Volume 134, Issue 4, Pages 1127-1136 (April 2008) Dramatically Increased Intestinal Absorption of Cholesterol Following Hypophysectomy Is Normalized by Thyroid Hormone  Cecilia Gälman, Ylva Bonde, Manuela Matasconi, Bo Angelin, Mats Rudling  Gastroenterology  Volume 134, Issue 4, Pages 1127-1136 (April 2008) DOI: 10.1053/j.gastro.2008.01.032 Copyright © 2008 AGA Institute Terms and Conditions

Figure 1 Effects of cholesterol/fat feeding (2% C) of intact and Hx rats (5/group) for 1 week. (A) Serum total cholesterol (solid bars) and triglycerides (open bars). *P < .01, **P < .001 vs intact. (B) Cholesterol lipoprotein profiles by FPLC. Lines show averages of all individuals/group. VLDL/CM, very-low-density lipoproteins/chylomicrons. (C) Hepatic total cholesterol (solid bars) and triglycerides (open bars). *P < .01, **P < .001 vs intact and Hx. (D) LDLR mRNA levels corrected for 18S of pooled liver samples from each group run in triplicate. (E) Cyp7a1 mRNA levels corrected for 18S. Solid bars show means of all individuals in each group Cyp7a1 protein corrected for β-actin (open bars) in liver membranes from each individual. (F) Microsomal Cyp7a1 enzymatic activity (solid bars) and serum levels of C4 (open bars). All results show means ± SEM. Gastroenterology 2008 134, 1127-1136DOI: (10.1053/j.gastro.2008.01.032) Copyright © 2008 AGA Institute Terms and Conditions

Figure 2 Cholesterol absorption in intact (n = 8) and Hx rats (n = 10). (A) Fecal dual-isotope measurement of intestinal cholesterol absorption. (B) Serum plant sterols. All results show mean ± SEM. *P < .05, **P < .01, ***P < .0001, t test vs intact. Gastroenterology 2008 134, 1127-1136DOI: (10.1053/j.gastro.2008.01.032) Copyright © 2008 AGA Institute Terms and Conditions

Figure 3 Effects of cholesterol absorption blockade on Hx rats during cholesterol/fat feeding. Hx rats (6/group) received standard chow or chow enriched with 0.4% or 2% cholesterol/10% corn oil ± ezetimibe (EZE) (3 mg/kg/day) for 1 week. (A) Cholesterol lipoprotein profiles by FPLC. Lines show averages from all individuals/group. (B) Enlargement of the cholesterol lipoprotein profiles for Hx rats on standard chow ± ezetimibe to clarify the reduction (P < .05) of lipoprotein cholesterol by EZE (solid lines, mean; broken lines, ±SEM). Gastroenterology 2008 134, 1127-1136DOI: (10.1053/j.gastro.2008.01.032) Copyright © 2008 AGA Institute Terms and Conditions

Figure 4 Effects of cholesterol absorption inhibition by ezetimibe on Hx rats during cholesterol/fat feeding. (A) Serum total cholesterol (shaded bars) and triglycerides (solid bars). (B) Serum plant sterols. (C) Serum C4 levels. (D) HMG-CoAR microsomal activity. (E) LDLR mRNA (shaded bars) and SREBP2 mRNA (solid bars) determined on all individuals corrected for 18S. All results are expressed as mean ± SEM. ***P < .001 vs Hx, ###P < .001 vs 0.4% chol, †††P < .001 vs 2% chol. Gastroenterology 2008 134, 1127-1136DOI: (10.1053/j.gastro.2008.01.032) Copyright © 2008 AGA Institute Terms and Conditions

Figure 5 Effects of hormonal substitution of Hx rats on serum plant sterols. Hx rats (n = 52) were substituted with GH (1.5 mg/kg/day), T4 (42 μg/kg/day), and cortisone (C) (400 μg/kg/day) for 1 week. Intact rats (I) (n = 5) and the Hx control group received subcutaneous injections of NaCl. †P < .05 vs intact; *P < .01, **P < .001 vs Hx; ns, not significant. Gastroenterology 2008 134, 1127-1136DOI: (10.1053/j.gastro.2008.01.032) Copyright © 2008 AGA Institute Terms and Conditions

Figure 6 Effect of hypophysectomy and substitution of Hx rats with T4 (42 μg/kg/day) for 1 week. Intact (n = 10), Hx (n = 10), and Hx rats treated with T4 (Hx+T4) (n = 9). (A) Cholesterol absorption was determined by the fecal dual-isotope method. (B) Intestinal ABCG5 (solid bars), ABCG8 (shaded bars), and NPC1L1 mRNA (open bars). (C). Hepatic ABCG5 (open bars) and ABCG8 (solid bars) mRNA. (D) Assay of fecal neutral sterols cholesterol (solid bars), coprostanol (medium shaded), cholestanol (dark shaded), campesterol (open bars), and coprostane-3-one (light bars). Comparisons shown are intact vs Hx and Hx vs Hx+T4. *P < .01; **P < .001; ns, not significant. All results shown are means ± SEM. mRNA was determined in all individual animals. Gastroenterology 2008 134, 1127-1136DOI: (10.1053/j.gastro.2008.01.032) Copyright © 2008 AGA Institute Terms and Conditions

Figure 7 Hepatic ABCG5 mRNA (solid bars) and ABCG8 (open bars) in (A) experiment described in legend of Figure 6. ###P < .001 vs intact, *P < .05, ***P < .001 vs Hx. (B) Experiment on Hx animals only as described in legends of Figures 4 and 5. *P < .05, **P < .01, ***P < .001 vs Hx. (C) Experiment described in legends of Figures 1 and 2. ###P < .001 vs intact, ***P < .001 vs Hx. All results shown are means from all individuals/group ± SEM. Gastroenterology 2008 134, 1127-1136DOI: (10.1053/j.gastro.2008.01.032) Copyright © 2008 AGA Institute Terms and Conditions

Figure 8 Effect of T4 treatment (42 μg/kg/day for 8 days) of intact and Hx rats (intact [n = 9], intact+T4 [n = 8], Hx [n = 9], and Hx+T4 [n = 5]) on (A) biliary secretion of cholesterol and (B) expression of hepatic ABCG5 mRNA (solid bars) and ABCG8 (open bars) corrected for 18S. Results shown are means of all individuals/group ± SEM. #P < .05, ##P < .01, ###P < .001 vs intact; ***P < .001 vs Hx. Gastroenterology 2008 134, 1127-1136DOI: (10.1053/j.gastro.2008.01.032) Copyright © 2008 AGA Institute Terms and Conditions