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MiR-27a targets sFRP1 in hFOB cells to regulate proliferation, apoptosis and differentiation.

ABSTRACT: MicroRNAs (miRNAs) play a key role in the regulation of almost all the physiological and pathological processes, including bone metabolism. Recent studies have suggested that miR-27 might play a key role in osteoblast differentiation and bone formation. Increasing evidence indicates that the canonical Wnt signaling pathway contributes to different stages of bone formation. In this study, we identify miR-27a can promote osteoblast differentiation by repressing a new target, secreted frizzled-related proteins 1 (sFRP1) expression at the transcriptional level. Here, 21 candidate targets of miR-27a involved in canonical Wnt/?-catenin signaling were predicted, and a significant decrease in sFRP1 luciferase activity was observed both in 293T and MG63 cells co-transfected with the matched luciferase reporter constructs and miR-27a mimic. Furthermore, the presence of exogenous miR-27a significantly decreased sFRP1 mRNA and protein expression in hFOB1.19 cells during both proliferation and osteogenic differentiation. The over-expression of miR-27a or knockdown sFRP1 significantly increased the percentage of apoptotic hFOBs, the percentage of cells in the G2-M phase of the cell cycle and the expression of key osteoblastic markers, including ALP, SPP1, RUNX2 and ALP activity. Over-expression of miR-27a or knockdown of endogenous sFRP1 led to an accumulation of ?-catenin in hFOBs. In the present study, we demonstrate that miR-27a induced gene silencing effect is a vital mechanism contributing to bone metabolism in hFOB cells in vitro, which is partly affected by the post-transcriptional regulation of sFRP1, during osteoblast proliferation, apoptosis and differentiation.

SUBMITTER: Guo D

PROVIDER: S-EPMC3953332 | biostudies-literature | 2014

REPOSITORIES: biostudies-literature

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MiR-27a targets sFRP1 in hFOB cells to regulate proliferation, apoptosis and differentiation.

Guo Donggeng D Li Qiuxia Q Lv Qing Q Wei Qiujing Q Cao Shuangyan S Gu Jieruo J

PloS one 20140313 3

MicroRNAs (miRNAs) play a key role in the regulation of almost all the physiological and pathological processes, including bone metabolism. Recent studies have suggested that miR-27 might play a key role in osteoblast differentiation and bone formation. Increasing evidence indicates that the canonical Wnt signaling pathway contributes to different stages of bone formation. In this study, we identify miR-27a can promote osteoblast differentiation by repressing a new target, secreted frizzled-rela ...[more]

PMID: 24625818

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Project description:BackgroundThe expression of miR-140-5p increased in the brain tissue of a bilateral common carotid artery ligation model, while the overexpression of miR-140-5p significantly decreased the number of neurons. The luciferase report experiment in the previous study proved that miR-140-5p negatively regulated one of the potential targets of Prospero-related homeobox 1 (Prox1). Therefore, we want to investigate the effect of miR-140-5p on the proliferation and differentiation of neural stem cells (NSCs) and the underlying mechanism.MethodsPrimary NSCs were extracted from pregnant ICR mice aged 16-18 days and induced to differentiate. After transient transfection with miR-140-5p mimic and inhibitor into NSCs, the cells were divided into five groups: blank, mimic normal control, mimic, inhibitor normal control, and inhibitor. Cell Counting Kit-8 (CCK-8) and 5-Bromo-2-deoxyUridine (BrDU), Ki-67 were used, and the diameter of neural spheres was measured to observe proliferation ability 48 h later. Doublecortin (DCX), glial fibrillary acidic protein (GFAP), microtubule-associated proteins 2 (MAP-2), synapsin I (SYN1), and postsynaptic density protein-95 (PSD-95) were stained to identify the effect of miR-140-5p on the differentiation ability of NSCs into neural precursor cells, astrocytes, and neurons and the expression of synapse-associated proteins. The expression of miR-140-5p, Prox1, p-ERK1/2, and ERK1/2 was analyzed by real time quantitative polymerase chain reaction (RT-qPCR) and Western blot analysis.ResultsWhile the expression of miR-140-5p decreased after NSC differentiation (P<0.05), the results of CCK-8, BrDU, and Ki-67 staining showed no significant difference in cell viability and the percentage of NSCs with proliferation ability (P>0.05). However, the neural spheres were shorter in the miR-140-5p overexpression group (P<0.05) and the expression of DCX, MAP2, synapsin I, and PSD-95 decreased, while the expression of GFAP increased after differentiation in the mimic group (P<0.05). In addition, the expression of Prox1 decreased and the expression of p-ERK1/2 protein increased (P<0.05), but the expression of ERK1/2 showed no significant difference (P>0.05) in the miR-140-5p overexpression group.ConclusionsMiR-140-5p reduced the proliferation rate of NSCs, inhibited their differentiation into neurons, produced synapse-associated proteins, and promoted their differentiation into astrocytes. MiR-140-5p negatively regulated downstream target Prox1 and activated the ERK/MAPK signaling pathway.

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MiR-27a targets sFRP1 in hFOB cells to regulate proliferation, apoptosis and differentiation.

Publications

MiR-27a targets sFRP1 in hFOB cells to regulate proliferation, apoptosis and differentiation.

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