Background Redox signaling and energy metabolism are known to be involved in controlling the balance between self-renewal and proliferation/diferentiation of stem cells. In this study we investigated metabolic and redox changes occurring during in vitro human dental pulp stem cells (hDPSCs) osteoblastic (OB) diferentiation and tested on them the impact of the reactive oxygen species (ROS) signaling. Methods hDPSCs were isolated from dental pulp and subjected to alkaline phosphatase and alizarin red staining, q-RT-PCR, and western blotting analysis of diferentiation markers to assess achievement of osteogenic/odontogenic diferentiation. Moreover, a combination of metabolic fux analysis and confocal cyto-imaging was used to profle the metabolic phenotype and to evaluate the redox tone of hDPSCs. Results In diferentiating hDPSCs we observed the down-regulation of the mitochondrial respiratory chain complexes expression since the early phase of the process, confrmed by metabolic fux analysis, and a reduction of the basal intracellular peroxide level in its later phase. In addition, dampened glycolysis was observed, thereby indicating a lower energy-generating phenotype in diferentiating hDPSCs. Treatment with the ROS scavenger Trolox, applied in the early-middle phases of the process, markedly delayed OB diferentiation of hDPSCs assessed as ALP activity, Runx2 expression, mineralization capacity, expression of stemness and osteoblast marker genes (Nanog, Lin28, Dspp, Ocn) and activation of ERK1/2. In addition, the antioxidant partly prevented the inhibitory efect on cell metabolism observed following osteogenic induction.

Bioenergetic profile and redox tone modulate in vitro osteogenesis of human dental pulp stem cells: new perspectives for bone regeneration and repair

Consiglia Pacelli;Carmela Mazzoccoli;Giuseppe Capitanio;Giorgio Mori;Claudia Piccoli;Nazzareno Capitanio
2023-01-01

Abstract

Background Redox signaling and energy metabolism are known to be involved in controlling the balance between self-renewal and proliferation/diferentiation of stem cells. In this study we investigated metabolic and redox changes occurring during in vitro human dental pulp stem cells (hDPSCs) osteoblastic (OB) diferentiation and tested on them the impact of the reactive oxygen species (ROS) signaling. Methods hDPSCs were isolated from dental pulp and subjected to alkaline phosphatase and alizarin red staining, q-RT-PCR, and western blotting analysis of diferentiation markers to assess achievement of osteogenic/odontogenic diferentiation. Moreover, a combination of metabolic fux analysis and confocal cyto-imaging was used to profle the metabolic phenotype and to evaluate the redox tone of hDPSCs. Results In diferentiating hDPSCs we observed the down-regulation of the mitochondrial respiratory chain complexes expression since the early phase of the process, confrmed by metabolic fux analysis, and a reduction of the basal intracellular peroxide level in its later phase. In addition, dampened glycolysis was observed, thereby indicating a lower energy-generating phenotype in diferentiating hDPSCs. Treatment with the ROS scavenger Trolox, applied in the early-middle phases of the process, markedly delayed OB diferentiation of hDPSCs assessed as ALP activity, Runx2 expression, mineralization capacity, expression of stemness and osteoblast marker genes (Nanog, Lin28, Dspp, Ocn) and activation of ERK1/2. In addition, the antioxidant partly prevented the inhibitory efect on cell metabolism observed following osteogenic induction.
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11586/449100
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