging with EM (Ultrastructure of the diseased tissue).Fig. 2


Axial distribution of myosin binding protein-C is unaffected by mutations in human cardiac and skeletal muscle.

Vydyanath A, Gurnett CA, Marston S, Luther PK - J. Muscle Res. Cell. Motil. (2012)

Light microscopy analysis of muscle biopsy and cardiac myectomy samples taken at ×20 (LHS panels) and ×100 (RHS panels). Paragon staining of semi thin sections of a ss-W236R, b ss-Y856H, c MH1, d M4 and e M9 is shown; the stain highlights cellular components in dark blue and connective tissue pink. Sarcomeric striations are observed in all sections at ×100. Bar = 10 μm. (Color figure online)
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Related In: Results  -  Collection

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Fig2: Light microscopy analysis of muscle biopsy and cardiac myectomy samples taken at ×20 (LHS panels) and ×100 (RHS panels). Paragon staining of semi thin sections of a ss-W236R, b ss-Y856H, c MH1, d M4 and e M9 is shown; the stain highlights cellular components in dark blue and connective tissue pink. Sarcomeric striations are observed in all sections at ×100. Bar = 10 μm. (Color figure online)
Mentions: Histological examinations performed on 0.5 μm thick plastic sections of the MyBP-C DA-1 patient samples are shown in Fig. 2. The morphology of the samples was highlighted by Paragon stain, which renders cellular components dark blue and connective tissue such as collagen pink. At lower magnification the staining of the muscle fibres is seen in blue, and at 100×, the striation pattern due to the sarcomeres in the longitudinal orientation can be clearly recognized in all the samples (both cardiac and skeletal). Light micrographs recorded with 20× objective and 100× oil immersion objective are shown in Fig. 2. The skeletal mutants displayed variable levels of staining for connective tissue (Fig. 2a, b). The amount of connective tissue appears to be more abundant in ss-Y856H (Fig. 2b) compared to ss-W236R (Fig. 2a). The ss-W236R tissue sections also show signs of severe bending of the myofibrils in some regions (indicated by "*" symbol in Fig. 2a). This is not an effect of sample mishandling because the surrounding fibres are normal (marked with a + symbol). This type of misalignment was not obvious for ss-Y856H under the light microscope since most of the fibres in the sample were in the transverse orientation, but was more clearly observed while imaging with EM (Ultrastructure of the diseased tissue).Fig. 2

Bottom Line: Myosin binding protein-C (MyBP-C), a major thick filament associated sarcomeric protein, plays an important functional and structural role in regulating sarcomere assembly and crossbridge formation.Due to the difficulty of obtaining normal human tissue, we compared the distribution to the A-band structure in normal frog skeletal, rat cardiac muscle and in cardiac muscle of MyBP-C-deficient mice.Very similar overall profile averages were obtained from the C-zones in cardiac HCM samples and skeletal DA-1 samples with MyBP-C gene mutations, suggesting that mutations in MyBP-C do not alter its mean axial distribution along the thick filament.

View Article: PubMed Central - PubMed

Affiliation: Faculty of Medicine, National Heart and Lung Institute, Imperial College London, London, UK. a.vydyanath@imperial.ac.uk

ABSTRACT
Myosin binding protein-C (MyBP-C), a major thick filament associated sarcomeric protein, plays an important functional and structural role in regulating sarcomere assembly and crossbridge formation. Missing or aberrant MyBP-C proteins (both cardiac and skeletal) have been shown to cause both cardiac and skeletal myopathies, thereby emphasising its importance for the normal functioning of the sarcomere. Mutations in cardiac MyBP-C are a major cause of hypertrophic cardiomyopathy (HCM), while mutations in skeletal MyBP-C have been implicated in a disease of skeletal muscle-distal arthrogryposis type 1 (DA-1). Here we report the first detailed electron microscopy studies on human cardiac and skeletal tissues carrying MyBP-C gene mutations, using samples obtained from HCM and DA-1 patients. We have used established image averaging methods to identify and study the axial distribution of MyBP-C on the thick filament by averaging profile plots of the A-band of the sarcomere from electron micrographs of human cardiac and skeletal myopathy specimens. Due to the difficulty of obtaining normal human tissue, we compared the distribution to the A-band structure in normal frog skeletal, rat cardiac muscle and in cardiac muscle of MyBP-C-deficient mice. Very similar overall profile averages were obtained from the C-zones in cardiac HCM samples and skeletal DA-1 samples with MyBP-C gene mutations, suggesting that mutations in MyBP-C do not alter its mean axial distribution along the thick filament.

Show MeSH
Related in: MedlinePlus