Abstract

Background

The durative endoplasmic reticulum stress (ERS) and subsequent apoptosis contributes to the development and progression of Alzheimer’s disease (AD). MiR-326 can reduce pyruvate kinase M2 (PKM2) expression, leading to ERS. Whereas, lncRNA RPPH1 is able to increase dendritic spine density and protect hippocampal pyramidal neurons through targeting miR-326. Our study aims to investigate the regulation of lncRNA RPPH1 and miR-326/PKM2 on ERS and related apoptosis in AD.

Methods

SH-SY5Y cells treated with Aβ_25-35 were selected as an in vitro AD model. RPPH1 and miR-326 overexpression and silencing cells were established by transforming vectors. The expression of RPPH1 and miR-326 were detected by qRT-PCR. MTT, flow cytometric, intracellular calcium assay and Western blot were used to test the functions of RPPH1 and miR-326 in SH-SY5Y cell proliferation, apoptosis and ERS. Dual-luciferase assay was used to detect the interaction among RPPH1, miR-326 and PKM2.

Results

RPPH1 overexpression enhanced the viability of SH-SY5Y cells, and attenuated the apoptosis of of SH-SY5Y cells. Moreover, RPPH1 overexpression down-regulated ER stress related proteins such as GRP78, CHOP and cleaved caspase-12. Mechanistically, RPPH1 directly targeted miR-326, thereby counteracting its inhibitory effect on PKM2 expression, contributing to attenuation of apoptosis and ERS induced by Aβ_25-35.

Conclusion

Aβ_25-35-induced ERS and apoptosis in SH-SY5Y cells can be attenuated by lncRNA RPPH1 through regulating miR-326/PKM2 axis. This study provided therapeutic options for AD patients.

中文翻译：

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LncRNA RPPH1通过miR-326 / PKM2减轻Aβ25-35诱导的内质网应激和SH-SY5Y细胞凋亡

摘要

背景

持续的内质网应激（ERS）和随后的细胞凋亡有助于阿尔茨海默氏病（AD）的发展和进程。MiR-326可以减少丙酮酸激酶M2（PKM2）的表达，从而导致ERS。而lncRNA RPPH1能够通过靶向miR-326增加树突棘密度并保护海马锥体神经元。我们的研究旨在探讨lncRNA RPPH1和miR-326 / PKM2对ERS以及AD中相关凋亡的调控。

方法

用Aβ处理的SH-SY5Y细胞_25-35被选定作为体外AD模型。RPPH1和miR-326过表达和沉默细胞是通过转化载体建立的。用qRT-PCR检测RPPH1和miR-326的表达。MTT法，流式细胞术，细胞内钙含量测定和Western blot检测RPPH1和miR-326在SH-SY5Y细胞增殖，凋亡和ERS中的作用。使用双重荧光素酶检测来检测RPPH1，miR-326和PKM2之间的相互作用。

结果

RPPH1过表达增强了SH-SY5Y细胞的活力，并减弱了SH-SY5Y细胞的凋亡。此外，RPPH1过表达下调了内质网应激相关蛋白，例如GRP78，CHOP和裂解的caspase-12。机械地，RPPH1直接目标的miR-326，从而抵消在PKM2表达的抑制作用，促进细胞凋亡和ERSAβ引起的衰减_25-35。

结论

Aβ _25-35诱导ERS和SH-SY5Y细胞的细胞凋亡可以通过lncRNA RPPH1通过调节的miR-326 / PKM2轴线衰减。该研究为AD患者提供了治疗选择。