qRT-PCR and western blot assays were used to evaluate the gene and protein expression levels of HOTAIRM1 in glioma patient tissue samples and cell lines. The role of HOTAIRM1 in glioma cell progression and VM formation was explored using a series of function gain-and-loss experiments. RNA-binding protein immunoprecipitation (RIP), RNA pull-down, and mechanism experiments were conducted to assess the interaction between HOTAIRM1/METTL3/IGFBP2 axis. Furthermore, rescue assays were conducted to explore the regulatory function of HOTAIRM1/METTL3/IGFBP2 in glioma cell cellular processes and VM formation.
We found that HOTAIRM1 presented up-regulation in glioma tissues and cells and overexpression of HOTAIRM1 facilitated glioma cell proliferation, migration, invasion, and VM formation. Furthermore, overexpression of HOTAIRM1 promoted glioma tumor growth and VM formation capacity in tumor xenograft mouse model. Moreover, HOTAIRM1 was demonstrated to interact with IGFBP2 and positively regulated IGFBP2 expression. IGFBP2 was found to promote glioma cell malignancy and VM formation. Mechanistically, METTL3 was highly expressed in glioma tissues and cells and was bound with HOTAIRM1 which stabilized HOTAIRM1 expression. Rescue assays demonstrated that METTL3 silencing counteracted the impact of HOTAIRM1 on glioma cell malignancy and VM formation capacity.
HOTAIRM1, post-transcriptionally stabilized by METTL3, promotes VM formation in glioma via up-regulating IGFBP2 expression, which provides a new direction for glioma therapy.
方法:qRT-PCR和westernblot测定用于评估HOTAIRM1在神经胶质瘤患者组织样品和细胞系中的基因和蛋白表达水平。使用一系列功能增益和损失实验探索HOTAIRM1在神经胶质瘤细胞进展和VM形成中的作用。RNA结合蛋白免疫沉淀(RIP),RNA下拉,并进行了机制实验以评估HOTAIRM1/METTL3/IGFBP2轴之间的相互作用。此外,进行了挽救试验,以探讨HOTAIRM1/METTL3/IGFBP2在神经胶质瘤细胞过程和VM形成中的调节功能。
结果:我们发现HOTAIRM1在神经胶质瘤组织和细胞中呈现上调,HOTAIRM1的过表达促进神经胶质瘤细胞增殖,迁移,入侵,和VM形成。此外,HOTAIRM1的过表达促进了肿瘤异种移植小鼠模型中胶质瘤肿瘤的生长和VM形成能力。此外,HOTAIRM1被证明与IGFBP2相互作用并正调节IGFBP2的表达。发现IGFBP2促进神经胶质瘤细胞恶性化和VM形成。机械上,METTL3在神经胶质瘤组织和细胞中高表达,并与HOTAIRM1结合,从而稳定HOTAIRM1的表达。挽救试验表明,METTL3沉默抵消了HOTAIRM1对神经胶质瘤细胞恶性和VM形成能力的影响。
结论:HOTAIRM1被METTL3转录后稳定,通过上调IGFBP2表达促进神经胶质瘤中的VM形成,这为胶质瘤的治疗提供了新的方向。