LncRNA HOTAIR inhibited osteogenic differentiation of BMSCs by regulating Wnt/β-catenin pathway

J.-J. Shen, C.-H. Zhang, Z.-W. Chen, Z.-X. Wang, D.-C. Yang, F.-L. Zhang, K.-H. Feng

Two Department of Bone Trauma, Gansu Provincial Hospital of Traditional Chinese Medicine, Lanzhou, China. shjjun2008@163.com

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• Stem Cells

OBJECTIVE: This study aims to investigate whether HOX transcript antisense RNA (HOTAIR) can participate in the osteogenic differentiation of bone marrow mesenchymal stem cells (BMSCs) by regulating the Wnt/β-catenin pathway, thereby participating in the pathogenesis of osteoporosis.

PATIENTS AND METHODS: We detected the expression level of HOTAIR in 60 osteoporosis patients and 60 normal controls by Quantitative Real-Time Polymerase Chain Reaction (qRT-PCR). Meanwhile, BMSCs derived from human or rats were subjected to determination of HOTAIR level. Subsequently, the effects of HOTAIR on osteogenic differentiation were evaluated by the activity of Alkaline Phosphatase (ALP), Alizarin Red S (ARS) staining, ALP staining and osteogenic-specific gene expression. The expression level of proteins related to the Wnt/β-catenin was determined by Western blot, and ALP activity was detected by ALP activity determination kit and alizarin red staining after knockdown or overexpression of HOTAIR, as well as the treatment of DKK1 or the Wnt pathway antagonist. Finally, osteoporosis model in rats was established by ovariectomy (OVX). We examined protein levels of HOTAIR, β-catenin, CyclinD, C-myc, and Runx2 in rat bone tissues. Bone morphology was observed in each group as well.

RESULTS: The serum and BMSCs levels of HOTAIR in patients with osteoporosis were remarkably higher than that in normal people. Inhibition of HOTAIR induced increased ALP activity increased osteogenic marker genes and enhanced number of calcified nodules in BMSCs. However, the overexpression of HOTAIR exhibited the opposite effects. HOTAIR inhibited the expression level of Wnt/β-catenin pathway-related protein. Also, Wnt pathway antagonist DKK1 partially reversed the regulatory effects of HOTAIR on Wnt/β-catenin. DKK1 treatment markedly reduced the promotive effect of HOTAIR knockdown on ALP activity, ALP content and calcification ability of BMSCs. DKK1 administration in rats undergoing OVX showed worse bone morphology relative to controls. Protein levels of HOTAIR, β-catenin, CyclinD, C-myc and Runx2 remarkably downregulated in OVX rats administrated with DKK1.

CONCLUSIONS: HOTAIR inhibits osteoblast differentiation of rat BMSCs. The underlying mechanism of which may be related to the mediation of Wnt/β-catenin pathway.

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