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Fig. 4. Myoferlin null myoblasts show impaired fusion in vitro. (A) Myoblasts
isolated from littermate control and myoferlin null neonatal mice were plated
at equal densities and induced to differentiate. After 4 days of
differentiation, cells were fixed and stained with anti-desmin antibodies
(red) and Sytox nuclear dye (green). Scale bar: 50 µm. (B) Desmin is
expressed only in myogenic cells, so the efficiency of fusion was determined
by quantifying the number of singly nucleated desmin-positive cells (white
squares), those containing two to three (green squares) nuclei, and those
containing four or more nuclei (blue squares). The percentage of nuclei not
associated with desmin staining was equivalent in wild-type and myoferlin null
cultures, and these nuclei were not included when determining fusion
efficiency. Myoferlin null cultures displayed significantly more
desmin-positive single nuclei than did littermate control cultures. Myotubes
containing four or more nuclei were reduced in myoferlin null cultures. Eight
10x fields from each genotype were used for quantification, comprising
2390 wild-type and 2485 myoferlin null nuclei.
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