十字花科蔬菜衍生物對於分化之3T3-L1脂肪細胞內Aryl hydrocarbon receptor(AHR)與Nuclear factor E2–related factor 2(Nrf2)相關因子的影響

Effect of cruciferous vegetable derivatives on Aryl hydrocarbon receptor (AHR) and Nuclear factor E2–related factor 2 (Nrf2) associated factors during differentiation of 3T3-L1 adipocytes

戴秋玉(Chiu-Yu Tai)

指導教授 ： 陳玉華

臺北醫學大學/公共衛生暨營養學院/保健營養學研究所/碩士(2014年)

https://doi.org/10.6831/TMU.2014.00090

摘要

肥胖是由於體內過多能量進入脂肪組織中刺激脂肪細胞的數目增加與體積增大所造成。當脂肪細胞增大時，不僅增加三酸甘油酯的堆積，亦會促使脂肪組織內氧化壓力的增加，且伴隨著增加體內發炎反應及脂肪組織中血管新生作用。肥胖者脂肪組織中具高量 Aryl hydrocarbon receptor (AHR) 下游基因 cytochrome P450 (CYP) 1B1 以及 Nuclear factor E2 -related factor 2 (Nrf2) 下游基因 NAD(P)H:quinone oxidoreductase (NQO-1) 之表現量，且 AHR 及 Nrf2 與脂質堆積、氧化壓力、發炎反應及血管新生作用有關。AHR 之 ligands 包括飲食中十字花科蔬菜衍生物 indole‐3‐carbinol (I3C) 及 3,3′‐diindolylmethane (DIM)，I3C 及 DIM 具活化肝細胞內調節外來物質代謝相關酵素的轉錄因子AHR 及參與調控氧化壓力之轉錄因子 Nrf2 的作用，且 I3C 可降低高脂飲食肥胖小鼠之體重及腹部脂肪組織量。因此本次實驗主要使用 3T3-L1 前脂肪細胞，觀察於脂肪分化過程，I3C 與其代謝物 DIM 對脂肪合成、氧化壓力及相關發炎反應、血管新生之影響是否與 AHR、Nrf2 相關因子有關。結果顯示，低濃度 I3C (5 μM) 及 DIM (5 及10 μM) 可增加成熟脂肪細胞中油滴堆積量，而高濃度 I3C (70 μM) 卻可降低成熟脂肪細胞內油滴堆積、三酸甘油酯堆積量以及細胞內氧化壓力指標 malondialdehyde (MDA) 的濃度，且降低細胞內促脂肪分化 Peroxisome proliferator-activated receptor (PPAR)-γ、Nrf2 以及增加參與代謝毒性物質之AHR 及抗氧化作用之 NQO-1 蛋白質的表現量。DIM (5 - 25 μM) 隨著劑量增加，具有增加細胞內三酸甘油酯以及降低 MDA 濃度之趨勢，但與控制組並無顯著差異。然而，不同濃度的 I3C 與 DIM 皆不影響細胞培養液中 TNF-α 、 IL-6 、 NO 及 VEGF 、MMP-2 、 MMP-9濃度或酵素活性。總言之，高濃度 I3C (70 μM) 不影響氧化壓力相關發炎反應及血管新生作用，但可降低成熟脂肪細胞內三酸甘油酯堆積量以及 MDA 濃度，此影響可能與降低細胞內 PPAR-γ、Nrf2 蛋白質表現量，進而降低脂肪細胞脂質堆積量，以及增加 AHR、NQO-1 蛋白質表現量而增加脂肪細胞內抗氧化作用有關。

關鍵字

十字花科蔬菜 ； 3T3-L1 前脂肪細胞 ； 脂質堆積 ； 氧化壓力 ； Nuclear factor E2–related factor 2 ； Aryl hydrocarbon receptor

並列摘要

Obesity is an enlargement of adipose tissue to store excess energy. Enlargement of the adipocytes not only increase the storage of triglyceride (TG), but also increase oxidative stress in adipose tissue, and lead to increase the production of mediators involved in inflammation and angiogenesis. High expression of Nuclear factor E2 -related factor 2 (Nrf2) and its downstream NAD(P)H:quinone oxidoreductase (NQO-1) as well as Aryl hydrocarbon receptor (AHR) and its downstream cytochrome P450 (CYP) 1B1 were reported in adipose tissue of obese subjects. AHR and Nrf2 are associated with lipid accumulation, oxidative stress, inflammation and angiogenesis. Cruciferous vegetable derivatives, indole-3-carbinol (I3C) and 3,3′‐diindolylmethane (DIM), could increase the expression of AHR and Nrf2 in hepatoma cell line, and I3C was able to decrease body weight and fat accumulation in epididymal adipose tissue of diet-induce-obesity mice. This study investigated the effects of I3C and DIM on lipid accumulation, oxidative stress, inflammation and angiogenesis and the expression of AHR and Nrf2-associated factors wee also determined. The results showed that low concentration of I3C (5 μM) and DIM (5, 10 μM) significantly increased lipid droplets foemation, but high concentration of I3C (70 μM) significantly decreased lipid droplets, TG accumulation, and intracelluar malondialdehyde (MDA) content in mature adipocytes. I3C (70 μM) decreased peroxisome proliferator-activated receptor (PPAR)-γ and Nrf2, but enhanced AHR and NQO-1 protein expression. However, DIM showed no difference although tended to increase TG accumulation and decrease MDA content. Furthermore, both compounds had no effects on the production of NO, TNF-α, VEGF and MMP-2 , MMP-9 activity by the adipocytes. In conclusion, high concentration of I3C decreased TG accumulation and MDA content, and such effects were associated with decreased expression of PPAR-γ and Nrf2 proteins and increased AHR, NQO-1 protein expression in adipocytes.

並列關鍵字

Cruciferous vegetable derivatives ； 3T3-L1 ； lipid accumulation ； oxidative stress ； Nuclear factor E2–related factor 2 ； Aryl hydrocarbon receptor

參考文獻

葉志嶸, 張新儀, 潘文涵 (2008).臺灣近十二年來之肥胖與代謝症候群之變遷趨勢由NAHSIT1993-1996到2005-2008. 台灣營養健康調查.

Anderton, M.J., Manson, M.M., Verschoyle, R.D., Gescher, A., Lamb, J.H., Farmer, P.B., Steward, W.P., and Williams, M.L. (2004). Pharmacokinetics and tissue disposition of indole-3-carbinol and its acid condensation products after oral administration to mice. Clin Cancer Res 10, 5233-5241.

Armani, A., Mammi, C., Marzolla, V., Calanchini, M., Antelmi, A., Rosano, G.M., Fabbri, A., and Caprio, M. (2010). Cellular models for understanding adipogenesis, adipose dysfunction, and obesity. J Cell Biochem 110, 564-572.

Armstrong, L.C., and Bornstein, P. (2003). Thrombospondins 1 and 2 function as inhibitors of angiogenesis. Matrix Biol 22, 63-71.

Bartness, T.J., Vaughan, C.H., and Song, C.K. (2010). Sympathetic and sensory innervation of brown adipose tissue. Int J Obes (Lond) 34 Suppl 1, S36-42.