Fig. 6. Combined activities of Hhip and Ptch during pancreas development and endocrine cell development. Hematoxylin/Eosin staining (A-D) shows a stepwise decline in pancreatic epithelial size and branching associated with the loss of Hhip (Hip1 in figure) and Ptch (Ptch1 in figure) alleles (throughout the figure dorsal is towards the top and anterior towards the left). Morphometric analysis was used to outline and quantify pancreatic epithelium (A-D,K, blue Hhip^+/+;Ptch^+/+, n=5; green, Hhip^+/+;Ptch^+/–, n=3; red, Hhip^–/–;Ptch^+/+, n=3; yellow, Hhip^–/–;Ptch^+/–; n=4; #no significant difference, *P<0.05, **P<0.01). Staining for glucagon-(red; E-G) and Isl1-(brown; H-J) expressing cells within the pancreatic epithelium and quantification of both cell types (L, green, Hhip^+/+;Ptch^+/– n=3; red, Hhip^–/–;Ptch^+/+ n=3; yellow, Hhip^–/–;Ptch^+/– n=3; #no significant difference, *P<0.05; Glucagon, Mann-Whitney test; Isl1, Student's t-test) reveals additive requirements of Hhip and Ptch for endocrine cell differentiation. Arrowheads indicate Isl1-positive cells within pancreatic epithelium. Yellow cell in E correspond to autofluorescing erythrocytes.