Xu Enjie, Huang Zhen, Zhu Kunpeng, Hu Jianping, Ma Xiaolong, Wang Yongjie, Zhu Jiazhuang, Zhang Chunlin
Department of Orthopedic Surgery, Shanghai Tenth People's Hospital, School of Medicine, Tongji University, Shanghai 200072, PR China; Institute of Bone Tumor Affiliated to Tongji University School of Medicine, Shanghai 200072, PR China.
Department of Orthopedic Surgery, Shanghai Tenth People's Hospital, School of Medicine, Tongji University, Shanghai 200072, PR China; Institute of Bone Tumor Affiliated to Tongji University School of Medicine, Shanghai 200072, PR China.
Cell Signal. 2025 Jan;125:111501. doi: 10.1016/j.cellsig.2024.111501. Epub 2024 Nov 4.
Osteosarcoma (OS) cells commonly suffer from hypoxia and dedifferentiation, resulting in poor prognosis. We plan to identify the role of hypoxia on dedifferentiation and the associated cellular signaling.
We performed sphere formation assays and determined spheroid cells as dedifferentiated cells by detecting stem cell-like markers. RNAi assay was used to explore the relationship between hypoxia inducible factor 1 subunit alpha (HIF1A) and platelet derived growth factor receptor beta (PDGFRB). We obtained PDGFRB knockdown and overexpression cells through lentiviral infection experiments and detected the expression of PDGFRB, p-PDGFRB, focal adhesion kinase (FAK), p-FAK, phosphorylated myosin light chain 2 (p-MLC2), and ras homolog family member A (RhoA) in each group. The effects of PDGFRB on cytoskeleton rearrangement and cell adhesion were explored by immunocytochemistry. Wound-healing experiments, transwell assays, and animal trials were employed to investigate the effect of PDGFRB on OS cell metastasis both in vitro and in vivo.
Dedifferentiated OS cells were found to exhibit high expression of HIF1A and PDGFRB, and HIF1A upregulated PDGFRB, subsequently activated RhoA, and increased the phosphorylation of MLC2. PDGFRB also enhanced the phosphorylation of FAK. The OS cell morphology and vinculin distribution were altered by PDGFRB. PDGFRB promoted cell dedifferentiation and had a significant impact on the migration and invasion abilities of OS cells in vitro. In addition, PDGFRB increased pulmonary metastasis of OS cells in vivo.
Our results demonstrated that HIF1A up-regulated PDGFRB under hypoxic conditions, and PDGFRB regulated the actin cytoskeleton, a process likely linked to the activation of RhoA and the phosphorylation of, thereby promoting OS dedifferentiation and pulmonary metastasis.
骨肉瘤（OS）细胞常处于缺氧和去分化状态，导致预后不良。我们计划确定缺氧在去分化中的作用以及相关的细胞信号传导。
我们进行了成球实验，并通过检测干细胞样标志物将球状体细胞确定为去分化细胞。采用RNA干扰实验探讨缺氧诱导因子1α亚基（HIF1A）与血小板衍生生长因子受体β（PDGFRB）之间的关系。通过慢病毒感染实验获得PDGFRB敲低和过表达细胞，并检测每组中PDGFRB、磷酸化PDGFRB、粘着斑激酶（FAK）、磷酸化FAK、磷酸化肌球蛋白轻链2（p-MLC2）和Ras同源家族成员A（RhoA）的表达。通过免疫细胞化学研究PDGFRB对细胞骨架重排和细胞粘附的影响。采用伤口愈合实验、Transwell实验和动物实验研究PDGFRB对OS细胞体外和体内转移的影响。
发现去分化的OS细胞中HIF1A和PDGFRB高表达，HIF1A上调PDGFRB，随后激活RhoA，并增加MLC2的磷酸化。PDGFRB还增强了FAK的磷酸化。PDGFRB改变了OS细胞的形态和纽蛋白分布。PDGFRB促进细胞去分化，并对体外OS细胞的迁移和侵袭能力有显著影响。此外，PDGFRB增加了OS细胞在体内的肺转移。
我们的结果表明，在缺氧条件下HIF1A上调PDGFRB，PDGFRB调节肌动蛋白细胞骨架，这一过程可能与RhoA的激活和磷酸化有关，从而促进OS去分化和肺转移。
