Patients with Tangier disease exhibit extremely low plasma HDL concentrations resulting from mutations in the ATP-binding cassette, sub-family A, member 1 (ABCA1) protein. ABCA1 controls the rate-limiting step in HDL particle assembly by mediating efflux of cholesterol and phospholipid from cells to lipid-free apoA-I, which forms nascent HDL particles. ABCA1 is widely expressed; however, the specific tissues involved in HDL biogenesis are unknown. To determine the role of the liver in HDL biogenesis, we generated mice with targeted deletion of the second nucleotide-binding domain of Abca1 in liver only (Abca1 -L/-L ). Abca1 -L/-L mice had total plasma and HDL cholesterol concentrations that were 19% and 17% those of wild-type littermates, respectively. In vivo catabolism of HDL apoA-I from wild-type mice or human lipid-free apoA-I was 2-fold higher in Abca1 -L/-L mice compared with controls due to a 2-fold increase in the catabolism of apoA-I by the kidney, with no change in liver catabolism. We conclude that in chow-fed mice, the liver is the single most important source of plasma HDL. Furthermore, hepatic, but not extrahepatic, Abca1 is critical in maintaining the circulation of mature HDL particles by direct lipidation of hepatic lipid-poor apoA-I, slowing its catabolism by the kidney and prolonging its plasma residence time.

Patients with Tangier disease exhibit extremely low plasma HDL concentrations resulting from mutations in the ATP-binding cassette, sub-family A, member 1 (ABCA1) protein. ABCA1 controls the rate-limiting step in HDL particle assembly by mediating efflux of cholesterol and phospholipid from cells to lipid-free apoA-I, which forms nascent HDL particles. ABCA1 is widely expressed; however, the specific tissues involved in HDL biogenesis are unknown. To determine the role of the liver in HDL biogenesis, we generated mice with targeted deletion of the second nucleotide-binding domain of Abca1 in liver only (Abca1 -L/-L ). Abca1 -L/-L mice had total plasma and HDL cholesterol concentrations that were 19% and 17% those of wild-type littermates, respectively. In vivo catabolism of HDL apoA-I from wild-type mice or human lipid-free apoA-I was 2-fold higher in Abca1 -L/-L mice compared with controls due to a 2-fold increase in the catabolism of apoA-I by the kidney, with no change in liver catabolism. We conclude that in chow-fed mice, the liver is the single most important source of plasma HDL. Furthermore, hepatic, but not extrahepatic, Abca1 is critical in maintaining the circulation of mature HDL particles by direct lipidation of hepatic lipid-poor apoA-I, slowing its catabolism by the kidney and prolonging its plasma residence time.

Objectives-The aim of this study was to determine the role of ATP binding cassette transporter A1 (ABCA1) on generation of different-sized nascent HDLs. Methods and Results-HEK293 cells stably-transfected with ABCA1 (HEK293-ABCA1) or non-transfected (control) cells were incubated with lipid free 125 I-apoA-I for 24 hours. Incubation of apoA-I with HEK293-ABCA1 cells, but not control cells, led to the formation of heterogeneous-sized, pre-␤ migrating nascent HDL subpopulations (pre-␤1 to -4) that varied in size (7.1 to 15.7 nm), lipid, and apoA-I content. Kinetic studies suggested that all subpopulations were formed simultaneously, with no evidence for a precursor-product relationship between smaller and larger-sized particles. When isolated nascent pre-␤ HDLs (pre-␤1 to -4) were added back to HEK293-ABCA1 cells, their ability to bind to ABCA1 and efflux lipid was severely compromised. Heat-denaturation of pre-␤1 HDL resulted in partial recovery of ABCA1 binding, suggesting that initial interaction of apoA-I with ABCA1 results in a constrained conformation of apoA-I that decreases subsequent binding. Key Words: ATP binding cassette transporter A1 Ⅲ apolipoprotein AI Ⅲ high density lipoprotein T he inverse relationship between plasma high-density lipoprotein (HDL) cholesterol concentration and the risk of premature atherosclerotic vascular diseases has generated interest in understanding the steps involved in HDL assembly and catabolism. HDLs are proposed to be antiatherogenic because of their ability to accept excess cellular cholesterol in peripheral tissues and transport it to the liver in a process denoted as reverse cholesterol transport (RCT). 1 HDLs are classified into two subpopulations based on electrophoretic mobility on agarose gels: ␣-HDL (90% to 95% total HDL in plasma) and pre-␤ HDL (5% to 10%). 2 Heterogeneity is found among pre-␤ and ␣-HDLs. Using 2-dimensional gel electrophoresis (2D-gel), several pre-␤ and ␣-HDL subpopulations have been identified as a function of increasing size. 3 Although heterogeneity among pre-␤ and ␣-HDLs is well-documented, little is known about the mechanism of formation and metabolism of these pre-␤ and ␣-HDL subpopulations. Conclusions-InteractionABCA1 is critical for maintaining normal plasma HDL levels and nascent HDL biogenesis. A critical role for ABCA1 in HDL metabolism was demonstrated in patients with Tangier disease, a genetic disorder in which the ABCA1 gene is mutated. 4 -6 These patients have plasma HDL-cholesterol and apoA-I concentrations Ͻ5% of normal, accumulation of CE in macrophages, and increased risk of atherosclerosis. 7,8 Fibroblasts from these patients also have a significant reduction in lipid efflux to apoA-I compared with controls, suggesting ABCA1 was essential for mediating transport of intracellular lipid to apoA-I. 9 Additionally, ABCA1 knockout mice recapitulate the Tangier disease phenotype, verifying the role of ABCA1 in cellular lipid homeostasis and plasma HDL maintenance. 10 Plasma pre-␤ HDLs exist as several subpopulations...

Background Hip fracture affects more than 1.6 million persons worldwide and causes substantial changes in body composition, function, and strength. Usual care (UC) has not successfully restored function to most patients, and prior research has not identified an effective restorative program. Our objective was to determine whether a yearlong home-based exercise program initiated following UC could be administered to older patients with hip fracture and improve outcomes. Methods A randomized controlled trial of 180 community dwelling female patients with hip fracture, 65 years and older, randomly assigned to intervention (n=91) or UC (n=89). Patients were recruited within 15 days of fracture from 3 Baltimore-area hospitals from November 1998 through September 2004. Follow-up assessments were conducted at 2, 6, and 12 months after fracture. The Exercise Plus Program was administered by exercise trainers that included supervised and independently performed aerobic and resistive exercises with increasing intensity. Main outcome measures included bone mineral density of the contralateral femoral neck. Other outcomes included time spent and kilocalories expended in physical activity using the Yale Physical Activity Scale, muscle mass and strength, fat mass, activities of daily living, and physical and psychosocial functioning. The effect of intervention for each outcome was estimated by the difference in outcome trajectories 2 to 12 months after fracture. Results More than 80% of participants received trainer visits, with the majority receiving more than 3 quarters (79%) of protocol visits. The intervention group reported more time spent in exercise activity during follow-up (P<.05). Overall, small effect sizes of 0 to 0.2 standard deviations were seen for bone mineral density measures, and no significant patterns of time-specific between-group differences were observed for the remaining outcome measures. Conclusion Patients with hip fracture who participate in a yearlong, in-home exercise program will increase activity level compared with those in UC; however, no significant changes in other targeted outcomes were detected.

We compared the in vivo metabolism of pre ␤ HDL particles isolated by anti-human apolipoprotein A-I (apoA-I) immunoaffinity chromatography (LpA-I) in human apoA-I transgenic (hA-I Tg) mice with that of lipid-free apoA-I (LFA-I) and small LpA-I. After injection, pre ␤ LpA-I were removed from plasma more rapidly than were LFA-I and small LpA-I. Pre ␤ LpA-I and LFA-I were preferentially degraded by kidney compared with liver; small LpA-I were preferentially degraded by the liver. Five minutes after tracer injection, 99% of LFA-I in plasma was found to be associated with medium-sized (8.6 nm) HDL, whereas only 37% of pre ␤ tracer remodeled to medium-sized HDL. Injection of pre ␤ LpA-I doses into C57Bl/6 recipients resulted in a slower plasma decay compared with hA-I Tg recipients and a greater proportion ( Ͼ 60%) of the pre ␤ radiolabel that was associated with medium-sized HDL. Pre ␤ LpA-I contained one to four molecules of phosphatidylcholine per molecule of apoA-I, whereas LFA-I contained less than one. We conclude that pre ␤ LpA-I has two metabolic fates in vivo, rapid removal from plasma and catabolism by kidney or remodeling to medium-sized HDL, which we hypothesize is determined by the amount of lipid associated with the pre ␤ particle and the particle's ability to bind to medium-sized HDL.

We investigated the in vivo metabolic fate of preb HDL particles in human apolipoprotein A-I transgenic (hA-I Tg ) mice. Pre-b HDL tracers were assembled by incubation of [125 I]tyramine cellobiose-labeled apolipoprotein A-I (apoA-I) with HEK293 cells expressing ABCA1. Radiolabeled pre-b HDLs of increasing size (pre-b1, -2, -3, and -4 HDLs) were isolated by fast-protein liquid chromatography and injected into hA-I Tg -recipient mice, after which plasma decay, in vivo remodeling, and tissue uptake were monitored. Pre-b2, -3, and -4 had similar plasma die-away rates, whereas pre-b1 HDL was removed 7-fold more rapidly. Radiolabel recovered in liver and kidney 24 h after tracer injection suggested increased (P , 0.001) liver and decreased kidney catabolism as pre-b HDL size increased. In plasma, pre-b1 and -2 were rapidly (,5 min) remodeled into larger HDLs, whereas pre-b3 and -4 were remodeled into smaller HDLs. Pre-b HDLs were similarly remodeled in vitro with control or LCAT-immunodepleted plasma, but not when incubated with phospholipid transfer protein knockout plasma. Our results suggest that initial interaction of apoA-I with ABCA1 imparts a unique conformation that partially determines the in vivo metabolic fate of apoA-I, resulting in increased liver and decreased kidney catabolism as pre-b HDL particle size

The fetus has a high requirement for cholesterol and synthesizes cholesterol at elevated rates. Recent studies suggest that fetal cholesterol also can be obtained from exogenous sources. The purpose of the current study was to examine the transport of maternal cholesterol to the fetus and determine the mechanism responsible for any cholesterol-driven changes in transport. Studies were completed in pregnant hamsters with normal and elevated plasma cholesterol concentrations. Cholesterol feeding resulted in a 3.1-fold increase in the amount of LDL-cholesterol taken up by the fetus and a 2.4-fold increase in the amount of HDL-cholesterol taken up. LDL-cholesterol was transported to the fetus primarily by the placenta, and HDL-cholesterol was transported by the yolk sac and placenta. Several proteins associated with sterol transport and efflux, including those induced by activated liver X receptor, were expressed in hamster and human placentas: NPC1, NPC1L1, ABCA2, SCP-x, and ABCG1, but not ABCG8. NPC1L1 was the only protein increased in hypercholesterolemic placentas. Thus, increasing maternal lipoprotein-cholesterol concentrations can enhance transport of maternal cholesterol to the fetus, leading to 1) increased movement of cholesterol down a concentration gradient in the placenta, 2) increased lipoprotein secretion from the yolk sac (shown previously), and possi- Cholesterol is essential for normal fetal development. Possibly the most noted role of cholesterol is as a structural component of membranes. Sterols are also the precursor for bile acids, steroid hormones, and oxysterols, which are all synthesized by the fetus and regulate various cellular processes. Cholesterol is also required for activation of sonic hedgehog (Shh), a protein involved in brain development, and for propagation of the Shh signal (1, 2). Cholesterol is obtained endogenously by de novo synthesis and exogenously by transfer of maternal cholesterol to the fetus. Interestingly, maternal plasma triglyceride and cholesterol concentrations increase during pregnancy in humans (3, 4), possibly an adaptation to maternal and fetal needs. However, whether these changes in maternal lipoprotein concentrations result in increased transport of maternal cholesterol to the fetus is unclear.Since the fetus does not come in direct contact with the maternal circulation, maternal cholesterol destined for the fetus must initially be taken up by the placenta and yolk sac prior to transport and delivery to the fetus. Indeed, the placenta and yolk sac take up maternal LDL-and HDLcholesterol at relatively elevated rates compared with other peripheral tissues (5). LDL is taken up by the LDL receptor (LDLR), which is expressed abundantly in the placenta but expressed at low levels, if at all, in the yolk sac (5, 6). LDL-derived sterol is transported to the lysosome/endosome pathway where the ester bond is hydrolyzed (7). The unesterified cholesterol is then transported by NiemannThese studies were supported by grants HD34089 (LAW) and GM31651 (FS) from the...

Over the past few years, new experimental approaches have reinforced the awareness among investigators that the heterogeneity of HDL particles indicates significant differences in production and catabolism of HDL particles. Recent kinetic studies have suggested that small HDL, containing two apolipoprotein A-I molecules per particle, are converted in a unidirectional manner to medium HDL or large HDL, containing three or four apolipoprotein A-I molecules per particle, respectively. Conversion appears to occur in close physical proximity with cells and not while HDL particles circulate in plasma. The medium and large HDL are terminal particles in HDL metabolism with large HDL, and perhaps medium HDL, being catabolized primarily by the liver. These novel kinetic studies of HDL subfraction metabolism are compelling in-vivo data that are consistent with the proposed role of HDL in reverse cholesterol transport.