LncRNA-MIAT promotes neural cell autophagy and apoptosis in ischemic stroke by up-regulating REDD1

Abstract

Background: Ischemic stroke (IS) accounts for 80% of stroke incidence, which has an impact on the life quality of patients. Long non-coding RNA (LncRNA), a class of non-coding transcripts greater than 200 nucleotidesin length, has been extensively studied in cerebrovascular diseases. Myocardial infarction associated transcript (MIAT) is highly expressed in nervous system. Therefore this study aims to explore the role of LncRNA MIAT in IS and to clarify its underlying mechanism, providing therapeutic value for the treatment of IS.

Methods: The neurological function of rats was evaluated by neurological deficit score. Triphenyltetrazolium chloride (TTC) staining was used to detect infarct area in brain tissues. Quantitative real-time polymerase chain reaction (qRT-PCR) was performed to examine the expression of MIAT. Western blotting was used to detect the expressions of REDD1, p-mTOR, autophagy-related proteins LC3 and p62, and apoptotic-related proteins Bax, cleaved-caspase3, Bcl-2. Flow cytometry was applied to examine neuronal cell apoptosis. RNA pull-down and RIP assay was used to verify the binding of MIAT and REDD1. The level of REDD1 ubiquitination was detected by ubiquitination and Co-immunoprecipitation (Co-IP) assay.

Results: The expressions of MIAT and REDD1 were increased in IS rats and oxygen-glucose deprivation/reoxygenation (OGD/R)-induced PC12 cell injury. After interference with si-MIAT, the results of flow cytometry showed that the rate of apoptosis was reduced. Western blotting results showed that the expression of LC3II/LC3I, Bax, and cleaved-caspase3 was decreased, while the expression of p-mTOR, p62, and Bcl-2 was increased. RNA pull-down and RIP assay found the binding relationship between MIAT and REDD1, and interference with si-MIAT down-regulated the expression of REDD1. The level of REDD1 ubiquitination was increased and the expression of REDD1 was decreased after interference with si-MIAT in PC12 cells. Co-IP results showed that interference with si-MIAT enhanced the binding ability of CUL4A-DDB1 and REDD1.

Conclusion: Altogether, MIAT promotes autophagy and apoptosis of neural cells and aggravates IS by up-regulating the expression of REDD1.

Keywords: Apoptosis; Autophagy; Ischemic stroke; LncRNA-MIAT; REDD1.