A siRNA mediated hepatic dpp4 knockdown affects lipid, but not glucose metabolism in diabetic mice


Autoři: Sven Wolfgang Görgens aff001;  Kerstin Jahn-Hofmann aff001;  Dinesh Bangari aff002;  Sheila Cummings aff002;  Christiane Metz-Weidmann aff001;  Uwe Schwahn aff001;  Paulus Wohlfart aff001;  Matthias Schäfer aff001;  Maximilian Bielohuby aff001
Působiště autorů: Sanofi-Aventis Deutschland GmbH, Industriepark Hoechst, Frankfurt am Main, Germany aff001;  Sanofi, Global Discovery Pathology, Translational In-vivo Models Framingham, MA, United States of America aff002
Vyšlo v časopise: PLoS ONE 14(12)
Kategorie: Research Article
doi: 10.1371/journal.pone.0225835

Souhrn

Systemic inhibition of dipeptidyl peptidase 4 (dpp4) represents an effective and established treatment option for type 2 diabetes (T2D). The current study investigated in mice if a liver selective knock-down of dpp4 by therapeutic siRNAs could be a novel, similarly effective treatment option for T2D. Furthermore, the potential effects on hepatic steatosis, inflammation and lipid metabolism were investigated after hepato-selective knock-down of dpp4. The knock-down efficiency and IC50 values of siRNAs targeting dpp4 were analyzed in PC3 cells. In two independent studies, either db/db mice or C57BL/6J mice were injected intravenously with a liposomal formulation of siRNAs targeting either dpp4 or a non-targeting control, followed by metabolically characterization. In comparator groups, additional cohorts of mice were treated with an oral dpp4 inhibitor. In both animal studies, we observed a robust knock-down (~75%) of hepatic dpp4 with a potent siRNA. Hepatic dpp4 knockdown did not significantly affect glucose metabolism or circulating incretin concentrations in both animal studies. However, in obese and diabetic db/db mice hepatic steatosis was reduced and hepatic mRNA expression of acaca, scd1, fasn and pparg was significantly lower after siRNA treatment. Systemic inhibition of the enzymatic dpp4 activity by an oral dpp4 inhibitor significantly improved glucose handling in db/db mice but did not affect hepatic endpoints. These data demonstrate that a targeted reduction of dpp4 expression in the liver may not be sufficient to improve whole-body glucose metabolism in obese and diabetic mice but may improve hepatic lipid metabolism.

Klíčová slova:

Fatty liver – Glucose metabolism – Inflammation – Mouse models – Obesity – Oral glucose suppression test – Small interfering RNAs – Steatosis


Zdroje

1. Hopsu-Havu VK, Glenner GG. A new dipeptide naphthylamidase hydrolyzing glycyl-prolyl-beta-naphthylamide. Histochemie. 1966;7(3):197–201. Epub 1966/01/01. doi: 10.1007/bf00577838 5959122.

2. Rohrborn D, Wronkowitz N, Eckel J. DPP4 in Diabetes. Front Immunol. 2015;6:386. Epub 2015/08/19. doi: 10.3389/fimmu.2015.00386 26284071; PubMed Central PMCID: PMC4515598.

3. Nauck M, Stockmann F, Ebert R, Creutzfeldt W. Reduced incretin effect in type 2 (non-insulin-dependent) diabetes. Diabetologia. 1986;29(1):46–52. Epub 1986/01/01. doi: 10.1007/bf02427280 3514343.

4. Mulvihill EE, Varin EM, Gladanac B, Campbell JE, Ussher JR, Baggio LL, et al. Cellular Sites and Mechanisms Linking Reduction of Dipeptidyl Peptidase-4 Activity to Control of Incretin Hormone Action and Glucose Homeostasis. Cell Metab. 2017;25(1):152–65. Epub 2016/11/15. doi: 10.1016/j.cmet.2016.10.007 27839908.

5. Marguet D, Baggio L, Kobayashi T, Bernard AM, Pierres M, Nielsen PF, et al. Enhanced insulin secretion and improved glucose tolerance in mice lacking CD26. Proc Natl Acad Sci U S A. 2000;97(12):6874–9. Epub 2000/05/24. doi: 10.1073/pnas.120069197 10823914; PubMed Central PMCID: PMC18768.

6. Conarello SL, Li Z, Ronan J, Roy RS, Zhu L, Jiang G, et al. Mice lacking dipeptidyl peptidase IV are protected against obesity and insulin resistance. Proc Natl Acad Sci U S A. 2003;100(11):6825–30. Epub 2003/05/16. doi: 10.1073/pnas.0631828100 12748388; PubMed Central PMCID: PMC164531.

7. Sell H, Bluher M, Kloting N, Schlich R, Willems M, Ruppe F, et al. Adipose dipeptidyl peptidase-4 and obesity: correlation with insulin resistance and depot-specific release from adipose tissue in vivo and in vitro. Diabetes Care. 2013;36(12):4083–90. Epub 2013/10/17. doi: 10.2337/dc13-0496 24130353; PubMed Central PMCID: PMC3836153.

8. Lamers D, Famulla S, Wronkowitz N, Hartwig S, Lehr S, Ouwens DM, et al. Dipeptidyl peptidase 4 is a novel adipokine potentially linking obesity to the metabolic syndrome. Diabetes. 2011;60(7):1917–25. Epub 2011/05/20. doi: 10.2337/db10-1707 21593202; PubMed Central PMCID: PMC3121429.

9. Varin EM, Mulvihill EE, Beaudry JL, Pujadas G, Fuchs S, Tanti JF, et al. Circulating Levels of Soluble Dipeptidyl Peptidase-4 Are Dissociated from Inflammation and Induced by Enzymatic DPP4 Inhibition. Cell Metab. 2019;29(2):320–34 e5. Epub 2018/11/06. doi: 10.1016/j.cmet.2018.10.001 30393019.

10. Baumeier C, Schluter L, Saussenthaler S, Laeger T, Rodiger M, Alaze SA, et al. Elevated hepatic DPP4 activity promotes insulin resistance and non-alcoholic fatty liver disease. Mol Metab. 2017;6(10):1254–63. Epub 2017/10/17. doi: 10.1016/j.molmet.2017.07.016 29031724; PubMed Central PMCID: PMC5641684.

11. Baumeier C, Saussenthaler S, Kammel A, Jahnert M, Schluter L, Hesse D, et al. Hepatic DPP4 DNA Methylation Associates With Fatty Liver. Diabetes. 2017;66(1):25–35. Epub 2016/12/22. doi: 10.2337/db15-1716 27999105.

12. Ghorpade DS, Ozcan L, Zheng Z, Nicoloro SM, Shen Y, Chen E, et al. Hepatocyte-secreted DPP4 in obesity promotes adipose inflammation and insulin resistance. Nature. 2018;555(7698):673–7. Epub 2018/03/22. doi: 10.1038/nature26138 29562231; PubMed Central PMCID: PMC6021131.

13. Wu SY, McMillan NA. Lipidic systems for in vivo siRNA delivery. AAPS J. 2009;11(4):639–52. Epub 2009/09/17. doi: 10.1208/s12248-009-9140-1 19757082; PubMed Central PMCID: PMC2782074.

14. Brachs S, Winkel AF, Tang H, Birkenfeld AL, Brunner B, Jahn-Hofmann K, et al. Inhibition of citrate cotransporter Slc13a5/mINDY by RNAi improves hepatic insulin sensitivity and prevents diet-induced non-alcoholic fatty liver disease in mice. Mol Metab. 2016;5(11):1072–82. Epub 2016/11/08. doi: 10.1016/j.molmet.2016.08.004 27818933; PubMed Central PMCID: PMC5081411.

15. Wohlfart P, Lin J, Dietrich N, Kannt A, Elvert R, Herling AW, et al. Expression patterning reveals retinal inflammation as a minor factor in experimental retinopathy of ZDF rats. Acta Diabetol. 2014;51(4):553–8. Epub 2014/01/31. doi: 10.1007/s00592-013-0550-2 24477469; PubMed Central PMCID: PMC4127441.

16. Bielohuby M, Popp S, Bidlingmaier M. A guide for measurement of circulating metabolic hormones in rodents: Pitfalls during the pre-analytical phase. Mol Metab. 2012;1(1–2):47–60. Epub 2012/01/01. doi: 10.1016/j.molmet.2012.07.004 24024118; PubMed Central PMCID: PMC3757653.

17. Bielohuby M, Bidlingmaier M, Schwahn U. Control of (pre)-analytical aspects in immunoassay measurements of metabolic hormones in rodents. Endocr Connect. 2018;7(4):R147–R59. Epub 2018/03/16. doi: 10.1530/EC-18-0035 29540488; PubMed Central PMCID: PMC5881432.

18. Lorenzer C, Dirin M, Winkler AM, Baumann V, Winkler J. Going beyond the liver: progress and challenges of targeted delivery of siRNA therapeutics. J Control Release. 2015;203:1–15. Epub 2015/02/11. doi: 10.1016/j.jconrel.2015.02.003 25660205.

19. Yi HS. Implications of Mitochondrial Unfolded Protein Response and Mitokines: A Perspective on Fatty Liver Diseases. Endocrinol Metab (Seoul). 2019;34(1):39–46. Epub 2019/03/27. doi: 10.3803/EnM.2019.34.1.39 30912337; PubMed Central PMCID: PMC6435852.

20. Oslowski CM, Urano F. Measuring ER stress and the unfolded protein response using mammalian tissue culture system. Methods Enzymol. 2011;490:71–92. Epub 2011/01/27. doi: 10.1016/B978-0-12-385114-7.00004-0 21266244; PubMed Central PMCID: PMC3701721.

21. Varin EM, Mulvihill EE, Beaudry JL, Pujadas G, Fuchs S, Tanti JF, et al. Circulating Levels of Soluble Dipeptidyl Peptidase-4 Are Dissociated from Inflammation and Induced by Enzymatic DPP4 Inhibition. Cell metabolism. 2018. Epub 2018/11/06. doi: 10.1016/j.cmet.2018.10.001 30393019.

22. Wang XM, Holz LE, Chowdhury S, Cordoba SP, Evans KA, Gall MG, et al. The pro-fibrotic role of dipeptidyl peptidase 4 in carbon tetrachloride-induced experimental liver injury. Immunol Cell Biol. 2017;95(5):443–53. Epub 2016/12/03. doi: 10.1038/icb.2016.116 27899813.

23. Nakamura Y, Tsuji M, Hasegawa H, Kimura K, Fujita K, Inoue M, et al. Anti-inflammatory effects of linagliptin in hemodialysis patients with diabetes. Hemodial Int. 2014;18(2):433–42. Epub 2014/01/11. doi: 10.1111/hdi.12127 24405885.

24. Jo CH, Kim S, Park JS, Kim GH. Anti-Inflammatory Action of Sitagliptin and Linagliptin in Doxorubicin Nephropathy. Kidney Blood Press Res. 2018;43(3):987–99. Epub 2018/06/19. doi: 10.1159/000490688 29913457.

25. Rohrborn D, Bruckner J, Sell H, Eckel J. Reduced DPP4 activity improves insulin signaling in primary human adipocytes. Biochem Biophys Res Commun. 2016;471(3):348–54. Epub 2016/02/14. doi: 10.1016/j.bbrc.2016.02.019 26872429.

26. Michurina SV, Ishenko IJ, Klimontov VV, Archipov SA, Myakina NE, Cherepanova MA, et al. Linagliptin alleviates fatty liver disease in diabetic db/db mice. World J Diabetes. 2016;7(19):534–46. Epub 2016/11/30. doi: 10.4239/wjd.v7.i19.534 27895822; PubMed Central PMCID: PMC5107713.

27. Klein T, Fujii M, Sandel J, Shibazaki Y, Wakamatsu K, Mark M, et al. Linagliptin alleviates hepatic steatosis and inflammation in a mouse model of non-alcoholic steatohepatitis. Med Mol Morphol. 2014;47(3):137–49. Epub 2013/09/21. doi: 10.1007/s00795-013-0053-9 24048504.

28. Kern M, Kloting N, Niessen HG, Thomas L, Stiller D, Mark M, et al. Linagliptin improves insulin sensitivity and hepatic steatosis in diet-induced obesity. PLoS One. 2012;7(6):e38744. Epub 2012/07/05. doi: 10.1371/journal.pone.0038744 22761701; PubMed Central PMCID: PMC3382200.

29. Kern M, Kloting N, Mark M, Mayoux E, Klein T, Bluher M. The SGLT2 inhibitor empagliflozin improves insulin sensitivity in db/db mice both as monotherapy and in combination with linagliptin. Metabolism. 2016;65(2):114–23. Epub 2016/01/17. doi: 10.1016/j.metabol.2015.10.010 26773934.

30. Moon YA, Liang G, Xie X, Frank-Kamenetsky M, Fitzgerald K, Koteliansky V, et al. The Scap/SREBP pathway is essential for developing diabetic fatty liver and carbohydrate-induced hypertriglyceridemia in animals. Cell Metab. 2012;15(2):240–6. Epub 2012/02/14. doi: 10.1016/j.cmet.2011.12.017 22326225; PubMed Central PMCID: PMC3662050.

31. Zhang YL, Hernandez-Ono A, Siri P, Weisberg S, Conlon D, Graham MJ, et al. Aberrant hepatic expression of PPARgamma2 stimulates hepatic lipogenesis in a mouse model of obesity, insulin resistance, dyslipidemia, and hepatic steatosis. J Biol Chem. 2006;281(49):37603–15. Epub 2006/09/15. doi: 10.1074/jbc.M604709200 16971390.

32. Wronkowitz N, Gorgens SW, Romacho T, Villalobos LA, Sanchez-Ferrer CF, Peiro C, et al. Soluble DPP4 induces inflammation and proliferation of human smooth muscle cells via protease-activated receptor 2. Biochim Biophys Acta. 2014;1842(9):1613–21. Epub 2014/06/15. doi: 10.1016/j.bbadis.2014.06.004 24928308.

33. Romacho T, Vallejo S, Villalobos LA, Wronkowitz N, Indrakusuma I, Sell H, et al. Soluble dipeptidyl peptidase-4 induces microvascular endothelial dysfunction through proteinase-activated receptor-2 and thromboxane A2 release. J Hypertens. 2016;34(5):869–76. Epub 2016/02/20. doi: 10.1097/HJH.0000000000000886 26895560.

34. Ohnuma K, Uchiyama M, Yamochi T, Nishibashi K, Hosono O, Takahashi N, et al. Caveolin-1 triggers T-cell activation via CD26 in association with CARMA1. J Biol Chem. 2007;282(13):10117–31. Epub 2007/02/09. doi: 10.1074/jbc.M609157200 17287217.


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