Department of Orthopedic surgery, Second Hospital of Lanzhou University, Lanzhou, China.
Gansu Provincial Hospital of Traditional Chinese Medicine, Lanzhou, China.
J Biochem Mol Toxicol. 2024 Sep;38(9):e23846. doi: 10.1002/jbt.23846.
As a subclass of noncoding RNAs, circular RNA play an important role in tumour development. The aim of this study was to investigate the role of circ_0004674 in osteosarcoma glycolysis and the molecular mechanism of its regulation. We examined the expression of circ_0004674, miR-140-3p, TCF4 and glycolysis-related proteins (including HK2, PKM2, GLUT1 and LDHA) in osteosarcoma cells and tissues by quantitative reverse transcription-polymerase chain reaction and immunoblotting (Western blot analysis). The role of circ_0004674, miR-140-3p and TCF4 in the proliferation, apoptosis, migration and invasion of OS cells was examined using CCK8 assay, Apoptosis assay, Wound healing assay, Transwell migration and Matrigel invasion assay. The interaction of circ_0004674/miR-140-3p and miR-1543/TCF4 was also analysed using a dual luciferase reporter assay. Finally, the glycolytic process was assessed by glucose uptake assays and lactate production measurements. The results showed that the expression of circ_0004674 and TCF4 was significantly higher in MG63 and U2OS cells compared to hFOB1.19 cells, while the expression of miR-140-3p was downregulated. Silencing of circ_0004674 gene significantly inhibited the proliferation, migration and invasion of cancer cells and promoted apoptosis of cancer cells. Experiments such as dual luciferase reporter analysis showed that circ_0004674 regulates the expression of glycolysis-related proteins through the miR-140-3p/TCF4 pathway, and inhibition of this gene attenuated the depletion of glucose content and the production of lactate in cancer cells. Furthermore, inhibition of miR-140-3p or overexpression of TCF could reverse the phenotypic changes in cancer cells induced by circ_0004674 silencing. In summary, this study elucidated the specific function and potential mechanisms of circ_0004674 in osteosarcoma glycolysis. The findings demonstrate that miR-140-3p and TCF4 function respectively as a tumor suppressor gene and an oncogene in osteosarcoma. Notably, they influence glycolysis and associated pathways, regulating osteosarcoma proliferation. Therefore, circ_0004674 promotes osteosarcoma glycolysis and proliferation through the miR-140-3p/TCF4 pathway, enhancing the malignant behaviour of tumours, and it is expected to be a potential molecular target for osteosarcoma treatment.
作为非编码 RNA 的一个子类，环状 RNA 在肿瘤发展中发挥重要作用。本研究旨在探讨 circ_0004674 在骨肉瘤糖酵解中的作用及其调控的分子机制。我们通过定量逆转录聚合酶链反应和免疫印迹（Western blot 分析）检测骨肉瘤细胞和组织中 circ_0004674、miR-140-3p、TCF4 和糖酵解相关蛋白（包括 HK2、PKM2、GLUT1 和 LDHA）的表达。通过 CCK8 测定、凋亡测定、划痕愈合测定、Transwell 迁移和 Matrigel 侵袭测定检测 circ_0004674、miR-140-3p 和 TCF4 在 OS 细胞增殖、凋亡、迁移和侵袭中的作用。还通过双荧光素酶报告基因测定分析 circ_0004674/miR-140-3p 和 miR-1543/TCF4 的相互作用。最后，通过葡萄糖摄取测定和乳酸产生测定评估糖酵解过程。结果显示，与 hFOB1.19 细胞相比，MG63 和 U2OS 细胞中 circ_0004674 和 TCF4 的表达明显升高，而 miR-140-3p 的表达下调。circ_0004674 基因沉默显著抑制癌细胞的增殖、迁移和侵袭，促进癌细胞凋亡。双荧光素酶报告分析等实验表明，circ_0004674 通过 miR-140-3p/TCF4 通路调节糖酵解相关蛋白的表达，该基因的抑制减弱了癌细胞中葡萄糖含量的消耗和乳酸的产生。此外，抑制 miR-140-3p 或过表达 TCF 可以逆转 circ_0004674 沉默诱导的癌细胞表型变化。总之，本研究阐明了 circ_0004674 在骨肉瘤糖酵解中的特定功能和潜在机制。研究结果表明，miR-140-3p 和 TCF4 分别作为骨肉瘤中的抑癌基因和癌基因发挥作用。值得注意的是，它们影响糖酵解和相关途径，调节骨肉瘤的增殖。因此，circ_0004674 通过 miR-140-3p/TCF4 通路促进骨肉瘤糖酵解和增殖，增强肿瘤的恶性行为，有望成为骨肉瘤治疗的潜在分子靶点。
