Aerobic capacity and exercise mediate protection against hepatic steatosis via enhanced bile acid metabolism

Abstract

High cardiorespiratory fitness and exercise show evidence of altering bile acid (BA) metabolism and are known to protect or treat diet-induced hepatic steatosis, respectively. Here, we tested the hypothesis that high intrinsic aerobic capacity and exercise both increase hepatic BA synthesis measured by the incorporation of ²H₂O. We also leveraged mice with inducible liver-specific deletion of Cyp7a1 (LCyp7a1KO), which encodes the rate-limiting enzyme for BA synthesis, to test if exercise-induced BA synthesis is critical for exercise to reduce hepatic steatosis. The synthesis of hepatic BA, cholesterol, and de novo lipogenesis was measured in rats bred for either high (HCR) vs. low (LCR) aerobic capacity consuming acute and chronic high-fat diets. HCR rats had increased synthesis of cholesterol and certain BA species in the liver compared to LCR rats. We also found that chronic exercise with voluntary wheel running (VWR) (4 weeks) increased newly synthesized BAs of specific species in male C57BL/6J mice compared to sedentary mice. Loss of Cyp7a1 resulted in fewer new BAs and increased liver triglycerides compared to controls after a 10-week high-fat diet. Additionally, exercise via VWR for 4 weeks effectively reduced hepatic triglycerides in the high-fat diet-fed control male and female mice as expected; however, exercise in LCyp7a1KO mice did not lower liver triglycerides in either sex. These results show that aerobic capacity and exercise increase hepatic BA metabolism, which may be critical for combatting hepatic steatosis.

Keywords: Cyp7a1; cholesterol synthesis; de novo lipogenesis; liver; metabolism; non-alcoholic fatty liver disease.

Figure 1.

Serum Bile Acid Composition. A. Serum bile acid composition and total bile acids from rats during a 1-week study (n=8). B. Serum bile acid composition and total bile acids from rats during a 20-week study (n=10).

Figure 2.

Intestinal and Fecal Bile Acid Content and Fecal Energy Loss. A. Intestinal bile acid measurements from rats during the 1-week study (n=8). B. Intestinal bile acid measurements from rats during the 20-week study (n=10). C. Fecal bile acid content from rats during the 1-week study (n=16). D. Fecal bile acid content from rats during the 20-week study (n=10). E. Fecal energy loss from rats during the 1-week study (n=16). F. Fecal energy loss from rats during the 20-week study (n=10). Data represented as means ± SEM. îndicates effect of strain within diet (^p<0.05, ^^p<0.01, ^^^p<0.001).

Figure 3.

DNL, Cholesterol Synthesis, and Bile Acid synthesis. A. DNL as measured by ²H incorporation into % newly synthesized hepatic palmitate from rats during the 1-week study (n=8). B. Cholesterol synthesis as measured by ²H incorporation into % newly synthesized hepatic cholesterol from rats during a 1-week study (n=8). C. Bile acid synthesis as measured by ²H incorporation into % newly synthesized T-αMCA, T-βMCA, T-CA, T-CDCA, and T-DCA from rats during a 1-week study (n=6–8). D. Bile acid synthesis as measured by ²H incorporation into % newly synthesized T-αMCA, T-βMCA, T-CA, T-CDCA, and T-DCA from rats during a 20-week study (n=10). Data represented as means ± SEM. *indicates effect of diet within strain and fed state (*p<0.05, **p<0.01, ***p<0.001); îndicates effect of strain within diet and fed state (^p<0.05, ^^p<0.01, ^^^p<0.001); ‡indicates effect of fed state within strain and diet (‡p<0.05, ‡‡p<0.01, ‡‡‡p<0.001).

Figure 4.

Cholesterol and Bile Acid Synthesis Gene Expression in HCR and LCR rats during a 1-week study. A. Gene expression for the cholesterol synthesis protein, HMG-CoA reductase (HMGCR). B. Gene expression for the rate-limiting protein in bile acid synthesis, Cyp7a1. C. Gene expression for a mitochondrial protein involved in the bile acid synthetic pathway, Cyp27a1. D. Gene expression for the protein responsible for determining bile acid pool composition, Cyp8b1. E. Gene expression for a protein in the alternative bile acid synthetic pathway, Cyp7b1. F. Gene expression for the Bile Acid-CoA:Amino Acid N-Acyltransferase (BAT) enzyme which controls the conjugation of bile acids to an amino acid synthesis (BAAT). G. Gene expression for the hepatic nuclear receptor involved in redundant feedback regulation of bile acids, FXR (NR1H4). H. Gene expression for a hepatic receptor involved in bile acid feedback from the intestines, FGFR4. I. Gene expression for a transcription factor that promotes cholesterol synthesis, SREBP-2. J. Gene expression for a mitochondrial protein involved in the bile acid synthetic pathway, SREBF1. K. Gene expression for the transcriptional co-activator peroxisome gamma co-activator 1 alpha (PGC1α), a master regulator of mitochondrial biogenesis and genes involved in energy metabolism (PGC1α). L. Gene expression for a transcription factor that helps regulate fatty acid oxidation in the liver, PPARα (PPARα). Data represented as normalized gene expression values with units as log-transformed counts per million (means ± SEM; n=4). *Indicates effect of diet within strain and fed state (*p<0.05, **p<0.01, ***p<0.001); îndicates effect of strain within diet and fed state (^p<0.05, ^^p<0.01, ^^^p<0.001); ‡indicates effect of fed state within strain and diet (‡p<0.05, ‡‡p<0.01, ‡‡‡p<0.001).

Figure 5.

Bile Acid Synthesis Measures in VWR Mice. Data shows bile acid synthesis as measured by ²H incorporation into % newly synthesized A. T-CA. B. T-αMCA. C. T-βMCA. D. T-CDCA and E. T-DCA. Measurements from mice (n=8) on a HFD (Control) that either remained sedentary (SED) or were given running wheels (VWR) for 4 weeks. Data represented as means ± SEM. *indicates effect of diet within running or sedentary condition (*p<0.05, **p<0.01, ***p<0.001); îndicates effect of running or sedentary condition within diet (^p<0.05, ^^p<0.01, ^^^p<0.001).

Figure 6.

Liver triglyceride and bile acid content in liver-specific Cyp7a1 knockout mice with VWR. A. Liver Cyp7a1 gene expression. B. Liver triglyceride content. C. Representative hematoxylin and eosin stains. D. Total liver bile synthesis. E. Liver T-CA bile acid synthesis. F. Liver T-αMCA bile acid synthesis. G. Liver T-βMCA bile acid synthesis. H. Liver T-CDCA bile acid synthesis. I. Liver T-DCA bile acid synthesis. Data represented as means ± SEM, (n=6–8). * Indicates main effect of LCyp7a1KO within sex (*P<0.05), # indicates main effect of VWR within sex (P<0.05), ε indicates an LCyp7a1KO and VWR interaction within sex, ^ P<0.05 vs. indicated group.