Figure 5: Adamts9^del/+ eyes have aberrant and intra-corneal nests of cells arising from epithelium-mesenchyme transformation of lens epithelium.

(a,b) E16.5, P0 and P21 Adamts9^+/+ and Adamts9^del/+ eyes were stained with H&E (a) or an anti-collagen IV antibody (red) and the nuclei were counterstained with DAPI (white) (b). At E16.5, a disorganized lens epithelium (arrowhead) was associated with weaker collagen IV staining in Adamts9^del/+ eyes. At P0 and P21, aberrant nests of cells surrounded by collagen IV-positive ECM were observed only in the Adamts9^del/+ eyes (arrow). Higher magnification of these cells in P21 Adamts9^del/+ eyes is shown (inset). These images represent at least 3 eyes analyzed at each time point. L = lens, C = cornea. Scale bar = 25 μm. (c) P0 and P21 Adamts9^del/+ and Adamts9^+/+ eyes were stained using antibodies directed against α-smooth muscle actin (α-SMA) (red), E-cadherin (red) or N-cadherin (red) and nuclei were stained by DAPI (nucleus: white). At P0 and P21, the ectopic cells in Adamts9^del/+ corneas stained positive for α-SMA, E-cadherin and N-cadherin (arrows) whereas no α-SMA expression was observed in the lens or the cornea of Adamts9^+/+ mice, and E-cadherin was distinctly expressed by the lens and corneal epithelium. The corneal endothelium of Adamts9^+/+ mice strongly expressed N-cadherin, but also low levels of E-cadherin. C = cornea, E = corneal endothelium, L = lens. (d) Adamts9^del/+ and Adamts9^+/+ mice were bred with mT/mG; Wnt1-Cre mice for lineage tracing of neural crest-derived cells (green). In adult Adamts9^del/+; mT/mG; Wnt1-Cre eyes (lower panel), ectopic cells are red (arrow) and thus, unlike corneal keratocytes and corneal endothelium (green, see mT/mG; Wnt1-Cre cornea in upper panel), they do not arise from the neural crest. Scale bar = 25 μm.