Two adjacent domains (141–150 and 151–160) of apoE covalently linked to a class A amphipathic helical peptide exhibit opposite atherogenic effects

Abstract

Objective We recently described anti-atherogenic properties of the dual domain peptide Ac-hE18A-NH2 derived by covalently linking the heparin binding domain 141-150 of apoE to 18A, a class A amphipathic helical peptide. In this paper we have compared the properties of Ac-hE18A-NH2 with the non-heparin binding 151-160 region of apoE linked to 18A (Ac-nhE18A-NH2). Methods and Results Both peptides were highly helical in solution and in association with lipids. Ac-hE18A-NH2 and not Ac-nhE18A-NH2 enhanced uptake of low density lipoprotein (LDL) in HepG2 cells. While Ac-hE18A-NH2 retarded the electrophoretic mobility of LDL, Ac-nhE18A-NH2 slightly enhanced mobility. Ac-hE18A-NH2 reduced monocyte association with endothelial cells, while Ac-nhE18A-NH2 increased it. Ac-hE18A-NH2 also reduced lipid hydroperoxide content of LDL while Ac-nhE18A-NH2 increased it. A single administration of Ac-hE18A-NH2 (100 μg/mouse) into apoE null mice dramatically reduced cholesterol (from 600 mg/dL to 180 mg/dL at five min. and to 60 mg/dL at five h) while Ac-nhE18A-NH2 had no effect. Administration (100μg/mouse/day, three days a week) into apoE null mice for six weeks showed Ac-hE18A-NH2 group having a moderate aortic sinus lesion reduction compared with the control group (−15.1%), while the Ac-nhE18A-NH2 administered group had increased lesion area (+33.0% vs controls and 36.1% vs Ac-hE18A-NH2). Plasma from mice administered Ac-hE18A-NH2 for six weeks showed a significant reduction in plasma cholesterol and triglyceride levels and increase in paraoxonase-1 (PON-1) activity compared to controls, while Ac-nhE18A-NH2 caused no change in plasma cholesterol and decreased PON-1 activity. Conclusion It is proposed that Ac-hE18A-NH2 reduced lesion progression in apoE null mice due to its anti-inflammatory and lipoprotein clearing properties, while Ac-nhE18A-NH2 exhibited pro-atherogenic effects.

A novel hemoglobin-binding peptide reduces cell-free hemoglobin in murine hemolytic anemia

“…These clearance data are similar to those reported in previous studies (17,20), where the hE peptide was linked to 18A to form the lipoprotein-clearing peptide hE-18A. Additionally, it has been previously demonstrated that peptides containing the hE sequence traffic to the liver (40). Consistent with these findings, FAM-labeled hEHb-B10 was detected in the livers of treated mice by fluorescent histological examination (Fig.…”

“…During peptide synthesis, Hb-B10 was covalently linked to a small fragment of ApoE (hE, residues 141-150) that has been shown to facilitate both the uptake of lipoproteins by small peptides and drug transport by liposomes via endocytic clearance through the ubiquitous heparan sulfate proteoglycan-associated pathway (14,47,51). Although modification of any small peptide might affect function, we found that the pharmacokinetic profile and liver localization of hE-Hb-B10 were very similar to previous reports for other hE-conjugated peptides (17,20,40).…”

“…In addition, both hE-Hb-B10 and hE-18A are taken up by the liver, and the plasma clearance kinetics of both peptides are similar (17,40). These data imply a common uptake and clearance mechanism for hE18A and hE-Hb-B10 via the heparan-sulfate proteoglycan-associated pathway of the liver (17,20).…”

“…Fluorescence in other tissues (the aorta, brain, lung, kidney, and spleen) was indistinguishable from background fluorescence. This suggests that hE-Hb-B10 likely traffics CF-Hb to the liver for clearance, similar to other hE-linked peptides (17,40).…”

Sidedness of interfacial arginine residues and anti-atherogenicity of apolipoprotein A-I mimetic peptides

“…At the end of the study protocol, animals were anesthetized using ketamine-xylazine and blood was collected by cardiac puncture. Quantifi cation of lesion area was performed as described previously ( 16,17 ). All animal studies were approved by the Institutional Animal Care and Use Committee of the University of Alabama at Birmingham.…”