Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2018 Sep;16(3):1814-1824.
doi: 10.3892/etm.2018.6388. Epub 2018 Jul 2.

Downregulation of microRNA-1 attenuates glucose-induced apoptosis by regulating the liver X receptor α in cardiomyocytes

Yongxia Cheng  1 , Wei Zhao  2 , Xiaodong Zhang  3 , Lixin Sun  4 , Heran Yang  5 , Ying Wang  2 , Yong Cao  1 , Yanhui Chu  6 , Guibo Liu  2
Affiliations

Downregulation of microRNA-1 attenuates glucose-induced apoptosis by regulating the liver X receptor α in cardiomyocytes

Yongxia Cheng et al. Exp Ther Med. 2018 Sep.

Abstract

Diabetic cardiomyopathy (DCM) is characterized by abnormal myocardial structure or performance. It has been suggested that microRNA-1 (miR-1) may be abnormally expressed in the hearts of patients with diabetes. In the present study, the role of miR-1 in glucose-induced apoptosis and its underlying mechanism of action was investigated in rat cardiomyocyte H9C2 cells. Cells were transfected with anti-miR-1 or miR-1-overexpression plasmids and the expression of miR-1 and liver X receptor α (LXRα) were determined by reverse transcription-quantitative polymerase chain reaction analysis. The proportion of apoptotic cells was determined using an Annexin-V-FITC apoptosis detection kit and the mitochondrial membrane potential (ΔΨ) was measured following staining with rhodamine 123. In addition, the expression of apoptosis-associated proteins was measured by western blot analysis. The results demonstrated that expression of miR-1 was significantly increased, whereas the expression of LXRα was significantly decreased in H9C2 cells following treatment with glucose. miR-1 knockdown significantly inhibited apoptosis, increased the ΔΨ and suppressed the cleavage of poly (adenosine diphosphate-ribose) polymerase, caspase-3 and caspase-9. It also significantly downregulated the expression of Bcl-2 and upregulated the expression of Bax. In addition, it was demonstrated that miR-1 regulates LXRα; transfection with anti-miR-1 significantly increased the expression of LXRα. Furthermore, treatment of cells with the LXR agonist GW3965 inhibited apoptosis in glucose-induced anti-miR-1 cells. These results suggest a novel function of miR-1: The regulation of cardiomyocyte apoptosis via LXRα, and provide novel insights into regarding the complex mechanisms involved in DCM.

Keywords: apoptosis; cardiomyocytes; diabetic cardiomyopathy; liver X receptor α; microRNA-1.

PubMed Disclaimer

Figures

Figure 1.
Figure 1.
Expression of miR-1 and LXRα in glucose-induced H9C2 cells. The expression of (A) miR-1 and (B) LXRa mRNA in glucose-induced H9C2 cells. (C) The expression of LXRα protein in glucose-induced H9C2 cells. (D) The expression of miR-1 in H9C2 cells following transfection with anti-miR-1 and NC plasmids. (E) The proportion of apoptotic cells in groups transfected with NC, anti-miR-1 and subsequently treated with or without glucose. **P<0.01 vs. H9C2; &&P<0.01 vs. NC; ##P<0.01 vs. NC+Glucose. NC, negative control; miR, microRNA; LXRα, liver X receptor α; FITC, fluorescein isothiocyanate; PI, propidium iodide.
Figure 2.
Figure 2.
Effect of miR-1 silencing on glucose-induced apoptosis. (A) Effect of miR-1 silencing on glucose-induced mitochondrial dysfunction in H9C2 cells. (B) Quantification of fluorescence intensity. (C) The expression of apoptosis-associated proteins, including Bcl-2, Bax, total/cleaved caspase-3, total/cleaved caspase-9 and total/cleaved PARP in H9C2 cells. (D) Quantitative analysis of western blot analysis results. **P<0.01 vs. H9C2; #P<0.05 and ##P<0.01 vs. NC+Glucose. NC, negative control; Bcl-2, B cell lymphoma-2; PARP, poly (adenosine diphosphate-ribose) polymerase; miR, microRNA.
Figure 3.
Figure 3.
miR-1 regulates the expression of LXRα in glucose-induced H9C2 cells. Silencing of miR-1 upregulated the (A) mRNA and (B) protein expression of LXRα in H9C2 cells as determined by RT-qPCR and western blot analysis, respectively. The expression of LXRα protein in H9C2 cells transfected with (C) anti-miR-1 or (D) miR-1 and NC plasmids was determined by western blot analysis. The expression of LXRα mRNA in H9C2 cells transfected with (E) anti-miR-1 or (F) miR-1 was determined by RT-qPCR. **P<0.01 vs. H9C2; &&P<0.01 vs. NC; #P<0.05 and ##P<0.01 vs. NC+Glucose. RT-qPCR, reverse transcription-quantitative polymerase chain reaction; miR, microRNA; LXRα, liver X receptor α; NC, negative control.
Figure 4.
Figure 4.
Effect of GW3965 on apoptosis in H9C2 cells. The apoptotic inhibitory effect of anti-miR-1 was amplified by GW3965 in glucose-induced H9C2 cells. (A) GW3965 inhibited the glucose-induced apoptosis in a dose-dependent manner in H9C2 cells, as determined by FACS analysis. (B) The results of FACS were quantified. (C) Quantification of the percentage of apoptotic cells following transfection with anti-miR-1 or NC plasmids. (D) Following transfection, and treatment with glucose or GW3965, apoptotic cells were detected by FACS analysis. **P<0.01 vs. H9C2; §§P<0.01, §P<0.05 vs. Glucose; #P<0.05 and ##P<0.01 vs. NC+Glucose; $P<0.05 vs. anti-miR-1+Glucose. miR, microRNA; NC, negative control; FACS, fluorescence-activated cell sorting; FITC, fluorescein isothiocyanate; PI, propidium iodide.
Figure 5.
Figure 5.
Effect of GW3965 on the expression of apoptosis-associated proteins. (A) The ΔΨ was detected by fluorescence-activated cell sorting analysis following staining with rhodamine 123 in H9C2 cells and (B) fluorescence intensity was quantified. (C) The expression of apoptosis-associated proteins in H9C2 cells was determined by western blot analysis and (D) the results were quantified. **P<0.01 vs. H9C2; #P<0.05 and ##P<0.01 vs. NC+Glucose; $P<0.05 and $$P<0.01 vs. anti-miR-1+Glucose. ΔΨ, mitochondrial membrane potential; miR, microRNA; NC, negative control.
Figure 6.
Figure 6.
miR-1 silencing increases the expression of SREBP-1c in glucose-induced H9C2 cells. The (A) mRNA and (B) protein expression of SREBP-1c was determined in glucose-induced H9C2 cells transfected with or without anti-miR-1 or NC plasmids. **P<0.01 vs. H9C2; ##P<0.01 vs. NC+Glucose. SREBP-1c, sterol regulatory element-binding transcription factor 1c; NC, negative control; miR, microRNA.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Benjamin EJ, Blaha MJ, Chiuve SE, Cushman M, Das SR, Deo R, de Ferranti SD, Floyd J, Fornage M, Gillespie C, et al. Heart disease and stroke statistics-2017 update: A report from the american heart association. Circulation. 2017;135:e146–e603. doi: 10.1161/CIR.0000000000000485. - DOI - PMC - PubMed
    1. Mishra PK, Ying W, Nandi SS, Bandyopadhyay GK, Patel KK, Mahata SK. Diabetic cardiomyopathy: An immunometabolic perspective. Front Endocrinol (Lausanne) 2017;8:72. doi: 10.3389/fendo.2017.00072. - DOI - PMC - PubMed
    1. Gersh BJ, Sliwa K, Mayosi BM, Yusuf S. Novel therapeutic concepts: The epidemic of cardiovascular disease in the developing world: Global implications. Eur Heart J. 2010;31:642–648. doi: 10.1093/eurheartj/ehq030. - DOI - PubMed
    1. Aneja A, Tang WH, Bansilal S, Garcia MJ, Farkouh ME. Diabetic cardiomyopathy: Insights into pathogenesis, diagnostic challenges, and therapeutic options. Am J Med. 2008;121:748–757. doi: 10.1016/j.amjmed.2008.03.046. - DOI - PubMed
    1. Shivalkar B, Dhondt D, Goovaerts I, Van Gaal L, Bartunek J, Van Crombrugge P, Vrints C. Flow mediated dilatation and cardiac function in type 1 diabetes mellitus. Am J Cardiol. 2006;97:77–82. doi: 10.1016/j.amjcard.2005.07.111. - DOI - PubMed