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KLF2 regulates dental pulp-derived stem cell differentiation through the induction of mitophagy and altering mitochondrial metabolism.

ABSTRACT: To define the regulatory role of Kruppel-like factor 2 (KLF2) during osteoblast (OB) differentiation of dental pulp-derived stem cell (DPSC)s, herein, we show that the levels of KLF2 and autophagy-related molecules were significantly increased in differentiated cells. Gain-of-function and loss-of-function approaches of KLF2 confirmed that KLF2 modulated autophagic and OB differentiation-related molecules. In addition, knockdown of the autophagic molecule (ATG7 or BECN1) in DPSCs resulted in reduced levels of KLF2 and OB differentiation-related molecules. Conversely, the induction of autophagy increased levels of KLF2 and OB differentiation-related molecules. Moreover, OB differentiation induced mitophagy and mitochondrial membrane potential-related molecules. In addition, OB differentiation reduced the generation of total and mitochondrial ROS productions and induced intracellular Ca2+ production. Measurements of glycolysis and oxidative phosphorylation simultaneously in live cells revealed that OB differentiation decreased the oxygen consumption rate, which is an indicator of mitochondrial respiration and reduced the level of ATP production. Furthermore, flux analysis also revealed that OB differentiation increased the extracellular acidification rate (ECAR) in the non-glycolytic acidification, and the glycolytic capacity conditions, increasing the lactate production and reducing the metabolic activity of the cells. Thus, a metabolic shift from mitochondrial respiration to the glycolytic pathway was observed during OB differentiation. Finally, chromatin immunoprecipitation (ChIP) analysis confirmed that the KLF2 and active epigenetic marks (H3K27Ac and H3K4me3) were upregulated in the promoter region of ATG7 during OB differentiation. These results provide evidence that the mitophagy process is important during OB differentiation, and KLF2 critically regulates it.

SUBMITTER: Maity J

PROVIDER: S-EPMC7417940 | biostudies-literature | 2020 Jul

REPOSITORIES: biostudies-literature

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KLF2 regulates dental pulp-derived stem cell differentiation through the induction of mitophagy and altering mitochondrial metabolism.

Maity Jyotirindra J Deb Moonmoon M Greene Carl C Das Hiranmoy H

Redox biology 20200703

To define the regulatory role of Kruppel-like factor 2 (KLF2) during osteoblast (OB) differentiation of dental pulp-derived stem cell (DPSC)s, herein, we show that the levels of KLF2 and autophagy-related molecules were significantly increased in differentiated cells. Gain-of-function and loss-of-function approaches of KLF2 confirmed that KLF2 modulated autophagic and OB differentiation-related molecules. In addition, knockdown of the autophagic molecule (ATG7 or BECN1) in DPSCs resulted in redu ...[more]

PMID: 32777717

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Project description:The preservation of biological and physiological vitality as well as the formation of dentin are among the main tasks of human dental pulp for a life time. Odontoblastic differentiation of human dental pulp stem cells (hDPSCs) exhibits the capacity of dental pulp regeneration and dentin complex rebuilding. Exploration of the mechanisms regulating differentiation and proliferation of hDPSCs may help to investigate potential clinical applications. BTB and CNC homology 1 (BACH1) is a transcription repressor engaged in the regulation of multiple cellular functions. This study aimed to investigate the effects of BACH1 on the proliferation and odontoblastic differentiation of hDPSCs in vitro. hDPSCs and pulpal tissues were obtained from extracted human premolars or third molars. The distribution of BACH1 was detected by immunohistochemistry. The mRNA and protein expression of BACH1 were examined by qRT-PCR and Western blot analysis. BACH1 expression was regulated by stable lentivirus-mediated transfection. Cell proliferation and cell cycle were assessed by cell counting kit-8 assay, 5-Ethynyl-2'-deoxyuridine assay and flow cytometry. The expression of mineralization markers, alkaline phosphatase (ALP) activity and alizarin red S staining were conducted to assess the odontoblastic differentiation ability. BACH1 expression was stronger in the odontoblast layer than in the cell rich zone. The total and nuclear protein level of BACH1 during odontoblastic differentiation was downregulated initially and then upregulated gradually. Knockdown of BACH1 greatly inhibited cell proliferation, arrested cell cycle, upregulated the heme oxygenase-1 (HO-1) expression and attenuated ALP activity, decreased calcium deposits and downregulated the expression of mineralization markers. Treatment of Tin-protoporphyrin IX, an HO-1 inhibitor, failed to rescue the impaired odonto/osteogenic differentiation capacity. Overexpression of BACH1 increased cell proliferation, ALP activity and the expression of mineralization markers. Our findings suggest that BACH1 is an important regulator of the proliferation and odontoblastic differentiation of hDPSCs in vitro. Manipulation of BACH1 expression may provide an opportunity to promote the regenerative capacity of hDPSCs.

Dataset Information

KLF2 regulates dental pulp-derived stem cell differentiation through the induction of mitophagy and altering mitochondrial metabolism.

Publications

KLF2 regulates dental pulp-derived stem cell differentiation through the induction of mitophagy and altering mitochondrial metabolism.

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