Interaction of free apolipoproteins with macrophages. Formation of high density lipoprotein-like lipoproteins and reduction of cellular cholesterol.

The properties of the mature and pro-forms of recombinant apolipoprotein A-I (apoA-I) were compared with those of apoA-I isolated from human plasma. When the synthesis and secretion of pro-and mature forms of apoA-I from a baculovirus/insect cell expression system were compared in parallel experiments, the amount of the pro-form of apoA-I synthesized and secreted was severalfold higher than that of the mature form of apoA-I. A comparison of the properties of the pro-and mature forms of recombinant apoA-I and human plasma apoA-I showed no difference between all three in their secondary structure, their ability to self-associate, lipid-binding capacity, lecithin: cholesterol acyltransferase activation, and binding to the phospholipid transfer protein. The properties of reconstituted high density lipoprotein (HDL) particles formed from the proteins and their ability to promote cholesterol and phospholipid efflux from human skin fibroblasts were also similar. However, their ability to bind to plasma HDL subfractions differed, because twice as much proapoA-I associated with pre ␤ 1 -HDL and pre ␤ 2 -HDL subfractions compared with both mature recombinant and plasma apoA-I. Correspondingly, the amount of proapoA-I in ␣ -HDL subfractions, especially in ␣ 1 -HDL and ␣ 2 -HDL, was decreased. We conclude that while the propeptide of apoA-I is required for the effective synthesis and secretion of apoA-I, cleavage of this peptide is a requisite for the effective interconversion of HDL subfractions. -Sviridov, D., L. E. Pyle, M. Jauhiainen, C. Ehnholm, and N. H. Fidge. Deletion of the propeptide of apolipoprotein A-I reduces protein expression, but stimulates effective conversion of pre ␤ -high density lipoprotein to ␣ -high density lipoprotein.

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Deletion of the propeptide of apolipoprotein A-I reduces protein expression but stimulates effective conversion of preβ-high density lipoprotein to α-high density lipoprotein

Sviridov

Pyle

Jauhiainen

et al. 2000

“…3-6). It is interesting that the rate of microsolubilization of plasma membrane FC and PL by apoA-I is enhanced when the cells are cholesterol-enriched (14,(24)(25)(26). However, the underlying mechanism of membrane microsolubilization is apparently independent of the level of cholesterol in fibroblasts (K. L. Gillotte, S. Lund-Katz, G. H. Rothblat, and M. C. Phillips, unpublished results).…”

Section: Pre-␤-hdl-mediated Membrane Microsolubilizationmentioning

confidence: 98%

“…In contrast to the situation with efflux of cellular FC to fully lipidated apoA-I particles where the aqueous diffusion mechanism is largely understood, the mechanism of efflux mediated by incompletely lipidated apolipoprotein molecules is not well defined. It is known that lipid-free apolipoproteins access both cellular FC and phospholipid (PL) (23)(24)(25)(26)(27). Although various mechanisms of simultaneous or sequential removal of these lipids have been proposed, this aspect of the pathway has not been precisely investigated.…”

mentioning

confidence: 99%

Removal of cellular cholesterol by pre-β-HDL involves plasma membrane microsolubilization

Gillotte

Davidson

Lund‐Katz

et al. 1998

High density lipoprotein (HDL) is able to remove unesterified cholesterol from peripheral cells in the process of reverse cholesterol transport by an aqueous diffusion mechanism as well as by an apolipoprotein (apo)-mediated process. The aqueous diffusion mechanism is understood but the molecular mechanism of lipid-poor pre-␤ -HDL-(apo-) mediated cholesterol removal is not known. Measurements of the initial rates of efflux of unesterified cholesterol and phospholipid from human fibroblasts to lipid-free, human apoA-I showed that both lipids are released from the cells during a 10-min incubation with apoA-I. The concentration-dependence of efflux of the lipids is the same ( K m ‫؍‬ 0.4 and 0.6 g apoA-I/ml for cholesterol and phospholipid flux, respectively), suggesting a membrane microsolubilization process. A finite pool of about 1% of the plasma membrane cholesterol is accessible for release by solubilization; the limited size of this cholesterol pool is not due to a lack of availability of apoA-I, but rather to the restricted amount of phospholipid that is removed from the plasma membrane.Plasma membrane domains may be involved in membrane microsolubilization, but caveolar cholesterol seems not to be specifically accessed in this process. Membrane microsolubilization is the process by which pre-␤ 1-HDL removes cell cholesterol in the first step of reverse cholesterol transport. When apoA-I is present in the extracellular space, the relative contributions of cholesterol efflux by membrane microsolubilization and by aqueous diffusion are determined by the degree of lipidation of the apoA-I molecules.-Gillotte, K.

“…This type of cholesterol efflux occurs independently of LCAT and in parallel with phospholipid efflux. It is sensitive to the treatment of cells with proteases and modulated by activation or inhibition of protein kinase C (4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15)(16). Therefore, it was suggested that cholesterol efflux mediated by lipid-free apoA-I depends on specific protein-cell surface interactions.…”

mentioning

confidence: 99%

Plasma and fibroblasts of Tangier disease patients are disturbed in transferring phospholipids onto apolipoprotein A-I

Eckardstein

Chirazi

Schuler-Lüttmann

et al. 1998

Plasmas of patients with Tangier disease (TD) lack lipid-rich ␣ -HDL which, in normal plasma, constitutes the majority of high density lipoprotein (HDL). Residual amounts of apolipoprotein (apo)A-I in TD plasma occur as lipid-poor or even lipid-free pre ␤ -HDL. By contrast to normal plasma, TD plasma does not convert pre ␤ -HDL into ␣ -HDL. Moreover, fibroblasts of TD patients were found to be defective in secreting cholesterol or phospholipids in the presence of lipid-free apoA-I. We have therefore hypothesized that both defective conversion of pre ␤ -HDL into ␣ -HDL and defective lipid efflux from TD cells onto lipidfree apoA-I result from a disturbance in phospholipid transfer occurring in both cellular and extracellular compartments. To test this hypothesis we established an assay that measures the activity of plasma, cells, and cell culture media to transfer radiolabeled phosphatidylcholine (PC), phosphatidylethanolamine (PE), and phosphatidylinositol (PI) from vesicles onto apoA-I, apoA-II, albumin, or reconstituted HDL. Plasmas, HDL, and lipoprotein-depleted plasma of normolipidemic probands as well as cell homogenates and culture media of normal fibroblasts were active at 37 Њ C but not at 4 Њ C in transferring radiolabeled PC, PI, and PE dose-and time-dependently onto either lipid-free apoA-I or reconstituted HDL. Transfer of glycerophospholipids onto apoA-II was much lower than onto apoA-I; transfer onto albumin was close to background. Compared to ten normolipidemic plasmas and four apoA-I-deficient plasmas, plasmas of six TD patients were significantly reduced by 40-50% in their glycerophospholipid transfer activities. Compared to eight normal fibroblast cell lines, homogenates and culture media of four TD fibroblast cell lines were reduced by 40-50% and 30-35%, respectively, in their activity to transfer PC, PI, or PE onto apoA-I. Our data suggest that in TD the same mechanism underlies both defective conversion of pre ␤ -HDL into ␣ -HDL and impaired efflux of cellular lipids, namely a defective phospholipid transfer.von Eckardstein, A