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
. 2016 Jun 29;11(6):e0158038.
doi: 10.1371/journal.pone.0158038. eCollection 2016.

Protective Effect of Irisin on Atherosclerosis via Suppressing Oxidized Low Density Lipoprotein Induced Vascular Inflammation and Endothelial Dysfunction

Affiliations

Protective Effect of Irisin on Atherosclerosis via Suppressing Oxidized Low Density Lipoprotein Induced Vascular Inflammation and Endothelial Dysfunction

Yuzhu Zhang et al. PLoS One. .

Abstract

Irisin, a newly discovered myokine, is considered as a promising candidate for the treatment of metabolic disturbances and cardiovascular diseases. In the present study, we used two animal models, apolipoprotein E-deficient mice fed on a high-cholesterol diet and a mouse carotid partial ligation model to test the anti-atherosclerotic effect of irisin. Irisin treatment (0.5 μg/g body weight/day) significantly reduced the severity of aortic atherosclerosis in apolipoprotein E-deficient mice fed on a high-cholesterol diet and suppressed carotid neointima formation in a carotid partial ligation model. It was associated with decreased inflammation and cell apoptosis in aortic tissues. In addition, in a cell culture model, irisin restored ox-LDL-induced human umbilical vein endothelial cell dysfunction by reducing the levels of inflammatory genes via inhibiting the reactive oxygen species (ROS)/ p38 MAPK/ NF-κB signaling pathway activation and inhibiting cell apoptosis via up-regulating Bcl-2 and down-regulating Bax and caspase-3 expression. Our study demonstrated that irisin significantly reduced atherosclerosis in apolipoprotein E-deficient mice via suppressing ox-LDL-induced cell inflammation and apoptosis, which might have a direct therapeutic effect on atherosclerotic diseases.

PubMed Disclaimer

Conflict of interest statement

Competing Interests: The authors have declared that no competing interests exist.

Figures

Fig 1
Fig 1. Influence of irisin on atherosclerotic lesion formation in Apo E-deficient mice.
Apo E-deficient mice were treated with irisin for the last 8 weeks of a 12-week HCD feeding program (n = 8). (A) The atherosclerotic lesions in en face-prepared aorta were identified with Oil-Red O staining. (B) Lesion area (%) was expressed as percentage of atherosclerotic area/total area of the aorta. (C) The lipid-rich atherosclerotic lesions in aortic sinus were identified with Oil-Red O staining. (D) Quantification of atherosclerotic lesion areas in aortic root sections. *P < 0.05 vs. control, the data was expressed as the mean ± SEM.
Fig 2
Fig 2. Influence of irisin on neointima formation in partial ligated carotid arteries.
Apo E-deficient mice were treated with or without irisin for 28 days after carotid artery partial ligation (n = 5). (A) Scheme of partial ligation of the left carotid artery is shown. The left external carotid artery (ECA), internal carotid artery (ICA), and occipital artery (OA) were ligated with 6–0 silk suture. After ligation, flow was maintained only in the superior thyroid artery (STA). (B) Cross-sections of carotid arteries were identified with hematoxylin–eosin staining. (C) Quantification of the neointima areas of the partially ligated carotid arteries is shown. (D) Degree of neointima formation 28 days after carotid ligation was calculated according to N/M ratio. *P < 0.05 vs. control. The data was expressed as the mean ± SEM.
Fig 3
Fig 3. Influence of irisin on the inflammatory infiltration in partial ligated carotid arteries.
Representative pictures of immunohistochemistry staining of CD3 (A), and CD68 (B) in partial ligated carotid arteries are shown. Brown staining exhibits positive area, while blue represents counterstaining with hematoxylin. (C) Quantification of positive staining areas in carotid artery sections is shown. *P < 0.05 vs. control, n = 5 per group. The data was expressed as the mean ± SEM.
Fig 4
Fig 4. Influence of irisin on the inflammatory genes in partial ligated carotid arteries.
(A) The mRNA expressions of IL-6, MCP-1, ICAM-1, and VCAM-1 in carotid arteries after partial ligation were analyzed by qPCR. The mRNA levels were normalized to that of β-actin. (B) The concentrations of IL-6, MCP-1, ICAM-1, and VCAM-1 protein in plasma were measured by ELISA. *P < 0.05 vs. WT mice, # P < 0.05 vs. Apo E-deficient mice treated with NS, n = 5 per group. The data was expressed as the mean ± SEM.
Fig 5
Fig 5. Effect of irisin on ox-LDL-induced inflammation in HUVECs.
(A) HUVECs were exposed to different concentrations of ox-LDL (20–100 μg/mL) for 24h, and the cell viability was assessed through an MTT assay. (B) HUVECs were treated with different concentrations of irisin (10, 20, and 40 nM) with or without 80 μg/mL ox-LDL for 24 h, and the cell viability was assessed through an MTT assay. (C) HUVECs were incubated with irisin and ox-LDL for 24 h. The expression levels of ICAM-1, VCAM-1, MCP-1, and IL-6 were determined using qRT-PCR. The mRNA levels were normalized to that of β-actin. (D) HUVECs were cultured with irisin for 1 h, followed by stimulation with ox-LDL for 30 min. The expression levels of p-p38 MAPK, p-NF-κB p65, and NF-κB p65 were analyzed by western blot. Total p38 and GAPDH were used as a loading control respectively. (E) The expression levels of NF-κB p65 in the nucleus and cytosol were analyzed by western blot. (F) Densitometric analysis of the related bands was performed. (G) HUVECs were exposed to ox-LDL and/or irisin for 24 h. Intracellular ROS levels were measured using DCFH-DA. The measured fluorescence values were expressed as a percentage of the fluorescence in control cells. The data were expressed as the mean ± SEM of three independent experiments. *P < 0.05 vs. control, # P < 0.05 vs. ox-LDL -treated group.
Fig 6
Fig 6. Irisin inhibited EC apoptosis in partial ligated carotid arteries and ox-LDL-induced HUVECs.
(A) Sections from the mice carotid arteries were labeled by TUNEL to detect apoptotic cells and counterstained with DAPI to detect nuclei. The yellow dotted lines indicated the lumen perimeter of the vessel. (B) Quantification of TUNEL-positive cells in the plaque. (C) HUVECs were incubated with irisin and ox-LDL for 24 h. Apoptosis of ox-LDL-exposed HUVECs after treatment with irisin was detected by using flow cytometry. (D) The apoptotic rate was determined by calculating the ratio of Annexin-V-positive and Annexin-V/PI-double positive cells to total cells. The data were expressed as the mean ± SEM of three independent experiments. *P < 0.05 vs. control, # P < 0.05 vs. ox-LDL -treated group.
Fig 7
Fig 7. Effect of irisin on Bax, Bcl-2, and caspase-3 protein levels in ox-LDL-induced HUVECs.
HUVECs were incubated with irisin and ox-LDL for 24 h. (A) The expression levels of caspase-3, Bax, and Bcl-2 were analyzed by western blot. GAPDH was used as a loading control. (B) Densitometric analysis of the related bands was performed. The data were expressed as the mean ± SEM of three independent experiments. *P < 0.05 vs. control, # P < 0.05 vs. ox-LDL -treated group.

Similar articles

Cited by

References

    1. Lusis AJ. Atherosclerosis. Nature. 2000;407(6801):233–41. 10.1038/35025203 - DOI - PMC - PubMed
    1. Peters SA, den Ruijter HM, Bots ML, Moons KG. Improvements in risk stratification for the occurrence of cardiovascular disease by imaging subclinical atherosclerosis: a systematic review. Heart (British Cardiac Society). 2012;98(3):177–84. Epub 2011/11/19. 10.1136/heartjnl-2011-300747 . - DOI - PubMed
    1. Stefano R, Massimiliano C, Marina C, Rossella M, Chiara P, Alessio L, et al. A score including ADAM17 substrates correlates to recurring cardiovascular event in subjects with atherosclerosis. Atherosclerosis. 2015;239(2):459–64. Epub 2015/02/18. 10.1016/j.atherosclerosis.2015.01.029 . - DOI - PubMed
    1. Libby P. Inflammation in atherosclerosis. Nature. 2002;420(6917):868–74. - PubMed
    1. Pober JS, Min W, Bradley JR. Mechanisms of endothelial dysfunction, injury, and death. Annual review of pathology. 2009;4:71–95. Epub 2008/08/30. 10.1146/annurev.pathol.4.110807.092155 . - DOI - PubMed

MeSH terms

Grants and funding

This work was supported by the Tai shan Scholar Fund, the National Natural Science Foundation of China [81570407], and the Major State Basic Research Development Program of China (973 Program) [2015CB553600].