Detection and imaging of non-contractile inclusions and sarcomeric anomalies in skeletal muscle by second harmonic generation combined with two-photon excited fluorescence
Journal
Journal of Structural Biology
Journal Volume
162
Journal Issue
3
Pages
500-508
Date Issued
2008
Author(s)
Ralston E.
Swaim B.
Czapiga M.
Pittis M.G.
Bembi B.
Schwartz O.
Plotz P.
Raben N.
Abstract
The large size of the multinucleated muscle fibers of skeletal muscle makes their examination for structural and pathological defects a challenge. Sections and single fibers are accessible to antibodies and other markers but imaging of such samples does not provide a three-dimensional view of the muscle. Regrettably, bundles of fibers cannot be stained or imaged easily. Two-photon microscopy techniques overcome these obstacles. Second harmonic generation (SHG) by myosin filaments and two-photon excited fluorescence (2PEF) of mitochondrial and lysosomal components provides detailed structural information on unstained tissue. Furthermore, the infrared exciting light can penetrate several layers of muscle fibers and the minimal processing is particularly valuable for fragile biopsies. Here we demonstrate the usefulness of SHG, combined with 2PEF, to reveal enlarged lysosomes and accumulations of non-contractile material in muscles from the mouse model for the lysosomal storage disorder Pompe disease (PD), and in biopsies from adult and infant PD patients. SHG and 2PEF also detect sarcomeric defects that may presage the loss of myofibrils in atrophying muscle and signify loss of elasticity. The combination of SHG and 2PEF should be useful in the analysis and diagnosis of a wide range of skeletal muscle pathologies.
SDGs
Other Subjects
myosin; article; controlled study; fluorescence; glycogen storage disease type 2; imaging; lysosome; microscopy; mouse; muscle biopsy; muscle fibril; muscle malformation; nonhuman; photon; priority journal; skeletal muscle; Adult; alpha-Glucosidases; Animals; Autophagy; Glycogen Storage Disease Type II; Humans; Infant; Infant, Newborn; Mice; Mice, Knockout; Microscopy, Fluorescence; Mitochondria; Muscle Contraction; Muscle, Skeletal; Sarcomeres
Type
journal article