QUICK SEARCH:   [advanced] Author: Keyword(s):
Home     Help     Feedback     Subscriptions     Archive     Search     Table of Contents

First published online 20 January 2004
doi: 10.1242/jcs.00911

This Article Summary Full Text Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Yurchenco, P. D. Articles by Li, S. Search for Related Content PubMed PubMed Citation Articles by Yurchenco, P. D. Articles by Li, S. Social Bookmarking                         What's this?

Loss of basement membrane, receptor and cytoskeletal lattices in a laminin-deficient muscular dystrophy

¹ Department of Pathology & Laboratory Medicine, UMDNJ-Robert Wood Johnson Medical School, Piscataway, NJ 08854, USA
² Howard Hughes Medical Institute, Department of Physiology & Biophysics, University of Iowa College of Medicine, Iowa City, IA 52242, USA

View larger version (116K): [in a new window]   Fig. 1. Laminin (Lm) and -dystroglycan (DG) in normal skeletal muscle. (a-d) Confocal immunofluorescence images of the surface of normal teased muscle fibers immunostained with laminin 2-G domain (a, FITC secondary, green) and -dystroglycan (b, IIH6, Cy3, red) antibodies (a-c, fixed and detergent treated). Laminin and dystroglycan striated patterns colocalize (arrows in a), with less intense laminin fluorescence between striations. (d) Similar fiber image with laminin-dystroglycan costained fiber treated with detergent following laminin immunostaining. (e-h) Peripheral nuclei (e, laminin; f, laminin (green) and -actinin (red); g, oblique optical section showing nucleus protruding beyond plane of sarcolemma (dystroglycan, red; -actinin, green); h, laminin, green; propidium iodide, red). Peripheral nuclei are sandwiched between -actinin of the sarcomere (which coincides with the major costameric circumferential striations) and plasma membrane dystroglycan. (i-j) Periodicity of orthogonally oriented longitudinal optical sections (i, confocal image, laminin-2/4 above and -dystroglycan below; j, deconvoluted widefield image, laminin 2G above and dystroglycan below) of normal fibers. Arrows indicate positions of Z-discs. (k-o) Widefield images reveal repeating pattern of laminin 2G epitope on internal (k,l) and external (m,n) basement membrane surfaces. (k) An oblique view of an internal basement membrane lying between adjacent fibers contained within a computed three-dimensional pixel (voxel) array prepared from 0.25 µm serial deconvoluted optical sections. (l) Location of internal basement membranes within optical cross-section through three adjacent fibers (arrows). (m-o) Images of fixed teased fiber prepared without detergent and stained with laminin 2G-specific antibody following (m) and before (n) deconvolution, and incubated only with secondary antibody (o). Costameric elements (Z, M, L striations) and location of peripheral nucleus (n) indicated in m.

View larger version (97K): [in a new window]   Fig. 2. Defects in the surface organization of laminin and dystroglycan in dy2J muscle. Confocal images of teased dy2J dystrophic muscle fibers (b-e: b, merged image; c, laminin-2-G; d, -dystroglycan). A wild-type control fibril (merged image) is shown in a. (e) A deconvoluted widefield cross-sectional view of dystrophic basement membrane zone (dystroglycan, red; laminin, green; merge, yellow). Note extensive (but not complete) effacement of rectilinear distributions for surface-distributed basement membrane, receptor and cytoskeletal components. Bars, 2 µm in e.

View larger version (101K): [in a new window]   Fig. 3. Defects in the surface organization of basement membrane, receptor and cytoskeletal components in dy2J muscle. Confocal images of wild-type (wt) and dystrophic (dy2J) muscle fibers. Muscle fibers immunostained to detect laminin-2 (Lm) and corresponding -actinin (a), dystrophin (Dys) and corresponding dystroglycan (DG), integrin ß1D, vinculin (Vn), nidogen (Nd), type IV collagen (Col4) and perlecan (Perl). Nonimmune (wild-type) controls for integrin (C1), vinculin (C2), dystrophin/basement membrane components (C3) and dystroglycan (C4) included. Note the extensive but incomplete effacement of rectilinear distributions for surface-distributed basement membrane, receptor and cytoskeletal components with preservation of the sarcomeric -actinin cross-band architecture. Arrows in top row panel indicate locations of peripheral nuclei which overlie the sarcomeres. Bar, 2 µm.

View larger version (102K): [in a new window]   Fig. 4. Grading of normal and dy2J surface architectures. Three classes of surface architecture were defined from the dystroglycan immunofluorescence patterns. Class 1, seen in normal muscle, corresponds to a regular rectilinear pattern. Class 2, largely present in dystrophic muscle, consists of a less regular lattice with splaying and with frequent loss of the finer circumferential and longitudinal lattice struts. Class 3 is a dystrophic pattern characterized by substantial loss of the rectilinear pattern with extensive inter-array clearing (shown) or near-complete pattern effacement. Corresponding paired images with 1.5x magnified insets for dystroglycan shown.

View larger version (35K): [in a new window]   Fig. 5. Age-dependent transitions in dy2J surface architecture. The orthogonal pattern in 6.5-week and 11-week-old dy2J/dy2J mice and their normal littermates was scored. Number of determinations (n, image fields) indicated in panels. The class 2 and 3 changes are characteristics of dystrophic muscle, developing after 6.5 weeks.

View larger version (121K): [in a new window]   Fig. 6. Loss of costameric lattice in dy2J fibers with central and peripheral nuclei. Confocal images of teased muscle fibers from an 11-week-old dy2J dystrophic mouse stained with DAPI (blue) to detect nuclei and with IIH6 antibody (red) to detect -dystroglycan. Cross-sectional views (a,b; d,e) reconstructed from serial optical 0.5 µm sections taken approximately at positions indicated by yellow lines in en face views of upper surface (c,f). Red vertical lines indicate lateral borders of fiber in c. Note dystrophic pattern seen in myofiber containing central nuclei (a-c) and peripheral nuclei (d-f).

View larger version (173K): [in a new window]   Fig. 7. Surface distribution of laminin-2/4 and -dystroglycan in mdx dystrophic muscle. Confocal images of teased muscle isolated from an 11-week-old mdx/mdx mouse. The pattern of laminin-2/4 (Lm2) revealed prominent Z bands with variable reductions of M- and longitudinal striations, and more prominent selective loss of the -dystroglycan (DG) inter-Z minor circumferential M bands. The furrows are sites of capillaries removed from the sarcolemma.

View larger version (80K): [in a new window]   Fig. 8. Model of normal and dystrophic costameric patterns. Sarcolemmal basement membrane (bm), plasma membrane (pm), cortical cytoskeleton (cc) and underlying sarcomeres (sm) are shown. Normal (wild-type, wt) basement membrane, although continuous, is condensed into circumferential Z (major) and M (minor) bands and longitudinal (L) striations that correspond to the same pattern elements seen with ß1-integrin and dystroglycan receptors and cytoskeletal vinculin and dystrophin. The basement membrane, receptor and cytoskeletal dy2J lattice-like patterns become generally effaced while the receptor (dystroglycan, this study) and cytoskeletal (spectrin) (Williams and Bloch, 1999) M and L elements of mdx muscle are lost, with less-pronounced changes in the laminin pattern.

CiteULike

Complore

Connotea

Del.icio.us

Digg

Reddit

Technorati

Twitter