Genetics and molecular biology: phospholipid transfer protein in atherogenesis

Human phospholipid transfer protein (PLTP) is a 476 amino acid secreted protein that catalyzes the transfer of phospholipids and vitamin E between lipoprotein particles [1,2•,3•]. PLTP plays a key role in the formation and homeostasis of high-density lipoprotein (HDL) particles; it transfers phospholipids from very-low-density lipoprotein (VLDL) surface remnants to HDL particles, it facilitates the interconversion of HDL subclasses, and generates lipid-poor preβ-HDL particles. PLTP is synthesized by various cell types but its primary function is in the plasma where it is transported in association with apolipoproteins. Plasma PLTP exists in two forms: a high activity form that co-purifies with apolipoprotein (apo) E and a low activity form associated with apoA-I. It was reported recently that both apoE and apoA-IV can activate PLTP [4•].

PLTP has both pro- and antiatherogenic characteristics. Preβ-HDL particles, a major product of PLTP catalysis, are required as acceptors for the atheroprotective cholesterol efflux pathway [5]. Studies in human subjects and genetically engineered mice indicate that plasma PLTP activity is proatherogenic. Human subjects with cardiovascular disease (CVD) have higher plasma PLTP activity compared with normal subjects [6]. Similarly, obesity and type 2 diabetes are coincident with high PLTP activity [7,8]. Overexpression of PLTP in LDLR^−/− mice results in a reduction in plasma HDL and apoA-I levels, an increase in VLDL secretion, and an increase in susceptibility to atherosclerosis despite lower apoB-containing lipoproteins [9]. Plasma HDL levels are also decreased in PLTP knock-out mice [10]. These mice, however, show a dramatic reduction in apoB synthesis and a significantly reduced susceptibility to diet-induced atherosclerosis.

Several recent studies were aimed at better understanding the pro- and antiatherogenic mechanisms of PLTP activity. The current consensus is that systemic or plasma PLTP is proatherogenic, but macrophage PLTP is atheroprotective. Support for a proatherogenic role of plasma PLTP was obtained from a recent study investigating the relationship between PLTP activity and various inflammatory markers in humans with CVD [11•]. PLTP activity was positively and independently correlated with the main inflammatory marker C-reactive protein (CRP). Positive association of PLTP was also seen with other inflammation markers including serum amyloid A protein (SAA) and interleukin-6. Cardiovascular risk factors including body mass index, insulin resistance, apoE (VLDL), and apoB levels were directly correlated with PLTP activity as well as with CRP and SAA levels. These results clearly indicate that PLTP activity is important in both inflammation and CVD. Another marker of CVD, coronary artery intima-media thickness (IMT), was found to be independently associated with PLTP activity in diabetics, but not in nondiabetic subjects [12•]. ApoE may play a key role in the pathophysiological effects of PLTP. In diabetic subjects, the level of active plasma PLTP appeared to be modulated by apoE [13•]. PLTP activity showed a strong positive correlation with plasma apoB and apoE levels, but was inversely related to apoA-I levels [13•]. As the active form of PLTP associates with apoE and the inactive form with apoA-I, these results suggest that apoE may be capable of converting inactive PLTP into the active form. The relationship between apoE and PLTP was further investigated in diabetic subjects of the Diabetes Atorvastatin Lipid Intervention (DALI) study [14•]. Atorvastatin treatment significantly reduced plasma apoE levels with a concomitant decrease in plasma PLTP activity. In a multivariate model, the change in apoE levels was the only determinant of the reduction in PLTP activity. Thus, apoE levels appear to modulate plasma PLTP activity.

The main function of PLTP is to modify and modulate HDL levels. Thus, plasma apoA-I and HDL levels are reduced in both PLTP over-expressing transgenic mice and in PLTP knock-out mice [9,10]. As the liver is a primary source of plasma apoA-I, the hepatic synthesis and secretion of apoA-I was compared in wild-type and PLTP-deficient mice (PLTP-KO) [15••]. The level of apoA-I synthesis was not compromised in hepatocytes from PLTP knock-out mice. The lipidation of apoA-I was, however, defective in PLTP-KO hepatocytes such that the secreted particle contained fewer phospholipids and an altered composition of phosphatidylcholine, sphingomyelin, and fatty acyl groups. This phospholipid-deficient apoA-I particle was less stable than the normally lipidated apoA-I particle derived from wild-type hepatocytes. Thus, the reduced HDL level in PLTP-deficient mice is due to increased degradation of apoA-I. Previous studies have already shown that stimulation of a proteolytic pathway is responsible for degradation of hepatic apoB in PLTP-deficient mice [16•].

The lower susceptibility to atherosclerosis seen in PLTP-deficient mice despite lower HDL levels is explained by the effect of peripheral PLTP at the cellular level. PLTP is responsible for the formation of preβ-HDL particles that serve as acceptors for ABCA1-mediated cholesterol efflux from macrophages [5]. The expression of both PLTP and ABCA1 in macrophages is induced by cholesterol-loading, a process mediated by the liver-X receptor [17]. Such coregulation suggests possible cooperative functions for PLTP and ABCA1. PLTP has previously been shown to bind to and stabilize cell surface ABCA1 [18]. Recently, a comparison of the cholesterol efflux capacities of peritoneal macrophages from wild-type and PLTP knock-out mice showed that lipid-free apoA-I was inefficient at removing cholesterol from cholesterol-loaded PLTP knock-out macrophages [19••]. Drastic upregulation of ABCA1, however, by treatment with cAMP abolished the difference between the efflux capacities of wild-type and PLTP knock-out macrophages. The nature of the cholesterol acceptor also contributed to the extent of efflux, as plasma from wild-type mice had a greater efflux potential than plasma from PLTP knock-out mice.

To examine the atheroprotective function of macrophage PLTP further, Valenta et al. [20] used bone marrow transplantation techniques to generate macrophage-specific deficiency of PLTP. In the LDLR^−/− background, macrophage PLTP deficiency caused a significant increase in aortic lesion area without affecting plasma cholesterol levels. PLTP-deficient macrophages also showed a reduced antioxidative potential and increased oxidative stress as indicated by a dramatic decrease in cellular vitamin E content and a corresponding increase in intracellular hydroperoxides [20]. Thus, various mechanisms may exert the atheroprotective function of macrophage PLTP: cholesterol efflux may be enhanced due to the formation of preβ-HDL particles as cholesterol acceptors, PLTP may stabilize ABCA1, or the accumulation of vitamin E may lead to reduced oxidative capacity of macrophages.

Abbreviations

apo

apolipoprotein

CRP

C-reactive protein

CVD

cardiovascular disease

HDL

high-density lipoprotein

IMT

intima-media thickness

LDL

low-density lipoprotein

LDLR

low-density lipoprotein receptor

PLTP

phospholipid transfer protein

VLDL

very-low-density lipoprotein