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Title Page

Contents

논문요약 6

INTRODUCTION 8

METHODS 10

RESULTS 13

Identification of MyHC Isoforms 13

Distribution of MyHC Isoforms along the Length of Global and Orbital Layers in each extraocular muscle 14

Quantitation of MyHC Isoforms in Global and Orbital Layers via immunohistochemical staining in each of the extraocular muscles 16

DISCUSSION 17

MyHC identifications 17

Distribution of MyHC isoforms in the global layer vs. orbital layer 19

Longitudinal variation 20

Differences between extraocular muscles 22

REFERENCES 24

ABSTRACT 34

List of Figures

Figure 1. SDS-PAGE of human superior rectus, medial rectus, lateral rectus, inferior rectus, and inferior oblique muscle (MyHC emb/IIx, MyHCIIa, Unidentified band1, MyHCeom, MyHCI, Unidentified band2). Because MyHCemb and MyHCIIx migrated together, they were counted together when... 27

Figure 2. Silver stain (left) and immunoblot of human superior rectus (SR) and medial rectus (MR) muscle labeled with A4.1025 antibody (middle) and SR muscle labeled with N2.261 antibody (Right). A4.1025 antibody labeled all MyHC isoforms and N2.261 antibody labeled MyHCeom in addition to MyHCI... 28

Figure 3. Top: Superior rectus muscle was cut into more than 12 pieces as shown. Bottom: Two gels onto which homogenates of the sections prepared from the global and orbital layers of superior rectus muscle were loaded. 29

Figure 4. Relative amounts of each MyHC in serial sections from the global and orbital layers of superior rectus muscles were determined by scanning with visible light and densitometry. 30

Figure 5. Photomicrograph of two sections, from the origin (left) and midbelly (right) portions of the same superior rectus muscle immunostained with anti-MyHCIIa antibody. Notice that large areas in the global layer of the midbelly section are practically unstained by anti-MyHCIIa. OL, orbital layer; GL, global... 31

Figure 6. Photomicrograph of nine sections, from superior rectus muscle immunostained with anti-all MyHC, anti-MyHCI+IIa+eom, anti-neonatal MyHC, anti-MyHCI, anti-MyHCeom, anti-MyHCIIa, anti-MyHCIIb, anti-MyHCIIx, and anti-MyHCemb antibody. OL, orbital layer; GL, global layer.... 32

Figure 7. Photomicrographs of sections of the insertion (left), midbelly (middle), and origin (right) portions of the same superior rectus muscle immunostained with anti-MyHCeom. Notice that anti-MyHCeom stained fibers in both layers, and the staining intensity was higher in the longitudinal central portion of both... 33

초록보기

Myosin isoform expression in human extraocular muscle

Purpose: To determine the distribution of myosin heavy chain (MyHC) isoforms along the length of global and orbital layers of human extraocular muscles (EOM), and compare the distribution of MyHC isoforms between EOMs.

Methods: Human EOMs were assessed via SDS-PAGE, immunoblotting, and immunocytochemistry, using antibodies against MyHC isoforms. Serial samples of orbital and global layers of EOMs were processed with gel electrophoresis, and relative amounts of each MyHC isoform in each sample were determined via scanning densitometry.

Results: Four prominent MyHC isoforms, and two additional MyHC isoforms at very low levels, were expressed. Four prominent MyHC isoforms, MyHCemb/IIx, MyHCIIa, MyHCeom, and MyHCI, were expressed along the entire length of the global and orbital layers with marked gradients in the levels along the length of both layers. In the rectus muscles and inferior oblique muscle, MyHCeom was found to be in higher levels in the central regions, which made up 47% to 54% of the total MyHC; however, the levels of MyHCeom were found to be in lower levels in the proximal and distal ends of the muscle (17% to 25% and 8% to 18%, respectively). In those regions, IIa isoforms predominate. In the superior oblique muscle, the changes in the relative levels of MyHC isoforms along the lengths of the EOMs were smaller than the levels in the rectus muscles.

Conclusions: The present study entailed a quantitative analysis of MyHC isoform expression along the entire length of both layers in human EOMs and between EOMs. Further studies, will be required in order to correlate the expression of myosin with fiber physiology.

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