Fig. 5

Histological and immunohistochemical evaluation of recellularized vascular grafts. Histological evaluation of the seeded, statically cultivated and immediately examined vascular graft (rTEVG-s) with H&E staining revealed multiple, morphologically round cells discontinuously attached to the basal membrane of the vascular graft (A). Further immunohistochemical staining with important endothelial cell markers such as CD31 (B), CD34 (C), eNOS (D) and vWF (E) revealed the majority of the cells were of endothelial phenotype. A few cells also stained positive for CD90, indicating a much lower number of hMSC dispersed within the hEPC (F). Histological evaluation of the dynamically cultivated vascular grafts (rTEVG-10d) displayed morphologically elongated cells due to introduction of gradually increasing medium flow and shear stress (G). Further characterization with CD31 (H), CD34 (I), eNOS (J) and vWF (K) revealed that while the cells stained strongly for eNOS, CD31 and vWF, staining with CD34, a marker of immature EPC was much weaker, a finding which was interpreted to point to cell maturation. Moreover, staining with CD90 antibody revealed scarce hMSC dispersed and attached to the basal membrane (L). Lastly, histological characterization of the vascular grafts perfused over 14 days (rTEVG-14d) revealed morphologically irregular cells (M), which stained slightly positive for CD31 (N) and vWF (O) and did not stain for CD34 (P), eNOS (Q) and CD90 (R), findings in line with the cell culture parameters, that pointed to excessive cell detachment and death after the 10th day of cultivation. Scale bar represents 50 μm