Fig 4. Shp2 deletion did not prevent the neural crest from giving rise to the periocular mesenchyme.
(A) In E10.5 En1-Cre;Fgf8f/f;Fgf15-/- embryos, Fgf8 was ablated in the midbrain-hindbrain junction, where FGF signaling response gene Etv5 was also down regulated, indicating a loss of FGF signaling. Nonetheless, lacrimal gland budded at E15.5 was unaffected. Arrow and arrowhead: Fgf8 and Etv5 expressions in the midbrain-hindbrain junction. Asterisks: lacrimal gland bud. (B) The migrating neural crest marked by Sox10 expression was reduced in Wnt1-Cre;Shp2f/f and Wnt1-Cre; Mek1f/f;Mek2-/- mutants, but rescued in Wnt1-Cre;Shp2f/f;LSL-KrasG12D embryos. Arrow and arrowhead: Sox10 positive neural crest cells in the periocular mesenchyme. e: eye. (C) Deletion of p53 in Shp2 mutants failed to prevent aberrant apoptosis in the branchial arches and in the periocular mesenchyme shown by lysotracking (upper panel) and cleaved caspase 3 staining (middle panel), respectively. Lacrimal gland budding was not rescued in the Shp2/p53 double mutants (bottom panel). Arrow: lysotracker staining in the branchial arch. Arrowhead: apoptotic cells in the periocular mesenchyme. Asterisk: developing lacrimal gland bud. (D) Lineage tracing by crossing Wnt1-Cre mice with R26R reporter mice showed that Shp2 ablation did not prevent neural crest cells from populating the periocular mesenchyme after E13.5. Arrow: Xgal-stained neural crest cells. (E) Periocular mesenchyme markers Pitx2 and Foxc1 were unperturbed in Shp2 mutants.