Figure 2. Intestines from Ire1b^−/− Mice Secrete More Lipids with Chylomicrons.

Ire1b^+/+ (+/+; white bars) and Ire1b^−/− (−/−; black bars) mice were fed a high-cholesterol diet for 2 weeks and fed 1 μCi of [³H]cholesterol or [³H]triolein as well as 0.1 mg of cholesterol in 15 μl of olive oil.

(A and C) After 2 hr, plasma was used to measure radioactivity.

(B and D) Liver slices were digested with OptiSolv and used for radioactivity determinations.

(E–J) In another experiment, Ire1b^+/+ and Ire1b^−/− mice fasted overnight were injected intraperitoneally with poloxamer 407 (30 μg/mouse). After 1 hr, mice were fed 1 μCi [³H]cholesterol and 0.1 mg unlabeled cholesterol. Plasma was collected after 2 hr and used to measure radioactivity (E) as well as cholesterol (G) and triglyceride (I) mass in total plasma, HDL, and non-HDL lipoproteins. Plasma was separated by gel filtration to determine lipids in different lipoproteins (F, H, and J).

(K) Enterocytes were isolated from cholesterol-fed male Ire1b^+/+ and Ire1b^−/− mice and incubated with 1 μCi/ml of [³H]cholesterol for different lengths of time to study uptake. At each time point, cellular lipids were extracted with isopropanol and counted.

(L) To study secretion, cells were labeled with [³H]cholesterol for 1 hr, washed, and chased for various lengths of time in the presence of 1.2 mM oleic acid. Radioactivity was measured in conditioned media.

(M and N) To identify lipoproteins carrying cholesterol, enterocytes from cholesterol-fed male Ire1b^+/+ and Ire1b^−/− mice were labeled for 1 hr with 1 μCi/ml of [³H]cholesterol, washed, and chased for 2 hr in the presence of 1.2 mM oleic acid-containing micelles. Conditioned media were used to measure radioactivity (M) or for density gradient ultracentrifugation. Fractions were collected from the top and assayed for radioactivity (N).

Each measurement was performed in triplicate with n = 3 mice per group. Values are mean ± SD. *p < 0.05, **p < 0.01, ***p < 0.001 versus Ire1b^+/+.