目标:由骨髓中的常见祖细胞产生,在骨髓间充质干细胞(BMSC)发育过程中,脂肪生成和成骨密切相关,但相互排斥。先前的研究表明,形态变化可以通过调节RhoA活性影响多能BMSCs对脂肪和成骨细胞谱系的早期承诺。RhoA途径调节肌动蛋白聚合以促进球状肌动蛋白(G-肌动蛋白)掺入丝状肌动蛋白(F-肌动蛋白)。在这样做的时候,与肌钙蛋白相关的转录因子(MRTF)与结合的G-肌动蛋白解离并进入细胞核以共同激活血清反应因子(SRF)靶基因表达。在这项研究中,我们研究了MRTFA/SRF是否在RhoA通路的下游起作用,从而调节小鼠BMSC的定型.
方法:使用Micro-CT在MRTFAKO小鼠中研究了敲除MRTFA对骨骼稳态的影响,QPCR和蛋白质印迹分析。为了确定MRTFA如何影响调节BMSC命运决定的机制,在体外分析来自WT和MRTFAKO小鼠的原代骨髓基质细胞以及C3H10T1/2细胞系。
结果:全球MRTFAKO小鼠的整体体重较低,股骨和胫骨的长度较短,股骨的骨量显着减少。与WT同窝相比,从KO小鼠分离的BMSC显示出增加的脂肪生成和减少的骨生成。KO老鼠,尤其是女性,随着年龄的增长出现骨质减少,高脂肪饮食增强了这一点。MRTFA或SRF的过表达增强CH310T1/2细胞系中的骨生成。Sca1(+),与WT对应物相比,来自KO骨髓的CD45(-)细胞表达较低量的平滑肌肌动蛋白(SMA)和TAZ/YAP靶基因。
结论:本研究通过调节BMSCs成脂分化和成骨分化之间的平衡,将MRTFA鉴定为一种新型的骨骼稳态调节因子。我们建议MRTFA通过维持BMSCs中SMA的产生来促进TAZ/YAP的成骨活性。
OBJECTIVE: Arising from common progenitors in the bone marrow, adipogenesis and osteogenesis are closely associated yet mutually exclusive during bone marrow mesenchymal stem cell (BMSC) development. Previous studies have shown that morphological changes can affect the early commitment of pluripotent BMSCs to the adipose versus osteoblastic lineage via modulation of RhoA activity. The RhoA pathway regulates actin polymerization to promote the incorporation of globular actin (G-actin) into filamentous actin (F-actin). In doing so, myocardin-related transcription factors (MRTFs) dissociate from bound G-actin and enter the nucleus to co-activate serum response factor (SRF) target gene expression. In this study, we investigated whether MRTFA/SRF is acting downstream of the RhoA pathway to regulate BMSC commitment in mice.
METHODS: The effects of knocking out MRTFA on skeletal homeostasis was studied in MRTFA KO mice using micro-CT, QPCR and western blot assays. To determine how MRTFA affects the mechanisms regulating BMSC fate decisions, primary bone marrow stromal cells from WT and MRTFA KO mice as well as C3H10T1/2 cell lines were analyzed in vitro.
RESULTS: Global MRTFA KO mice have lower whole body weight, shorter femoral and tibial lengths as well as significantly decreased bone mass in their femurs. BMSCs isolated from the KO mice show increased adipogenesis and reduced osteogenesis when compared to WT littermates. KO mice, particularly females, develop osteopenia with age, and this was enhanced by a high fat diet. Over-expression of MRTFA or SRF enhances osteogenesis in CH310T1/2 cell lines. Sca1(+), CD45(-) cells from KO marrow express lower amounts of smooth muscle actin (SMA) and TAZ/YAP target genes compared to WT counterparts.
CONCLUSIONS: This study identified MRTFA as a novel regulator of skeletal homeostasis by regulating the balance between adipogenic and osteogenic differentiation of BMSCs. We propose that MRTFA promotes the osteogenic activity of TAZ/YAP by maintaining SMA production in BMSCs.