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Novel Apo E-Derived ABCA1 Agonist Peptide (CS-6253) Promotes Reverse Cholesterol Transport and Induces Formation of preβ-1 HDL In Vitro.

Hafiane A, Bielicki JK, Johansson JO, Genest J - PLoS ONE (2015)

Bottom Line: We have previously reported the effects of compound ATI-5261 on its ability to replicate many functions of native apo A-I in the process of HDL biogenesis.CS-6253 significantly increases cholesterol efflux in murine macrophages and in human THP-1 macrophage-derived foam cells expressing ABCA1.When incubated with human plasma CS-6253 was also found to bind with HDL and LDL and promoted the transfer of cholesterol from HDL to LDL predominantly.

View Article: PubMed Central - PubMed

Affiliation: Cardiovascular Research Laboratories Laboratory, Research Institute of the McGill University Health Centre, Montréal, Québec H4A 3J1, Canada.

ABSTRACT
Apolipoprotein (apo) mimetic peptides replicate some aspects of HDL function. We have previously reported the effects of compound ATI-5261 on its ability to replicate many functions of native apo A-I in the process of HDL biogenesis. ATI-5261 induced muscle toxicity in wild type C57Bl/6 mice, increased CPK, ALT and AST and increase in triglyceride (Tg) levels. Aromatic phenylalanine residues on the non-polar face of ATI-5261, together with positively charged arginine residues at the lipid-water interface were responsible for these effects. This information was used to create a novel analog (CS-6253) that was non-toxic. We evaluated this peptide designed from the carboxyl terminus of apo E, in its ability to mimic apo A-I functionality. Our data shows that the lipidated particles generated by incubating cells overexpressing ABCA1 with lipid free CS-6253 enhances the rate of ABCA1 lipid efflux with high affinity interactions with native ABCA1 oligomeric forms and plasma membrane micro-domains. Interaction between ABCA1 and lipid free CS-6253 resulted in formation of nascent HDL-CS-6253 particles that are actively remodeled in plasma. Mature HDL-CS-6253 particles deliver cholesterol to liver cells via SR-BI in-vitro. CS-6253 significantly increases cholesterol efflux in murine macrophages and in human THP-1 macrophage-derived foam cells expressing ABCA1. Addition of CS-6253 to plasma dose-dependently displaced apo A-I from α-HDL particles and led to de novo formation of preβ-1 HDL that stimulates ABCA1 dependent cholesterol efflux efficiently. When incubated with human plasma CS-6253 was also found to bind with HDL and LDL and promoted the transfer of cholesterol from HDL to LDL predominantly. Our data shows that CS-6253 mimics apo A-I in its ability to promote ABCA1-mediated formation of nascent HDL particles, and enhances formation of preβ-1 HDL with increase in the cycling of apo A-I between the preβ and α-HDL particles in-vitro. These mechanisms are potentially anti-atherogenic.

No MeSH data available.


Related in: MedlinePlus

Competitive binding of CS-6253 peptide to ABCA1 cells expressing ABCA1.BHK-ABCA1 cells were plated in 24-well plates and stimulated for 20 h. Cells were then incubated with 2 μg/ml of 125I-apo A-I for 2 h at 37°C with increasing molar concentrations relative to apo A-I of either CS-6253 peptide, ATI-5261, human apo E, and unlabelled apo A-I (0, 0.0035, 0.017, 0.035, 0.071, 0.17, 0.35, 1.79 μM). Cells were then washed rapidly three times with ice-cold PBS/BSA and then PBS alone. 125I-apo A-I cell associated was determined as described under “Experimental Procedures.” The values shown represent the mean ± S.D from triplicate wells. The 100% of control value measured in the absence of competitors was 0.21 ng of apo A-I /μg cell protein. Values of IC50 shown were determined using the Graph Pad Prism 6 software.
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pone.0131997.g003: Competitive binding of CS-6253 peptide to ABCA1 cells expressing ABCA1.BHK-ABCA1 cells were plated in 24-well plates and stimulated for 20 h. Cells were then incubated with 2 μg/ml of 125I-apo A-I for 2 h at 37°C with increasing molar concentrations relative to apo A-I of either CS-6253 peptide, ATI-5261, human apo E, and unlabelled apo A-I (0, 0.0035, 0.017, 0.035, 0.071, 0.17, 0.35, 1.79 μM). Cells were then washed rapidly three times with ice-cold PBS/BSA and then PBS alone. 125I-apo A-I cell associated was determined as described under “Experimental Procedures.” The values shown represent the mean ± S.D from triplicate wells. The 100% of control value measured in the absence of competitors was 0.21 ng of apo A-I /μg cell protein. Values of IC50 shown were determined using the Graph Pad Prism 6 software.

Mentions: Competition studies were performed to determine the ability of CS-6253 to compete for the binding of 125I-apo A-I to ABCA1 in stimulated BHK-ABCA1. Lipid free CS-6253 inhibited the binding of 125I-apo A-I to ABCA1 as efficiently as apo A-I and apo E, i.e. (IC50 = 0.035±1.66 μM) for apo A-I, (0.057±1.53 μM) for apo E and (0.072±1.41 μM) for CS-6253 (Fig 3). Control experiments were conducted to examine whether the apparent decrease in cell binding of the labeled apo A-I may be attributable to the 125I-apo A-I binding to different competitor particles instead of the cells, as described in experimental procedures (data not shown). No significant amount of 125I-apo A-I was found associated with CS-6253, supporting the results shown in Fig 3. Control data generating IC50 were performed in the same experiment for CS-6253 and ATI-5261, as previously published (7).


Novel Apo E-Derived ABCA1 Agonist Peptide (CS-6253) Promotes Reverse Cholesterol Transport and Induces Formation of preβ-1 HDL In Vitro.

Hafiane A, Bielicki JK, Johansson JO, Genest J - PLoS ONE (2015)

Competitive binding of CS-6253 peptide to ABCA1 cells expressing ABCA1.BHK-ABCA1 cells were plated in 24-well plates and stimulated for 20 h. Cells were then incubated with 2 μg/ml of 125I-apo A-I for 2 h at 37°C with increasing molar concentrations relative to apo A-I of either CS-6253 peptide, ATI-5261, human apo E, and unlabelled apo A-I (0, 0.0035, 0.017, 0.035, 0.071, 0.17, 0.35, 1.79 μM). Cells were then washed rapidly three times with ice-cold PBS/BSA and then PBS alone. 125I-apo A-I cell associated was determined as described under “Experimental Procedures.” The values shown represent the mean ± S.D from triplicate wells. The 100% of control value measured in the absence of competitors was 0.21 ng of apo A-I /μg cell protein. Values of IC50 shown were determined using the Graph Pad Prism 6 software.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4514675&req=5

pone.0131997.g003: Competitive binding of CS-6253 peptide to ABCA1 cells expressing ABCA1.BHK-ABCA1 cells were plated in 24-well plates and stimulated for 20 h. Cells were then incubated with 2 μg/ml of 125I-apo A-I for 2 h at 37°C with increasing molar concentrations relative to apo A-I of either CS-6253 peptide, ATI-5261, human apo E, and unlabelled apo A-I (0, 0.0035, 0.017, 0.035, 0.071, 0.17, 0.35, 1.79 μM). Cells were then washed rapidly three times with ice-cold PBS/BSA and then PBS alone. 125I-apo A-I cell associated was determined as described under “Experimental Procedures.” The values shown represent the mean ± S.D from triplicate wells. The 100% of control value measured in the absence of competitors was 0.21 ng of apo A-I /μg cell protein. Values of IC50 shown were determined using the Graph Pad Prism 6 software.
Mentions: Competition studies were performed to determine the ability of CS-6253 to compete for the binding of 125I-apo A-I to ABCA1 in stimulated BHK-ABCA1. Lipid free CS-6253 inhibited the binding of 125I-apo A-I to ABCA1 as efficiently as apo A-I and apo E, i.e. (IC50 = 0.035±1.66 μM) for apo A-I, (0.057±1.53 μM) for apo E and (0.072±1.41 μM) for CS-6253 (Fig 3). Control experiments were conducted to examine whether the apparent decrease in cell binding of the labeled apo A-I may be attributable to the 125I-apo A-I binding to different competitor particles instead of the cells, as described in experimental procedures (data not shown). No significant amount of 125I-apo A-I was found associated with CS-6253, supporting the results shown in Fig 3. Control data generating IC50 were performed in the same experiment for CS-6253 and ATI-5261, as previously published (7).

Bottom Line: We have previously reported the effects of compound ATI-5261 on its ability to replicate many functions of native apo A-I in the process of HDL biogenesis.CS-6253 significantly increases cholesterol efflux in murine macrophages and in human THP-1 macrophage-derived foam cells expressing ABCA1.When incubated with human plasma CS-6253 was also found to bind with HDL and LDL and promoted the transfer of cholesterol from HDL to LDL predominantly.

View Article: PubMed Central - PubMed

Affiliation: Cardiovascular Research Laboratories Laboratory, Research Institute of the McGill University Health Centre, Montréal, Québec H4A 3J1, Canada.

ABSTRACT
Apolipoprotein (apo) mimetic peptides replicate some aspects of HDL function. We have previously reported the effects of compound ATI-5261 on its ability to replicate many functions of native apo A-I in the process of HDL biogenesis. ATI-5261 induced muscle toxicity in wild type C57Bl/6 mice, increased CPK, ALT and AST and increase in triglyceride (Tg) levels. Aromatic phenylalanine residues on the non-polar face of ATI-5261, together with positively charged arginine residues at the lipid-water interface were responsible for these effects. This information was used to create a novel analog (CS-6253) that was non-toxic. We evaluated this peptide designed from the carboxyl terminus of apo E, in its ability to mimic apo A-I functionality. Our data shows that the lipidated particles generated by incubating cells overexpressing ABCA1 with lipid free CS-6253 enhances the rate of ABCA1 lipid efflux with high affinity interactions with native ABCA1 oligomeric forms and plasma membrane micro-domains. Interaction between ABCA1 and lipid free CS-6253 resulted in formation of nascent HDL-CS-6253 particles that are actively remodeled in plasma. Mature HDL-CS-6253 particles deliver cholesterol to liver cells via SR-BI in-vitro. CS-6253 significantly increases cholesterol efflux in murine macrophages and in human THP-1 macrophage-derived foam cells expressing ABCA1. Addition of CS-6253 to plasma dose-dependently displaced apo A-I from α-HDL particles and led to de novo formation of preβ-1 HDL that stimulates ABCA1 dependent cholesterol efflux efficiently. When incubated with human plasma CS-6253 was also found to bind with HDL and LDL and promoted the transfer of cholesterol from HDL to LDL predominantly. Our data shows that CS-6253 mimics apo A-I in its ability to promote ABCA1-mediated formation of nascent HDL particles, and enhances formation of preβ-1 HDL with increase in the cycling of apo A-I between the preβ and α-HDL particles in-vitro. These mechanisms are potentially anti-atherogenic.

No MeSH data available.


Related in: MedlinePlus