Note to users. If you're seeing this message, it means that your browser cannot find this page's style/presentation instructions -- or possibly that you are using a browser that does not support current Web standards. Find out more about why this message is appearing, and what you can do to make your experience of our site the best it can be.

Subscribe

Logo for

Science 331 (6024): 1621-1624

Copyright © 2011 by the American Association for the Advancement of Science

FGF19 as a Postprandial, Insulin-Independent Activator of Hepatic Protein and Glycogen Synthesis

Serkan Kir1, Sara A. Beddow2, Varman T. Samuel2, Paul Miller3,*, Stephen F. Previs3,*, Kelly Suino-Powell4, H. Eric Xu4, Gerald I. Shulman2,5, Steven A. Kliewer1,6,{dagger}, and David J. Mangelsdorf1,7,{dagger}

1 Department of Pharmacology, University of Texas Southwestern Medical Center, 6001 Forest Park Road, Dallas, TX 75390, USA.
2 Department of Internal Medicine, Yale University School of Medicine, New Haven, CT 06510, USA.
3 Department of Nutrition, Case Western Reserve University, Cleveland, Ohio, 44106, USA.
4 Laboratory of Structural Sciences, Van Andel Research Institute, 333 Bostwick Avenue Northeast, Grand Rapids, MI 49503, USA.
5 Howard Hughes Medical Institute, Yale University School of Medicine, New Haven, CT 06510, USA.
6 Department of Molecular Biology, University of Texas Southwestern Medical Center, Dallas, TX 75390, USA.
7 Howard Hughes Medical Institute, University of Texas Southwestern Medical Center, Dallas, TX 75390, USA.


Figure 1
View larger version (36K):
[in this window]
[in a new window]

 
Fig. 1. Stimulation by FGF19 of signaling pathways that regulate protein synthesis in liver. (A to C) Mice fasted overnight were injected intravenously (i.v.) with vehicle or 1 mg of FGF19 protein per kg of body weight. This concentration is based on the optimal systemic dose to observe the physiologic effects of FGF19 on bile acid metabolism. The animals were killed 1 hour after the injections. Proteins from liver homogenates were separated by SDS–polyacrylamide gel electrophoresis (SDS-PAGE) and identified by Western blotting with the indicated antibodies. Results represent triplicate experiments. (D and E) Overnight serum-starved HepG2 cells were starved for amino acids in Hank’s buffered salt solution medium for 1 hour. Vehicle, wortmannin (200 nM), rapamycin (20 nM), U0126 (10 μM), or BI-D1870 (10 μM) was added for a further 1-hour treatment. The cells were treated with vehicle or 250 ng/ml FGF19 and harvested after 30 min. Proteins were identified by Western blotting with the indicated antibodies. BI-D1870 treatment blocks the negative-feedback effect of p90RSK on ERK, which results in increased basal phosporylation of ERK and p90RSK. Numbers below blots represent fold-change relative to the vehicle group. Asterisk (*) marks nonspecific band (C).

 

Figure 2
View larger version (11K):
[in this window]
[in a new window]

 
Fig. 2. Increased rates of global protein synthesis and albumin synthesis in mouse liver treated with FGF19. (A) Mice fasted overnight received 0.5 ml 2H2O intraperitoneally (i.p.); 90 min later, the animals were refed or kept fasted for 6 hours and killed (n = 10). Protein samples were hydrolyzed, and 2H labeling of alanine was determined by mass spectrometry. (B and C) Mice fed ad libitum received 0.5 ml 2H2O; 90 min later (at 6 p.m.), vehicle or 1 mg/kg FGF19 was injected subcutaneously. The next morning (8 a.m.), animals were injected again with the same dose and, 6 hours later, were killed (n = 10). Protein samples were hydrolyzed, and 2H labeling of alanine was determined by mass spectrometry. For albumin synthesis, 2H incorporation into plasma albumin was measured in the same way. (D) Over a 3-day period, mice (n = 6) received vehicle or 1 mg/kg FGF19 subcutaneously 3 times at 6 p.m. and once on the day they were killed at 8 a.m. Then, 6 hours after the last injection, the livers were harvested. Plasma albumin levels were determined with a Vitros 250 instrument. Values are means ± SEM. Statistics by two-tailed t test. *P < 0.05, **P < 0.005.

 

Figure 3
View larger version (23K):
[in this window]
[in a new window]

 
Fig. 3. FGF19 inhibits GSK3 signaling to increase liver glycogen in mice. (A) Mice fasted overnight were treated i.v. with vehicle or 1 mg/kg FGF19 and killed 10 min later. Proteins from liver homogenates were separated by SDS-PAGE and identified by Western blotting with the indicated antibodies. Results represent triplicate experiments. (B) The ability of glycogen synthase in the homogenates of the same livers to incorporate radiolabeled uridine 5'-diphosphate–glucose into glycogen in the absence and presence of glucose-6-phosphate was measured, and the ratio was shown as glycogen synthase activity (n = 3). (C) Mice fed ad libitum were injected subcutaneously with vehicle or 1 mg/kg FGF19 at 6 p.m. and the next morning at 8 a.m. Then, 6 hours after the last injection, the animals were killed, and liver weight and glycogen content were determined (n = 6). (D) Liver glycogen content was determined in wild-type and Fgf15–/– mice fed ad libitum (n = 5). (E) Oral glucose tolerance test in wild-type and Fgf15–/– mice (n = 6). Values are means ± SEM. Asterisks (*) refer to differences between wild-type and Fgf15–/– groups; number signs (#) refer to differences between Fgf15–/– and Fgf15–/– plus FGF19 groups. Statistics by two-tailed t test. *P < 0.05, **P < 0.005, #P < 0.05, ##P < 0.005.

 

Figure 4
View larger version (21K):
[in this window]
[in a new window]

 
Fig. 4. FGF19-induced glycogen synthesis is independent of insulin. (A) Overnight serum-starved HepG2 cells were pretreated with vehicle, wortmannin (200 nM), or BI-D1870 (10 μM) for 1 hour. The cells were lysed 30 min after vehicle or FGF19 (250 ng/ml) treatment. Proteins were identified by Western blotting with the indicated antibodies. Numbers below blots represent fold-change relative to the vehicle group. (B and C) Mice were treated i.p. with STZ (175 mg/kg). Eight days later, diabetic animals were chosen and treated with vehicle or 1 mg/kg FGF19 i.p. at 6 p.m. for seven consecutive days, and killed 6 hours after the last injection at 8 a.m.(n = 5 to 9). Liver glycogen content and plasma insulin levels were determined. *P < 0.05 is between control and STZ-vehicle groups; #P < 0.05 is between STZ-vehicle and STZ-FGF19 groups. (D) Three-hour hyperglycemic clamp study was performed on rats fasted overnight (n = 5 to 7). Animals were continuously infused with insulin and somatostatin to maintain low levels of insulin and glucagon and variably infused with glucose to maintain hyperglycemia. Net glycogen synthesis was determined by assessing the glycogen content in the clamped animals subtracted by the glycogen content of unclamped animals that were killed after the same duration of fasting. (E) Insulin and FGF19 act through different signaling pathways to coordinate overlapping but distinct postprandial responses in liver. (B, C, and D) Values are means ± SEM. Statistics by two-tailed t test. *P < 0.05, ***P < 0.0005.

 


To Advertise     Find Products


Science Signaling. ISSN 1937-9145 (online), 1945-0877 (print). Pre-2008: Science's STKE. ISSN 1525-8882