Lee Eugene, Park Seo-Young, Moon Jae-Yeon, Ko Ji-Yun, Kim Tae Kyung, Im Gun-Il
Research Institute for Integrative Regenerative Biomedical Engineering, Dongguk University, Goyang, Republic of Korea.
Department of Orthopaedics, Dongguk University Ilsan Hospital, Goyang, Republic of Korea.
J Bone Miner Res. 2022 Jul;37(7):1382-1399. doi: 10.1002/jbmr.4565. Epub 2022 May 23.
Poor survival of grafted cells is the major impediment of successful cell-based therapies for bone regeneration. Implanted cells undergo rapid death in an ischemic environment largely because of hypoxia and metabolic stress from glucose deficiency. Understanding the intracellular metabolic processes and finding genes that can improve cell survival in these inhospitable conditions are necessary to enhance the success of cell therapies. Thus, the purpose of this study was to investigate changes of metabolic profile in glucose-deprived human bone marrow stromal/stem cells (hBMSCs) through metabolomics analysis and discover genes that could promote cell survival and osteogenic differentiation in a glucose-deprived microenvironment. Metabolomics analysis was performed to determine metabolic changes in a glucose stress metabolic model. In the absence of glucose, expression levels of all metabolites involved in glycolysis were significantly decreased than those in a glucose-supplemented state. In glucose-deprived osteogenic differentiation, reliance on tricarboxylic acid cycle (TCA)-predicted oxidative phosphorylation instead of glycolysis as the main mechanism for energy production in osteogenic induction. By comparing differentially expressed genes between glucose-deprived and glucose-supplemented hBMSCs, NR2F1 (Nuclear Receptor Subfamily 2 Group F Member 1) gene was discovered to be associated with enhanced survival and osteogenic differentiation in cells under metabolic stress. Small, interfering RNA (siRNA) for NR2F1 reduced cell viability and osteogenic differentiation of hBMSCs under glucose-supplemented conditions whereas NR2F1 overexpression enhanced osteogenic differentiation and cell survival of hBMSCs in glucose-deprived osteogenic conditions via the protein kinase B (AKT)/extracellular signal-regulated kinase (ERK) pathway. NR2F1-transfected hBMSCs significantly enhanced new bone formation in a critical size long-bone defect of rats compared with control vector-transfected hBMSCs. In conclusion, the results of this study provide an understanding of the metabolic profile of implanted cells in an ischemic microenvironment and demonstrate that NR2F1 treatment may overcome this deprivation by enhancing AKT and ERK regulation. These findings can be utilized in regenerative medicine for bone regeneration. © 2022 American Society for Bone and Mineral Research (ASBMR).
移植细胞的低存活率是基于细胞的骨再生治疗取得成功的主要障碍。植入的细胞在缺血环境中会迅速死亡，这主要是由于缺氧以及葡萄糖缺乏导致的代谢应激。了解细胞内代谢过程并找到能够在这些恶劣条件下提高细胞存活率的基因，对于提高细胞治疗的成功率至关重要。因此，本研究的目的是通过代谢组学分析来研究葡萄糖缺乏的人骨髓基质/干细胞（hBMSC）的代谢谱变化，并发现能够在葡萄糖缺乏的微环境中促进细胞存活和成骨分化的基因。进行代谢组学分析以确定葡萄糖应激代谢模型中的代谢变化。在缺乏葡萄糖的情况下，糖酵解中所有代谢物的表达水平均比葡萄糖补充状态下显著降低。在葡萄糖缺乏的成骨分化过程中，依靠三羧酸循环（TCA）预测的氧化磷酸化而非糖酵解作为成骨诱导中能量产生的主要机制。通过比较葡萄糖缺乏和葡萄糖补充的hBMSC之间的差异表达基因，发现NR2F1（核受体亚家族2组F成员1）基因与代谢应激下细胞的存活率提高和成骨分化有关。针对NR2F1的小干扰RNA（siRNA）在葡萄糖补充条件下降低了hBMSC的细胞活力和成骨分化，而NR2F1过表达通过蛋白激酶B（AKT）/细胞外信号调节激酶（ERK）途径增强了葡萄糖缺乏的成骨条件下hBMSC的成骨分化和细胞存活。与对照载体转染的hBMSC相比，NR2F1转染的hBMSC在大鼠临界尺寸长骨缺损中显著增强了新骨形成。总之，本研究结果提供了对缺血微环境中植入细胞代谢谱的理解，并表明NR2F1治疗可能通过增强AKT和ERK调节来克服这种缺乏。这些发现可用于骨再生的再生医学。©2022美国骨与矿物质研究学会（ASBMR）
