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Nogo-A Regulates the Fate of Human Dental Pulp Stem Cells toward Osteogenic, Adipogenic, and Neurogenic Differentiation


Lai, Chai Foong; Shen, Juliet; Balic, Anamaria; Pagella, Pierfrancesco; Schwab, Martin E; Mitsiadis, Thimios A (2022). Nogo-A Regulates the Fate of Human Dental Pulp Stem Cells toward Osteogenic, Adipogenic, and Neurogenic Differentiation. Cells, 11(21):3415.

Abstract

Human teeth are highly innervated organs that contain a variety of mesenchymal stem cell populations that could be used for cell-based regenerative therapies. Specific molecules are often used in these treatments to favorably modulate the function and fate of stem cells. Nogo-A, a key regulator of neuronal growth and differentiation, is already used in clinical tissue regeneration trials. While the functions of Nogo-A in neuronal tissues are extensively explored, its role in teeth still remains unknown. In this work, we first immunohistochemically analyzed the distribution of Nogo-A protein in the dental pulp of human teeth. Nogo-A is localized in a variety of cellular and structural components of the dental pulp, including odontoblasts, fibroblasts, neurons and vessels. We also cross-examined Nogo expression in the various pulp cell clusters in a single cell RNA sequencing dataset of human dental pulp, which showed high levels of expression in all cell clusters, including that of stem cells. We then assessed the role of Nogo-A on the fate of human dental pulp stem cells and their differentiation capacity in vitro. Using immunostaining, Alizarin Red S, Nile Red and Oil Red O staining we showed that Nogo-A delayed the differentiation of cultured dental pulp stem cells toward the osteogenic, adipogenic and neurogenic lineages, while addition of the blocking anti-Nogo-A antibody had opposite effects. These results were further confirmed by qRT-PCR, which demonstrated overexpression of genes involved in osteogenic (RUNX2, ALP, SP7/OSX), adipogenic (PPAR-γ2, LPL) and neurogenic (DCX, TUBB3, NEFL) differentiation in the presence of the anti-Nogo-A antibody. Conversely, the osteogenic and adipogenic genes were downregulated by Nogo-A. Taken together, our results show that the functions of Nogo-A are not restricted to neuronal cells but are extended to other cell populations, including dental pulp stem cells. We show that Nogo-A regulates their fates toward osteogenic, adipogenic and neurogenic differentiation, thus indicating its potential use in clinics.

Abstract

Human teeth are highly innervated organs that contain a variety of mesenchymal stem cell populations that could be used for cell-based regenerative therapies. Specific molecules are often used in these treatments to favorably modulate the function and fate of stem cells. Nogo-A, a key regulator of neuronal growth and differentiation, is already used in clinical tissue regeneration trials. While the functions of Nogo-A in neuronal tissues are extensively explored, its role in teeth still remains unknown. In this work, we first immunohistochemically analyzed the distribution of Nogo-A protein in the dental pulp of human teeth. Nogo-A is localized in a variety of cellular and structural components of the dental pulp, including odontoblasts, fibroblasts, neurons and vessels. We also cross-examined Nogo expression in the various pulp cell clusters in a single cell RNA sequencing dataset of human dental pulp, which showed high levels of expression in all cell clusters, including that of stem cells. We then assessed the role of Nogo-A on the fate of human dental pulp stem cells and their differentiation capacity in vitro. Using immunostaining, Alizarin Red S, Nile Red and Oil Red O staining we showed that Nogo-A delayed the differentiation of cultured dental pulp stem cells toward the osteogenic, adipogenic and neurogenic lineages, while addition of the blocking anti-Nogo-A antibody had opposite effects. These results were further confirmed by qRT-PCR, which demonstrated overexpression of genes involved in osteogenic (RUNX2, ALP, SP7/OSX), adipogenic (PPAR-γ2, LPL) and neurogenic (DCX, TUBB3, NEFL) differentiation in the presence of the anti-Nogo-A antibody. Conversely, the osteogenic and adipogenic genes were downregulated by Nogo-A. Taken together, our results show that the functions of Nogo-A are not restricted to neuronal cells but are extended to other cell populations, including dental pulp stem cells. We show that Nogo-A regulates their fates toward osteogenic, adipogenic and neurogenic differentiation, thus indicating its potential use in clinics.

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Item Type:Journal Article, refereed, original work
Communities & Collections:04 Faculty of Medicine > Center for Dental Medicine > Institute of Oral Biology
04 Faculty of Medicine > Institute for Regenerative Medicine (IREM)
Dewey Decimal Classification:610 Medicine & health
Scopus Subject Areas:Life Sciences > General Biochemistry, Genetics and Molecular Biology
Language:English
Date:28 October 2022
Deposited On:29 Dec 2022 11:53
Last Modified:28 Apr 2024 01:41
Publisher:MDPI Publishing
ISSN:2073-4409
OA Status:Gold
Free access at:PubMed ID. An embargo period may apply.
Publisher DOI:https://doi.org/10.3390/cells11213415
PubMed ID:36359811
  • Content: Published Version
  • Language: English
  • Licence: Creative Commons: Attribution 4.0 International (CC BY 4.0)