doi: 10.1161/01.ATV.0000193889.65915.f9
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© 2005 American Heart Association, Inc.
Editorials |
Apolipoprotein AV
Low Concentration, High Impact
From the Departments of General Internal Medicine, Endocrinology and Metabolism (P.C.N.R., K.W.v.D., L.M.H.), Human Genetics (K.W.v.D.), and Cardiology (L.M.H.), Leiden University Medical Center, and TNO-Quality of Life, Department of Biomedical Research (P.C.N.R., L.M.H.), Gaubius Laboratory, Leiden, The Netherlands.
Correspondence to Patrick C.N. Rensen, PhD, Leiden University Medical Center, Department of Endocrinology and Metabolism, C4-R81, Albinusdreef 2, P.O. Box 9600, 2300 RC Leiden, The Netherlands. E-mail P.C.N.Rensen{at}lumc.nl
Hypertriglyceridemia is an independent risk factor for coronary heart disease.1 Apolipoproteins that have been shown to affect plasma triglyceride (TG) levels include apolipoprotein E (apoE) and the C-apolipoproteins (apoCI, apoCII, and apoCIII). The recently identified apoAV is the newest member of this family of apolipoproteins affecting plasma TG levels.2,3 ApoAV is a 39-kDa protein (343 amino acids) that is expressed exclusively by the liver. Plasma TG concentrations in mice were 4-fold increased on deficiency of the endogenous apoa5 gene and were decreased by 65% on expression of the human APOA5 gene.2 Subsequent studies have shown that adenovirus-mediated transfer of murine apoa5 to mice resulted in a dose-dependent reduction of plasma TG.4,5 These studies have thus established an important role of apoAV in TG homeostasis.
See page 2573
Several groups have studied the mechanism underlying the effect of apoAV on TG metabolism. ApoAV has been proposed to affect TG levels by both an intra- and extracellular effect: (1) apoAV may inhibit the hepatic secretion of VLDL,5,6 and (2) apoAV may facilitate the clearance of TG from plasma.5,7,8 Evidence for these two mechanisms has been mainly obtained from mouse models that overexpress apoAV.
In this issue of Atherosclerosis, Thrombosis, and Vascular Biology, Grosskopf et al9 extend these studies on the mechanisms underlying the effects of apoAV on TG metabolism, by using apoAV-deficient (apoa5–/–) mice. From a diverse set of experiments, the elevation of TG in apoa5–/– mice is attributed to the disturbance of two pathways: (1) reduction in lipoprotein remnant generation caused by a reduced LPL-mediated lipolysis of VLDL-TG and decreased plasma LPL and HL levels, and (2) reduction in the hepatic uptake of lipoprotein core remnants, caused by a reduced affinity for the LDL receptor. Grosskopf et al9 observed that apoAV deficiency did not affect the production rate of VLDL-TG.
Weinberg et al6 initially proposed that apoAV may retard VLDL assembly, based on the fact that apoAV is very hydrophobic and displays slow binding kinetics at hydrophobic interfaces. Transfection of COS-1 cells with apoAV demonstrated that apoAV was poorly secreted and remained largely associated with the endoplasmatic reticulum, which may result in modifying the lipidation of apoB100.6 In line with these data, we demonstrated that relatively low hepatic overexpression of murine apoa5 (ie, 10-fold) in wild-type C57Bl/6 mice decreased the VLDL-TG secretion rate by 30% without affecting the number of VLDL particles produced by the liver.5 This effect appeared specific for apoAV, because adenovirus-mediated expression of apoCI has no effect (P.C.N.R., K.W.v.D., L.M.H., unpublished observations, 2005), and apoE severely increases VLDL-TG production.10 However, the effect of apoAV overexpression on VLDL-TG production may be dependent on the genetic background of the strain, because overexpression of apoa5 in hyperlipidemic APOE2-knockin mice and APOE*3-Leiden transgenic mice did not affect VLDL-TG production despite a marked reduction of plasma TG.11 Similarly, VLDL-TG production rates were reported to be unaltered in APOA5 transgenic mice.7 Grosskopf et al9 now also report that the VLDL-TG production in apoa5–/– mice is unaffected. Thus, whether apoAV has a physiological relevance for modulating the lipidation of apoB100 clearly needs further investigation.
Although apoAV may be retained to some extent in hepatocytes, apoAV is also secreted into the plasma. ApoAV has been found associated with apoB-containing lipoproteins (ie, chylomicrons and VLDL) and HDL, with an equal distribution between VLDL and HDL in fasting serum.12 However, the total plasma concentrations are very low as compared with other apolipoproteins (ie, 125 to 180 ng/mL).12,13 Because the clearance of TG from plasma is highly dependent on hydrolysis by lipoprotein lipase (LPL), studies have been undertaken to address the role of apoAV in the regulation of LPL activity. Initially, we5 and others7 have demonstrated that apoAV enhances LPL-mediated TG hydrolysis in a dose-dependent manner in vitro, but only in the presence of the LPL coactivator apoCII.5 In addition, we showed that low doses of apoAV can reverse the inhibition of LPL activity by apoCIII.5 In line with these data, the LPL-dependent clearance of VLDL-like emulsion particles and chylomicrons from plasma was strongly accelerated in mice transfected with a murine apoa5-expressing adenovirus, with an increased uptake of TG-derived fatty acids observed in tissues that express LPL such as muscle and adipose tissue.5 Although Biacore studies have suggested that apoAV may exert its LPL-stimulatory action by direct physical interaction with LPL,7 subsequent studies from Merkel et al8 and Lookene et al14 have indicated that such an LPL-activating action of apoAV is likely to involve enhanced apoAV-mediated binding of VLDL and chylomicron particles to heparan sulfate proteoglycans (HSPG). Whether apoAV directly interacts with LPL and/or HSPG in vivo remains to be determined, but the mechanism needs to reconcile that very low concentrations of apoAV are already sufficient to accelerate the turnover of chylomicrons and VLDL by stimulating LPL-mediated TG hydrolysis.
The current manuscript of Grosskopf et al9 shows that the effect of apoAV on LPL-mediated TG clearance is also a main factor explaining the hypertriglyceridemia in apoAV-deficient mice. However, instead of attributing this to a direct effect of apoAV on LPL activity, it is postulated that the elevated TG levels result from as much as 87% decreased post-heparin LPL levels. Interestingly, heterozygous LPL-deficient mice, which have 43% lower LPL levels, already show 3-fold increased plasma TG levels.15 Therefore, if such a tremendous effect of apoAV-deficiency on LPL levels indeed occurs, this may already fully explain the observed hypertriglyceridemia in apoa5–/– mice. As additional LPL-regulating mechanisms in apoa5–/– mice, a decreased lipolysis of apoAV-deficient VLDL by soluble LPL is reported, which may be related to a 2-fold increased content of apoCs (as judged from apoC:apoB ratio). The authors further characterize the effect of apoAV deficiency on hepatic remnant removal mechanisms by evaluating the hepatic uptake of radiolabeled rat chylomicrons and their remnants in vivo and ex vivo. These studies suggest that apoAV deficiency may retard the clearance of TG-rich lipoproteins by the liver, either by a decreased affinity of lipoproteins for the LDL receptor and/or a decreased initial binding of lipoproteins to proteoglycans. However, at least under in vivo conditions, a diminished hepatic clearance of core remnants may well be the direct consequence of decreased LPL-mediated lipoprotein remodeling. The proposed mechanisms underlying the TG-lowering effect of apoAV, reported thus far, are summarized in the accompanying Figure.
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In addition to the well-documented effect of apoAV on VLDL-TG metabolism, evidence accumulates that apoAV influences HDL metabolism. Grosskopf et al9 observe that apoa5–/– mice have 41% increased HDL-cholesterol levels, which corroborates our earlier findings that overexpression of apoAV dose-dependently decreases HDL-cholesterol levels.5 It is intriguing to speculate on the mechanisms underlying the effect of apoAV on HDL. Could apoAV also increase the activity of other plasma proteins involved in HDL metabolism, such as hepatic lipase (HL), endothelial lipase (EL), lecithin cholesterol acyltransferase (LCAT), and/or phospholipid transfer protein (PLTP) leading to an enhanced catabolism of HDL? If so, would these effects still be apparent in species that express CETP, like humans?
Regardless of the precise mechanisms underlying these effects of apoAV on the metabolism of chylomicrons, VLDL and HDL, an increasing number of human studies have revealed strong associations between single nucleotide APOA5 polymorphisms and TG levels.16,17 Recent observations indeed suggest a direct TG-lowering role of the apoAV protein itself. For example, apoAV deficiency caused by a nonsense mutation (Q145X) has recently been described in subjects with severe hypertriglyceridemia.18 In addition, haplotype analysis indicates that the APOA5*3 haplotype, which is strongly associated with hypertriglyceridemia in multiple independent populations, is characterized by a single S19W mutation, and thus likely constitutes the functional and causative polymorphism. Moreover, the S19W polymorphism affects the signal peptide of the apoAV precursor, and this has recently been shown to result in an altered conformation of this peptide and a reduced protein secretion from Huh7 cells.19
The availability of sandwich ELISAs12,13 which can be used to quantify apoAV levels in plasma now opens the possibility to validate the postulated mechanisms by which apoAV affects VLDL (and HDL) levels in humans. Initial studies have shown a negative correlation between plasma levels of apoAV and TG in normolipidemic individuals.12,13 However, it has to be awaited whether the hypertriglyceridemia, associated with the various APOA5 polymorphisms, is correlated with either increased apoAV levels (in case of apoAV mutants with impaired function) or decreased apoAV levels (eg, in case of the APOA5*3 haplotype, which is expected to result in lower levels of otherwise functional apoAV). Large epidemiologic studies will obviously be needed to clearly define the relation between apoAV concentration on the one hand and hypertriglyceridemia and coronary heart disease risk on the other hand.
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