Revascularization of human dental pulp using tissue engineering approaches

Advancement in stem cell technology allows for many therapeutic opportunities, including the treatment of previously intractable conditions. Revascularization of dental pulp tissue, and specifically angiogenic differentiation of Human Dental Pulp Stem Cells, is of great interest due to the crucial role of this process not only in dental pulp regeneration, but also in wound healing and in regenerative medicine in general. One of the challenges of tissue engineering is the ability to provide sufficient blood supply for engineered tissue and organs in the first phase after transplantation. The present study investigated the potential use of HDPSCs in revascularization of dental pulp in vitro as well as in vivo using Matrigel basement membrane as 3D scaffold and compared this data to that of stem cells isolated from dental pulp tissue using the stem cell marker Stro-1. Initially these cells were cultured under angiogenic condition (EGM-2) for cell differentiation and treated with VEGF. The angiogenic potential of human dental pulp stromal/stem cells was investigated at gene and protein level by qRT-PCR and immunohistochemical analysis of appropriate angiogenic markers. Moreover we monitored the differentiation of these cells by confocal and light microscopy. In the second part of the present study we investigated the ability of these cells to differentiate and form vascular tissue in an appropriate animal model. Two in vivo models were used; HDPSCs or Stro-1 +CD45- cells were suspended in Matrigel and injected into the root canal space of human tooth sections and implanted subcutaneously into immunocompromised mice for 3 weeks. The other model was the Matrigel plug assay, which is widely used for angiogenesis studies. qRT-PCR and Immunohistochemistry studies indicated that CD31 and VEGFR-2 were upregulated in HDPSCs and stro-t- CD45- cells in monolayer cultures, and all angiogenic markers (CD31, CD34, vWF, and VEGFR-2) in Matrigel cultures were upregulated as well following treatment with VEGF in endothelial cells growth medium-2. In 3D Matrigel culture, cells were also able to form tube like network structures. These results were confirmed by in vivo study, in which we were able to regenerate vascular like tissue which contained red blood cells in both in vivo models. This data indicated that these vessels are functional when compared to normal vascular tissue in both human and mice. In conclusion, the present study confirmed that HDPSCs and Stro-1 +CD45- cells were induced to express angiogenic markers in vitro and can be recruited in the formation of vascular tissue in a tooth section as well as Matrigel plug constructs in immunocompromised mice. This technique can be used in the future to revascularize dental pulp which will enhance the survival rate of traumatized teeth.EThOS - Electronic Theses Online ServiceGBUnited Kingdo