Date Published: October 14, 2019
Publisher: Sociedade Brasileira para o Desenvolvimento da Pesquisa em
Author(s): Jian-Min Xiao, Ji-Jia Wang, Li-Li Sun.
To reveal the function of miR-134 in myocardial ischemia.
Real-time PCR and western blotting were performed to measure the expression
of miR-134, nitric oxide synthase 3 (NOS3) and apoptotic-associated
proteins. Lactic dehydrogenase (LDH) assay, cell counting kit-8 (CCK-8),
Hoechst 33342/PI double staining and flow cytometry assay were implemented
in H9c2 cells, respectively. MiR-134 mimic/inhibitor was used to regulate
miR-134 expression. Bioinformatic analysis and luciferase reporter assay
were utilized to identify the interrelation between miR-134 and NOS3. Rescue
experiments exhibited the role of NOS3. The involvement of PI3K/AKT was
assessed by western blot analysis.
MiR-134 was high regulated in the myocardial ischemia model, and miR-134
mimic/inhibitor transfection accelerated/impaired the speed of cell
apoptosis and attenuated/exerted the cell proliferative prosperity induced
by H/R regulating active status of PI3K/AKT signaling. LDH activity was also
changed due to the different treatments. Moreover, miR-134 could target NOS3
directly and simultaneously attenuated the expression of NOS3.
Co-transfection miR-134 inhibitor and pcDNA3.1-NOS3 highlighted the
inhibitory effects of miR-134 on myocardial H/R injury.
This present work puts insights into the crucial effects of the miR-134/NOS3
axis in myocardial H/R injury, delivering a potential therapeutic technology
Myocardial infarction (MI) is one of the leading causes for high mortality worldwide
and usually poses a huge threat to human health 1 . It refers to a kind of acute coronary syndrome induced by abnormal blood
flow in the heart 2 , 3 . Prior studies have validated the importance of reperfusion as an
efficacious strategy against MI 4 . However, the conduction of reperfusion often results in a myocardial
ischemia-reperfusion (I/R) injury by diverse physiological processes 5 . Thus, how to prevent cardiomyocytes from I/R injury is the key for
treatment of myocardial ischemia and other related diseases.
It has been extensively studied that H/R can trigger damage to the membrane of
cardiomyocytes, further leading to release of cellular enzymes. Thus, the detection
of LDH activity contributes to confirm the degree of myocardial damage 14 , 15 . In addition, considering the important significance of skeletal myoblasts
in heart diseases, we selected H9c2 cells with better stability and reproducibility
instead of primary rat cardiomyocytes 16 , 17 . MiR-134 is a well-described microRNA in brain, and it has been widely used
to modulate the development and physiology of brain 18 . According to previous researches, we observed that miR-134 is located in
hippocampal neurons in rats and interacts with memory formation through controlling
CREB protein expression level 19 , 20 . In the study of Rong et al . 21 , miR-134 was presumed to be a peripheral biomarker for diagnosis of bipolar
disorder because of its altered level in blood. This seems to be prompting us that
miR-134 was involved in brain-related diseases by manipulating physiological
reactions correlated with blood. As evidence, it has been verified that a reduction
of miR-134 can moderate ischemic injury 10 . Mice were also utilized to demonstrate the effect of miR-134 in the
cerebral ischemic cortex and similar results were obtained 22 . Based on the extensive literature about the involvement of miRNAs in
myocardial I/R injury treatment and the impact of miR-134 in ischemic disorders, we
turned to explore the effects of miR-134 against myocardial H/R injury 22 – 24 . As expected, we found that miR-134 was increased after H/R by means of H9c2
cells and the conduction of flow cytometry assay and CCK-8 analysis showed us that
miR-134 mimic/inhibitor can strengthen/alleviate cardiomyocyte injury led by H/R.
LDH activity was also regulated by a diverse expression level of miR-134.
The deletion of miR-134 could attenuate myocardial H/R injury modulating the PI3K/Akt
pathway and targeting NOS3, thus weakening apoptotic activity and elevating
proliferative ability. These results will prove useful in expanding our knowledge of
how to treat myocardial I/R injury effectively.