Objectives: To evaluate the effect of SHED-CM on the proliferation, differentiation, migration ability, cell death, gene expression and production of VEGF of HUVEC and in a rodent orthotopic dental pulp regeneration.
Methods: Three culture media [M199, DMEM/Ham's F12 and DMEM/Ham's F12 conditioned by SHEDs] were used as experimental groups. SHED-CM was prepared maintaining confluent cells in culture without serum for 3 days. The proliferation and cell death marker of HUVECs were assessed using flow cytometry. The capacity of formation of vascular-like structures was analyzed in cells grown over Matrigel in hypoxic condition. HUVECs migration was followed using the scratch test. VEGF-A expression in HUVECs was assessed using real time RT-qPCR; and VEGF synthesis with ELISA test. SHED-CM was also applied in rodent ortotopic model of dental pulp regeneration in rats. The formed tissue was submitted to histological and immunohistochemical analyses.
Results: SHED-CM promoted significantly lower expression of 7AAD in HUVECs; whereas the expression of the Ki67 was similar in all groups. The vascular-like structures were observed in all groups. Migration of SHED-CM group was faster than DMEM/Ham's F12. SHED-CM induced similar expression of VEGF-A than M199, and higher than DMEM/Ham's F12. SHED-CM induced significantly higher VEGF synthesis than other media. SHED-CM induced formation of a vascularized connective tissue inside the root canal.
Conclusion: The study showed that SHEDs release angiogenic and cytoprotective factors, which are of great importance for tissue engineering.
Clinical Significance: SHED-CM could be an option to the use of stem cells in tissue engineering.
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| http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6488540 | PMC |
| http://dx.doi.org/10.1016/j.heliyon.2019.e01560 | DOI Listing |
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