Fig. 3.

Wnt3 increases the number of agrin-induced AChR clusters in muscle cells in vitro. Myotubes derived from the C2C12 cell line were exposed to control or Wnt3 conditioned medium overnight in the absence or presence of agrin. (A) Wnt3 alone does not affect the number, size, or distribution of AChR clusters (Upper). However, Wnt3 significantly increases the number of agrin-induced AChR aggregates compared with agrin alone (Lower). (Scale bar, 40 μm.) (B) Quantification shows that Wnt3 enhances the activity of agrin by ≈50% (***, P < 0.001, t test). Data are expressed as the number of AChR clusters per mm² myotube area. (C) Myotube differentiation is not affected, as no difference in the area of myotubes was observed in the various treatments. (D) Quantification shows that, in the presence of agrin, Wnt3 increases the average cluster area by ≈60% (***, P < 0.001, t test). (E) Analyses of size distribution reveals that Wnt3 decreases the percentage of small clusters (*, P < 0.05), does not alter the proportion of medium-sized clusters (P > 0.05), but does increase the proportion of large clusters compared with agrin alone (*, P < 0.05).