Degradation of Phospholipid Transfer Protein (PLTP) and PLTP-generated Pre-β-high Density Lipoprotein by Mast Cell Chymase Impairs High Affinity Efflux of Cholesterol from Macrophage Foam Cells

Abstract

Human atherosclerotic lesions contain mast cells filled with the neutral protease chymase. Here we studied the effect of human chymase on (i) phospholipid transfer protein (PLTP)-mediated phospholipid (PL) transfer activity, and (ii) the ability of PLTP to generate pre-␤-high density lipoprotein (HDL). Immunoblot analysis of PLTP after incubation with chymase for 6 h revealed, in addition to the original 80-kDa band, four specific proteolytic fragments of PLTP with approximate molecular masses of 70, 52, 48, and 31 kDa. This specific pattern of PLTP degradation remained stable for at least 24 h of incubation with chymase. Such proteolyzed PLTP had reduced ability (i) to transfer PL from liposome donor particles to acceptor HDL 3 particles, and (ii) to facilitate the formation of pre-␤-HDL. However, when PLTP was incubated with chymase in the presence of HDL 3 , only one major cleavage product of PLTP (48 kDa) was generated, and PL transfer activity was almost fully preserved. Moreover, chymase effectively depleted the pre-␤-HDL particles generated from HDL 3 by PLTP and significantly inhibited the high affinity component of cholesterol efflux from macrophage foam cells. These results suggest that the mast cells in human atherosclerotic lesions, by secreting chymase, may prevent PLTP-dependent formation of pre-␤-HDL particles from HDL 3 and so impair the anti-atherogenic function of PLTP.Reverse cholesterol transport is a physiologically important route for removal of excess cholesterol from the membranes of peripheral cells and its transport to the liver for secretion into the bile. This process is of special importance for macrophages, which, having no ability to regulate incoming cholesterol, are readily transformed into cholesterol-loaded foam cells. Several factors, such as apolipoproteins, lipolytic enzymes, lipid transfer proteins, and lipoprotein receptors, are involved in reverse cholesterol transport (1). Initially, efflux of cellular cholesterol is promoted by extracellular cholesterol acceptors. The primary and most efficient acceptors of cholesterol seem to be small discoidal lipid-poor pre-␤-migrating high density lipoproteins (pre-␤-HDL) 1 (2). These particles interact with the cell membrane through a microsolubilization process that leads to uptake first of phospholipids and then of cholesterol from the membrane (3). This process appears to be controlled by the ATP-binding cassette transporter protein 1 on the macrophage cell membrane (4), its transcription being regulated in part by the orphan nuclear receptor LXR. Activation of LXR in macrophages increases not only ATP-binding cassette transporter A1 but also the expression of genes encoding ABCG1 and apoE (5, 6), which may be also involved in efflux of cholesterol toward HDL. Recently, it was shown (7) that LXR activation in macrophages also up-regulates phospholipid transfer protein (PLTP) expression, but this up-regulation does not have any direct effect on cholesterol efflux from mouse peritoneal macrophages.PLTP contributes to the remodelin...

Depletion of Pre-β-high Density Lipoprotein by Human Chymase Impairs ATP-binding Cassette Transporter A1- but Not Scavenger Receptor Class B Type I-mediated Lipid Efflux to High Density Lipoprotein

“…6). These data, using a system in which ABCA1-dependent efflux dominates, confirmed that the ability of HDL 3 to stimulate cholesterol efflux from cholesterol-loaded macrophages is increased after PLTP treatment (39). More importantly, using HDL 3 preparations with graded pre-␤ content it could be demonstrated that the rate of cholesterol efflux from the cells varied in parallel with the content of pre-␤ particles in HDL 3 (Fig.…”

“…To increase the content of pre-␤ particles, HDL 3 was preincubated for 24 h at 37°C in buffer alone or with PLTP, demonstrated previously to promote the size conversion of HDL particles and to generate pre-␤-HDL (38). As reported previously (39), PLTP treatment of HDL 3 for 24 h increased the content of pre-␤-HDL in the HDL 3 preparation (in this experiment from 9 to 18%; Fig. 5, A and B, left).…”

“…In line with this observation we have found that the apoA-I molecules on the surface of the globular ␣-migrating HDL remain biologically active also in terms of HDL remodeling. Indeed, chymase cleavage of apoA-I in the ␣-migrating HDL does not abolish its ability to generate PLTP-dependent pre-␤-migrating particles (39) or to undergo cholesterol esterification by lecithin-cholesterol acyltransferase (44).…”

Association of cholesteryl ester transfer protein with HDL particles reduces its proteolytic inactivation by mast cell chymase

“…Furthermore, studies in vitro have demonstrated that chymase specifically depletes preb-HDL particles (26), thus blocking cholesterol efflux mediated by the ABCA1 pathway (27). In contrast, limited proteolysis of HDL 3 particles by chymase does not impair their ability to activate LCAT (28) or to act as phospholipid acceptors in PLTP-mediated phospholipid transfer reactions (29). These observations reveal that the spherical a-HDL particles primarily remain functional even after being exposed to chymase.…”

“…These observations reveal that the spherical a-HDL particles primarily remain functional even after being exposed to chymase. Chymase also degrades PLTP, thereby reducing its phospholipid transfer activity (29). However, it is not known whether chymase is able to degrade CETP, its structurally closely related lipid transfer protein.…”