Inhibition of LncRNA-HRIM Increases Cell Viability by Regulating Autophagy Levels During Hypoxia/Reoxygenation in Myocytes

Abstract

Backgrund/Aims: Ischemia reperfusion (I/R) promotes the severity of cardiomyocyte injury. Long noncoding RNAs (LncRNAs) are key regulators in cardiovascular diseases. However, the association between LncRNAs and myocardial I/R injury has not been thoroughly characterized to date. We attempted to clarify the potential biological role of a LncRNA (E230034O05Rik), which we named hypoxia/reoxygenation (H/R) injury-related factor in myocytes (HRIM), by investigating the differential expression of LncRNAs between groups of myocytes exposed to either a normal level of oxygen or to H/R. Methods: Microarray analysis was used to determine analyze the global differential expression of LncRNAs in H9c2 myocytes exposed either to a normal level of oxygen or to H/R. Target LncRNA levels were further verified in vitro and ex vivo by real-time polymerase chain reaction (qPCR). Cell viability was analyzed using the Cell Counting Kit-8 assay. Autophagy levels were confirmed by Western blotting, transmission electron microscopy, and autophagic double-labeled (mRFP-GFP-LC3) adenovirus analyses. Results: Gene expression profiling revealed that 797 LncRNAs and 1898 mRNAs were differentially expressed in the H/R group compared with the normal oxygen group. Among these LncRNAs and mRNAs, 6 upregulated LncRNAs and 2 downregulated LncRNAs in the H/R group were selected and further validated by qPCR in vitro and ex vivo. Additionally, LncRNA-HRIM was inhibited by specific siRNAs in H9c2 myocytes exposed to H/R. The inhibition of LncRNA-HRIM by siRNA prevented cell death by suppressing excessive autophagic activity in myocytes, This finding suggests a detrimental role of LncRNA-HRIM in the regulation of I/R injury. Conclusions: LncRNAs are involved in H/R injury of H9c2 myocytes. Inhibition of LncRNA-HRIM increased cell viability by reducing autophagy in myocytes during H/R.

Long noncoding RNAs: Important participants and potential therapeutic targets for myocardial ischaemia reperfusion injury

“…Studies in rodent models have shown that I/R causes dysregulated lncRNA expressions in cardiac tissue. [10][11][12] For instance, Liu 11 with a similar trend of changes observed in both of the cell culture and the cardiac tissue. Collectively, these results support the notion that I/R causes a dysregulation of the expression of cardiac lncRNAs.…”

“…Among these dysregulated lncRNAs, 12 have been confirmed to be related to genes encoding for apoptosis‐related proteins such as STAT3 and IL1R1 10 . Huang et al have explored the effect of I/R on lncRNA expressions inH9c2 myocytes and Langendorff‐perfused hearts from rats, showing that 797 lncRNAs and 1,898 mRNAs are differentially expressed after I/R challenge, 11 with a similar trend of changes observed in both of the cell culture and the cardiac tissue. Collectively, these results support the notion that I/R causes a dysregulation of the expression of cardiac lncRNAs.…”

“…Studies in rodent models have shown that I/R causes dysregulated lncRNA expressions in cardiac tissue 10‐12 . For instance, Liu et al 12 have found that 151 lncRNAs are differentially expressed (with 64 upregulated and 87 downregulated) in I/R‐treated myocardium from rats.…”

Non-coding RNAs modulate autophagy in myocardial ischemia-reperfusion injury: a systematic review

“…The global differential expression of lncRNAs analyzed by microarray analysis showed that 797 lncRNAs were differentially expressed in the H/R group. Therein, the repression lncRNA-HRIM via specific siRNAs protected cells from death through diminishing autophagy in H9c2 myocytes during H/R [ 80 ]. LncRNA NEAT1 associated with the development of various diseases was upregulated in peripheral blood and mouse cardiomyocytes during MI, which markedly increased the proliferation and migration of cardiomyocytes.…”

Noncoding RNAs in Cardiac Autophagy following Myocardial Infarction

“…HRIM silencing prevented death of cells by suppressing the autophagic activity in H/R-treated cells. However, the target genes of this lncRNA and the detailed mechanism of its autophagic effect need to be elucidated [42]. Other lncRNAs were highly expressed in diabetic murine heart and contributed to I/R injury by regulating autophagy.…”