Transplantation of Human Neonatal Foreskin Stromal Cells in Ex Vivo Organotypic Cultures of Embryonic Chick Femurs

We have previously reported that human neonatal foreskin stromal cells (hNSSCs) promote angiogenesis in vitro and in chick embryo chorioallantoic membrane (CAM) assay in vivo. To examine the in vivo relevance of this observation, we examined in the present study the differentiation potential of hNSSCs in ex vivo organotypic cultures of embryonic chick femurs. Isolated embryonic chick femurs (E10 and E11) were cultured for 10 days together with micro-mass cell pellets of hNSSCs, human umbilical vein endothelial cells (HUVEC) or a combination of the two cell types. Changes in femurs gross morphology and integration of the cells within the femurs were investigated using standard histology and immunohistochemistry. After 10 days, the femurs that were cultured in the presence of hNSSCs alone or hNSSC + HUVEC cells grew longer, exhibited thicker diaphysis and an enlarged epiphyseal region compared to control femurs cultured in the absence of cells. Analysis of cell-femur interactions, revealed intense staining for CD31 and enhanced deposition of collagen rich matrix along the periosteum in femurs cultured with hNSSCs alone or hNSSCs + HUVEC and the most pronounced effects were observed in hNSSC + HUVEC cultures. Our data suggest that organotypic cultures can be employed to test the differentiation potential of stem cells and demonstrate the importance of stem cell interaction with 3D-intact tissue microenvironment for their differentiation.