Apolipoprotein A-V is present in bile and its secretion increases with lipid absorption in Sprague-Dawley rats.
Zhang Linda S,Sato Hirokazu,Yang Qing,Ryan Robert O,Wang David Q-H,Howles Philip N,Tso Patrick
American journal of physiology. Gastrointestinal and liver physiology
Apolipoprotein (apo) A-V is a protein synthesized only in the liver that dramatically modulates plasma triglyceride levels. Recent studies suggest a novel role for hepatic apoA-V in regulating the absorption of dietary triglycerides, but its mode of action on the gut remains unknown. The aim of this study was to test for apoA-V in bile and to determine whether its secretion is regulated by dietary lipids. After an overnight recovery, adult male Sprague-Dawley bile fistula rats indeed secreted apoA-V into bile at a constant rate under fasting conditions. An intraduodenal bolus of intralipid (n = 12) increased the biliary secretion of apoA-V but not of other apolipoproteins, such as A-I, A-IV, B, and E. The lipid-induced increase of biliary apoA-V was abolished under conditions of poor lymphatic lipid transport, suggesting that the stimulation is regulated by the magnitude of lipids associated with chylomicrons transported into lymph. We also studied the secretion of apoA-V into bile immediately following bile duct cannulation. Biliary apoA-V increased over time (∼6-fold increase at hour 16, n = 8) but the secretions of other apolipoproteins remained constant. Replenishing luminal phosphatidylcholine and taurocholate (n = 9) only enhanced apoA-V secretion in bile, suggesting that the increase was not due to depletion of phospholipids or bile salts. This is the first study to demonstrate that apoA-V is secreted into bile, introducing a potential route of delivery of hepatic apoA-V to the gut lumen. Our study also reveals the uniqueness of apoA-V secretion into bile that is regulated by mechanisms different from other apolipoproteins.
10.1152/ajpgi.00227.2015
Structure-function relationships in reconstituted HDL: Focus on antioxidative activity and cholesterol efflux capacity.
Cukier Alexandre M O,Therond Patrice,Didichenko Svetlana A,Guillas Isabelle,Chapman M John,Wright Samuel D,Kontush Anatol
Biochimica et biophysica acta. Molecular and cell biology of lipids
AIMS:High-density lipoprotein (HDL) contains multiple components that endow it with biological activities. Apolipoprotein A-I (apoA-I) and surface phospholipids contribute to these activities; however, structure-function relationships in HDL particles remain incompletely characterised. METHODS:Reconstituted HDLs (rHDLs) were prepared from apoA-I and soy phosphatidylcholine (PC) at molar ratios of 1:50, 1:100 and 1:150. Oxidative status of apoA-I was varied using controlled oxidation of Met112 residue. HDL-mediated inactivation of PC hydroperoxides (PCOOH) derived from mildly pre-oxidized low-density lipoprotein (LDL) was evaluated by HPLC with chemiluminescent detection in HDL+LDL mixtures and re-isolated LDL. Cellular cholesterol efflux was characterised in RAW264.7 macrophages. RESULTS:rHDL inactivated LDL-derived PCOOH in a dose- and time-dependent manner. The capacity of rHDL to both inactivate PCOOH and efflux cholesterol via ATP-binding cassette transporter A1 (ABCA1) increased with increasing apoA-I/PC ratio proportionally to the apoA-I content in rHDL. Controlled oxidation of apoA-I Met112 gradually decreased PCOOH-inactivating capacity of rHDL but increased ABCA1-mediated cellular cholesterol efflux. CONCLUSIONS:Increasing apoA-I content in rHDL enhanced its antioxidative activity towards oxidized LDL and cholesterol efflux capacity via ABCA1, whereas oxidation of apoA-I Met112 decreased the antioxidative activity but increased the cholesterol efflux. These findings provide important considerations in the design of future HDL therapeutics. Non-standard abbreviations and acronyms: AAPH, 2,2'-azobis(-amidinopropane) dihydrochloride; ABCA1, ATP-binding cassette transporter A1; apoA-I, apolipoprotein A-I; BHT, butylated hydroxytoluene; CV, cardiovascular; EDTA, ethylenediaminetetraacetic acid; HDL-C, high-density lipoprotein cholesterol; LOOH, lipid hydroperoxides; Met(O), methionine sulfoxide; Met112, methionine 112 residue; Met86, methionine 86 residue; oxLDL, oxidized low-density lipoprotein; PBS, phosphate-buffered saline; PC, phosphatidylcholine; PL, phospholipid; PCOOH, phosphatidylcholine hydroperoxide; PLOOH, phospholipid hydroperoxide.
10.1016/j.bbalip.2017.05.010
A multiple ascending dose study of CSL112, an infused formulation of ApoA-I.
Easton Rachael,Gille Andreas,D'Andrea Denise,Davis Roslyn,Wright Samuel D,Shear Charles
Journal of clinical pharmacology
CSL112 is apoA-I purified from human plasma and reconstituted with phosphatidylcholine (PC) to form high-density lipoprotein (HDL)-particles suitable for infusion. CSL112 is in development for the potential treatment of acute coronary syndromes (ACS) by optimizing cholesterol efflux. This study assesses the pharmacokinetics (PK), safety and tolerability of CSL112. Repeat doses of CSL112 or placebo were administered intravenously once- (3.4 g or 6.8 g) or twice-weekly (3.4 g) to healthy subjects in a placebo-controlled, randomized (3 CSL112: 1 placebo), ascending-dose study (NCT01281774). Twenty-seven subjects received CSL112 and nine received placebo. Study endpoints included plasma apoA-I and PC concentrations and specific PK parameters. CSL112 infusions immediately produced robust increases in apoA-I concentration in a dose-proportional manner, reaching levels higher than observed with currently available or investigational HDL products. After infusion of CSL112, apoA-I levels remained above baseline for approximately 3 days. Multiple infusions of CSL112 were safe and well tolerated with no evidence of major organ toxicity or immunogenicity. CSL112 may provide a novel option to rapidly transport cholesterol from atherosclerotic plaque to the liver and reduce early recurrent events following ACS. The data presented here support continued clinical development of CSL112 in patient populations.
10.1002/jcph.194
A single infusion of MDCO-216 (ApoA-1 Milano/POPC) increases ABCA1-mediated cholesterol efflux and pre-beta 1 HDL in healthy volunteers and patients with stable coronary artery disease.
Kallend D G,Reijers J A A,Bellibas S E,Bobillier A,Kempen H,Burggraaf J,Moerland M,Wijngaard P L J
European heart journal. Cardiovascular pharmacotherapy
AIMS:Apolipoprotein A-1 (ApoA-1), based on epidemiology, is inversely associated with cardiovascular (CV) events. Human carriers of the ApoA-1 Milano variant have a reduced incidence of CV disease. Regression of atherosclerotic plaque burden was previously observed on intravascular ultrasound (IVUS) with ETC-216, a predecessor of MDCO-216. MDCO-216, a complex of dimeric ApoA-1 Milano and 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine, is being developed to reduce atherosclerotic plaque burden and CV events. We investigated the efficacy and safety of a single infusion of MDCO-216 in healthy volunteers and in patients with coronary artery disease (CAD). METHODS AND RESULTS:Twenty-four healthy volunteers and 24 patients with documented CAD received a 2-h infusion of MDCO-216 in a randomized, placebo controlled, single ascending dose study. Five cohorts of healthy volunteers and four cohorts of CAD patients received ApoA-1 Milano doses ranging from 5 to 40 mg/kg. Subjects were followed for 30 days. Dose-dependent increases in ApoA-1, phospholipid, and pre-beta 1 HDL and decreases in ApoE were observed. Prominent and sustained increases in triglyceride, and decreases in HDL-C, endogenous ApoA-1 and ApoA-II occurred at doses >20 mg/kg and profound increases in ABCA1-mediated cholesterol efflux were observed. Other lipid and lipoprotein parameters were generally unchanged. MDCO-216 was well tolerated. CONCLUSIONS:MDCO-216-modulated lipid parameters profoundly increased ABCA1-mediated cholesterol efflux and was well tolerated. These single-dose data support further development of this agent for reducing atherosclerotic disease and subsequent CV events.
10.1093/ehjcvp/pvv041
High-Density Lipoprotein Subfractions and Cholesterol Efflux Capacities After Infusion of MDCO-216 (Apolipoprotein A-IMilano/Palmitoyl-Oleoyl-Phosphatidylcholine) in Healthy Volunteers and Stable Coronary Artery Disease Patients.
Kempen Herman J,Asztalos Bela F,Moerland Matthijs,Jeyarajah Elias,Otvos James,Kallend David G,Bellibas S Eralp,Wijngaard Peter L J
Arteriosclerosis, thrombosis, and vascular biology
OBJECTIVE:To determine effects of single ascending doses of MDCO-216 on high-density lipoprotein (HDL) subfractions in relation to changes in cholesterol efflux capacity in healthy volunteers and in patients with stable angina pectoris. APPROACH AND RESULTS:Doses of 5- (in volunteers only), 10-, 20-, 30-, and 40-mg/kg MDCO-216 were infused during 2 hours, and plasma and serum were collected during 30 days. Plasma levels of HDL subfractions were assessed by 2-dimensional gel electrophoresis, immunoblotting, and image analysis. Lipoprotein particle concentrations and sizes were also assessed by proton nuclear magnetic resonance ((1)H-NMR). There was a rapid dose-dependent increase of total apolipoprotein A-I (apoA-I) in pre-β1, α-1, and α-2 HDL levels and decrease in α-3 and α-4 HDL. Using a selective antibody apoA-IMilano was detected in the large α-1 and α-2 HDL on all doses and at each time point. ApoA-IMilano was also detected at the α-4 position but only at high doses. (1)H-NMR analysis similarly showed a rapid and dose-dependent shift from small- to large-sized HDL particles. The increase of basal and ATP-binding cassette transporter A1-mediated efflux capacities reported previously correlated strongly and independently with the increase in pre-β1-HDL and α-1 HDL, but not with that in α-2 HDL. CONCLUSIONS:On infusion, MDCO-216 rapidly eliminates small HDL and leads to formation of α-1 and α-2 HDL containing both wild-type apoA-I and apoA-IMilano. In this process, endogenous apoA-I is liberated appearing as pre-β1-HDL. In addition to pre-β1-HDL, the newly formed α-1 HDL particle containing apoA-I Milano may have a direct effect on cholesterol efflux capacity.
10.1161/ATVBAHA.115.307052
Effect of repeated apoA-IMilano/POPC infusion on lipids, (apo)lipoproteins, and serum cholesterol efflux capacity in cynomolgus monkeys.
Kempen Herman J,Gomaraschi Monica,Bellibas S Eralp,Plassmann Stephanie,Zerler Brad,Collins Heidi L,Adelman Steven J,Calabresi Laura,Wijngaard Peter L J
Journal of lipid research
MDCO-216, a complex of dimeric recombinant apoA-IMilano (apoA-IM) and palmitoyl-oleoyl-phosphatidylcholine (POPC), was administered to cynomolgus monkeys at 30, 100, and 300 mg/kg every other day for a total of 21 infusions, and effects on lipids, (apo)lipoproteins, and ex-vivo cholesterol efflux capacity were monitored. After 7 or 20 infusions, free cholesterol (FC) and phospholipids (PL) were strongly increased, and HDL-cholesterol (HDL-C), apoA-I, and apoA-II were strongly decreased. We then measured short-term effects on apoA-IM, lipids, and (apo)lipoproteins after the first or the last infusion. After the first infusion, PL and FC went up in the HDL region and also in the LDL and VLDL regions. ApoE shifted from HDL to LDL and VLDL regions, while ApoA-IM remained located in the HDL region. On day 41, ApoE levels were 8-fold higher than on day 1, and FC, PL, and apoE resided mostly in LDL and VLDL regions. Drug infusion quickly decreased the endogenous cholesterol esterification rate. ABCA1-mediated cholesterol efflux on day 41 was markedly increased, whereas scavenger receptor type B1 (SRB1) and ABCG1-mediated effluxes were only weakly increased. Strong increase of FC is due to sustained stimulation of ABCA1-mediated efflux, and drop in HDL and formation of large apoE-rich particles are due to lack of LCAT activation.
10.1194/jlr.M033779
Mechanism of the defect in cholesteryl ester clearance from macrophages of atherosclerosis-susceptible White Carneau pigeons.
Yancey P G,St Clair R W
Journal of lipid research
Recent studies from our laboratory (Yancey, P.G., and R. W. St. Clair. 1992. Arterioscler. Thromb. 12: 1291-1304) have shown that cultured peritoneal macrophages from White Carneau (WC) pigeons clear cholesteryl esters at a slower rate than do macrophages from Show Racer (SR) pigeons (9% per 24 h vs. 42% per 24 h, respectively) when incubated in the presence of a cholesterol acceptor apo high density lipoprotein/phosphatidylcholine (apoHDL/PC) at concentrations that are not rate-limiting for cholesterol efflux. In the present studies we have examined some potential mechanisms for this difference in cholesterol efflux. The desorption of [3H]cholesterol from the plasma membranes of non-cholesterol-loaded cells was log-linear with half-times of 24-31 h, and was not different for WC and SR macrophages. As this rate of cholesterol desorption was 2- to 3-times faster than the rate of cholesteryl ester clearance, it is unlikely to be rate-limiting. In cells loaded with cholesteryl esters, the re-esterification of cholesterol was equally low in both WC and SR macrophages when incubated with apoHDL/PC. The addition of the acyl-CoA:cholesterol acyltransferase (ACAT) inhibitor, Sandoz 58,035, reduced esterification approximately 50%, but did not affect the clearance of cholesteryl ester mass from either WC or SR macrophages. Thus, differences in cholesteryl re-esterification could not explain the differences in cholesteryl ester clearance. The rate of hydrolysis of radiolabeled cellular cholesteryl esters was log-linear for up to 36 h in WC and SR macrophages. The half-time for hydrolysis of cholesteryl ester was 89 h in WC macrophages, compared to only 37 h in the SR macrophages, and paralleled the rate of clearance of cholesteryl esters in these cells (133 h and 51 h, respectively). Cyclic AMP stimulated cholesterol ester hydrolysis in WC macrophages and increased by 5-fold the clearance of cholesteryl esters in the presence of apoHDL/PC and Sandoz 58,035. These data are consistent with the conclusion that the rate-limiting step in the clearance of cholesteryl esters from pigeon macrophages is the hydrolysis of cholesteryl esters, and that the slow rate of cholesteryl ester clearance in WC macrophages is due, at least in part, to a defect in cholesteryl ester hydrolysis. It is tempting to speculate that this defect in cholesteryl ester hydrolysis may play a role in the difference in susceptibility to atherosclerosis between WC and SR pigeons.
Hepatic uptake and metabolism of phosphatidylcholine associated with high density lipoproteins.
Robichaud Julie C,van der Veen Jelske N,Yao Zemin,Trigatti Bernardo,Vance Dennis E
Biochimica et biophysica acta
BACKGROUND:Phosphatidylcholine (PC) is the predominant phospholipid associated with high density lipoproteins (HDL). Although the hepatic uptake of cholesteryl esters from HDL is well characterized, much less is known about the fate of PC associated with HDL. Thus, we investigated the uptake and subsequent metabolism of HDL-PC in primary mouse hepatocytes. METHODS AND RESULTS:The absence of scavenger receptor-BI resulted in a 30% decrease in cellular incorporation of [(3)H]PC whereas [(3)H]cholesteryl ether uptake was almost completely abolished. Although endocytosis is not involved in the uptake of cholesteryl esters from HDL, we demonstrate that HDL internalization accounts for 40% of HDL-PC uptake. Extracellular remodeling of HDL by secretory phospholipase A(2) significantly enhances HDL lipid uptake. HDL-PC taken up by hepatocytes is partially converted to triacylglycerols via PC-phospholipase C-mediated hydrolysis of PC and incorporation of diacylglycerol into triacylglycerol. The formation of triacylglycerol is independent of scavenger receptor-BI and occurs in extralysosomal compartments. CONCLUSIONS AND GENERAL SIGNIFICANCE:These findings indicate that HDL-associated PC is incorporated into primary hepatocytes via a pathway that differs significantly from that of HDL-cholesteryl ester, and shows that HDL-PC is more than a framework molecule, as evidenced by its partial conversion to hepatic triacylglycerol.
10.1016/j.bbagen.2009.02.010