Fig. 5

Characterization and seeding of ROEC in decellularized scaffolds. a ROEC growing over irradiated feeder layer. Scale bar: 100 µm. b Colony forming efficiency of 100 and 300 ROEC stained with Rhodamine B. Scale bar: 1.5 cm. c Growth rate of representative ROEC as total number of cells at different days of culture. d–f Immunofluorescence of ROEC for the expression of CK14 and CK13 (d), Ki67 and E-cadherin (e) and CK14 and p63 (f). Nuclei were counterstained with DAPI. Scale bar: 66 µm. g Hematoxylin and eosin (H&E) staining, immunostaining for CK14 and Ki67 in sections of scaffolds seeded with ROEC and cultured in static conditions for 3 days. Scale bar: 100 µm. h Immunofluorescence staining for Ki67 and E-cadherin, CK14 and CK13, and ZO-1 after 14 days of culture. Nuclei were counterstained with DAPI. Scale bar: 50 µm. i Immunofluorescence staining of a native rat oesophagus for Ki67 and E-cadherin, CK14, CK13 and ZO-1. Nuclei were counterstained with DAPI. Scale bar: 50 µm. j Bioluminescence images of a representative scaffold seeded with Luc⁺ZsGreen⁺ROEC and cultured in dynamic conditions for 3 days. k Graph of the average radiance measured from the whole scaffold at different time points. l Hematoxylin and eosin staining in a section of scaffold seeded with ROEC and cultured in dynamic conditions for 3 days. Scale bar: 100 µm. m–p Immunostaining for p63 (m), E-cadherin (n), CK14 (o) and Ki67 (p) in sections of scaffolds seeded with ROEC and cultured in dynamic conditions for 3 days. Nuclei were counterstained with DAPI. Insets show staining specificity and localization. Scale bar: 100 µm