Neurology India
menu-bar5 Open access journal indexed with Index Medicus
  Users online: 6601  
 Home | Login 
About Editorial board Articlesmenu-bullet NSI Publicationsmenu-bullet Search Instructions Online Submission Subscribe Videos Etcetera Contact
  Navigate Here 
 Resource Links
  »  Similar in PUBMED
 »  Search Pubmed for
 »  Search in Google Scholar for
 »Related articles
  »  Article in PDF (2,133 KB)
  »  Citation Manager
  »  Access Statistics
  »  Reader Comments
  »  Email Alert *
  »  Add to My List *
* Registration required (free)  

  In this Article
 »  Abstract
 » Introduction
 » Case Report
 » Discussion
 » Acknowledgment
 »  References
 »  Article Figures

 Article Access Statistics
    PDF Downloaded48    
    Comments [Add]    
    Cited by others 2    

Recommend this journal


Table of Contents    
Year : 2012  |  Volume : 60  |  Issue : 5  |  Page : 510-511

Cytoskeletal and extracellular matrix alterations in limb girdle muscular dystrophy 2I muscle fibers

1 Institute of Molecular Genetic-CNR, Rizzoli Orthopedic Institute, Bologna, Italy
2 Department of Rizzoli-Sicily, Rizzoli Orthopedic Institute, Bologna, Italy
3 Department of Rizzoli-Sicily, Physical Medicine and Rehabilitation, Rizzoli Orthopedic Institute, Bagheria, Italy

Date of Submission20-Jul-2012
Date of Decision19-Aug-2012
Date of Acceptance19-Sep-2012
Date of Web Publication03-Nov-2012

Correspondence Address:
Luciano Merlini
Physical Medicine and Rehabilitation, Department of Rizzoli-Sicily, Rizzoli Orthopedic Institute, SS 113 km 246, 90011 Bagheria
Login to access the Email id

Source of Support: Grant. No. GUP11007 Telethon Italy, Conflict of Interest: None

DOI: 10.4103/0028-3886.103200

Rights and Permissions

 » Abstract 

In this detailed muscle biopsy study of a patient with molecularly confirmed diagnosis of limb-girdle muscular dystrophy 2I (LGMD2I) we show some new data, that is the presence of altered expression pattern of costamere components as integrin α7B and integrin β1D associated with vinculin costameric derangement and basal lamina ultrastructural abnormalities as detachments and discontinuities suggesting that different cellular compartments are involved in LGMD2I and the altered basement membrane-plasmalemma-cytoskeleton binding can underlie muscle degeneration.

Keywords: Fukutin-related protein gene, integrin α7B, itegrin β1D, limb-girdle muscular dystrophy, vinculin costameric derangement

How to cite this article:
Sabatelli P, Pellegrini C, Faldini C, Merlini L. Cytoskeletal and extracellular matrix alterations in limb girdle muscular dystrophy 2I muscle fibers. Neurol India 2012;60:510-1

How to cite this URL:
Sabatelli P, Pellegrini C, Faldini C, Merlini L. Cytoskeletal and extracellular matrix alterations in limb girdle muscular dystrophy 2I muscle fibers. Neurol India [serial online] 2012 [cited 2020 Jul 6];60:510-1. Available from:

 » Introduction Top

Fukutin-related protein (FKRP) is a putative glycosyltransferase localized in the Golgi apparatus. [1] Pathogenic mutations in the FKRP gene cause both the more severe congenital muscular dystrophy Type 1C (MDC1C) and the milder limb-girdle muscular dystrophy Type 2I (LGMD2I) which are associated with dystrophic changes and secondary basal lamina muscle proteins' reduction, particularly of laminin α2 chain and alpha-dystroglycan, independently of mutation type or clinical severity. [2] To gain insight into the LGMD2I pathogenetic mechanism we studied a muscle biopsy of a patient affected by LGMD2I carrying heterozygous mutations in the FKRP gene, by electron microscopy and immunohistochemistry. We found alterations in basal lamina composition and organization, associated with vinculin costameric derangement [Figure 1].
Figure 1: Simplified model of the costameres. The costameres are transverse structures organized in a rib-like pattern, which lie at the sarcolemma over the Z and M lines of nearby myofibrils. This model depicts the relationship of costameres with basal lamina, plasma membrane and cytoskeleton proteins. Two sets of plasma membrane proteins connect the contractile apparatus to the costameres at the sarcolemma: One set via the dystroglycan complex (á-DG andβ-DG) and the other via the integrins (á7β1)

Click here to view

 » Case Report Top

This 37-year-old male had a normal birth and psychomotor milestones and started toe walking at 2 years of age. He experienced a slow but steady progressive weakness. He lost the ability to rise from the floor at the age of 27, and to climb stairs 3 years later, and at the age of 37 years, he needed support to walk outdoors. Examination showed loss of muscle bulk and weakness in the axial, shoulder, hip and thigh muscles. He had normal intelligence, a very mild equinus cavus deformity of the foot but no other contractures. Creatine kinase was 21 times normal. Cardiac examination showed dilated cardiomyopathy, Class II according to the New York Heart Association (NYHA) functional classification, a left ventricular hypertrophy with an ejection fraction of 30%. Forced vital capacity was 51%.

Direct sequencing of the FKRP gene identified 2 mutations: The common c826C>A missense mutation resulting in a p. 276Leu>Ile, and a previously unreported missense mutation c.1074T>C resulting in a p. 359Trp>Arg. This latter mutation was not found in a panel of 100 individuals.

The muscle biopsy showed increased fiber size variability, several internal nuclei, mild connective tissue proliferation, rare necrosis, and some regenerating fibers. The immunohistochemical analysis on frozen sections was performed as previously described. [3] α-dystroglycan glyco-epitope (α-DG) was absent in most of the fibers as detected by VIA4-1 labeling [Figure 2]c. Laminin α2 chain was moderately reduced in numerous fibers [Figure 2]d while laminin-y1 (Chemicon) showed a normal pattern (not shown). Double labeling with anti-laminin α2 chain and anti-integrin α7B antibodies (rabbit polyclonal, gift of E. Engvall) showed an altered expression of both proteins in the same areas of the sarcolemma [Figure 3]c and d. Similar results were obtained with double labeling of anti-integrin β1D-anti-vinculin (SIGMA) antibodies [Figure 3]g and h. The distribution of vinculin and sarcomeric α-actinin (SIGMA) was examined by confocal laser microscopy in longitudinal sections, in areas that provided an en face view of the sarcolemma. [4] In control muscle fibers, vinculin was detected at the costameres, with a labeling pattern characterized by alternating wide and thinner rib-like structures [Figure 4]a-c. In muscle fibers from the LGMD2I patient, vinculin was missing at the thinner longitudinal domains and apparently normally localized at the wider line [Figure 4]f-h. In contrast, α-actinin pattern was comparable to control [Figure 4]d, e, i and l.
Figure 2: Immunofluorescence analysis of α -dystroglycan (a and c) and laminin α2 chain (b and d) in serial sections of muscle biopsy from control (a and b) and LGMD2I patient (c and d) Bar 30 μm

Click here to view
Figure 3: Double labeling of laminin α 2 chain (a and c) and integrin α 7B (b and d) and, integrin β1D (e and g) and vinculin antibodies (f and h) in control (a and b, e and f) and muscle fibers from LGMD2I patient (c and d, g and h) Bar 30 μm

Click here to view
Figure 4: Confocal microscopy analysis of vinculin (a and b, f and g) and sarcomeric α-actinin (d, e, i and l) in longitudinal sections of normal (a-e) and LGMD2I muscle fibers [f- h] Bar 15 μm

Click here to view

The electron microscopy study revealed striking sarcolemmal alterations in non-necrotic muscle fibers, consisting of basal lamina discontinuities [[Figure 5]b, asterisks] and detachments from the plasmalemma [[Figure 5]b, arrows]. In correspondence to these basal lamina detachments the subcortical cytoskeleton appeared disrupted. At this level the dense plaques, cytoskeletal structures linking the contractile apparatus to the plasmalemma were absent [Figure 5]b, arrowheads]. Focal aspects of dense plaque disorganization were also detected in areas where basal lamina was apparently well preserved [[Figure 5]c-d, arrowheads]. The myofibrillar organization of sarcomeres adjacent to areas lacking dense plaques was apparently unaffected.
Figure 5: Electron microscopy analysis of control (a) and LGMD2I muscle biopsies (b-d). Bar 250 nm

Click here to view

 » Discussion Top

In this study we report alterations of basal lamina and costameres in a muscle biopsy from a patient affected by LGMD2I caused by heterozygous mutations in the FKRP gene involving the common p. 276Leu>Ileu and a novel missense mutation. This patient, in spite of the early onset of weakness, had a mild LGMD phenotype (with mild cardiorespiratory compromise, minimal contractures and normal intelligence) much different from the severe congenital muscular dystrophy 1C (MDC1C) due to defects in FKRP. [5] α-DG was undetectable in most of the muscle fibres, possibly as a consequence of its altered glycosylation pattern. In addition, several α-DG deficient fibers showed reduction of laminin α2 chain.

The most noticeable molecular pathology of FKRP-related diseases is the secondary defects in glycosylation of α-DG and laminin α2 chain reduction. [5],[6] Although there is no evidence that fukutin-related protein functions as a glycosyltransferase, [5] sequence analysis showed that FKRP contains a consensus putative catalytic DxD motif commonly found in many glycosyltransferases in the Golgi apparatus. [7] As a consequence of FKRP mutations, hypo-glycosylated α-DG fails to bind to components of the extra-cellular matrix, affecting laminins' deposition in the formation of basement membranes.

Basal lamina alterations have previously been described in muscular dystrophy with secondary deficiency of α-DG and/or laminin α2 chain: Duchenne muscular dystrophy (DMD), [8] Fukuyama congenital muscular dystrophy (FCMD), [9] and Walker-Warburg syndrome (WWS). [3] Thus, α-dystroglycan and laminin α2 chain deficiency detected in muscle fibers of the LGMD2I patient could underlie the structural defect of muscle fibre basal lamina. Integrin α7β1 and laminin α2 chain co-distribute at the sarcolemma in a costameric pattern with subcortical cytoskeleton-associated proteins as talin and vinculin. We reported here an altered expression pattern of costamere components, as integrin α7β1 and loss of the thinner transversal domain of vinculin. Electron microscopy analysis confirmed the costamere alterations: the dense plaques, vinculin-rich structures associated with the sub-cortical cytoskeleton of muscle membrane, appeared disarranged or absent at discrete sites. These alterations have been detected in non-necrotic muscle fibers and may represent an early event in muscle degeneration in LGMD2I muscle fibers. Costameres play a role in maintaining the stability of the myofibril network and this altered binding is known to result in muscle weakness and cellular death. [10] Our data, although limited to one patient, may indicate that different cellular compartments are involved in LGMD2I and, the altered basement membrane-plasmalemma-cytoskeleton binding can underlie muscle degeneration.

 » Acknowledgment Top

The financial support of Telethon - Italy (Grant No. GUP11007) is gratefully acknowledged.

 » References Top

1.Alhamidi M, Kjeldsen Buvang E, Fagerheim T, Brox V, Lindal S, Van Ghelue M, et al. Fukutin-related protein resides in the Golgi cisternae of skeletal muscle fibres and forms disulfide-linked homodimers via an N-terminal interaction. PLoS One 2011;6:e22968.  Back to cited text no. 1
2.Muntoni F, Torelli S, Brockington M. Muscular dystrophies due to glycosylation defects. Neurotherapeutics 2008;5:627-32.  Back to cited text no. 2
3.Sabatelli P, Columbaro M, Mura I, Capanni C, Lattanzi G, Maraldi NM, et al. Extracellular matrix and nuclear abnormalities in skeletal muscle of a patient with Walker-Warburg syndrome caused by POMT1 mutation. Biochim Biophys Acta 2003;1638:57-62.  Back to cited text no. 3
4.Di Giaimo R, Riccio M, Santi S, Galeotti C, Ambrosetti DC, Melli M. New insights into the molecular basis of progressive myoclonus epilepsy: A multiprotein complex with cystatin B. Hum Mol Genet 2002;11:2941-50.  Back to cited text no. 4
5.Godfrey C, Foley AR, Clement E, Muntoni F. Dystroglycanopathies: Coming into focus. Curr Opin Genet Dev 2011;21:278-85.  Back to cited text no. 5
6.Yamamoto LU, Velloso FJ, Lima BL, Fogaca LL, de Paula F, Vieira NM, et al. Muscle protein alterations in LGMD2I patients with different mutations in the Fukutin-related protein gene. J Histochem Cytochem 2008;56:995-1001.  Back to cited text no. 6
7.Esapa CT, McIlhinney RA, Blake DJ. Fukutin-related protein mutations that cause congenital muscular dystrophy result in ER-retention of the mutant protein in cultured cells. Hum Mol Genet 2005;14:295-305.  Back to cited text no. 7
8.Carpenter S, Karpati G. Duchenne muscular dystrophy: Plasma membrane loss initiates muscle cell necrosis unless it is repaired. Brain 1979;102:147-61.  Back to cited text no. 8
9.Matsubara S, Mizuno Y, Kitaguchi T, Isozaki E, Miyamoto K, Hirai S. Fukuyama-type congenital muscular dystrophy: Close relation between changes in the muscle basal lamina and plasma membrane. Neuromuscul Disord1999;9:388-98.  Back to cited text no. 9
10.Bloch RJ, Reed P, O'Neill A, Strong J, Williams M, Porter N, et al. Costameres mediate force transduction in healthy skeletal muscle and are altered in muscular dystrophies. J Muscle Res Cell Motil 2004;25:590-92.  Back to cited text no. 10


  [Figure 1], [Figure 2], [Figure 3], [Figure 4], [Figure 5]

This article has been cited by
1 Mitochondrial Alterations and Oxidative Stress in an Acute Transient Mouse Model of Muscle Degeneration
Renjini Ramadasan-Nair,Narayanappa Gayathri,Sudha Mishra,Balaraju Sunitha,Rajeswara Babu Mythri,Atchayaram Nalini,Yashwanth Subbannayya,Hindalahalli Chandregowda Harsha,Ullas Kolthur-Seetharam,Muchukunte Mukunda Srinivas Bharath
Journal of Biological Chemistry. 2014; 289(1): 485
[Pubmed] | [DOI]
2 Mitochondrial alterations and oxidative stress in an acute transient mouse model of muscle degeneration: Implications for Muscular dystrophy and related muscle pathologies
Ramadasan-Nair, R., Gayathri, N., Mishra, S., Kolthur-Seetharam, U., Bharath, M.M.S.
Journal of Biological Chemistry. 2014; 289(1): 485-509


Print this article  Email this article
Online since 20th March '04
Published by Wolters Kluwer - Medknow