MiR-27b Impairs Adipocyte Differentiation of Human Adipose Tissue-Derived Mesenchymal Stem Cells by Targeting LPL

Abstract

Background/Aims: In this study, the molecular mechanisms of miR-27b and lipoprotein lipase (LPL) that regulate human adipose-derived mesenchymal stem cells (hASCs) adipogenic differentiation were detected. Methods: Microarray analysis was applied to screen for differentially expressed miRNAs and mRNA during hASCs adipocyte differentiation induction. MiR-27b and LPL were found to have abnormal expression. Then, a dual luciferase reporter assay was employed to validate the targeting relationship between miR-27b and LPL. We also utilized qRT-PCR, western blot, cellular immunofluorescence and an oil red O staining assay to analyze the regulation of miR-27b and LPL during adipogenic differentiation. Results: The microarray analysis demonstrated that, during adipogenic differentiation, miR-27b was down-regulated, while LPL was up-regulated but tended to become stable 14 days after induction. A dual luciferase reporter assay confirmed the negative targeting regulatory relationship between miR-27b and LPL. After overexpressing and silencing miR-27b, LPL was found to be reversely regulated by miR-27b according to qRT-PCR and western blot. The fat-formation-related biomarkers CCAAT-enhancer binding protein α (c/EBPα) and peroxisome proliferator-activated receptors γ (PPARγ) had decreasing levels after over-expressing miR-27b or knockdown of LPL followed by adipogenic differentiation. Meanwhile, the oil red O staining assay revealed that the accumulation of lipid droplets decreased. There was no change in the expression of c/EBPα, PPARγ, or lipid droplet accumulation when overexpressing miR-27b and LPL. Conclusion: During the adipogenic differentiation of hASCs, miR-27b expression decreased, and LPL expression increased. The abnormal expression of miR-27b and LPL effectively regulated the adipogenic differentiation of hASCs.

Adipogenesis, Osteogenesis, and Chondrogenesis of Human Mesenchymal Stem/Stromal Cells: A Comparative Transcriptome Approach

“…One of the miR-30 target is the RUNX2 (osteogenesis transcription factor) which could be, at least in part, responsible for miR-30 positive effects on adipocyte differentiation ( Zaragosi et al, 2011 ). Another study using microarray analysis found 32 miRNAs differentially expressed during adipogenesis, among them is miR27b which was downregulated, while lipoprotein lipase (LPL) mRNA was up-regulated ( Hu et al, 2018 ). miR-27b targeted LPL and inhibits adipogenic differentiation ( Hu et al, 2018 ).…”

“…For differentiation induction, several protocols have been established to promote adipogenesis, chondrogenesis and osteogenesis which includes the usage of different culture media ( Table 1 ), passage number and confluency. In the manuscripts analyzed in this review, the passage at which differentiation was induced ranged from 2-3 ( Kulterer et al, 2007 ; Gu et al, 2017 ; Huang et al, 2017 ; Hu et al, 2018 ; Shaik et al, 2019 ) to 15 ( Ambele et al, 2016 ). Regarding confluency, some groups reported that the differentiation was induced with subconfluent cells (up to 90% confluence) ( Hu et al, 2018 ; Yi et al, 2019 ), others used confluent cultures ( Zaragosi et al, 2011 ; Luan et al, 2015 ; Xu et al, 2016 ; Min et al, 2019 ), or even a few days after the cells reached confluence ( Scheideler et al, 2008 ).…”

“…In the manuscripts analyzed in this review, the passage at which differentiation was induced ranged from 2-3 ( Kulterer et al, 2007 ; Gu et al, 2017 ; Huang et al, 2017 ; Hu et al, 2018 ; Shaik et al, 2019 ) to 15 ( Ambele et al, 2016 ). Regarding confluency, some groups reported that the differentiation was induced with subconfluent cells (up to 90% confluence) ( Hu et al, 2018 ; Yi et al, 2019 ), others used confluent cultures ( Zaragosi et al, 2011 ; Luan et al, 2015 ; Xu et al, 2016 ; Min et al, 2019 ), or even a few days after the cells reached confluence ( Scheideler et al, 2008 ). Confluency and passaging may affect the differentiation potential of MSC ( Wall et al, 2007 ; Safwani et al, 2014 ; Abo-Aziza and Zaki, 2017 ; Noda et al, 2019 ), although some groups did not find this correlation ( Kulterer et al, 2007 ).…”

Effects of PSMA1 on the differentiation and lipid deposition of bovine preadipocytes

“…LPL and C/EBPα have important roles in preadipocyte differentiation and cellular lipid accumulation. The expression of these two genes begins with lipogenesis and induces lipoprotein storage and lipid metabolism (Bauer et al, 2015;Rozovski et al, 2016;Glatz and Luiken, 2017;Hu et al, 2018). Consistent with this, the expression of PSMA1 is potentially associated with LXRα, which is the upstream regulator of LPL and C/EBPα (Zhang et al, 2001;Steffensen et al, 2002;Kim et al, 2007b;Han et al, 2009).…”

Up-Regulated MicroRNA-27b Promotes Adipocyte Differentiation via Induction of Acyl-CoA Thioesterase 2 Expression

“…Several studies have indicated that miR-27b plays an important role in lipid metabolism and adipogenesis [24,25]. It has been reported that miR-27b is down-regulated in in vitro cultured adipocytes and adipose tissue in obesity-related models and that miR-27b impairs adipocyte differentiation in 3T3-L1 cells and human adipose-derived mesenchymal stem cells via its associations with PPARγ and C/EBPα [24][25][26][27]. In zebrafish, depletion of miR-27b has shown increases in PPARγ, C/EBPα, and SREBP-1c, resulting in the enhancement of adipocyte hyperplasia [28].…”