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CC-Chemokine Ligand 2 (CCL2) Suppresses High Density Lipoprotein (HDL) Internalization and Cholesterol Efflux via CC-Chemokine Receptor 2 (CCR2) Induction and p42/44 Mitogen-activated Protein Kinase (MAPK) Activation in Human Endothelial Cells *

View Article: PubMed Central - PubMed

ABSTRACT

High density lipoprotein (HDL) has been proposed to be internalized and to promote reverse cholesterol transport in endothelial cells (ECs). However, the mechanism underlying these processes has not been studied. In this study, we aim to characterize HDL internalization and cholesterol efflux in ECs and regulatory mechanisms. We found mature HDL particles were reduced in patients with coronary artery disease (CAD), which was associated with an increase in CC-chemokine ligand 2 (CCL2). In cultured primary human coronary artery endothelial cells and human umbilical vein endothelial cells, we determined that CCL2 suppressed the binding (4 °C) and association (37 °C) of HDL to/with ECs and HDL cellular internalization. Furthermore, CCL2 inhibited [3H]cholesterol efflux to HDL/apoA1 in ECs. We further found that CCL2 induced CC-chemokine receptor 2 (CCR2) expression and siRNA-CCR2 reversed CCL2 suppression on HDL binding, association, internalization, and on cholesterol efflux in ECs. Moreover, CCL2 induced p42/44 mitogen-activated protein kinase (MAPK) phosphorylation via CCR2, and p42/44 MAPK inhibition reversed the suppression of CCL2 on HDL metabolism in ECs. Our study suggests that CCL2 was elevated in CAD patients. CCL2 suppressed HDL internalization and cholesterol efflux via CCR2 induction and p42/44 MAPK activation in ECs. CCL2 induction may contribute to impair HDL function and form atherosclerosis in CAD.

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Related in: MedlinePlus

CCL2 inhibited [3H]cholesterol efflux to HDL/apoA1 in ECs.A, basal cholesterol efflux to HDL/apoA1 in ECs. HCAECs and HUVECs were loaded with [3H]cholesterol (1 Ci/ml) for 24 h. Efflux was initiated by BSA alone and BSA plus 20 μg/ml apoA1 or 50 μg/ml HDL for 2 h. The radioactivity of the medium and cells was measured with a liquid scintillation counter. The cholesterol efflux was expressed as the percentage of counts in the medium relative to the total counts for the medium and cells together. B, dose response of CCL2 on cholesterol efflux to HDL. Cells were initiated by 50 μg/ml human HDL 2 h after treating with increasing doses (0, 20, 40, and 80 ng/ml) of CCL2. The final cholesterol efflux was calculated as the percentage of total [3H]cholesterol released into the medium after subtraction of the values obtained in the absence of HDL. Other procedures are the same as A. C, time course of CCL2 on cholesterol efflux to HDL. Cells were incubated with 50 μg/ml human HDL 2 h after treating with 40 ng/ml CCL2 for indicate times (0, 12, 18, and 24 h). Others were the same as A. D, dose response of CCL2 on cholesterol efflux to apoA1. Cells were initiated by 20 μg/ml human apoA1 2 h after CCL2 treated with increasing doses. Other procedures were the same as A. E, time course of CCL2 on cholesterol efflux to apoA1. Cells were incubated by 20 μg/ml human apoA1 2 h after CCL2 treatment for different times. Other procedures were the same as A. The results were expressed as mean ± S.D. (n = 3). The points represent the averages of three values. #, p < 0.001 compared with the BSA alone in HCAEC; ##, p < 0.001 compared with the BSA alone in HUVEC. *, p < 0.05; **, p < 0.01; ***, p < 0.001 compared with the untreated cells.
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Figure 2: CCL2 inhibited [3H]cholesterol efflux to HDL/apoA1 in ECs.A, basal cholesterol efflux to HDL/apoA1 in ECs. HCAECs and HUVECs were loaded with [3H]cholesterol (1 Ci/ml) for 24 h. Efflux was initiated by BSA alone and BSA plus 20 μg/ml apoA1 or 50 μg/ml HDL for 2 h. The radioactivity of the medium and cells was measured with a liquid scintillation counter. The cholesterol efflux was expressed as the percentage of counts in the medium relative to the total counts for the medium and cells together. B, dose response of CCL2 on cholesterol efflux to HDL. Cells were initiated by 50 μg/ml human HDL 2 h after treating with increasing doses (0, 20, 40, and 80 ng/ml) of CCL2. The final cholesterol efflux was calculated as the percentage of total [3H]cholesterol released into the medium after subtraction of the values obtained in the absence of HDL. Other procedures are the same as A. C, time course of CCL2 on cholesterol efflux to HDL. Cells were incubated with 50 μg/ml human HDL 2 h after treating with 40 ng/ml CCL2 for indicate times (0, 12, 18, and 24 h). Others were the same as A. D, dose response of CCL2 on cholesterol efflux to apoA1. Cells were initiated by 20 μg/ml human apoA1 2 h after CCL2 treated with increasing doses. Other procedures were the same as A. E, time course of CCL2 on cholesterol efflux to apoA1. Cells were incubated by 20 μg/ml human apoA1 2 h after CCL2 treatment for different times. Other procedures were the same as A. The results were expressed as mean ± S.D. (n = 3). The points represent the averages of three values. #, p < 0.001 compared with the BSA alone in HCAEC; ##, p < 0.001 compared with the BSA alone in HUVEC. *, p < 0.05; **, p < 0.01; ***, p < 0.001 compared with the untreated cells.

Mentions: First, we observed that HCAECs had a high basal rate of [3H]cholesterol efflux to BSA compared with HUVECs, and 50 μg/ml HDL increased the [3H]cholesterol efflux from HCAECs and HUVECs by 54.1 and 76.5%, respectively. Nevertheless, 20 μg/ml apoA1 only increased the [3H]cholesterol efflux from HCAECs by 71.7% but did not mediate cholesterol efflux from HUVEC (Fig. 2A). These findings were similar to O'Connell et al. (21), and the reason for this observation may be the low ABCA1 expression levels in HUVECs.


CC-Chemokine Ligand 2 (CCL2) Suppresses High Density Lipoprotein (HDL) Internalization and Cholesterol Efflux via CC-Chemokine Receptor 2 (CCR2) Induction and p42/44 Mitogen-activated Protein Kinase (MAPK) Activation in Human Endothelial Cells *
CCL2 inhibited [3H]cholesterol efflux to HDL/apoA1 in ECs.A, basal cholesterol efflux to HDL/apoA1 in ECs. HCAECs and HUVECs were loaded with [3H]cholesterol (1 Ci/ml) for 24 h. Efflux was initiated by BSA alone and BSA plus 20 μg/ml apoA1 or 50 μg/ml HDL for 2 h. The radioactivity of the medium and cells was measured with a liquid scintillation counter. The cholesterol efflux was expressed as the percentage of counts in the medium relative to the total counts for the medium and cells together. B, dose response of CCL2 on cholesterol efflux to HDL. Cells were initiated by 50 μg/ml human HDL 2 h after treating with increasing doses (0, 20, 40, and 80 ng/ml) of CCL2. The final cholesterol efflux was calculated as the percentage of total [3H]cholesterol released into the medium after subtraction of the values obtained in the absence of HDL. Other procedures are the same as A. C, time course of CCL2 on cholesterol efflux to HDL. Cells were incubated with 50 μg/ml human HDL 2 h after treating with 40 ng/ml CCL2 for indicate times (0, 12, 18, and 24 h). Others were the same as A. D, dose response of CCL2 on cholesterol efflux to apoA1. Cells were initiated by 20 μg/ml human apoA1 2 h after CCL2 treated with increasing doses. Other procedures were the same as A. E, time course of CCL2 on cholesterol efflux to apoA1. Cells were incubated by 20 μg/ml human apoA1 2 h after CCL2 treatment for different times. Other procedures were the same as A. The results were expressed as mean ± S.D. (n = 3). The points represent the averages of three values. #, p < 0.001 compared with the BSA alone in HCAEC; ##, p < 0.001 compared with the BSA alone in HUVEC. *, p < 0.05; **, p < 0.01; ***, p < 0.001 compared with the untreated cells.
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Figure 2: CCL2 inhibited [3H]cholesterol efflux to HDL/apoA1 in ECs.A, basal cholesterol efflux to HDL/apoA1 in ECs. HCAECs and HUVECs were loaded with [3H]cholesterol (1 Ci/ml) for 24 h. Efflux was initiated by BSA alone and BSA plus 20 μg/ml apoA1 or 50 μg/ml HDL for 2 h. The radioactivity of the medium and cells was measured with a liquid scintillation counter. The cholesterol efflux was expressed as the percentage of counts in the medium relative to the total counts for the medium and cells together. B, dose response of CCL2 on cholesterol efflux to HDL. Cells were initiated by 50 μg/ml human HDL 2 h after treating with increasing doses (0, 20, 40, and 80 ng/ml) of CCL2. The final cholesterol efflux was calculated as the percentage of total [3H]cholesterol released into the medium after subtraction of the values obtained in the absence of HDL. Other procedures are the same as A. C, time course of CCL2 on cholesterol efflux to HDL. Cells were incubated with 50 μg/ml human HDL 2 h after treating with 40 ng/ml CCL2 for indicate times (0, 12, 18, and 24 h). Others were the same as A. D, dose response of CCL2 on cholesterol efflux to apoA1. Cells were initiated by 20 μg/ml human apoA1 2 h after CCL2 treated with increasing doses. Other procedures were the same as A. E, time course of CCL2 on cholesterol efflux to apoA1. Cells were incubated by 20 μg/ml human apoA1 2 h after CCL2 treatment for different times. Other procedures were the same as A. The results were expressed as mean ± S.D. (n = 3). The points represent the averages of three values. #, p < 0.001 compared with the BSA alone in HCAEC; ##, p < 0.001 compared with the BSA alone in HUVEC. *, p < 0.05; **, p < 0.01; ***, p < 0.001 compared with the untreated cells.
Mentions: First, we observed that HCAECs had a high basal rate of [3H]cholesterol efflux to BSA compared with HUVECs, and 50 μg/ml HDL increased the [3H]cholesterol efflux from HCAECs and HUVECs by 54.1 and 76.5%, respectively. Nevertheless, 20 μg/ml apoA1 only increased the [3H]cholesterol efflux from HCAECs by 71.7% but did not mediate cholesterol efflux from HUVEC (Fig. 2A). These findings were similar to O'Connell et al. (21), and the reason for this observation may be the low ABCA1 expression levels in HUVECs.

View Article: PubMed Central - PubMed

ABSTRACT

High density lipoprotein (HDL) has been proposed to be internalized and to promote reverse cholesterol transport in endothelial cells (ECs). However, the mechanism underlying these processes has not been studied. In this study, we aim to characterize HDL internalization and cholesterol efflux in ECs and regulatory mechanisms. We found mature HDL particles were reduced in patients with coronary artery disease (CAD), which was associated with an increase in CC-chemokine ligand 2 (CCL2). In cultured primary human coronary artery endothelial cells and human umbilical vein endothelial cells, we determined that CCL2 suppressed the binding (4 &deg;C) and association (37 &deg;C) of HDL to/with ECs and HDL cellular internalization. Furthermore, CCL2 inhibited [3H]cholesterol efflux to HDL/apoA1 in ECs. We further found that CCL2 induced CC-chemokine receptor 2 (CCR2) expression and siRNA-CCR2 reversed CCL2 suppression on HDL binding, association, internalization, and on cholesterol efflux in ECs. Moreover, CCL2 induced p42/44 mitogen-activated protein kinase (MAPK) phosphorylation via CCR2, and p42/44 MAPK inhibition reversed the suppression of CCL2 on HDL metabolism in ECs. Our study suggests that CCL2 was elevated in CAD patients. CCL2 suppressed HDL internalization and cholesterol efflux via CCR2 induction and p42/44 MAPK activation in ECs. CCL2 induction may contribute to impair HDL function and form atherosclerosis in CAD.

No MeSH data available.


Related in: MedlinePlus