Biomedical subjects

Lita Freeman

Publications and source records attributed to Lita Freeman.

9 recordsLinked to original sources

PubMed2006

Lipolytic and ligand-binding functions of hepatic lipase protect against atherosclerosis in LDL receptor-deficient mice.

To elucidate the separate contributions of the lipolytic versus ligand-binding functions of hepatic lipase (HL) to lipoprotein metabolism and atherosclerosis, and to investigate the role of the low density lipoprotein receptor (LDLr) in these processes, we compared mice expressing catalytically active HL (HL-WT) with mice expressing inactive HL (HL-S145G) in a background lacking endogenous HL and the LDLr (LDLr-KOxHL-KO). HL-WT and HL-S145G reduced (P < 0.05 for all) cholesterol (55% vs. 20%), non-HDL-cholesterol (63% vs. 22%), and apolipoprotein B (apoB; 34% vs. 16%) by enhancing the catabolism of autologous (125)I-apoB-intermediate density lipoprotein (IDL)/LDL (fractional catabolic rate in day(-1): 6.07 +/- 0.25, LDLr-KOxHL-WT; 4.76 +/- 0.30, LDLr-KOxHL-S145G; 3.70 +/- 0.13, LDLr-KOxHL-KO); HL-WT had a greater impact on the concentration, composition, particle size, and catabolism of apoB-containing lipoproteins (apoB-Lps) and HDL. Importantly, consistent with the changes in apoB-Lps, atherosclerosis in LDLr-KOxHL-KO mice fed a regular chow diet (RCD) was reduced by both HL-WT and HL-S145G (by 71% and 51% in cross-sectional analysis, and by 85% and 67% in en face analysis; P < 0.05 for all). These data identify physiologically relevant but distinct roles for the lipolytic versus ligand-binding functions of HL in apoB-Lp metabolism and atherosclerosis and demonstrate that their differential effects on these processes are mediated by changes in catabolism via non-LDLr pathways. These changes, evident even in the presence of apoE, establish an antiatherogenic role of the ligand-binding function of HL in LDLr-deficient mice.

Animals↗

PubMed2004

Hepatic lipase, lipoprotein metabolism, and atherogenesis.

The role of hepatic lipase as a multifunctional protein that modulates lipoprotein metabolism and atherosclerosis has been extensively documented over the last decade. Hepatic lipase functions as a lipolytic enzyme that hydrolyzes triglycerides and phospholipids present in circulating plasma lipoproteins. Hepatic lipase also serves as a ligand that facilitates lipoprotein uptake by cell surface receptors and proteoglycans, thereby directly affecting cellular lipid delivery. Recently, another process by which hepatic lipase modulates atherogenic risk has been identified. Bone marrow transplantation studies demonstrate that hepatic lipase present in aortic lesions markedly alters aortic lesion formation even in the absence of changes in plasma lipids. These multiple functions of hepatic lipase, which facilitate not only plasma lipid metabolism but also cellular lipid uptake, can be anticipated to have a major and complex impact on atherogenesis. Consistently, human and animal studies support proatherogenic and antiatherogenic roles for hepatic lipase. The concept of hepatic lipase as mainly a lipolytic enzyme that reduces atherogenic risk has evolved into that of a complex protein with multiple functions that, depending on genetic background and sites of expression, can have a variable effect on atherosclerosis.

Animals↗

PubMed2004

Hepatic ABCG5 and ABCG8 overexpression increases hepatobiliary sterol transport but does not alter aortic atherosclerosis in transgenic mice.

The individual roles of hepatic versus intestinal ABCG5 and ABCG8 in sterol transport have not yet been investigated. To determine the specific contribution of liver ABCG5/G8 to sterol transport and atherosclerosis, we generated transgenic mice that overexpress human ABCG5 and ABCG8 in the liver but not intestine (liver G5/G8-Tg) in three different genetic backgrounds: C57Bl/6, apoE-KO, and low density lipoprotein receptor (LDLr)-KO. Hepatic overexpression of ABCG5/G8 enhanced hepatobiliary secretion of cholesterol and plant sterols by 1.5-2-fold, increased the amount of intestinal cholesterol available for absorption and fecal excretion by up to 27%, and decreased the accumulation of plant sterols in plasma by approximately 25%. However, it did not alter fractional intestinal cholesterol absorption, fecal neutral sterol excretion, hepatic cholesterol concentrations, or hepatic cholesterol synthesis. Consequently, overexpression of ABCG5/G8 in only the liver had no effect on the plasma lipid profile, including cholesterol, HDL-C, and non-HDL-C, or on the development of proximal aortic atherosclerosis in C57Bl/6, apoE-KO, or LDLr-KO mice. Thus, liver ABCG5/G8 facilitate the secretion of liver sterols into bile and serve as an alternative mechanism, independent of intestinal ABCG5/G8, to protect against the accumulation of dietary plant sterols in plasma. However, in the absence of changes in fractional intestinal cholesterol absorption, increased secretion of sterols into bile induced by hepatic overexpression of ABCG5/G8 was not sufficient to alter hepatic cholesterol balance, enhance cholesterol removal from the body or to alter atherogenic risk in liver G5/G8-Tg mice. These findings demonstrate that overexpression of ABCG5/G8 in the liver profoundly alters hepatic but not intestinal sterol transport, identifying distinct roles for liver and intestinal ABCG5/G8 in modulating sterol metabolism.

ATP Binding Cassette Transporter, Subfamily G, Mem↗

PubMed2003

Histone tail-independent chromatin binding activity of recombinant cohesin holocomplex.

Cohesin, an SMC (structural maintenance of chromosomes) protein-containing complex, governs several important aspects of chromatin dynamics, including the essential chromosomal process of sister chromatid cohesion. The exact mechanism by which cohesin achieves the bridging of sister chromatids is not known. To elucidate this mechanism, we reconstituted a recombinant cohesin complex and investigated its binding to DNA fragments corresponding to natural chromosomal sites with high and low cohesin occupancy in vivo. Cohesin displayed uniform but nonspecific binding activity with all DNA fragments tested. Interestingly, DNA fragments with high occupancy by cohesin in vivo showed strong nucleosome positioning in vitro. We therefore utilized a defined model chromatin fragment (purified reconstituted dinucleosome) as a substrate to analyze cohesin interaction with chromatin. The four-subunit cohesin holocomplex showed a distinct chromatin binding activity in vitro, whereas the Smc1p-Smc3p dimer was unable to bind chromatin. Histone tails and ATP are dispensable for cohesin binding to chromatin in this reaction. A model for cohesin association with chromatin is proposed.

Adenosine Triphosphate↗

PubMed2003

Study of ABCA1 function in transgenic mice.

The ATP-binding cassette transporter A1 (ABCA1), identified in 1999 as the gene defective in Tangier disease, promotes efflux of cellular cholesterol from macrophages and other peripheral tissues to apolipoprotein acceptors. These ABCA1-mediated processes are anticipated to have antiatherogenic properties, prompting the development of pharmacological agents that increase ABCA1 gene expression as well as the establishment of ABCA1-transgenic mouse lines. Preliminary studies of ABCA1-Tg mice seem to validate the selection of this transporter as a therapeutic target for the treatment of low HDL syndromes and cardiovascular disease but have also raised new questions regarding the function of ABCA1. In particular, the relative contribution of hepatic and peripheral ABCA1 to plasma HDL levels and to reverse cholesterol transport, as well as the potential role of ABCA1 in modulating the plasma concentrations of the apolipoprotein B-containing lipoproteins and protecting against atherosclerosis, seem to be promising areas of investigation. The present review summarizes the most recent studies and discusses insights provided by these transgenic mouse models.

ATP Binding Cassette Transporter 1↗

PubMed2003

Hepatic lipase expression in macrophages contributes to atherosclerosis in apoE-deficient and LCAT-transgenic mice.

Hepatic lipase (HL) has a well-established role in lipoprotein metabolism. However, its role in atherosclerosis is poorly understood. Here we demonstrate that HL deficiency raises the proatherogenic apoB-containing lipoprotein levels in plasma but reduces atherosclerosis in lecithin cholesterol acyltransferase (LCAT) transgenic (Tg) mice, similar to results previously observed with HL-deficient apoE-KO mice. These findings suggest that HL has functions that modify atherogenic risk that are separate from its role in lipoprotein metabolism. We used bone marrow transplantation (BMT) to generate apoE-KO and apoE-KO x HL-KO mice, as well as LCAT-Tg and LCAT-Tg x HL-KO mice, chimeric for macrophage HL gene expression. Using in situ RNA hybridization, we demonstrated localized production of HL by donor macrophages in the artery wall. We found that expression of HL by macrophages enhances early aortic lesion formation in both apoE-KO and LCAT-Tg mice, without changing the plasma lipid profile, lipoprotein lipid composition, or HL and lipoprotein lipase activities. HL does, however, enhance oxidized LDL uptake by peritoneal macrophages. These combined data demonstrate that macrophage-derived HL significantly contributes to early aortic lesion formation in two independent mouse models and identify a novel mechanism, separable from the role of HL in plasma lipoprotein metabolism, by which HL modulates atherogenic risk in vivo.

Animals↗

PubMed2002

Role of the hepatic ABCA1 transporter in modulating intrahepatic cholesterol and plasma HDL cholesterol concentrations.

The current model for reverse cholesterol transport proposes that HDL transports excess cholesterol derived primarily from peripheral cells to the liver for removal. However, recent studies in ABCA1 transgenic mice suggest that the liver itself may be a major source of HDL cholesterol (HDL-C). To directly investigate the hepatic contribution to plasma HDL-C levels, we generated an adenovirus (rABCA1-GFP-AdV) that targets expression of mouse ABCA1-GFP in vivo to the liver. Compared with mice injected with control AdV, infusion of rABCA1-GFP-AdV into C57Bl/6 mice resulted in increased expression of mouse ABCA1 mRNA and protein in the liver. ApoA-I-dependent cholesterol efflux was increased 2.6-fold in primary hepatocytes isolated 1 day after rABCA1-GFP-AdV infusion. Hepatic ABCA1 expression in C57Bl/6 mice (n = 15) raised baseline levels of TC, PL, FC, HDL-C, apoE, and apoA-I by 150-300% (P < 0.05 all). ABCA1 expression led to significant compensatory changes in expression of genes that increase hepatic cholesterol, including HMG-CoA reductase (3.5-fold), LDLr (2.1-fold), and LRP (5-fold) in the liver. These combined results demonstrate that ABCA1 plays a key role in hepatic cholesterol efflux, inducing pathways that modulate cholesterol homeostasis in the liver, and establish the liver as a major source of plasma HDL-C.

ATP Binding Cassette Transporter 1↗

PubMed2002

Comparative genome analysis of potential regulatory elements in the ABCG5-ABCG8 gene cluster.

The excretion of sterols from the liver and intestine is regulated by the ABCG5 and ABCG8 transporters. To identify potential regulatory elements, 152 kb of the human ABCG5-ABCG8 gene cluster was sequenced and comparative genome analysis was performed. The two genes are oriented in a head-to-head configuration and are separated by a 374-bp intergenic region, which is highly conserved among several species. Using a reporter construct, the intergenic region was found to act as a bidirectional promoter. A conserved GATA site in the intergenic region was shown by site-directed mutagenesis to act as a repressor for the ABCG5 promoter. The intergenic region was also shown to be partially responsive to treatment by LXR agonists. In summary, several potential regulatory elements were found for the ABCG5 and ABCG8 genes, and the intergenic region was found to act as a bidirectional promoter.

Animals↗

PubMed2002

Identification of mouse and human macrophages as a site of synthesis of hepatic lipase.

Hepatic lipase (HL) is synthesized by the liver and is also present in steroidogenic tissues. As both a lipolytic enzyme and a ligand that facilitates the cellular uptake of lipoproteins, HL plays a major role in lipoprotein metabolism and may modulate atherogenic risk. However, HL has not been directly implicated in lesion development. In the present study we demonstrate that HL is also synthesized by mouse and human macrophages. Northern analysis and real time RT-PCR showed that HL mRNA is present in mouse peritoneal macrophages, RAW-264.7, and IC-21 cells. The levels of HL mRNA in mouse peritoneal macrophages were approximately 10-30% that of mouse liver. HL protein was identified by Western blot analyses in human monocyte-derived macrophages, THP, RAW-264.7, and mouse peritoneal macrophages following fractionation by heparin-sepharose affinity chromatography. These combined findings establish that HL is synthesized de novo by macrophages as well as liver, and raises the possibility that HL may have a direct role in the pathogenesis of atherosclerosis.

Animals↗