Figure 2

Fate mapping analyses of neural crest cell (NCC) migration in control and NCC-specific integrin-linked kinase (ILK) knockout (NKO) embryos. (A-B’) β-Galactosidase (β-gal) staining at E8.5 (5 somites) shows a slightly increased number of NCCs in the first pharyngeal arch and premature migration of postotic NCCs ((B,B’), arrow, inset). (C-D’) The caudal stream of postotic NCCs in NKO mutant mice seems wider compared to control embryos at E9.0 ((C,D), arrow) and there is significant accumulation of NCCs in the cranial region, the pharyngeal arches and circumpharyngeal region (C,D), some of which have prematurely migrated into aortic sac (as) and the outflow tract (OFT) (C’,D’). (E,F”) Compared to control littermates at E10.5 (E’,E”), there are significantly increased NKO mutant NCCs in the pharyngeal arches 3 to 6. NKO mutant NCCs migrate beyond the conotruncal junction and populate entire layers of the OFT ((F’,F”) red arrowhead). (G-M) immunostaining with phosphohistone H3 (Ph3) antibody revealed a similar rate of proliferation in NCCs (tdTomato+) in the dorsal neural tube and surrounding mesenchyme at E8.5 (S6) (G-H’), and in NCC derived cells (tdTomato+) within the OFTs at E11 (I,J) and E13 (K-L’). Note that, in NKO mutant embryos at E8.5 (S6) (H,H’), there were a significantly increased number of cranial NCCs migrating into paraxial mesenchyme (H), and premature postotic NCC migration (H’). It should also be noted that the sections shown in (G) and (H) are tilted. NKO mutant hearts at E13 exhibit enlarged CAT compared to control littermates ((K,L), aortic arch/distal OFT; (K’,L’) proximal OFT) (aa aortic arch, ao aorta, pa pulmonary artery, tg trigeminal neural crest). Quantitative analysis of Ph3+/tdTomato + cells (mitotic index) (M). (N-O’) Neural tube explants (ex) cultures show abnormal migration of NKO mutant NCCs (O,O’) compared to control NCCs. Panels (N’) and (O’), phase contrast and tdTomato Red merging pictures. (P) Quantitative analysis of migration index/areas of NKO mutant and control NCCs.