Energy Metabolism and Lipidome Are Highly Regulated during Osteogenic Differentiation of Dental Follicle Cells

Abstract

Dental follicle cells (DFCs) are stem/progenitor cells of the periodontium and give rise to alveolar osteoblasts. However, understanding of the molecular mechanisms of osteogenic differentiation, which is required for cell-based therapies, is delimited. This study is aimed at analyzing the energy metabolism during the osteogenic differentiation of DFCs. Human DFCs were cultured, and osteogenic differentiation was induced by either dexamethasone or bone morphogenetic protein 2 (BMP2). Previous microarray data were reanalyzed to examine pathways that are regulated after osteogenic induction. Expression and activity of metabolic markers were evaluated by western blot analysis and specific assays, relative amount of mitochondrial DNA was measured by real-time quantitative polymerase chain reaction, the oxidative state of cells was determined by a glutathione assay, and the lipidome of cells was analyzed via mass spectrometry (MS). Moreover, osteogenic markers were analyzed after the inhibition of fatty acid synthesis by 5-(tetradecyloxy)-2-furoic acid or C75. Pathway enrichment analysis of microarray data revealed that carbon metabolism was amongst the top regulated pathways after osteogenic induction in DFCs. Further analysis showed that enzymes involved in glycolysis, citric acid cycle, mitochondrial activity, and lipid metabolism are differentially expressed during differentiation, with most markers upregulated and more markedly after induction with dexamethasone compared to BMP2. Moreover, the cellular state was more oxidized, and mitochondrial DNA was distinctly upregulated during the second half of differentiation. Besides, MS of the lipidome revealed higher lipid concentrations after osteogenic induction, with a preference for species with lower numbers of C-atoms and double bonds, which indicates a de novo synthesis of lipids. Concordantly, inhibition of fatty acid synthesis impeded the osteogenic differentiation of DFCs. This study demonstrates that energy metabolism is highly regulated during osteogenic differentiation of DFCs including changes in the lipidome suggesting enhanced de novo synthesis of lipids, which are required for the differentiation process.

Analysis of the phosphoproteome in human dental follicle cells during osteogenic differentiation

“…In conclusion, our study provides new information on the mechanisms of the later phase of osteogenic differentiation of DFCs. In connection with the results of our previous study [17], during this differentiation phase metabolic processes are induced in DFCs that increase oxidative stress that need to be regulated. It remains unclear to what extent the signaling pathways associated with the Rho-GTPases are involved in the differentiation and regulation of oxidative stress.…”

“…Recent studies found that lipid synthesis (for example, of cholesterol) and other metabolic processes are also involved in the later stages of osteogenic differentiation (14 days after induction) [16, 17]. Moreover, inhibition of fatty acid synthesis decreased the osteogenic differentiation potential of DFCs [17]. The present study also shows that oxidative stress is induced during osteogenic differentiation.…”

“…Interestingly, it is also known that during osteogenic differentiation, when the signaling pathways mentioned here are regulated in the DFCs, metabolic processes in the cell also change. Recent studies found that lipid synthesis (for example, of cholesterol) and other metabolic processes are also involved in the later stages of osteogenic differentiation (14 days after induction) [16, 17]. Moreover, inhibition of fatty acid synthesis decreased the osteogenic differentiation potential of DFCs [17].…”

“…Other molecular processes must be involved in the induction/regulation of mineralization and metabolic processes, both of which are induced 14 days after osteogenic differentiation [17]. In the present study, we therefore examined phosphorylated proteins in the proteome (phosphoproteome) of DFCs 14 days after induction of osteogenic differentiation and identified regulated proteins belonging to signaling pathways associated with the Rho‐GTPase cycle and apoptosis.…”

Matricellular Protein SMOC2 Potentiates BMP9-Induced Osteogenic Differentiation in Mesenchymal Stem Cells through the Enhancement of FAK/PI3K/AKT Signaling

“…Recently, induced pluripotent stem cells-derived MSC (iPSC-MSCs) has been proposed as an alternative resource. iPSCs generated with this pluripotent have numerous applications, including expansion to large scale cell numbers for tissue engineering and the development of cellular therapeutics [54][55][56]. Here, although MEFs cells derived from embryonic tissues show pluripotency, the exact mechanism remains to be investigated.…”