胰島素阻抗為第二型糖尿病之病理特徵，且伴隨著許多發病因子，如增加氧化壓力及粒線體失能。部分文獻證實出活性氧產生的壓力以及粒線體功能缺失皆參與發病過程，但其分子機制仍然不明。Glucose transporter 10，簡稱GLUT10，屬於第三類型葡萄糖運送家族，並且是由位於染色體上與糖尿病有關之20q12-13.1區域中的SLC2A10基因轉譯。 先前我們實驗室已證實在GLUT10大量表達的脂肪細胞中可藉由胰島素刺激移動至粒線體，此外，GLUT10為脫氫抗壞血酸運送子的功能可保護細胞免於由雙氧水產生之氧化壓力的傷害。在此篇研究中，我們想更加了解GLUT10和糖尿病的關係。我們使用高葡萄糖處理脂肪細胞模擬高血糖狀態。此處理不僅導致活性氧壓力增加、還會降低粒線體膜電位和耗氧量。另外，我們發現在這些脂肪細胞補充抗壞血酸不只可以降低活性氧的產生、還能增加粒線體膜功能，這些情況在加入胰島素刺激後有更大的改善。我的結果提供一個研究GLUT10和第二類型糖尿病關係的高葡萄糖壓力條件。

Insulin resistance is a characteristic feature of type 2 diabetes mellitus (T2DM) and is accompanied many pathological factors, such as increased oxidative stress and mitochondrial dysfunction. Some studies evidenced that reactive oxygen species (ROS) and mitochondrial dysfunction are involving in the pathological progress, but the molecular mechanisms are still unclear. Glucose transporter 10 (GLUT10) is a member of class III glucose transporter family and encoded by SLC2A10 gene which is located on chromosome 20q12-13.1 where was an association with type 2 diabetes. Previously, our lab has reported that GLUT10 transfers to mitochondrial under insulin stimulation in adipocytes where the GLUT10 is largely expressed. In addition, GLUT10 transports L-dehydroascorbic acid (DHA) in to mitochondria and against oxidative stress under H202-induced stress condition.
In this study, we aim to understand the association between GLUT10 and T2DM. We mimic hyperglycemic condition using high glucose treating adipocytes. The treatment leads to increase ROS stress, collapse mitochondrial membrane potential, and decrease oxygen consumption rate. In addition, we found that replenishment with ascorbic acid can not only reduce the intracellular ROS production, but also increase mitochondrial function in adipocytes. Furthermore, treated these adipocytes with insulin can further improve ROS stress and mitochondrial function. My results provide a high glucose stress condition to study the correlation between GLUT10 and insulin resistance in adipocytes.

References
1. Zhao, F. Q. and Keating, A. F. (2007) Functional properties and genomics of glucose transporters. Current genomics. 8, 113-128
2. Uldry, M. and Thorens, B. (2004) The SLC2 family of facilitated hexose and polyol transporters. Pflugers Archiv : European journal of physiology. 447, 480-489
3. Joost, H. G. and Thorens, B. (2001) The extended GLUT-family of sugar/polyol transport facilitators: nomenclature, sequence characteristics, and potential function of its novel members (review). Molecular membrane biology. 18, 247-256
4. Joost, H. G., Bell, G. I., Best, J. D., Birnbaum, M. J., Charron, M. J., Chen, Y. T., Doege, H., James, D. E., Lodish, H. F., Moley, K. H., Moley, J. F., Mueckler, M., Rogers, S., Schurmann, A., Seino, S. and Thorens, B. (2002) Nomenclature of the GLUT/SLC2A family of sugar/polyol transport facilitators. American journal of physiology. Endocrinology and metabolism. 282, E974-976
5. Augustin, R. (2010) The protein family of glucose transport facilitators: It's not only about glucose after all. IUBMB life. 62, 315-333
6. McVie-Wylie, A. J., Lamson, D. R. and Chen, Y. T. (2001) Molecular cloning of a novel member of the GLUT family of transporters, SLC2a10 (GLUT10), localized on chromosome 20q13.1: a candidate gene for NIDDM susceptibility. Genomics. 72, 113-117
7. Dawson, P. A., Mychaleckyj, J. C., Fossey, S. C., Mihic, S. J., Craddock, A. L. and Bowden, D. W. (2001) Sequence and functional analysis of GLUT10: a glucose transporter in the Type 2 diabetes-linked region of chromosome 20q12-13.1. Molecular genetics and metabolism. 74, 186-199
8. Wood, I. S., Hunter, L. and Trayhurn, P. (2003) Expression of Class III facilitative glucose transporter genes (GLUT-10 and GLUT-12) in mouse and human adipose tissues. Biochemical and biophysical research communications. 308, 43-49
9. Coucke, P. J., Willaert, A., Wessels, M. W., Callewaert, B., Zoppi, N., De Backer, J., Fox, J. E., Mancini, G. M., Kambouris, M., Gardella, R., Facchetti, F., Willems, P. J., Forsyth, R., Dietz, H. C., Barlati, S., Colombi, M., Loeys, B. and De Paepe, A. (2006) Mutations in the facilitative glucose transporter GLUT10 alter angiogenesis and cause arterial tortuosity syndrome. Nature genetics. 38, 452-457
10. Lee, Y. C., Huang, H. Y., Chang, C. J., Cheng, C. H. and Chen, Y. T. (2010) Mitochondrial GLUT10 facilitates dehydroascorbic acid import and protects cells against oxidative stress: mechanistic insight into arterial tortuosity syndrome. Human molecular genetics. 19, 3721-3733
11. Sullivan, F. (1999) Current medical diagnosis and treatment 1998. Bmj. 318, 743A
12. Das, S. K. and Elbein, S. C. (2006) The Genetic Basis of Type 2 Diabetes. Cellscience. 2, 100-131
13. Ripsin, C. M., Kang, H. and Urban, R. J. (2009) Management of blood glucose in type 2 diabetes mellitus. American family physician. 79, 29-36
14. Houstis, N., Rosen, E. D. and Lander, E. S. (2006) Reactive oxygen species have a causal role in multiple forms of insulin resistance. Nature. 440, 944-948
15. Maddux, B. A., See, W., Lawrence, J. C., Jr., Goldfine, A. L., Goldfine, I. D. and Evans, J. L. (2001) Protection against oxidative stress-induced insulin resistance in rat L6 muscle cells by mircomolar concentrations of alpha-lipoic acid. Diabetes. 50, 404-410
16. Furukawa, S., Fujita, T., Shimabukuro, M., Iwaki, M., Yamada, Y., Nakajima, Y., Nakayama, O., Makishima, M., Matsuda, M. and Shimomura, I. (2004) Increased oxidative stress in obesity and its impact on metabolic syndrome. Journal of Clinical Investigation. 114, 1752-1761
17. Ikemura, M., Nishikawa, M., Hyoudou, K., Kobayashi, Y., Yamashita, F. and Hashida, M. (2010) Improvement of Insulin Resistance by Removal of Systemic Hydrogen Peroxide by PEGylated Catalase in Obese Mice. Mol Pharmaceut. 7, 2069-2076
18. Smeitink, J., van den Heuvel, L. and DiMauro, S. (2001) The genetics and pathology of oxidative phosphorylation. Nature reviews. Genetics. 2, 342-352
19. Valko, M., Leibfritz, D., Moncol, J., Cronin, M. T., Mazur, M. and Telser, J. (2007) Free radicals and antioxidants in normal physiological functions and human disease. The international journal of biochemistry & cell biology. 39, 44-84
20. Anderson, E. J., Lustig, M. E., Boyle, K. E., Woodlief, T. L., Kane, D. A., Lin, C. T., Price, J. W., 3rd, Kang, L., Rabinovitch, P. S., Szeto, H. H., Houmard, J. A., Cortright, R. N., Wasserman, D. H. and Neufer, P. D. (2009) Mitochondrial H2O2 emission and cellular redox state link excess fat intake to insulin resistance in both rodents and humans. The Journal of clinical investigation. 119, 573-581
21. Mogensen, M., Sahlin, K., Fernstrom, M., Glintborg, D., Vind, B. F., Beck-Nielsen, H. and Hojlund, K. (2007) Mitochondrial respiration is decreased in skeletal muscle of patients with type 2 diabetes. Diabetes. 56, 1592-1599
22. Hammarstedt, A., Jansson, P. A., Wesslau, C., Yang, X. and Smith, U. (2003) Reduced expression of PGC-1 and insulin-signaling molecules in adipose tissue is associated with insulin resistance. Biochemical and biophysical research communications. 301, 578-582
23. Bonnard, C., Durand, A., Peyrol, S., Chanseaume, E., Chauvin, M. A., Morio, B., Vidal, H. and Rieusset, J. (2008) Mitochondrial dysfunction results from oxidative stress in the skeletal muscle of diet-induced insulin-resistant mice. The Journal of clinical investigation. 118, 789-800
24. Rudich, A., Tirosh, A., Potashnik, R., Hemi, R., Kanety, H. and Bashan, N. (1998) Prolonged oxidative stress impairs insulin-induced GLUT4 translocation in 3T3-L1 adipocytes. Diabetes. 47, 1562-1569
25. Krauss, S., Zhang, C. Y., Scorrano, L., Dalgaard, L. T., St-Pierre, J., Grey, S. T. and Lowell, B. B. (2003) Superoxide-mediated activation of uncoupling protein 2 causes pancreatic beta cell dysfunction. Journal of Clinical Investigation. 112, 1831-1842
26. Inoguchi, T., Li, P., Umeda, F., Yu, H. Y., Kakimoto, M., Imamura, M., Aoki, T., Etoh, T., Hashimoto, T., Naruse, M., Sano, H., Utsumi, H. and Nawata, H. (2000) High glucose level and free fatty acid stimulate reactive oxygen species production through protein kinase C-dependent activation of NAD(P)H oxidase in cultured vascular cells. Diabetes. 49, 1939-1945
27. Tada-Oikawa, S., Hiraku, Y., Kawanishi, M. and Kawanishi, S. (2003) Mechanism for generation of hydrogen peroxide and change of mitochondrial membrane potential during rotenone-induced apoptosis. Life sciences. 73, 3277-3288
28. Smith, U. (2002) Impaired ('diabetic') insulin signaling and action occur in fat cells long before glucose intolerance - is insulin resistance initiated in the adipose tissue? Int J Obesity. 26, 897-904
29. Garvey, W. T., Maianu, L., Huecksteadt, T. P., Birnbaum, M. J., Molina, J. M. and Ciaraldi, T. P. (1991) Pretranslational Suppression of a Glucose Transporter Protein Causes Insulin Resistance in Adipocytes from Patients with Non-Insulin-Dependent Diabetes-Mellitus and Obesity. Journal of Clinical Investigation. 87, 1072-1081
30. Urakawa, H., Katsuki, A., Sumida, Y., Gabazza, E. C., Murashima, S., Morioka, K., Maruyama, N., Kitagawa, N., Tanaka, T., Hori, Y., Nakatani, K., Yano, Y. and Adachi, Y. (2003) Oxidative stress is associated with adiposity and insulin resistance in men. J Clin Endocr Metab. 88, 4673-4676
31. Ihara, Y., Toyokuni, S., Uchida, K., Odaka, H., Tanaka, T., Ikeda, H., Hiai, H., Seino, Y. and Yamada, Y. (1999) Hyperglycemia causes oxidative stress in pancreatic beta-cells of GK rats, a model of type 2 diabetes. Diabetes. 48, 927-932
32. Donath, M. Y., Gross, D. J., Cerasi, E. and Kaiser, N. (1999) Hyperglycemia-induced beta-cell apoptosis in pancreatic islets of Psammomys obesus during development of diabetes. Diabetes. 48, 738-744
33. Gao, C. L., Zhu, C., Zhao, Y. P., Chen, X. H., Ji, C. B., Zhang, C. M., Zhu, J. G., Xia, Z. K., Tong, M. L. and Guo, X. R. (2010) Mitochondrial dysfunction is induced by high levels of glucose and free fatty acids in 3T3-L1 adipocytes. Mol Cell Endocrinol. 320, 25-33
34. Li, X., Cobb, C. E., Hill, K. E., Burk, R. F. and May, J. M. (2001) Mitochondrial uptake and recycling of ascorbic acid. Archives of biochemistry and biophysics. 387, 143-153
35. Addabbo, F., Montagnani, M. and Goligorsky, M. S. (2009) Mitochondria and Reactive Oxygen Species. Hypertension. 53, 885-892
36. Mootha, V. K., Lindgren, C. M., Eriksson, K. F., Subramanian, A., Sihag, S., Lehar, J., Puigserver, P., Carlsson, E., Ridderstrale, M., Laurila, E., Houstis, N., Daly, M. J., Patterson, N., Mesirov, J. P., Golub, T. R., Tamayo, P., Spiegelman, B., Lander, E. S., Hirschhorn, J. N., Altshuler, D. and Groop, L. C. (2003) PGC-1alpha-responsive genes involved in oxidative phosphorylation are coordinately downregulated in human diabetes. Nature genetics. 34, 267-273
37. Palmeira, C. M., Rolo, A. P., Berthiaume, J., Bjork, J. A. and Wallace, K. B. (2007) Hyperglycemia decreases mitochondrial function: the regulatory role of mitochondrial biogenesis. Toxicology and applied pharmacology. 225, 214-220
38. Winkler, B. S. (1987) In vitro oxidation of ascorbic acid and its prevention by GSH. Biochimica et biophysica acta. 925, 258-264
39. Li, X., Cobb, C. E., Hill, K. E., Burk, R. F. and May, J. M. (2001) Mitochondrial uptake and recycling of ascorbic acid. Archives of biochemistry and biophysics. 387, 143-153
40. Li, X., Cobb, C. E. and May, J. M. (2002) Mitochondrial recycling of ascorbic acid from dehydroascorbic acid: dependence on the electron transport chain. Archives of biochemistry and biophysics. 403, 103-110
41. Liu, S., Okada, T., Assmann, A., Soto, J., Liew, C. W., Bugger, H., Shirihai, O. S., Abel, E. D. and Kulkarni, R. N. (2009) Insulin signaling regulates mitochondrial function in pancreatic beta-cells. PloS one. 4, e7983
42. Riehle, C., Wayment, B., Pires, K. M., Moreira, A. B., Theobald, H., Dong, X. C., White, M. F., Litwin, S. E. and Abel, E. D. (2008) Insulin Receptor Substrates (IRS) Signaling are Essential Regulators of Mitochondrial Function and Cardiomyocyte Survival. Circulation. 118, S444-S444
43. Govers, R., Coster, A. C. and James, D. E. (2004) Insulin increases cell surface GLUT4 levels by dose dependently discharging GLUT4 into a cell surface recycling pathway. Molecular and cellular biology. 24, 6456-6466
44. Kc, S., Carcamo, J. M. and Golde, D. W. (2005) Vitamin C enters mitochondria via facilitative glucose transporter 1 (Glut1) and confers mitochondrial protection against oxidative injury. FASEB journal : official publication of the Federation of American Societies for Experimental Biology. 19, 1657-1667