ZNF521 Represses Osteoblastic Differentiation in Human Adipose-Derived Stem Cells

Human adipose-derived stem cells (hADSCs) are multipotent mesenchymal cells that can differentiate into adipocytes, chondrocytes, and osteocytes. During osteoblastogenesis, the osteoprogenitor cells differentiate into mature osteoblasts and synthesize bone matrix components. Zinc finger protein 521 (ZNF521/Zfp521) is a transcription co-factor implicated in the regulation of hematopoietic, neural, and mesenchymal stem cells, where it has been shown to inhibit adipogenic differentiation. The present study is aimed at determining the effects of ZNF521 on the osteoblastic differentiation of hADSCs to clarify whether it can influence their osteogenic commitment. The enforced expression or silencing of ZNF521 in hADSCs was achieved by lentiviral vector transduction. Cells were cultured in a commercial osteogenic medium for up to 20 days. The ZNF521 enforced expression significantly reduced osteoblast development as assessed by the morphological and molecular criteria, resulting in reduced levels of collagen I, alkaline phosphatase, osterix, osteopontin, and calcium deposits. Conversely, ZNF521 silencing, in response to osteoblastic stimuli, induced a significant increase in early molecular markers of osteogenesis and, at later stages, a remarkable enhancement of matrix mineralization. Together with our previous findings, these results show that ZNF521 inhibits both adipocytic and osteoblastic maturation in hADSCs and suggest that its expression may contribute to maintaining the immature properties of hADSCs.

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Effect of CGRP-Adenoviral Vector Transduction on the Osteoblastic Differentiation of Rat Adipose-Derived Stem Cells

PLoS ONE ◽

10.1371/journal.pone.0072738 ◽

2013 ◽

Vol 8 (8) ◽

pp. e72738 ◽

Cited By ~ 15

Author(s):

Zhong Fang ◽

Qin Yang ◽

Wei Xiong ◽

Guang-hui Li ◽

Hui Liao ◽

...

Keyword(s):

Stem Cells ◽

Adenoviral Vector ◽

Osteoblastic Differentiation ◽

Adipose Derived Stem Cells ◽

Vector Transduction

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A NOTCH1/LSD1/BMP2 Co-Regulatory Network Mediated by miR-137 Negatively Regulates Osteogenesis of Human Adipose-Derived Stem Cells

10.21203/rs.3.rs-70186/v1 ◽

2020 ◽

Author(s):

Cong Fan ◽

Xiaohan Ma ◽

Yuejun Wang ◽

Longwei Lv ◽

Yuan Zhu ◽

...

Keyword(s):

Stem Cells ◽

Osteoblastic Differentiation ◽

Bone Morphogenetic Protein 2 ◽

Luciferase Reporter ◽

Adipose Derived Stem Cells ◽

Degree Relative ◽

Matrix Mineralization ◽

Differentiation Potential ◽

Lineage Differentiation ◽

Relative Expression Level

Abstract Background: MicroRNAs have been recognized as critical regulators for the osteoblastic lineage differentiation of human adipose-derived stem cells (hASCs). Previously, we have displayed that silencing of miR-137 enhances the osteoblastic differentiation potential of hASCs partly through the coordination of lysine-specific histone demethylase 1 (LSD1), bone morphogenetic protein 2 (BMP2), and mothers against decapentaplegic homolog 4 (SMAD4). However, still numerous molecules involved in the osteogenic regulation of miR-137 remain unknown. This study aimed to further elucidate the epigenetic mechanisms of miR-137 on the osteogenic differentiation of hASCs.Methods: hASCs transfected with miR-137 overexpression or knockdown lentiviruses were used to assess the osteogenic capacity by testing the alkaline phosphatase activity, matrix mineralization degree, relative expression level of osteogenesis-associated genes and ectopic osteogenesis in nude mice. Dual-luciferase reporter assay was performed to examine the targeting of the 3' untranslated region (3' UTR) of NOTCH1 by miR-137. Interrelationships of signaling pathways of NOTCH1-hairy and enhancer of split 1 (HES1), LSD1, and BMP2-SMAD4 were thoroughly investigated by separate knockdown of NOTCH1, LSD1, and BMP2.Results: We confirmed that miR-137 directly targeted the 3' UTR of NOTCH1 while positively regulated HES1. After knocking down NOTCH1 or BMP2 individually, we found that these two signals formed a positive feedback loop and activated LSD1. In addition, LSD1 knockdown induced the expression of NOTCH1 while suppressed HES1.Conclusions: Collectively, we proposed a NOTCH1/LSD1/BMP2 co-regulatory signaling network to elucidate the modulation of miR-137 on the osteoblastic differentiation of hASCs, thus providing mechanism-based rationale for miRNA-targeted therapy of bone defect.

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A NOTCH1/LSD1/BMP2 co-regulatory network mediated by miR-137 negatively regulates osteogenesis of human adipose-derived stem cells

Stem Cell Research & Therapy ◽

10.1186/s13287-021-02495-3 ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

Cong Fan ◽

Xiaohan Ma ◽

Yuejun Wang ◽

Longwei Lv ◽

Yuan Zhu ◽

...

Keyword(s):

Stem Cells ◽

Osteoblastic Differentiation ◽

Negative Control ◽

Bone Morphogenetic Protein 2 ◽

Luciferase Reporter ◽

Adipose Derived Stem Cells ◽

Differentiation Potential ◽

Lineage Differentiation ◽

Morphogenetic Protein ◽

Genes Expression

Abstract Background MicroRNAs have been recognized as critical regulators for the osteoblastic lineage differentiation of human adipose-derived stem cells (hASCs). Previously, we have displayed that silencing of miR-137 enhances the osteoblastic differentiation potential of hASCs partly through the coordination of lysine-specific histone demethylase 1 (LSD1), bone morphogenetic protein 2 (BMP2), and mothers against decapentaplegic homolog 4 (SMAD4). However, still numerous molecules involved in the osteogenic regulation of miR-137 remain unknown. This study aimed to further elucidate the epigenetic mechanisms of miR-137 on the osteogenic differentiation of hASCs. Methods Dual-luciferase reporter assay was performed to validate the binding to the 3′ untranslated region (3′ UTR) of NOTCH1 by miR-137. To further identify the role of NOTCH1 in miR-137-modulated osteogenesis, tangeretin (an inhibitor of NOTCH1) was applied to treat hASCs which were transfected with miR-137 knockdown lentiviruses, then together with negative control (NC), miR-137 overexpression and miR-137 knockdown groups, the osteogenic capacity and possible downstream signals were examined. Interrelationships between signaling pathways of NOTCH1-hairy and enhancer of split 1 (HES1), LSD1 and BMP2-SMADs were thoroughly investigated with separate knockdown of NOTCH1, LSD1, BMP2, and HES1. Results We confirmed that miR-137 directly targeted the 3′ UTR of NOTCH1 while positively regulated HES1. Tangeretin reversed the effects of miR-137 knockdown on osteogenic promotion and downstream genes expression. After knocking down NOTCH1 or BMP2 individually, we found that these two signals formed a positive feedback loop as well as activated LSD1 and HES1. In addition, LSD1 knockdown induced NOTCH1 expression while suppressed HES1. Conclusions Collectively, we proposed a NOTCH1/LSD1/BMP2 co-regulatory signaling network to elucidate the modulation of miR-137 on the osteoblastic differentiation of hASCs, thus providing mechanism-based rationale for miRNA-targeted therapy of bone defect.

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