Atherosclerosis (ath"er-o-skleh-RO'sis) comes from the Greek words athero (meaning gruel or paste) and sclerosis (hardness). It's the name of the process in which deposits of fatty substances, cholesterol, cellular waste products, calcium and other substances build up in the inner lining of an artery. This buildup is called plaque. It usually affects large and medium-sized arteries. Some hardening of arteries often occurs when people grow older. Plaques can grow large enough to significantly reduce the blood's flow through an artery. But most of the damage occurs when they become fragile and rupture. Plaques that rupture cause blood clots to form that can block blood flow or break off and travel to another part of the body. If either happens and blocks a blood vessel that feeds the heart, it causes a heart attack. If it blocks a blood vessel that feeds the brain, it causes a stroke. And if blood supply to the arms or legs is reduced, it can cause difficulty walking and eventually gangrene.

Atherosclerotic cardiovascular disease is the leading cause of mortality in industrialized nations, accounting for nearly 50% of all deaths. Risk factors for the development of atherosclerosis include both genetic and environmental factors. Numerous epidemiological studies have identified decreased high-density lipoprotein (HDL) cholesterol and increased low-density lipoprotein (LDL) cholesterol as major contributors to atherogenesis. Although the exact atheroprotective mechanism of HDL is not known, it is speculated that the reverse transport of cholesterol from arterial cells to the liver may be the cause.

The earliest atherosclerotic lesions, the fatty streak, are characterized by the accumulation of lipid-laden macrophages (foam cells) in the arterial wall. The Foam cells develop following the uptake up large amounts of oxidized LDL (oxLDL) by scavenger receptors such as CD36, SR-A and SR-BI. Unlike the LDL receptor, scavenger receptor expression is not subject to feedback inhibition by intracellular cholesterol levels. Initially, macrophage uptake oxLDL can be viewed as a physiological response designed to eliminate harmful extra cellular debris (apoptotic cells and oxLDL). It becomes pathological only when the cell is overwhelmed by the lipid load, and becomes trapped within the lesion. So there should be some balance between the lipid uptake by the scavenger receptors and the lipid disposal by the LXRs.

Understanding of the mechanisms that regulate both the accumulation and elimination of lipids in macrophages is central to understanding the development of the atherosclerotic plaque. Recent studies have shown that liver X receptors (LXRs) play as an important mediator of the effects of oxLDL on cellular gene expression, which are also central components of cellular response to lipid loading.

Liver X receptors (LXRs)

The livers X receptors (LXRs) are nuclear receptors activated by oxy sterols that are now recognized to play an important role in the control of lipid homeostasis. LXRs have been implicated in the regulation of cholesterol and fatty acid metabolism in multiple tissues, including liver and intestine, as well as in macrophages. The importance of these receptors in physiological lipid metabolism suggests that they may also influence the development of metabolic disorders such as hyper lipidemia and atherosclerosis. Strong support for this idea has been provided by recent studies that directly linked LXR activity to the pathogenesis of atherosclerosis. These observations identify the LXR pathway as an attractive target for intervention in cardiovascular disease.

The endogenous ligands for these receptors are intermediates or end products of metabolic pathways. The two LXRs, LXR alpha and LXR beta, are activated by physiological concentrations of oxidized derivatives of cholesterol, such as 22(R)-hydroxycholesterol, 27-hydroxycholesterol and 24(S), 25-epoxycholesterol. LXR alpha and LXR beta share considerable sequence homology (77% identity in DNA- and ligand-binding domains), and appear to respond to the same endogenous ligands. LXR? is expressed at high levels in liver, intestine, adipose tissue and macrophages, whereas LXR beta is expressed ubiquitously.

LXR target genes

ATP binding cassette (ABC) transporters are integral membrane proteins that couple ATP to the transport of various substrates across cellular membranes. One LXR target gene that has garnered a lot of attention for its role in cholesterol and phospholipids transport is ABCA1. The ABCA1 protein is critical for the first step in the reverse cholesterol transport pathway: the efflux of excess cellular cholesterol to apolipoprotein acceptors. The importance of ABCA1 in systemic cholesterol metabolism has become clear from the study of patients with Tangier disease, who carry loss-of-function mutations in this gene. Tangier disease is characterized by the absence of plasma HDL, and the accumulation of cholesterol ester in the reticuloendothelial cells of multiple tissues, including tonsils, thymus, lymph nodes and spleen. Cells from Tangier patients are defective in their ability to efflux cholesterol.

Three other ABC transporters have also been identified as LXR targets: ABCG1, ABCG5 and ABCG8. ABCG1 is also induced in macrophages by lipid loading and treatment with hydroxy sterols. The function of ABCG1 is unknown, but it has been proposed to play a role in cholesterol efflux, perhaps by working in concert with ABCA1. Another rare genetic disorder in man, sitosterolemia, involves mutations in the LXR target genes ABCG5 and ABCG8. Genetic deficiency of these transporters leads to abnormal absorption of sitosterols (plant sterols) and a hyper absorption of cholesterol.

Another LXR target gene that is important in cholesterol homeostasis is apoE. In contrast to ABCA1, regulation of apoE by LXR is tissue-specific, occurring only in macrophages and adiposities. ApoE mediates the hepatic uptake of very-low-density lipoprotein (VLDL) and chylomicronremnants, and can serve as an extra cellular acceptor for cholesterol in the ABCA1 efflux pathway. The beneficial nature of apoE production by macrophages is clear from several animal studies. Mice expressing apoE only in macrophages are protected against atherosclerosis, whereas those specifically lacking apoE expression in macrophages are more susceptible. Recently, the entire apoE/ apoCI/ apoCII/ apoCIV gene cluster was shown to respond to LXR activation in both human and murine macrophages. Thus, LXR induces the expression of multiple lipoproteins that could serve as cholesterol acceptors in the context of an atherosclerotic lesion.

LXR has also been shown to control expression of two lipoprotein-remodeling enzymes: the cholesterol ester transfer protein (CETP) and the phospholipids transfer protein (PLTP). CETP mediates the transfer of HDL cholesterol esters to apoB-containing particles for return to the liver. Triglycerides are transferred to HDL in exchange. This modification of HDL by CETP makes HDL more susceptible to hydrolysis by hepatic lipase at the hepatocyte surface, which is an important component of the regeneration of small HDL particles and free apoAI that can re circulate in the reverse cholesterol transport pathway. PLTP has been identified as a key modulator of HDL metabolism, and might also be involved in reverse cholesterol transport. In addition, PLTP has recently been shown to regulate VLDL secretion from the liver. PLTP-deficient mice exhibit decreased levels of VLDL and LDL in an apoE-deficient or apoB-transgenic background. Therefore, some actions of LXR agonists on HDL and VLDL levels (see below) are consistent with the known roles for PLTP in lipoprotein metabolism. In particular, it seems likely that the ability of LXR agonists to raise plasma VLDL and triglyceride levels may involve PLTP.

LXRs have also been implicated in the control of fatty acid metabolism. Mice carrying a targeted disruption in the LXRs gene were noted to be deficient in expression of SREBP-1c (sterol regulatory element binding protein), FAS (fatty acid synthase), SCD-1 (steroyl CoA desaturase I) and ACC (acyl CoA carboxylase), in addition to defects in cholesterol metabolism. Administration of synthetic LXR ligands to mice triggers induction of the lipogenic pathway, and elevates plasma triglyceride levels. The demonstration that the SREBP-1c promoter is a direct target of LXR provided a straightforward explanation for the ability of LXR ligands to induce hepatic lipogenesis. In addition to effects on SREBP-1c, direct actions of LXR on certain lipogenic genes, such as FAS and PLTP, are also likely to contribute to the ability of LXR agonists to cause hyper triglyceridemia. Clearly, the lipogenic activity of LXR agonists represents a significant obstacle to their development as drugs.

Conclusion

The ability of synthetic LXR ligands to promote cellular cholesterol efflux and to inhibit atherosclerosis in animal models makes them potentially attractive agents for the modulation of human lipid metabolism. Their lipogenic activity, however, is a major limitation. The current generation of LXR ligands is potent, but not particularly selective for LXR? or LXR?. From a drug development standpoint, the most desirable compound would appear to be one that is a strong inducer of ABCA1 and apoE expression, yet lacks activity on the SREBP-1c and FAS promoters.

To date, more than a dozen LXR target genes have been identified (see box). The majority of known LXR target genes appear to have one of two biological functions; firstly, removal of excess cholesterol through efflux, catabolism or decreased absorption; and secondly, synthesis of fatty acids. Alterations in cholesterol and fatty acid metabolism each have the potential to influence the development of cardiovascular disease. The established functions of many of these LXR targets have fueled speculation as to how their pharmacological regulation by LXR ligands might impact metabolic disease.