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Phillip E. Kish, Dan S. Kasprick, Donika D. Gallina, Alon Kahana; Early Proliferative Changes During Zebrafish Lateral Rectus Muscle Regeneration. Invest. Ophthalmol. Vis. Sci. 2011;52(14):1562.
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To examine the switch from quiescence to proliferation in a severely damaged zebrafish extraocular muscle (EOM).
Lateral rectus (LR) was myectomized, and the removed muscle was saved for analysis. BrdU pulse-labeling was used to identify the proliferating zone within the regenerating muscle. Fish were allowed to recover for 48 hours (hps), and then the residual muscle stump on the sclera was collected surgically. Fish were euthanized, and heads cryosectioned. Leica LMD6500 was used to collect non-regenerating and regenerating muscle. Microarray gene expression analysis was performed on collected tissues. Results were validated by PCR and in situ hybridization techniques.
Following near-total LR myectomy, zebrafish could regenerate a functional muscle over 10 days. BrdU labeling of proliferating cells demonstrated a proliferative zone marking the leading edge of the regenerating muscle. Microarray analysis 48 hrs post-injury revealed significant up-regulation of genes associated with cell proliferation, such as midkine-A, cyclins, cdks, and bmyb. Concurrently, genes associated with muscle identity, e.g. actin, myosin, were down-regulated. Differential interference contrast (DIC) light microscopy revealed loss of cell and tissue architecture that extended beyond the proliferative region.
Following injury, zebrafish EOMs repopulate via a proliferative burst focused on the tissues closest to the injury, marking a leading edge of repair. This burst is associated with down regulation of muscle markers, and up regulation of cell cycle markers. Analysis of these cells will help with identification of zebrafish EOM satellite cells, and lays the groundwork for further research on EOM regeneration following injury.
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