Novel Effects of the Acyl-Coenzyme A:Cholesterol Acyltransferase Inhibitor 58-035 on Foam Cell Development in Primary Human Monocyte–Derived Macrophages

Abstract

Abstract-We examined the effect of acyl-coenzyme A:cholesterol acyltransferase (ACAT) inhibitors on intracellular cholesterol stores in primary human monocyte-derived macrophages (HMMs) during foam cell formation. HMMs were exposed to acetylated low density lipoprotein (acLDL, 500 g protein per mL) with or without 58-035 (1 to 10 g/mL) or CI-976 (2 g/mL) for 2 to 48 hours. Total cholesterol (TC) and esterified cholesterol (EC) mass was significantly lower while unesterified cholesterol (UC) increased slightly in cells incubated with acLDL plus ACAT inhibitors. Sterol mass was also measured in cells coincubated with acLDL (500 g protein per mL) with or without 58-035 (2 g/mL), high density lipoprotein (HDL, 400 g protein per mL), or HDLϩ58-035 for 48 hours. TC and EC were 23% and 55% lower, respectively (PϽ0.0004), while UC was 11% higher (PϽ0.04) in cells incubated with acLDL plus 58-035. In contrast, coincubation with HDL alone did not significantly affect TC, EC, or UC mass compared with acLDL alone. The effect of 58-035 could not be explained by cytotoxicity, because adenine release, secreted lactate dehydrogenase, glucose utilization, and cell protein were similar in cells exposed to acLDL regardless of the presence of 58-035. We investigated several potential mechanisms for the decreased TC mass, including increased UC efflux and decreased acLDL binding and uptake. Efflux was measured in cells exposed to [1,2-3 H]cholesteryl oleate-labeled acLDL, unlabeled control acLDL, and native untreated acLDL (500 g protein per mL) with or without 58-035 (5 g/mL) for 24 or 48 hours. UC efflux increased in a time-dependent manner from cells exposed to acLDL plus 58-035 compared with cells exposed to acLDL alone (PϽ0.04). High-affinity binding was measured in cells exposed to 125 I-acLDL (5 g protein per mL) with or without excess unlabeled acLDL (100 or 500 g protein per mL) for 4 hours at 4°C. Specific acLDL binding, uptake, and total degradation were significantly lower when 58-035 was present during cholesterol enrichment compared with cells exposed to acLDL alone (PϽ0.001). Unlike the effects of ACAT inhibitors on foam cell formation in rodent macrophages, these compounds lowered TC accumulation in HMMs during foam cell formation by limiting the uptake of acLDL and enhancing UC efflux. They may offer promise as drug therapies for atherosclerosis.

Enrichment of Endoplasmic Reticulum with Cholesterol Inhibits Sarcoplasmic-Endoplasmic Reticulum Calcium ATPase-2b Activity in Parallel with Increased Order of Membrane Lipids

“…Thus, ER calcium depletion, which is an early event in macrophage FC loading, could be a key upstream event leading to macrophage death. Interestingly, Rodriguez and colleagues (63) have reported that macrophage-like cells derived from cultured human blood monocytes do not accumulate large amounts of FC and therefore, as expected, do not die when incubated with acetyl-LDL and an ACAT inhibitor. This finding is associated with a high level of cholesterol efflux and down-regulation of the scavenger receptor.…”

Neutral-Lipid Analysis Reveals Elevation of Acylglycerols and Lack of Cholesterol Esters in Plasmodium falciparum -Infected Erythrocytes

“…Although we failed to detect cholesterol esterification activity, PfDGAT is highly homologous to the ACAT and DGAT families (2,12), and therefore, we treated P. falciparum-infected red cells with Sandoz 58-035 for 2 h at a concentration (10 g/ml) known to inhibit 99% of ACAT activity in mammalian cells (14,17). However, we failed to induce any change in the incorporation of [ 3 H]oleic acid into either TAG or its intermediates or show any deleterious effects on plasmodial growth (data not shown).…”

Inhibitory Effect of Acyl-CoA: Cholesterol Acyltransferase Inhibitor-Low Density Lipoprotein Complex on Experimental Atherosclerosis.

“…The lipid (TC, FC, and TG) and protein concentrations of KV-LDL complex were similar to the corresponding concentrations of LDL. The KV-2920 concentration was 615.2 mg/ml, and the amount of KV-2920 contained in the KV-LDL complex was 800Ϯ40 mol/mol LDL (LDL molecular weight of 5.5ϫ10 5 based on the protein concentration). The electrophoresis pattern of the KV-LDL complex was the same as that of LDL (data not shown).…”

“…In fact, many studies have reported that an ACAT inhibitor prevents foam cell formation by macrophages and the development of atherosclerotic plaques. [5][6][7] However, most ACAT inhibitors have problems associated with low oral bioavailability and adrenal and/or hepatic toxicity. 8,9) In particular, adrenotoxicity is a major problem if an ACAT inhibitor is to be used in clinical situations.…”