Caracterização da diferenciação osteogênica in vitro a partir das células-tronco da polpa dentária de paciente com Neurofibromatose tipo 1

Abstract

Neurofibromatosis type 1 (NF1) is a genetic disorder characterized by hyperpigmentation of the skin (Café-au-lait spots) and neurofibromas and also has other clinical manifestations, among them the bone abnormalities, affecting up to 50% of patients. Animal models such as rodents and even Drosophila have been important in elucidating the molecular and cellular mechanisms resulting from NF1, but these models have limitations. Homozygous transgenic mice (NF1-/-) die in womb in 12º/14º days of gestation, whereas heterozygous mice (NF1 +/-), although viable and do not develop neither neurofibromas nor Café-au-lait spots in vivo. Thus, there is a need to develop new models. Stem cells are undifferentiated cells capable of self renew and differentiate into a specific cell lineage. They can be classified according to their origin into two main types: embryonic (ESCs) or adult (ASCs). The use of ASCs in research aiming at cell therapy and tissue bioengineer is important and provides advantages compared to ESCs, since their differentiation is more controlled and, when introduced into the body, is unlikely to produce tumors. Among the adult stem cells, we highlight the dental pulp stem cells (DPSCs). The DPSCs show a molecular profile similar to human embryonic cells, have the capacity to differentiate spontaneously under appropriate culture conditions, and have postnatal origin, being used to in vitro study of various diseases. We propose to use the DPSCs also as a model for in vitro NF1 study. The aim of this study is to compare the stem cells from the pulp of deciduous teeth of a patient with Neurofibromatosis Type 1, with dental pulp stem cells of healthy people, specifically evaluating the rate of proliferation and senescence of these cells, as well as osteogenic differentiation followed by calcium quantification. The analysis of cell proliferation suggests that the patient's DPSCs have a high proliferative rate relative to controls. The analysis of cellular senescence suggests that these cells enter in a delayed senescence compared to control cells. A pooled analysis of cellular proliferation and senescence shows that NF1 patient’s DPSCs already presents tumor profile. This can help us understand the formation of neurofibromas throughout the body extension. In order to answer whether the DPSCs are an excellent model for in vitro study, once the animal model has limitations, osteogenic differentiation and NF1 differentiated cells calcium quantification was performed. The results suggest low NF1 cells calcium production compared to control cells. Statistical analysis revealed significant differences between the samples. Our conclusion shows that DPSCs are an excellent model for in vitro study of several human diseases due to the potential that these cells present.