Abstract
BackgroundFibrous dysplasia (FD) is a bone marrow stromal cell (BMSC) disease caused by activating mutations of guanine nucleotide-binding protein alpha-stimulating activity polypeptide (GNAS) and is characterized by increased proliferative activity and disrupted osteogenesis of BMSCs. However, the molecular mechanisms regulating the pathophysiologic features of BMSCs in FD remain unknown. This study aimed to identify and verify the roles of the CREB1-miR-181a-5p regulatory loop in FD pathophysiology.MethodsMicroRNA (miRNA) sequencing analysis was used to identify the possible miRNAs implicated in FD. The proliferation, apoptosis, and osteogenic differentiation of BMSCs, as well as the osteoclast-induced phenotype, were measured and compared after exogenous miR-181a-5p transfection into FD BMSCs or miR-181a-5p inhibitor transfection into normal BMSCs. Chromatin immunoprecipitation and luciferase reporter assays were performed to verify the interactions between CREB1 and miR-181a-5p and their effects on the FD pathological phenotype.ResultsCompared to normal BMSCs, FD BMSCs showed decreased miR-181a-5p levels and exhibited increased proliferative activity, decreased apoptotic capacity, and impaired osteogenesis. FD BMSCs also showed a stronger osteoclast activation effect. miR-181a-5p overexpression reversed the pathophysiologic features of FD BMSCs, whereas miR-181a-5p suppression induced an FD-like phenotype in normal BMSCs. Mechanistically, miR-181a-5p was the downstream target of CREB1, and CREB1 was posttranscriptionally regulated by miR-181a-5p.ConclusionsOur study identifies that the interaction loop between CREB1 and miR-181a-5p plays a crucial role in regulating the pathophysiologic features of FD BMSCs. MiR-181a-5p may be a potential therapeutic target for the treatment of FD.
Highlights
Fibrous dysplasia (FD) is a bone marrow stromal cell (BMSC) disease caused by activating mutations of guanine nucleotide-binding protein alpha-stimulating activity polypeptide (GNAS) and is characterized by increased proliferative activity and disrupted osteogenesis of bone marrow stromal cells (BMSCs)
Identification of downregulated miR‐181a‐5p in FD BMSCs Aiming to screen the possible miRNAs implicated in FD, we measured the expression of miRNAs by miRNA sequencing and found that miR-181a-5p was expressed at significantly lower levels in FD BMSCs than in normal BMSCs (Fig. 1a), whereas miR-145-3p and miR-98-3p were downregulated 9.1 folds and 9 folds in FD BMSCs, respectively
We performed Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis of the 459 target genes, and the results showed that miR-181a-5p might be involved in several pathways, including the cyclic adenosine monophosphate (cAMP) signaling pathway, apoptosis and osteoclast differentiation (See figure on page.) Fig. 1 MiR-181a-5p is expressed at low levels in FD BMSCs. a MicroRNA sequencing analysis showed the differentially expressed miRNAs between FD BMSCs and BMSCs. b Quantitative real-time PCR analysis was used to detect miR-181a-5p, miR-145-3p, miR-98-3p and miR-92b-5p expression between FD BMSCs and BMSCs. c cAMP levels in BMSCs treated with cAMP or IBMX were analyzed by ELISA
Summary
Fibrous dysplasia (FD) is a bone marrow stromal cell (BMSC) disease caused by activating mutations of guanine nucleotide-binding protein alpha-stimulating activity polypeptide (GNAS) and is characterized by increased proliferative activity and disrupted osteogenesis of BMSCs. the molecular mechanisms regulating the pathophysiologic features of BMSCs in FD remain unknown. Fibrous dysplasia (FD) is a nonhereditary bone disease caused by GNAS gene mutation in bone marrow stromal cells (BMSCs); in FD, normal bone tissue is replaced by. Overaccumulation of cyclic adenosine monophosphate (cAMP) in FD due to guanine nucleotide-binding protein alpha-stimulating activity polypeptide (GNAS) mutation is the key cause of FD pathogenesis (Zhang et al 2012a; Riminucci et al 2010). Increasing evidence indicates that miRNAs participate in multiple biological processes in many diseases, and modulation of miRNAs has been used for the therapy of different disorders
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