Non-redundant roles for LXRalpha and LXRbeta in atherosclerosis susceptibility in low density lipoprotein receptor knockout mice

Abstract

The liver X receptors LXRalpha and LXRbeta play critical roles in maintaining lipid homeostasis by functioning as transcription factors that regulate genetic networks controlling the transport, catabolism, and excretion of cholesterol. The studies described in this report examine the individual anti-atherogenic activity of LXRalpha and LXRbeta and determine the ability of each subtype to mediate the biological response to LXR agonists. Utilizing individual knockouts of LXRalpha and LXRbeta in the Ldlr(-/-) background, we demonstrate that LXRalpha has a dominant role in limiting atherosclerosis in vivo. Functional studies in macrophages indicate that LXRalpha is required for a robust response to LXR ligands, whereas LXRbeta functions more strongly as a repressor. Furthermore, selective knockout of LXRalpha in hematopoietic cells and rescue experiments indicate that the anti-atherogenic activity of this LXR subtype is not restricted to macrophages. These studies indicate that LXRalpha plays a selective role in limiting atherosclerosis in response to hyperlipidemia.

Fig. 1.

Plasma lipid levels in double knockout mice. Ldlr ^−/−(n = 12), Ldlr ^−/−/Lxrα^−/− (n = 9), and Ldlr ^−/−/Lxrβ^−/− (n = 12) mice were fed a Western diet for 20 weeks, and plasma triglyceride (A) and total cholesterol (B) levels were determined.

Fig. 2.

Atherosclerosis in double knockout mice. Mice were fed a Western diet for 20 weeks, and atherosclerosis was quantitated by root section (A) and en face analysis of the aorta (B) as described in the “Methods.” (C) Representative Sudan IV stained aortas. *Significantly different from Ldlr ^−/−controls.

Fig. 3.

LXR agonist activity in double knockout mice. Mice were exposed to a Western diet for 12 weeks and dosed daily with vehicle or 10 mg/kg T090137. The Ldlr ^−/−/Lxrβ^−/− + T0901317 group was switched from 10 mg/kg to 3 mg/kg at week 4. After completion of the study plasma triglycerides (A) and total plasma cholesterol (B) were measured. *Significantly different from strain-matched vehicle controls. **Significantly different from vehicle-treated Ldlr^−/− mice. Atherosclerosis was measured by quantitation of root sections (C) and by en face analysis of the entire aorta (D). *Significantly different from strain-matched vehicle controls. **Significantly different from vehicle-treated Ldlr ^−/− mice.

Fig. 4.

LXR target gene expression in bone marrow derived macrophages. Bone marrow derived macrophages isolated from C57BL/6, Lxrα^−/−, Lxrβ^−/−, and Lxrα^−/−/Lxrβ^−/− mice were treated with vehicle or increasing concentrations of T0901317 for 48 h. After agonist treatment total RNA was isolated, and the mRNA levels of ABCA1 (A and D), ABCG1 (B and E), and SREBP1c (C and F) were determined by quantitative PCR. *Significantly different from C57BL/6 controls (P ≤ 0.05). ABCA1, ATP binding cassette transporter A1; ABCG1, ATP binding cassette transporter G1; SREBP, sterol-regulatory element binding protein.

Fig. 5.

LXRα activity is required in bone marrow derived and nonbone marrow cells. Recipient mice (as defined in the figure) were irradiated and reconstituted with bone marrow from mice of the appropriate genotype. Four weeks after recovery from the transplant, mice were exposed to the Western diet, and atherosclerosis was quantitated by en face analysis after an additional 8 weeks. *Statistically significant difference from Ldlr ^−/− to Ldlr ^−/− controls (see text for P values). **Statistically significant difference from Ldlr ^−/−/Lxrα^−/− to Ldlr ^−/−/Lxrα^−/− controls (see text for P values).