Question: 

We investigated the angiogenic potency of bone marrow stem cells (BMSCs) which were differentiated into endothelial cells by the application of growth factors into culture media.

Methods: 

We isolated BMSC by bone marrow aspiration of the mini pig femur. Bone marrow was filtered with a 100mm cell strainer to eliminate bone remains and tissue fragments. After centrifugation pellet was dissolved in PBS-EDTA solution and the mononuclear cells were isolated bydensitygradientcentrifugationwith histopaque. Thereafter cells were washed with media and centrifuged again for 5min. The pellet was resolved in medium containing endothelial cell growth factor supplement (10%). Medium was changed every second day. To characterize confluent cells, immunofluorescence staining was performed for the expression of endothelial nitric oxide synthase (eNOS) and CD 31 as double staining and for von Willebrandt factor (vWF) staining. Furthermore, the ability of cells to release NO was examined byspectrophotometricinvestigation. To investigate whether endothelial like cells could form vascular structure in a 3D culture, angiogenesis assays were performed.

Results: 

Isolated BMSC cultured with endothelial cell growth factor exhibited a typical cobblestone-like endothelial cell phenotype. Investigations by immunofluorescence staining showed that differentiated BMSCs expressed eNOS in the same manner like venous endothelial cells. They were also positive for the typical endothelial cell marker vWF. We showed that endothelial-like BMSCs were able to liberate NO and investigations using angiogenesis assay indicated formation of vascular structure after 3D cultivation in matrigel. These are essential features for the functionality of endothelial cells.

Conclusion: 

The reprogramming of isolated pig BMSCs into endothelial-like cells with endothelial characteristics such as angiogenic capability and NO release could be achieved by addition of endothelial cell growth factor. In future studies BMSCs could be a source for production of endothelial-like cells for application in tissue engineering.