Limits...
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

Apo A-I (left panels) and CS-6253 (right panels) desorb phospholipids species from raft and nonraft domains.BHK-ABCA1 cells were labeled with 3[H]choline for 48 h, followed by stimulation with mifepristone for 18–20h as described in “experimental procedures”. Cells were then incubated for 12h with apo A-I or CS-6253 (0.96 μM). After lipid extraction by Folch, 3[H]phosphatidyl choline (3[H]PC) (A and B), or 3[H]sphingomyelin (3[H]SM) (C and D) in each fraction was assessed for radioactivity. Radioactivity appearing in fractions corresponding to raft (1–5) and nonraft (6–10) material was pooled, and desorption of 3[H]PC (A and B inset) and 3[H]SM (C and D inset), from raft versus nonraft in the presence of apo A-I or CS-6253 was expressed as a percentage of control (100%, in the absence of acceptor). 3[H]PC and 3[H]SM distributions in sucrose gradient fractions after 12h incubation with apo A-I or CS-6253 were assessed by TLC respectively. Results shown are representative of three independent experiments. *P < 0.05 by Student's t-test.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4514675&req=5

pone.0131997.g004: Apo A-I (left panels) and CS-6253 (right panels) desorb phospholipids species from raft and nonraft domains.BHK-ABCA1 cells were labeled with 3[H]choline for 48 h, followed by stimulation with mifepristone for 18–20h as described in “experimental procedures”. Cells were then incubated for 12h with apo A-I or CS-6253 (0.96 μM). After lipid extraction by Folch, 3[H]phosphatidyl choline (3[H]PC) (A and B), or 3[H]sphingomyelin (3[H]SM) (C and D) in each fraction was assessed for radioactivity. Radioactivity appearing in fractions corresponding to raft (1–5) and nonraft (6–10) material was pooled, and desorption of 3[H]PC (A and B inset) and 3[H]SM (C and D inset), from raft versus nonraft in the presence of apo A-I or CS-6253 was expressed as a percentage of control (100%, in the absence of acceptor). 3[H]PC and 3[H]SM distributions in sucrose gradient fractions after 12h incubation with apo A-I or CS-6253 were assessed by TLC respectively. Results shown are representative of three independent experiments. *P < 0.05 by Student's t-test.

Mentions: We investigated the role of PM lipid microdomains in the formation of nHDL-CS-6253 at the cellular level. The PM levels of cholesterol and total phospholipids separated by sucrose fractionation were measured by spectroscopy. Addition of apo A-I or CS-6253 for 45 min, 6 h or 12 h to BHK-ABCA1 cells promotes significant lipid desorption from both raft and non-raft domains (S4 and S7 Figs). Incubation with CS-6253 was found to promote significantly cholesterol desorption from both rafts and non-rafts after 45 min respectively (20% CS-6253 vs 25% apo A-I) (S4 and S5 Figs,inset) and only from rafts after 6 h incubation (data not shown). No cholesterol removal was seen following 12 h incubation with CS-6253 in contrary to apo A-I (data not shown). Phospholipids desorption after 45 min incubation was similar for both apo A-I and CS-6253 (S6 and S7 Figs). Moreover, we sought to investigate the effect of CS-6253 (0.96 μM) on PM phospholipids species, i.e phosphatidylcholine (PC) and sphingomyelin (SM) after 12h incubation with cells and using apo A-I as control [35]. Incubation with apo A-I (0.96 μM) was found to promote less PC desorption in PM microdomains than CS-6253 with 18% significant reduction in raft fraction vs. control (p<0.05) (Fig 4A and 4B). In addition, SM were found to be desorbed from both non-raft and raft domains as shown by apo A-I and CS-6253 profile distributions (Fig 4C and 4D). This is consistent with quantification of radioactivity appearing in rafts (fractions 1 to 5) and non-rafts (fractions 6 to 10) expressed as percentage of control (100%, in the absence of apo A-I) for 3[H]PC and 3[H]SM (Fig 4A, 4B, 4C and 4Dinset).


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)

Apo A-I (left panels) and CS-6253 (right panels) desorb phospholipids species from raft and nonraft domains.BHK-ABCA1 cells were labeled with 3[H]choline for 48 h, followed by stimulation with mifepristone for 18–20h as described in “experimental procedures”. Cells were then incubated for 12h with apo A-I or CS-6253 (0.96 μM). After lipid extraction by Folch, 3[H]phosphatidyl choline (3[H]PC) (A and B), or 3[H]sphingomyelin (3[H]SM) (C and D) in each fraction was assessed for radioactivity. Radioactivity appearing in fractions corresponding to raft (1–5) and nonraft (6–10) material was pooled, and desorption of 3[H]PC (A and B inset) and 3[H]SM (C and D inset), from raft versus nonraft in the presence of apo A-I or CS-6253 was expressed as a percentage of control (100%, in the absence of acceptor). 3[H]PC and 3[H]SM distributions in sucrose gradient fractions after 12h incubation with apo A-I or CS-6253 were assessed by TLC respectively. Results shown are representative of three independent experiments. *P < 0.05 by Student's t-test.
© Copyright Policy
Related In: Results  -  Collection

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

pone.0131997.g004: Apo A-I (left panels) and CS-6253 (right panels) desorb phospholipids species from raft and nonraft domains.BHK-ABCA1 cells were labeled with 3[H]choline for 48 h, followed by stimulation with mifepristone for 18–20h as described in “experimental procedures”. Cells were then incubated for 12h with apo A-I or CS-6253 (0.96 μM). After lipid extraction by Folch, 3[H]phosphatidyl choline (3[H]PC) (A and B), or 3[H]sphingomyelin (3[H]SM) (C and D) in each fraction was assessed for radioactivity. Radioactivity appearing in fractions corresponding to raft (1–5) and nonraft (6–10) material was pooled, and desorption of 3[H]PC (A and B inset) and 3[H]SM (C and D inset), from raft versus nonraft in the presence of apo A-I or CS-6253 was expressed as a percentage of control (100%, in the absence of acceptor). 3[H]PC and 3[H]SM distributions in sucrose gradient fractions after 12h incubation with apo A-I or CS-6253 were assessed by TLC respectively. Results shown are representative of three independent experiments. *P < 0.05 by Student's t-test.
Mentions: We investigated the role of PM lipid microdomains in the formation of nHDL-CS-6253 at the cellular level. The PM levels of cholesterol and total phospholipids separated by sucrose fractionation were measured by spectroscopy. Addition of apo A-I or CS-6253 for 45 min, 6 h or 12 h to BHK-ABCA1 cells promotes significant lipid desorption from both raft and non-raft domains (S4 and S7 Figs). Incubation with CS-6253 was found to promote significantly cholesterol desorption from both rafts and non-rafts after 45 min respectively (20% CS-6253 vs 25% apo A-I) (S4 and S5 Figs,inset) and only from rafts after 6 h incubation (data not shown). No cholesterol removal was seen following 12 h incubation with CS-6253 in contrary to apo A-I (data not shown). Phospholipids desorption after 45 min incubation was similar for both apo A-I and CS-6253 (S6 and S7 Figs). Moreover, we sought to investigate the effect of CS-6253 (0.96 μM) on PM phospholipids species, i.e phosphatidylcholine (PC) and sphingomyelin (SM) after 12h incubation with cells and using apo A-I as control [35]. Incubation with apo A-I (0.96 μM) was found to promote less PC desorption in PM microdomains than CS-6253 with 18% significant reduction in raft fraction vs. control (p<0.05) (Fig 4A and 4B). In addition, SM were found to be desorbed from both non-raft and raft domains as shown by apo A-I and CS-6253 profile distributions (Fig 4C and 4D). This is consistent with quantification of radioactivity appearing in rafts (fractions 1 to 5) and non-rafts (fractions 6 to 10) expressed as percentage of control (100%, in the absence of apo A-I) for 3[H]PC and 3[H]SM (Fig 4A, 4B, 4C and 4Dinset).

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