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The biphasic effects of oxidized-low density lipoprotein on the vasculogenic function of endothelial progenitor cells.

Lin FY, Tsao NW, Shih CM, Lin YW, Yeh JS, Chen JW, Nakagami H, Morishita R, Sawamura T, Huang CY - PLoS ONE (2015)

Bottom Line: This study aimed to investigate the concentration-related effects of oxLDL on EPC functions and related angiogenesis, in vitro and in vivo.Higher concentrations of oxLDL (10-50 μg/mL) impaired EPC function via the activation of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase pathways and consequent inhibition of eNOS activity, which could be reversed by anti-oxidants (diphenylene iodonium and apocynin) and gp91phox siRNA.In conclusion, oxLDL has concentration-dependent biphasic effects on human late-outgrowth EPC tube formation in vitro and in vivo.

View Article: PubMed Central - PubMed

Affiliation: Department of Internal Medicine, School of Medicine, College of Medicine, Taipei Medical University, Taipei, Taiwan; Division of Cardiology, Department of Internal Medicine and Cardiovascular Research Center, Taipei Medical University Hospital, Taipei, Taiwan.

ABSTRACT
Late-outgrowth endothelial progenitor cells (EPCs) are stress-resistant and responsible for reparative functions in the cardiovascular system. Oxidized-LDL (oxLDL) plays a critical role in cardiovascular disease pathogenesis. However, it is largely unknown what the impacts of oxLDL are on late-outgrowth EPCs. This study aimed to investigate the concentration-related effects of oxLDL on EPC functions and related angiogenesis, in vitro and in vivo. In this study, early and late-outgrowth EPCs were generated from circulating human mononuclear cells. oxLDL may regulate EPC vasculogenic function via the lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1). Lower concentrations (5 μg/mL) of oxLDL can potentiate EPC tube formation in vitro and in vivo by activating eNOS mechanisms, which are mediated by p38 MAPK- and SAPK/JNK-related pathways. Higher concentrations of oxLDL (10-50 μg/mL) impaired EPC function via the activation of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase pathways and consequent inhibition of eNOS activity, which could be reversed by anti-oxidants (diphenylene iodonium and apocynin) and gp91phox siRNA. In conclusion, oxLDL has concentration-dependent biphasic effects on human late-outgrowth EPC tube formation in vitro and in vivo.

No MeSH data available.


Related in: MedlinePlus

Effects of oxLDL on tube formation in EPCs is mediated by LOX-1.(A and B) EPCs were pretreated with anti-SR-A, anti-SR-B1, or LOX-1 blocking antibodies for 1 hour prior to oxLDL treatment. The effect of oxLDL on EPC neovascularization was analyzed using an in vitro angiogenesis assay. (C) EPCs were treated for 12 hours with 0–50 μg/mL oxLDL, and membrane LOX-1, SR-A, and SR-B1 were analyzed by Western blot. α-tubulin protein levels were used as a loading control. (D) The graph shows the quantification of LOX-1, SR-A, and SR-B1 density in oxLDL-treated EPCs. Data are expressed as the mean ± SEM of three experiments. *p < 0.05 was compared to the control group in the same time treatment and considered significant.
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pone.0123971.g002: Effects of oxLDL on tube formation in EPCs is mediated by LOX-1.(A and B) EPCs were pretreated with anti-SR-A, anti-SR-B1, or LOX-1 blocking antibodies for 1 hour prior to oxLDL treatment. The effect of oxLDL on EPC neovascularization was analyzed using an in vitro angiogenesis assay. (C) EPCs were treated for 12 hours with 0–50 μg/mL oxLDL, and membrane LOX-1, SR-A, and SR-B1 were analyzed by Western blot. α-tubulin protein levels were used as a loading control. (D) The graph shows the quantification of LOX-1, SR-A, and SR-B1 density in oxLDL-treated EPCs. Data are expressed as the mean ± SEM of three experiments. *p < 0.05 was compared to the control group in the same time treatment and considered significant.

Mentions: LOX-1, SR-A and SR-B1 were originally identified on the membrane of vascular endothelial cells and EPCs. Therefore, we investigated whether the potential effects of oxLDL on EPC angiogenesis are mediated by these membrane receptors. Neovascularization was analyzed using an in vitro angiogenesis assay in EPCs incubated with rabbit anti-SR-A, rabbit anti-SR-B1, or LOX-1 blocking antibody (10 μg/mL) for 1 hour followed by 5 or 50 μg/mL of oxLDL treatment. Fig 2A shows that 10 μg/mL of LOX-1 blocking antibody, but not SR-A or SR-B1 blocked the promotion of EPC tube formation capacity under 5 μg/mL of oxLDL treatment. Similar to Fig 1A, addition of 10 μg/mL of LOX-1 blocking antibody may have significantly reduced the impaired tube formation capacity of EPCs treated with 50 μg/mL oxLDL. As a negative control in the competition assay, a nonspecific IgG2α isotype antibody was substituted for the specific antibodies (data not shown). Additionally, we also analyzed membrane LOX-1, SR-A, and SR-B1 production with Western blot of oxLDL-treated EPCs. As shown in Fig 2C, an oxLDL concentration of more than 10 μg/mL may elevate LOX-1 expression in EPCs; in contrast, neither SR-A nor SR-B1 was affected by oxLDL treatment.


The biphasic effects of oxidized-low density lipoprotein on the vasculogenic function of endothelial progenitor cells.

Lin FY, Tsao NW, Shih CM, Lin YW, Yeh JS, Chen JW, Nakagami H, Morishita R, Sawamura T, Huang CY - PLoS ONE (2015)

Effects of oxLDL on tube formation in EPCs is mediated by LOX-1.(A and B) EPCs were pretreated with anti-SR-A, anti-SR-B1, or LOX-1 blocking antibodies for 1 hour prior to oxLDL treatment. The effect of oxLDL on EPC neovascularization was analyzed using an in vitro angiogenesis assay. (C) EPCs were treated for 12 hours with 0–50 μg/mL oxLDL, and membrane LOX-1, SR-A, and SR-B1 were analyzed by Western blot. α-tubulin protein levels were used as a loading control. (D) The graph shows the quantification of LOX-1, SR-A, and SR-B1 density in oxLDL-treated EPCs. Data are expressed as the mean ± SEM of three experiments. *p < 0.05 was compared to the control group in the same time treatment and considered significant.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4446352&req=5

pone.0123971.g002: Effects of oxLDL on tube formation in EPCs is mediated by LOX-1.(A and B) EPCs were pretreated with anti-SR-A, anti-SR-B1, or LOX-1 blocking antibodies for 1 hour prior to oxLDL treatment. The effect of oxLDL on EPC neovascularization was analyzed using an in vitro angiogenesis assay. (C) EPCs were treated for 12 hours with 0–50 μg/mL oxLDL, and membrane LOX-1, SR-A, and SR-B1 were analyzed by Western blot. α-tubulin protein levels were used as a loading control. (D) The graph shows the quantification of LOX-1, SR-A, and SR-B1 density in oxLDL-treated EPCs. Data are expressed as the mean ± SEM of three experiments. *p < 0.05 was compared to the control group in the same time treatment and considered significant.
Mentions: LOX-1, SR-A and SR-B1 were originally identified on the membrane of vascular endothelial cells and EPCs. Therefore, we investigated whether the potential effects of oxLDL on EPC angiogenesis are mediated by these membrane receptors. Neovascularization was analyzed using an in vitro angiogenesis assay in EPCs incubated with rabbit anti-SR-A, rabbit anti-SR-B1, or LOX-1 blocking antibody (10 μg/mL) for 1 hour followed by 5 or 50 μg/mL of oxLDL treatment. Fig 2A shows that 10 μg/mL of LOX-1 blocking antibody, but not SR-A or SR-B1 blocked the promotion of EPC tube formation capacity under 5 μg/mL of oxLDL treatment. Similar to Fig 1A, addition of 10 μg/mL of LOX-1 blocking antibody may have significantly reduced the impaired tube formation capacity of EPCs treated with 50 μg/mL oxLDL. As a negative control in the competition assay, a nonspecific IgG2α isotype antibody was substituted for the specific antibodies (data not shown). Additionally, we also analyzed membrane LOX-1, SR-A, and SR-B1 production with Western blot of oxLDL-treated EPCs. As shown in Fig 2C, an oxLDL concentration of more than 10 μg/mL may elevate LOX-1 expression in EPCs; in contrast, neither SR-A nor SR-B1 was affected by oxLDL treatment.

Bottom Line: This study aimed to investigate the concentration-related effects of oxLDL on EPC functions and related angiogenesis, in vitro and in vivo.Higher concentrations of oxLDL (10-50 μg/mL) impaired EPC function via the activation of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase pathways and consequent inhibition of eNOS activity, which could be reversed by anti-oxidants (diphenylene iodonium and apocynin) and gp91phox siRNA.In conclusion, oxLDL has concentration-dependent biphasic effects on human late-outgrowth EPC tube formation in vitro and in vivo.

View Article: PubMed Central - PubMed

Affiliation: Department of Internal Medicine, School of Medicine, College of Medicine, Taipei Medical University, Taipei, Taiwan; Division of Cardiology, Department of Internal Medicine and Cardiovascular Research Center, Taipei Medical University Hospital, Taipei, Taiwan.

ABSTRACT
Late-outgrowth endothelial progenitor cells (EPCs) are stress-resistant and responsible for reparative functions in the cardiovascular system. Oxidized-LDL (oxLDL) plays a critical role in cardiovascular disease pathogenesis. However, it is largely unknown what the impacts of oxLDL are on late-outgrowth EPCs. This study aimed to investigate the concentration-related effects of oxLDL on EPC functions and related angiogenesis, in vitro and in vivo. In this study, early and late-outgrowth EPCs were generated from circulating human mononuclear cells. oxLDL may regulate EPC vasculogenic function via the lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1). Lower concentrations (5 μg/mL) of oxLDL can potentiate EPC tube formation in vitro and in vivo by activating eNOS mechanisms, which are mediated by p38 MAPK- and SAPK/JNK-related pathways. Higher concentrations of oxLDL (10-50 μg/mL) impaired EPC function via the activation of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase pathways and consequent inhibition of eNOS activity, which could be reversed by anti-oxidants (diphenylene iodonium and apocynin) and gp91phox siRNA. In conclusion, oxLDL has concentration-dependent biphasic effects on human late-outgrowth EPC tube formation in vitro and in vivo.

No MeSH data available.


Related in: MedlinePlus