Fig. 7.

Slit1 and Slit2 can attract primary muscle progenitors. Chemotaxis assays with primary costal muscle progenitors of the diaphragm in a collagen I matrix reveal that ∼50% of all cells are attracted by FGF2 (B) whereas cells are hardly attracted without any chemokine (A). In contrast, Slit1 (C) and Slit2 (D) attract almost one third of muscle progenitors. Exposure of muscle progenitors to a combination of Slit1 and Slit2 further enhances the attractive effect, although roughly one fifth of the cells turn towards repulsion (E). Quantification of migrating cell direction by determining the center of mass shows that FGF2, as well as Slit1 and Slit2 significantly attracted muscle progenitors, whereas the center of mass did not change with the combination of both Slit proteins. Quantitative PCR on E14.5 costal diaphragm muscles validates the expression Robo1 and Robo2 during muscle progenitor migration and fusion and reveals that Npn-1 is also upregulated in cells within the developing diaphragm muscle (G). Isolated primary muscle progenitors have a relatively homogenous size (H) and the majority of primary muscle progenitors are positive for the myogenic marker Desmin (H′). Transwell migration assay shows that Slit1 and Slit2, as well as the combination of both are strongly attractive when compared to the negative control without any ligand addition. FGF2, a known muscle progenitor attractant served as a positive control (I). Quantitative results are mean±s.e.m. (n=4 independent experiments). *P≤0.05, **P≤0.01, ***P≤0.001 (Mann–Whitney U-test). Axis scaling for A–E is in µm.