Inhibition of citrate cotransporter Slc13a5/mINDY by RNAi improves hepatic insulin sensitivity and prevents diet-induced non-alcoholic fatty liver disease in mice.
Non-alcoholic fatty liver disease is a world-wide health concern and risk factor for cardio-metabolic diseases. Citrate uptake modifies intracellular hepatic energy metabolism and is controlled by the conserved sodium-dicarboxylate cotransporter solute carrier family 13 member 5 (SLC13A5, mammalian homolog of INDY: mINDY). InDrosophila melanogaster andCaenorhabditis elegansINDY reduction decreased whole-body lipid accumulation. Genetic deletion ofSlc13a5in mice protected from diet-induced adiposity and insulin resistance. We hypothesized that inducible hepatic mINDY inhibition should prevent the development of fatty liver and hepatic insulin resistance.



Adult C57BL/6J mice were fed a Western diet (60% kcal from fat, 21% kcal from carbohydrate) ad libitum. Knockdown of mINDY was induced by weekly injection of a chemically modified, liver-selective siRNA for 8 weeks. Mice were metabolically characterized and the effect of mINDY suppression on glucose tolerance as well as insulin sensitivity was assessed with an ipGTT and a hyperinsulinemic-euglycemic clamp. Hepatic lipid accumulation was determined by biochemical measurements and histochemistry.



Within the 8 week intervention, hepatic mINDY expression was suppressed by a liver-selective siRNA by over 60%. mINDY knockdown improved hepatic insulin sensitivity (i.e. insulin-induced suppression of endogenous glucose production) of C57BL/6J mice in the hyperinsulinemic-euglycemic clamp. Moreover, the siRNA-mediated mINDY inhibition prevented neutral lipid storage and triglyceride accumulation in the liver, while we found no effect on body weight.



We show that inducible mINDY inhibition improved hepatic insulin sensitivity and prevented diet-induced non-alcoholic fatty liver disease in adult C57BL6/J mice. These effects did not depend on changes of body weight or body composition.
Department of Endocrinology, Diabetes and Nutrition, Center for Cardiovascular Research, Charité - University School of Medicine, Berlin, 10117, Germany; DZHK (German Center for Cardiovascular Research), Partner Site, Berlin, Germany. Electronic address: Sebastian.Brachs@charite.de.
Sebastian
S
Brachs
Sanofi-Aventis Deutschland GmbH, Industriepark Hoechst, Frankfurt am Main, 65926, Germany. Electronic address: Angelika.Winkel@sanofi.com.
Angelika F
AF
Winkel
Department of Endocrinology, Diabetes and Nutrition, Center for Cardiovascular Research, Charité - University School of Medicine, Berlin, 10117, Germany; DZHK (German Center for Cardiovascular Research), Partner Site, Berlin, Germany. Electronic address: Hui.Tang@charite.de.
Hui
H
Tang
Department of Endocrinology, Diabetes and Nutrition, Center for Cardiovascular Research, Charité - University School of Medicine, Berlin, 10117, Germany; DZHK (German Center for Cardiovascular Research), Partner Site, Berlin, Germany; Section of Metabolic Vascular Medicine, Medical Clinic III and Paul Langerhans Institute Dresden (PLID), a Member of the German Diabetes Center (DZD), Technische Universität, Dresden, 01307, Germany. Electronic address: Andreas.Birkenfeld@uniklinikum-dresden.de.
Andreas L
AL
Birkenfeld
Sanofi-Aventis Deutschland GmbH, Industriepark Hoechst, Frankfurt am Main, 65926, Germany. Electronic address: Bodo.Brunner@sanofi.com.
Bodo
B
Brunner
Sanofi-Aventis Deutschland GmbH, Industriepark Hoechst, Frankfurt am Main, 65926, Germany. Electronic address: Kerstin.Jahn-Hofmann@sanofi.com.
Kerstin
K
Jahn-Hofmann
Sanofi-Aventis Deutschland GmbH, Industriepark Hoechst, Frankfurt am Main, 65926, Germany. Electronic address: Daniel.Margerie@sanofi.com.
Daniel
D
Margerie
Sanofi-Aventis Deutschland GmbH, Industriepark Hoechst, Frankfurt am Main, 65926, Germany. Electronic address: Hartmut.Ruetten@sanofi.com.
Hartmut
H
Ruetten
Sanofi-Aventis Deutschland GmbH, Industriepark Hoechst, Frankfurt am Main, 65926, Germany. Electronic address: Dieter.Schmoll@sanofi.com.
Dieter
D
Schmoll
Department of Endocrinology, Diabetes and Nutrition, Center for Cardiovascular Research, Charité - University School of Medicine, Berlin, 10117, Germany; DZHK (German Center for Cardiovascular Research), Partner Site, Berlin, Germany. Electronic address: joachim.spranger@charite.de.
Joachim
J
Spranger
D000818:Animals; D002951:Citrates; D019343:Citric Acid; D029382:Dicarboxylic Acid Transporters / Q000502:physiology*; D004032:Diet; D007333:Insulin Resistance; D050356:Lipid Metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D065626:Non-alcoholic Fatty Liver Disease; D034622:RNA Interference; D027981:Symporters / Q000502:physiology*
D002951:Citrates; D029382:Dicarboxylic Acid Transporters; C487219:Slc13a5 protein, mouse; D027981:Symporters; D019343:Citric Acid
2-DG, 2-Deoxy-d-glucose; Citrate transport; EE, energy expenditure; EGP, endogenous glucose production; FA, fatty acids; FLD, fatty liver disease; GIR, glucose infusion rate; HE clamp, hyperinsulinemic-euglycemic clamp; HFD, high-fat diet; IEX, anion-exchange high-performance liquid chromatography; INDY, ‘I'm not dead Yet’; INDY/Slc13a5; Insulin resistance; KO, knockout; Lipid accumulation; ORO, oil red O; RER, respiratory exchange ratio; SCR, non-silencing scrambled control siRNA; SKM, skeletal muscle; Steatosis; T2D, type-2 diabetes; TCA, tricarboxylic acid; WAT, white adipose tissue; WD, western diet; e, epididymal; mINDY, Slc13a5/SLC13A5; p, perirenal; s, subcutaneous; siINDY, mINDY-specific siRNA; siRNA; solute carrier family 13, member 5