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Imaging of Intracellular and Extracellular ROS Levels in Atherosclerotic Mouse Aortas Ex Vivo: Effects of Lipid Lowering by Diet or Atorvastatin.

Ekstrand M, Gustafsson Trajkovska M, Perman-Sundelin J, Fogelstrand P, Adiels M, Johansson M, Mattsson-Hultén L, Borén J, Levin M - PLoS ONE (2015)

Bottom Line: Atorvastatin treatment did not affect lesion inflammation (aortic arch mRNA levels of CXCL 1, ICAM-1, MCP-1, TNF-α, VCAM, IL-6, and IL-1β) or cellular composition (smooth muscle cell, macrophage, and T-cell content).Our results suggest that within lesions, macrophages produce mainly intracellular ROS whereas smooth muscle cells produce extracellular ROS.Short term atorvastatin treatment, but not lipid lowering by diet, decreases ROS levels within established advanced lesions; this may help explain the lesion stabilizing and anti-inflammatory effects of long term statin treatment.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular and Clinical Medicine/Wallenberg Laboratory, University of Gothenburg and Sahlgrenska University Hospital, SE-413 45, Gothenburg, Sweden.

ABSTRACT

Objective: The first objective was to investigate if intracellular and extracellular levels of reactive oxygen species (ROS) within the mouse aorta increase before or after diet-induced lesion formation. The second objective was to investigate if intracellular and extracellular ROS correlates to cell composition in atherosclerotic lesions. The third objective was to investigate if intracellular and extracellular ROS levels within established atherosclerotic lesions can be reduced by lipid lowering by diet or atorvastatin.

Approach and results: To address our objectives, we established a new imaging technique to visualize and quantify intracellular and extracellular ROS levels within intact mouse aortas ex vivo. Using this technique, we found that intracellular, but not extracellular, ROS levels increased prior to lesion formation in mouse aortas. Both intracellular and extracellular ROS levels were increased in advanced lesions. Intracellular ROS correlated with lesion content of macrophages. Extracellular ROS correlated with lesion content of smooth muscle cells. The high levels of ROS in advanced lesions were reduced by 5 days high dose atorvastatin treatment but not by lipid lowering by diet. Atorvastatin treatment did not affect lesion inflammation (aortic arch mRNA levels of CXCL 1, ICAM-1, MCP-1, TNF-α, VCAM, IL-6, and IL-1β) or cellular composition (smooth muscle cell, macrophage, and T-cell content).

Conclusions: Aortic levels of intracellular ROS increase prior to lesion formation and may be important in initiation of atherosclerosis. Our results suggest that within lesions, macrophages produce mainly intracellular ROS whereas smooth muscle cells produce extracellular ROS. Short term atorvastatin treatment, but not lipid lowering by diet, decreases ROS levels within established advanced lesions; this may help explain the lesion stabilizing and anti-inflammatory effects of long term statin treatment.

No MeSH data available.


Related in: MedlinePlus

Intracellular ROS correlates with macrophage content and extracellular ROS with smooth muscle cell content in advanced atherosclerotic lesions Female Apoe-/- mice were fed Western diet to induce advanced atherosclerotic lesions in the aortic arch.(A) Intracellular and extracellular ROS were analyzed in the aortic arch (red area) (B) Smooth muscle cell and macrophage content was quantified in the aortic arch by analyzing sections from 4 different levels. (C) Section stained for macrophages (CD68: blue) and smooth muscle cells (α-actin: red). (D and E) Smooth muscle cell content in lesions correlated with extracellular (E) but not intracellular ROS (D). (F and G) Macrophage content in lesions correlated with intracellular (F) but not extracellular ROS (G). Linear regression (n = 25). NS (non significant).
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pone.0130898.g003: Intracellular ROS correlates with macrophage content and extracellular ROS with smooth muscle cell content in advanced atherosclerotic lesions Female Apoe-/- mice were fed Western diet to induce advanced atherosclerotic lesions in the aortic arch.(A) Intracellular and extracellular ROS were analyzed in the aortic arch (red area) (B) Smooth muscle cell and macrophage content was quantified in the aortic arch by analyzing sections from 4 different levels. (C) Section stained for macrophages (CD68: blue) and smooth muscle cells (α-actin: red). (D and E) Smooth muscle cell content in lesions correlated with extracellular (E) but not intracellular ROS (D). (F and G) Macrophage content in lesions correlated with intracellular (F) but not extracellular ROS (G). Linear regression (n = 25). NS (non significant).

Mentions: Both smooth muscle cells and macrophages produce ROS in atherosclerotic lesions [1–3]. However, it is not known if the cell types produce intracellular and/or extracellular ROS. To investigate this, we correlated intracellular and extracellular ROS levels with cellular composition of atherosclerotic lesions in the aortic arch (Fig 3). Ex vivo levels of intracellular and extracellular ROS were assessed in the aortic arch of female Apoe-/- mice fed Western diet for 3–9 weeks. After ROS analysis, lesion content of macrophages and smooth muscle cells was quantified. Intracellular ROS levels in atherosclerotic lesions showed significant positive correlation with macrophage content. Extracellular ROS levels showed significant positive correlation with lesion smooth muscle cell content. This indicates that macrophages within atherosclerotic lesions predominantly produce intracellular ROS and smooth muscle cells extracellular ROS.


Imaging of Intracellular and Extracellular ROS Levels in Atherosclerotic Mouse Aortas Ex Vivo: Effects of Lipid Lowering by Diet or Atorvastatin.

Ekstrand M, Gustafsson Trajkovska M, Perman-Sundelin J, Fogelstrand P, Adiels M, Johansson M, Mattsson-Hultén L, Borén J, Levin M - PLoS ONE (2015)

Intracellular ROS correlates with macrophage content and extracellular ROS with smooth muscle cell content in advanced atherosclerotic lesions Female Apoe-/- mice were fed Western diet to induce advanced atherosclerotic lesions in the aortic arch.(A) Intracellular and extracellular ROS were analyzed in the aortic arch (red area) (B) Smooth muscle cell and macrophage content was quantified in the aortic arch by analyzing sections from 4 different levels. (C) Section stained for macrophages (CD68: blue) and smooth muscle cells (α-actin: red). (D and E) Smooth muscle cell content in lesions correlated with extracellular (E) but not intracellular ROS (D). (F and G) Macrophage content in lesions correlated with intracellular (F) but not extracellular ROS (G). Linear regression (n = 25). NS (non significant).
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4476734&req=5

pone.0130898.g003: Intracellular ROS correlates with macrophage content and extracellular ROS with smooth muscle cell content in advanced atherosclerotic lesions Female Apoe-/- mice were fed Western diet to induce advanced atherosclerotic lesions in the aortic arch.(A) Intracellular and extracellular ROS were analyzed in the aortic arch (red area) (B) Smooth muscle cell and macrophage content was quantified in the aortic arch by analyzing sections from 4 different levels. (C) Section stained for macrophages (CD68: blue) and smooth muscle cells (α-actin: red). (D and E) Smooth muscle cell content in lesions correlated with extracellular (E) but not intracellular ROS (D). (F and G) Macrophage content in lesions correlated with intracellular (F) but not extracellular ROS (G). Linear regression (n = 25). NS (non significant).
Mentions: Both smooth muscle cells and macrophages produce ROS in atherosclerotic lesions [1–3]. However, it is not known if the cell types produce intracellular and/or extracellular ROS. To investigate this, we correlated intracellular and extracellular ROS levels with cellular composition of atherosclerotic lesions in the aortic arch (Fig 3). Ex vivo levels of intracellular and extracellular ROS were assessed in the aortic arch of female Apoe-/- mice fed Western diet for 3–9 weeks. After ROS analysis, lesion content of macrophages and smooth muscle cells was quantified. Intracellular ROS levels in atherosclerotic lesions showed significant positive correlation with macrophage content. Extracellular ROS levels showed significant positive correlation with lesion smooth muscle cell content. This indicates that macrophages within atherosclerotic lesions predominantly produce intracellular ROS and smooth muscle cells extracellular ROS.

Bottom Line: Atorvastatin treatment did not affect lesion inflammation (aortic arch mRNA levels of CXCL 1, ICAM-1, MCP-1, TNF-α, VCAM, IL-6, and IL-1β) or cellular composition (smooth muscle cell, macrophage, and T-cell content).Our results suggest that within lesions, macrophages produce mainly intracellular ROS whereas smooth muscle cells produce extracellular ROS.Short term atorvastatin treatment, but not lipid lowering by diet, decreases ROS levels within established advanced lesions; this may help explain the lesion stabilizing and anti-inflammatory effects of long term statin treatment.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular and Clinical Medicine/Wallenberg Laboratory, University of Gothenburg and Sahlgrenska University Hospital, SE-413 45, Gothenburg, Sweden.

ABSTRACT

Objective: The first objective was to investigate if intracellular and extracellular levels of reactive oxygen species (ROS) within the mouse aorta increase before or after diet-induced lesion formation. The second objective was to investigate if intracellular and extracellular ROS correlates to cell composition in atherosclerotic lesions. The third objective was to investigate if intracellular and extracellular ROS levels within established atherosclerotic lesions can be reduced by lipid lowering by diet or atorvastatin.

Approach and results: To address our objectives, we established a new imaging technique to visualize and quantify intracellular and extracellular ROS levels within intact mouse aortas ex vivo. Using this technique, we found that intracellular, but not extracellular, ROS levels increased prior to lesion formation in mouse aortas. Both intracellular and extracellular ROS levels were increased in advanced lesions. Intracellular ROS correlated with lesion content of macrophages. Extracellular ROS correlated with lesion content of smooth muscle cells. The high levels of ROS in advanced lesions were reduced by 5 days high dose atorvastatin treatment but not by lipid lowering by diet. Atorvastatin treatment did not affect lesion inflammation (aortic arch mRNA levels of CXCL 1, ICAM-1, MCP-1, TNF-α, VCAM, IL-6, and IL-1β) or cellular composition (smooth muscle cell, macrophage, and T-cell content).

Conclusions: Aortic levels of intracellular ROS increase prior to lesion formation and may be important in initiation of atherosclerosis. Our results suggest that within lesions, macrophages produce mainly intracellular ROS whereas smooth muscle cells produce extracellular ROS. Short term atorvastatin treatment, but not lipid lowering by diet, decreases ROS levels within established advanced lesions; this may help explain the lesion stabilizing and anti-inflammatory effects of long term statin treatment.

No MeSH data available.


Related in: MedlinePlus