Exosomal miR-100-5p inhibits osteogenesis of hBMSCs and angiogenesis of HUVECs by suppressing the BMPR2/Smad1/5/9 signalling pathway

Wu Yang,Yu Chen,Zijie Xu,Haobo Bai,Weiqian Jiang,Minkang Guo,Husun Qian,Yunfei Yang,Weiwen Zhu,Jian Zhang,Chengjie Lian,Tingmei Chen

doi:10.1186/s13287-021-02438-y

Abstract

BackgroundNontraumatic osteonecrosis of the femoral head (NONFH) is a common, progressive, and refractory orthopaedic disease. Decreased osteogenesis and angiogenesis are considered the main factors in the pathogenesis of NONFH. We aimed to figure out whether exosomes and exosomal miRNA from necrotic bone tissues of patients with NONFH are involved in the pathogenesis of NONFH and reveal the underlying mechanisms.MethodsRT-PCR and western blotting (WB) were used to detect the expression of osteogenic, adipogenic, and angiogenic markers. ALP staining and Alizarin Red S (ARS) staining were used to evaluate osteogenic differentiation of human bone marrow-derived mesenchymal stem cells (hBMSCs). Oil Red O staining was performed to assess the adipocyte deposition. A tube formation assay was used to study angiogenesis of human umbilical vascular endothelial cells (HUVECs). H&E staining and immunohistochemistry (IHC) staining were used to detect the effect of the NONFH exosomes in vivo. MicroRNA sequencing was conducted to identify potential regulators in the NONFH exosomes. The target relationship between miR-100-5p and BMPR2 was predicted and confirmed by a dual luciferase reporter assay and WB.ResultsThe NONFH exosomes reduced the osteogenic differentiation of hBMSCs and angiogenesis of HUVECs. In addition, the injection of the NONFH exosomes caused thinning and disruption of bone trabeculae in the femoral heads of rats. MiR-100-5p expression was upregulated in the NONFH exosomes and inhibited the osteogenesis of hBMSCs and angiogenesis of HUVECs by targeting BMPR2 and suppressing the BMPR2/SMAD1/5/9 signalling pathway. Silencing miR-100-5p expression rescued the reduction in osteogenesis and angiogenesis caused by the NONFH exosomes by activating the BMPR2/SMAD1/5/9 signalling pathway.ConclusionThe NONFH exosomal miR-100-5p can lead to NONFH-like damage by targeting BMPR2 and suppressing the BMPR2/SMAD1/5/9 signalling pathway, which may be involved in the pathophysiological mechanisms of nontraumatic osteonecrosis of the femoral head (NONFH).

Highlights

Nontraumatic osteonecrosis of the femoral head (NONFH) is a common, progressive, and refractory orthopaedic disease which usually results in substantial loss of function and inconvenience in the daily life of the patients [1,2,3]
nontraumatic osteonecrosis of the femoral head (NONFH) exosomes inhibited osteogenesis of human bone marrow-derived mesenchymal stem cells (hBMSCs) and angiogenesis of vessel endothelial cells (VECs), promoting adipogenesis of hBMSCs To investigate the effect of femoral neck fracture (FNF) exosomes and NONFH exosomes on the osteogenesis and adipogenesis of hBMSCs, we examined osteogenic and adipogenic markers, including OCN, OPN, Alkaline phosphatase (ALP), Runtrelated transcription factor 2 (RUNX2), collagen type 1, and Peroxisome proliferator-activated receptor γ (PPARγ), in the hBMSCs treated with Phosphate buffer saline (PBS), FNF exosomes, or NONFH exosomes for 7 days
MiR-100-5p inhibited osteogenesis of hBMSCs and angiogenesis of human umbilical vascular endothelial cells (HUVECs), promoting adipogenesis of hBMSCs As the results of miRNA sequencing and RT-qPCR showed that miR-100-5p expression was upregulated in the NONFH exosomes, we studied the effects of miR100-5p on hBMSCs and HUVECs

Summary

Introduction

Nontraumatic osteonecrosis of the femoral head (NONFH) is a common, progressive, and refractory orthopaedic disease which usually results in substantial loss of function and inconvenience in the daily life of the patients [1,2,3]. The main hypotheses about the pathogenesis of SANFH include the imbalance between osteogenesis and adipogenesis of BMSCs [5] and the impairment of vessel endothelial cells (VECs) [6]. BMSCs and VECs have strong proliferative potential, it is still difficult to reverse the disease progression of NONFH after pathogenic factors are removed. We wondered whether necrotic bone tissues release some signals to impair the self-repair of BMSCs and VECs. Nontraumatic osteonecrosis of the femoral head (NONFH) is a common, progressive, and refractory orthopaedic disease. We aimed to figure out whether exosomes and exosomal miRNA from necrotic bone tissues of patients with NONFH are involved in the pathogenesis of NONFH and reveal the underlying mechanisms

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Conclusion