| ORIGINAL ARTICLES |
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A Fluorescent Glucose Transport Assay for Screening SGLT2 Inhibitors in Endogenous SGLT2-Expressing HK-2 Cells |
| Yan-Ting Lu1,2, Xiu-Li Ma1,2, Yu-Hui Xu1, Jing Hu1,2, Fang Wang1,2, Wan-Ying Qin1,2, Wen-Yong Xiong1,3 |
1 State Key Laboratory of Phytochemistry and Plant Resources in West China, Kunming Institute of Botany, Chinese Academy of Sciences, Kunming 650201, China;
2 University of the Chinese Academy of Sciences, Beijing 100049, China;
3 Yunnan Key Laboratory of Natural Medicinal Chemistry, Kunming 650201, China |
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Abstract The sodium-dependent glucose transporters 2 (SGLT2) plays important role in renal reabsorption of urinal glucose back to plasma for maintaining glucose homeostasis. The approval of SGLT2 inhibitors for treatment of type 2 diabetes highlights the SGLT2 as a feasible and promising drug target in recent years. Current methods for screening SGLT2 inhibitors are complex, expensive and labor intensive. Particularly, these methods cannot directly measure nonradioactive glucose uptake in endogenous SGLT2-expressing kidney cells. In present work, human kidney cells, HK-2, was incubated with a fluorescent D-glucose derivant 2-[N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl) amino]-2-deoxy-D-glucose (2-NBDG) and the fluorescent intensity of 2-NBDG was employed to measure the amount of glucose uptake into the cells. By optimizing the passages of HK-2 cells, 2-NBDG concentration and incubation time, and by measuring glucose uptake treated by Dapagliflozin, a clinical drug of SGLT2 inhibitors, we successfully developed a new assay for measuring glucose uptake through SGLT2. The nonradioactive microplate and microscope-based high-throughput screening assay for measuring glucose can be a new method for screening of SGLT2 inhibitors and implied for other cell assays for glucose measurement extensively.
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| Keywords
SGLT2 inhibitor
2-NBDG
HK-2
Glucose uptake
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| Fund:This work was financially supported by Yunnan Provincial Science and Technology Department of China to WX (2017FA044 and 2013HA023), Ministry of Science and Technology of the People's Republic of China-The National Key Research and Development Program (2017YFC1700906). |
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Corresponding Authors:
Wen-Yong Xiong
E-mail: xiong.wenyong@mail.kib.ac.cn
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Issue Date: 28 January 2019
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1. V. Vallon, Annu. Rev. Med. 66, 255-270 (2015)
2. M.R. Evans, S.G. Wei, B.A. Posner, R.H. Unger, M.G. Roth, J. Biomol. Screen. 21, 325-332 (2016)
3. C. Levetan, Curr. Med. Res. Opin. 23, 945-952 (2007)
4. M.J. Meneses, B.M. Silva, M. Sousa, R. Sa, P.F. Oliveira, M.G. Alves, Curr. Pharm. Design 21, 3606-3620 (2015)
5. V. Vallon, S.C. Thomson, Diabetologia 60, 215-225 (2017)
6. R. Sabino-Silva, R.C. Mori, A. David-Silva, M.M. Okamoto, H.S. Freitas, U.F. Machado, Braz J Med Biol 43, 1019-1026 (2010)
7. I. Vrhovac, D. Balen Eror, D. Klessen, C. Burger, D. Breljak, O. Kraus, N. Radovic, S. Jadrijevic, I. Aleksic, T. Walles, C. Sauvant, I. Sabolic, H. Koepsell, Pflugers Archiv:Eur. J. Physiol 467, 1881-1898 (2015)
8. E.M. Wright, D.D. Loo, B.A. Hirayama, Physiol. Rev. 91, 733-794 (2011)
9. M.J. Jurczak, H.Y. Lee, A.L. Birkenfeld, F.R. Jornayvaz, D.W. Frederick, R.L. Pongratz, X. Zhao, G.W. Moeckel, V.T. Samuel, J.M. Whaley, G.I. Shulman, R.G. Kibbey, Diabetes 60, 890-898 (2011)
10. I. Dardi, T. Kouvatsos, S.A. Jabbour, Biochem. Pharmacol. 101, 27-39 (2016)
11. H. Fattah, V. Vallon, Drugs 78, 717-726 (2018)
12. J. Chen, F. Fan, J.Y. Wang, Y. Long, C.L. Gao, R.C. Stanton, Y. Xu, Sci. Rep. 7, 44128 (2017)
13. E. Ferrannini, E. Muscelli, S. Frascerra, S. Baldi, A. Mari, T. Heise, U.C. Broedl, H.J. Woerle, J. Clin. Investig. 124, 499-508 (2014)
14. R.M. Guthrie, Postgrad. Med. 125, 21-32 (2013)
15. J.R. Ehrenkranz, N.G. Lewis, C.R. Kahn, J. Roth, Diabetes/Metab. Res. Rev. 21, 31-38 (2005)
16. I. Fernandez-Ruiz, Nat. Rev. Cardiol. 15, 2-3 (2018)
17. S. Lim, R.H. Eckel, K.K. Koh, Atherosclerosis 272, 33-40 (2018)
18. J.F. Mosley 2nd, L. Smith, E. Everton, C. Fellner, Pharm. Ther. 40, 451-462 (2015)
19. Y. Fujita, N. Inagaki, J. Diabetes Investig. 5, 265-275 (2014)
20. M.J. Ryan, G. Johnson, J. Kirk, S.M. Fuerstenberg, R.A. Zager, B. Torok-Storb, Kidney Int. 45, 48-57 (1994)
21. J. Ye, J. Li, Y. Yu, Q. Wei, W. Deng, L. Yu, Regul. Pept. 161, 58-66 (2010)
22. J.X. Huang, G. Kaeslin, M.V. Ranall, M.A. Blaskovich, B. Becker, M.S. Butler, M.H. Little, L.H. Lash, M.A. Cooper, Pharmacol. Res. Perspect. 3, e00148 (2015)
23. F. Castaneda, R.K.H. Kinne, Mol. Cell. Biochem. 280, 91-98 (2005)
24. L. Sokoloff, M. Reivich, C. Kennedy, M.H. Desrosiers, C.S. Patlak, K.D. Pettigrew, O. Sakurada, M. Shinohara, J. Neurochem. 28, 897-916 (1977)
25. K. Yoshioka, K.B. Oh, M. Saito, Y. Nemoto, H. Matsuoka, Appl. Microbiol. Biotechnol. 46, 400-404 (1996)
26. C.H. Zou, Y.J. Wang, Z.F. Shen, J. Biochem. Biophys. Methods 64, 207-215 (2005)
27. V. Tsytsarev, K.I. Maslov, J. Yao, A.R. Parameswar, A.V. Demchenko, L.V. Wang, J. Neurosci. Methods 203, 136-140 (2012)
28. R. Santer, J. Calado, Clin. J. Am. Soc. Nephrol. 5, 133-141 (2010)
29. A.B. Blodgett, R.K. Kothinti, I. Kamyshko, D.H. Petering, S. Kumar, N.M. Tabatabai, Diabetes Technol. Ther. 13, 743-751 (2011)
30. A. Kanwal, S.P. Singh, P. Grover, S.K. Banerjee, Anal. Biochem. 429, 70-75 (2012)
31. H.C. Chang, S.F. Yang, C.C. Huang, T.S. Lin, P.H. Liang, C.J. Lin, L.C. Hsu, Mol. BioSyst. 9, 2010-2020 (2013)
32. S.H. Wu, C.H. Yao, C.J. Hsieh, Y.W. Liu, Y.S. Chao, J.S. Song, J.C. Lee, Eur. J. Pharm. Sci. 74, 40-44 (2015)
33. B.A. Hirayama, M.P. Lostao, M. PanayotovaHeiermann, D.D.F. Loo, E. Turk, E.M. Wright, Am. J. Physiol-Gastr. L 270, G919-G926 (1996)
34. S.W. Ball, J.R. Bailey, J.M. Stewart, C.M. Vogels, S.A. Westcott, Can. J. Physiol. Pharmacol. 80, 205-209 (2002)
35. R.G. O'Neil, L. Wu, N. Mullani, Mol. Imaging Biol. 7, 388-392 (2005)
36. K. Yamada, M. Saito, H. Matsuoka, N. Inagaki, Nat. Protoc. 2, 753-762 (2007)
37. S. Sato, J. Takeo, C. Aoyama, H. Kawahara, Bioorgan. Med. Chem. 15, 3445-3449 (2007)
38. H. Morita, J. Deguchi, Y. Motegi, S. Sato, C. Aoyama, J. Takeo, M. Shiro, Y. Hirasawa, Bioorg. Med. Chem. Lett. 20, 1070-1074 (2010) |
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