Zhang Yuhao, Liu Shiming, Wu Cheng, Gao Xin, Zhao Hongtao, Li Ou, Yang Kaichuang, Gao Faliang
Department of Neurosurgery, Cancer Center, Zhejiang Provincial People's Hospital (Affiliated People's Hospital), Hangzhou Medical College, No. 158, Shangtang Road, Gongshu District, Hangzhou City, 310000, Zhejiang Province, China.
General Surgery, Cancer Center, Department of Gastrointestinal and Pancreatic Surgery, Zhejiang Provincial People's Hospital (Affiliated People's Hospital), Hangzhou Medical College, No. 158, Shangtang Road, Gongshu District, Hangzhou City, Zhejiang Province, 310000, China.
Mol Neurobiol. 2025 Apr 15. doi: 10.1007/s12035-025-04875-9.
Our study aimed to explore the involvement of circ_0001583 in the progression of glioblastoma (GBM). The expression levels of CircRNA in GBM were examined using the GEO database, and quantification of circ_0001583 levels was performed through qRT-PCR in both GBM tissues and cell lines. Survival analysis was conducted using Kaplan-Meier plots. The effects of circ_0001583 knockdown on cell proliferation, invasion, and glycolysis were evaluated through functional assays and measurements of glycolytic activity. Bioinformatics and reporter assays revealed that miR-647 serves as a target for circ_0001583. Tumorigenesis following circ_0001583 knockdown was investigated in a nude mouse model utilizing U251 cells. Additionally, a co-culture system with normal human astrocytes (NHAs) and GBM-conditioned medium was employed to study circ_0001583 expression and its influence on cell proliferation. The presence of circ_0001583 in normal human astrocytes was assessed using PKH67 labeling and immunofluorescence techniques. Bioinformatics analyses revealed a notable increase in the expression of circ_0001583 in GBM, correlating with metastasis. The inhibition of circ_0001583 expression led to decreased viability, proliferation, and invasive potential of GBM cells. It was found that circ_0001583 targets miR-647, which is downregulated in GBM. Silencing circ_0001583 resulted in elevated levels of miR-647. CKAP2L, a target of miR-647, showed an overexpression in GBM cases. Additionally, a positive relationship was established between circ_0001583 and CKAP2L, contrasting with a negative association with miR-647. An increase in miR-647 expression led to a decrease in CKAP2L protein levels. Rescue assays further validated that circ_0001583 influences the functions of GBM cells through the miR-647/CKAP2L pathway. In vivo investigations demonstrated that the knockdown of circ_0001583 resulted in retarded tumor growth, enhanced levels of miR-647, and diminished expression of Ki-67, CKAP2L, HK2, and LDHA within tumors. Moreover, co-culture studies involving GBM-conditioned medium demonstrated an upregulation of circ_0001583 in NHAs, which promoted cell proliferation. Our data collectively suggest that exosomal circ_0001583 promotes glioblastoma progression via the miR-647/CKAP2L pathway.
我们的研究旨在探讨circ_0001583在胶质母细胞瘤（GBM）进展中的作用。使用GEO数据库检测GBM中CircRNA的表达水平，并通过qRT-PCR对GBM组织和细胞系中的circ_0001583水平进行定量。使用Kaplan-Meier图进行生存分析。通过功能测定和糖酵解活性测量评估circ_0001583敲低对细胞增殖、侵袭和糖酵解的影响。生物信息学和报告基因检测表明，miR-647是circ_0001583的靶点。利用U251细胞在裸鼠模型中研究circ_0001583敲低后的肿瘤发生情况。此外，采用正常人星形胶质细胞（NHA）与GBM条件培养基的共培养系统，研究circ_0001583的表达及其对细胞增殖的影响。使用PKH67标记和免疫荧光技术评估正常人星形胶质细胞中circ_0001583的存在情况。生物信息学分析显示，GBM中circ_0001583的表达显著增加，与转移相关。circ_0001583表达的抑制导致GBM细胞的活力、增殖和侵袭潜力降低。研究发现，circ_0001583靶向miR-647，而miR-647在GBM中表达下调。沉默circ_0001583导致miR-647水平升高。miR-647的靶点CKAP2L在GBM病例中呈过表达。此外，circ_0001583与CKAP2L之间呈正相关，与miR-647呈负相关。miR-647表达的增加导致CKAP2L蛋白水平降低。挽救实验进一步验证了circ_0001583通过miR-647/CKAP2L途径影响GBM细胞的功能。体内研究表明，circ_0001583的敲低导致肿瘤生长迟缓、miR-647水平升高，以及肿瘤内Ki-67、CKAP2L、HK2和LDHA的表达降低。此外，涉及GBM条件培养基的共培养研究表明，NHA中circ_0001583上调，促进细胞增殖。我们的数据共同表明，外泌体circ_0001583通过miR-647/CKAP2L途径促进胶质母细胞瘤进展。
