Limits...
Characterization of Proliferating Lesion ‐ Resident Cells During All Stages of Atherosclerotic Growth

View Article: PubMed Central - PubMed

ABSTRACT

Background: Monocyte recruitment leads to accumulation of macrophage foam cells and contributes to atherosclerotic lesion growth. Recent studies have reported that lesion‐resident macrophages can proliferate and represent a major cellular component during lesion development. This study was designed to assess whether the rate of macrophage proliferation changes during well‐established stages of lesion growth and to characterize other populations of proliferating cells within these lesions.

Methods and results: Using murine models of atherosclerosis (Apoe−/− and LDLr−/− mice) and human coronary artery lesions, in situ proliferation of lesion‐resident cells at different stages of growth was assessed by staining for Ki67 and bromodeoxyuridine (BrdU). In early lesions, close to half of all actively growing macrophages were proliferating in situ. BrdU pulse labeling allowed for accurate identification of in situ proliferating macrophages compared to those derived from monocyte recruitment. Local macrophage proliferation declined as lesions advanced. Interestingly, intimal inflammatory cell infiltrates containing proliferating T lymphocytes were identified during the active phase of lesion growth and correlated with apoptotic cell death. Inflammatory cell infiltrates were completely resolved in advanced lesions and replaced with the necrotic core.

Conclusions: Our findings indicate that atherosclerotic lesions contain locally proliferating macrophages primarily during early and intermediate stages of lesion growth. Furthermore, T‐lymphocyte‐enriched inflammatory cell infiltrates represent a novel subset of proliferating cells within the atherosclerotic lesion that correlate with apoptosis and precede the necrotic core. These findings have novel implications in understanding the pathogenesis of atherosclerosis and may implicate proliferating T lymphocytes as a contributing factor to lesion progression and stability.

No MeSH data available.


Related in: MedlinePlus

Detection of proliferating cells during murine and human atherosclerotic lesion growth. Ki67 immunohistochemistry and H&E staining in Apoe−/− mice on chow diet identified proliferating cells during various stages of lesion progression, ranging from initial macrophage infiltration to advanced lesions (A). In the initial and early stage, macrophages below the endothelium, as well as foam cells in fatty streaks, were Ki67 positive (Initial and Early, arrows). At 24 weeks of age, in lesions of the intermediate type with large necrotic cores, few Ki67‐positive cells were observed (Intermediate, arrows). At 55 weeks of age, lesions were mostly necrotic with very few Ki67‐positive cells (Advanced). Human lesions from coronary arteries stained with Ki67 contained some Ki67‐positive cells in areas rich in macrophages (B, arrows). Bar=50 μm. H&E indicates hematoxylin and eosin.
© Copyright Policy - creativeCommonsBy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC5015311&req=5

jah31705-fig-0001: Detection of proliferating cells during murine and human atherosclerotic lesion growth. Ki67 immunohistochemistry and H&E staining in Apoe−/− mice on chow diet identified proliferating cells during various stages of lesion progression, ranging from initial macrophage infiltration to advanced lesions (A). In the initial and early stage, macrophages below the endothelium, as well as foam cells in fatty streaks, were Ki67 positive (Initial and Early, arrows). At 24 weeks of age, in lesions of the intermediate type with large necrotic cores, few Ki67‐positive cells were observed (Intermediate, arrows). At 55 weeks of age, lesions were mostly necrotic with very few Ki67‐positive cells (Advanced). Human lesions from coronary arteries stained with Ki67 contained some Ki67‐positive cells in areas rich in macrophages (B, arrows). Bar=50 μm. H&E indicates hematoxylin and eosin.

Mentions: Paraffin sections of the aortic root from female Apoe−/− mice at different stages of atherosclerosis were stained with hematoxylin and eosin (H&E) to assess lesion growth and gross cellular morphology (Figure 1A, upper panel). Sections were also stained for Ki67, a well‐established marker of active cell division,22 to assess the presence of proliferating cells in these different stages of atherosclerosis (Figure 1A, lower panel). At 8 weeks of age, designated the initial stage, the first intimal macrophages were identified in the subendothelial space. At 12 weeks of age, designated the early lesion group, intimal fatty streaks consisting of lipid‐enriched macrophage foam cells were observed. Consistent with these findings, lipid‐enriched macrophage foam cells stained intensely for filipin, a marker of free cholesterol, and Oil Red O (ORO), a marker of neutral lipids (Figure S1). This early stage of lesion growth showed no evidence of apoptotic cell death nor the appearance of a lipid‐enriched necrotic core, consistent with previous findings.8 At 24 weeks of age, designated the intermediate stage of lesion growth, the intima consisted of a necrotic lipid core with cellular debris and a cellular/fibrous cap. Stretches of simple fatty streaks or individual intimal macrophages were also observed adjacent to these complex lesions (not shown). Filipin and ORO staining was markedly increased in both the necrotic regions and adjacent intact foam cells (Figure S1). At 55 weeks, designated the advanced stage of lesion growth, the lesions were relatively acellular as defined by lack of nuclei, comprised of only a few intact viable cells.


Characterization of Proliferating Lesion ‐ Resident Cells During All Stages of Atherosclerotic Growth
Detection of proliferating cells during murine and human atherosclerotic lesion growth. Ki67 immunohistochemistry and H&E staining in Apoe−/− mice on chow diet identified proliferating cells during various stages of lesion progression, ranging from initial macrophage infiltration to advanced lesions (A). In the initial and early stage, macrophages below the endothelium, as well as foam cells in fatty streaks, were Ki67 positive (Initial and Early, arrows). At 24 weeks of age, in lesions of the intermediate type with large necrotic cores, few Ki67‐positive cells were observed (Intermediate, arrows). At 55 weeks of age, lesions were mostly necrotic with very few Ki67‐positive cells (Advanced). Human lesions from coronary arteries stained with Ki67 contained some Ki67‐positive cells in areas rich in macrophages (B, arrows). Bar=50 μm. H&E indicates hematoxylin and eosin.
© Copyright Policy - creativeCommonsBy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC5015311&req=5

jah31705-fig-0001: Detection of proliferating cells during murine and human atherosclerotic lesion growth. Ki67 immunohistochemistry and H&E staining in Apoe−/− mice on chow diet identified proliferating cells during various stages of lesion progression, ranging from initial macrophage infiltration to advanced lesions (A). In the initial and early stage, macrophages below the endothelium, as well as foam cells in fatty streaks, were Ki67 positive (Initial and Early, arrows). At 24 weeks of age, in lesions of the intermediate type with large necrotic cores, few Ki67‐positive cells were observed (Intermediate, arrows). At 55 weeks of age, lesions were mostly necrotic with very few Ki67‐positive cells (Advanced). Human lesions from coronary arteries stained with Ki67 contained some Ki67‐positive cells in areas rich in macrophages (B, arrows). Bar=50 μm. H&E indicates hematoxylin and eosin.
Mentions: Paraffin sections of the aortic root from female Apoe−/− mice at different stages of atherosclerosis were stained with hematoxylin and eosin (H&E) to assess lesion growth and gross cellular morphology (Figure 1A, upper panel). Sections were also stained for Ki67, a well‐established marker of active cell division,22 to assess the presence of proliferating cells in these different stages of atherosclerosis (Figure 1A, lower panel). At 8 weeks of age, designated the initial stage, the first intimal macrophages were identified in the subendothelial space. At 12 weeks of age, designated the early lesion group, intimal fatty streaks consisting of lipid‐enriched macrophage foam cells were observed. Consistent with these findings, lipid‐enriched macrophage foam cells stained intensely for filipin, a marker of free cholesterol, and Oil Red O (ORO), a marker of neutral lipids (Figure S1). This early stage of lesion growth showed no evidence of apoptotic cell death nor the appearance of a lipid‐enriched necrotic core, consistent with previous findings.8 At 24 weeks of age, designated the intermediate stage of lesion growth, the intima consisted of a necrotic lipid core with cellular debris and a cellular/fibrous cap. Stretches of simple fatty streaks or individual intimal macrophages were also observed adjacent to these complex lesions (not shown). Filipin and ORO staining was markedly increased in both the necrotic regions and adjacent intact foam cells (Figure S1). At 55 weeks, designated the advanced stage of lesion growth, the lesions were relatively acellular as defined by lack of nuclei, comprised of only a few intact viable cells.

View Article: PubMed Central - PubMed

ABSTRACT

Background: Monocyte recruitment leads to accumulation of macrophage foam cells and contributes to atherosclerotic lesion growth. Recent studies have reported that lesion‐resident macrophages can proliferate and represent a major cellular component during lesion development. This study was designed to assess whether the rate of macrophage proliferation changes during well‐established stages of lesion growth and to characterize other populations of proliferating cells within these lesions.

Methods and results: Using murine models of atherosclerosis (Apoe−/− and LDLr−/− mice) and human coronary artery lesions, in situ proliferation of lesion‐resident cells at different stages of growth was assessed by staining for Ki67 and bromodeoxyuridine (BrdU). In early lesions, close to half of all actively growing macrophages were proliferating in situ. BrdU pulse labeling allowed for accurate identification of in situ proliferating macrophages compared to those derived from monocyte recruitment. Local macrophage proliferation declined as lesions advanced. Interestingly, intimal inflammatory cell infiltrates containing proliferating T lymphocytes were identified during the active phase of lesion growth and correlated with apoptotic cell death. Inflammatory cell infiltrates were completely resolved in advanced lesions and replaced with the necrotic core.

Conclusions: Our findings indicate that atherosclerotic lesions contain locally proliferating macrophages primarily during early and intermediate stages of lesion growth. Furthermore, T‐lymphocyte‐enriched inflammatory cell infiltrates represent a novel subset of proliferating cells within the atherosclerotic lesion that correlate with apoptosis and precede the necrotic core. These findings have novel implications in understanding the pathogenesis of atherosclerosis and may implicate proliferating T lymphocytes as a contributing factor to lesion progression and stability.

No MeSH data available.


Related in: MedlinePlus