Fig. 6.

Circulating l-4F increases RCT from macrophages and cholesterol efflux from lipoproteins in Tg112 mice. A: Macrophage RCT was determined in Tg112 mice compared with littermate controls (WT). J774 cells were grown in culture and loaded with acetylated LDL that had been labeled with ³H-cholesterol. Loaded macrophages were injected intraperitoneally (∼2 × 10⁶ cells and 1 μCi per mouse) into both WT and Tg112 mice (n = 4/group). After 8 h, the mice were euthanized, and lumen rinse was collected. Following chloroform/methanol extraction of the rinse, label was determined by scintillation counting in both the chloroform fraction (³H-cholesterol) and the methanol/water fraction (³H-bile acids). B: We determined the effect of intravenously administered l-4F on macrophage RCT. We proceeded as in A, except that the mice were intravenously administered either 25 mg/kg ¹⁴C-l-4F or vehicle at both t = 4 h and t = 7 h (n = 5/group). As positive control, 3 WT animals had received 1 week pretreatment with the LXR agonist T0901317; these were otherwise treated as vehicle controls. l-4F significantly increased RCT into the intestinal lumen for both Tg112 (* P = 0.01) and WT (** P = 0.001) mice. LXR agonist treatment likewise increased RCT (*** P = 0.03). Data are expressed as percent of the corresponding vehicle controls. C: ¹⁴C label was determined in the luminal PBS rinse of both the Tg112 and WT mice from B, and dpm were converted to μg ¹⁴C-l-4F. D: In order to investigate the mechanism by which 4F modulates RCT, we repeated the basic experiments of A and B except that we used HDL and LDL rather than macrophages as the cholesterol source. Human LDL and HDL were prelabeled with ³H-cholesterol. The lipoproteins were then premixed with either ¹⁴C-l-4F or vehicle, and 100 μg HDL cholesterol + 100 μg LDL-C + 25 mg/kg ¹⁴C-l-4F (or vehicle) was injected via tail vein into both Tg112 and WT mice (n = 7 mice/group). At t = 3 h, the mice were given a second injection of either 25 mg/kg ¹⁴C-l-4F or vehicle. At t = 4 h the mice were euthanized, and radioactivity was determined as before. ¹⁴C-l-4F significantly increased cholesterol efflux into the SI lumen in both Tg112 (* P = 0.004) and WT (** P = 0.001) mice. The effect in Tg112 mice strongly suggests that l-4F can upregulate cholesterol efflux via the TICE pathway. In this experiment, however, l-4F also significantly increased cholesterol efflux in WT mice compared with Tg112 mice (*** P = 0.04), suggesting that l-4F also affects cholesterol efflux through the hepatobiliary pathway in this context. Error is reported as SEM.