本研究探討綠茶唲茶素中表沒食子酸酯型唲茶素酸酯(簡寫為EGCG)調控第一型內皮素(簡寫為ET-1 )在短時間(0.5小時)、中長時間(2小時)、長時間(6小時)刺激脂肪細胞攝入葡萄糖的訊息機制。利用3T3-L1脂肪細胞，實驗中發現，在ET-1處理0.5小時和2小時的環境下，預先處理PI3K的抑制劑LY294002及wortmannin時，EGCG無法抑制由ET-1所誘導的葡萄糖攝取作用，然而在前處理ERK/MAPK的抑制劑PD98059及U0126的組別中則無此效應，因此推測EGCG會使ET-1在短時間處理時所減弱葡萄糖攝取的作用與PI3K/AKT此訊息路徑相關。另一方面也發現，當脂肪細胞在已有JAK2的抑制劑AG490(50 ?M )的環境下，ET-1處理0.5小時、2小時的組別中會發現葡萄糖攝取有增加的趨勢，因此可推論EGCG的作用會與JAK2/STAT-3訊息相關。在脂肪細胞裡鑑定到一個EGCG接受子[已知為67-kDa laminin接受子 (67LR)]。在本研究中，預先處理67 LR 抗體時無論是在處理0.5、2、6小時ET-1的環境下，皆呈現EGCG無法抑制ET-1促進葡萄糖攝取的現象，因此推論EGCG會透過67LR來抑制由ET-1所誘導的葡萄糖攝取。另外本實驗中發現在ET-1處理0.5小時的環境下，預先處理抗氧化劑NAC時脂肪細胞本身及由ET-1所誘導的葡萄糖攝取的能力皆有增加的趨勢，因而推測NAC在短時間影響脂肪細胞本身及ET-1所促進的葡萄糖攝取作用與ROS的生成相關。而在預先處理AMPK抑制劑compound C的組別中，無論是在處理0.5、2、6小時ET-1的環境下，皆呈現EGCG無法抑制ET-1促進葡萄糖攝取的現象，因此推論EGCG會透過AMPK來抑制由ET-1所誘導的葡萄糖攝取現象。而當ET-1處理2小時和6小時，EGCG會藉由AMPK的路徑去減弱ET-1所促使的GLUT-1蛋白表現，進而減少葡萄糖攝取的作用。 This study investigated the possible mechanism involved in EGCG modulation of ET-1-stimulated glucose uptake in 3T3-L1 adipocytes. We found that pretreatment with the PI3K inhibitors, such as LY294002 and wortmannin, inhibited EGCG-induced decreases on the acutely ET-1 stimulated glucose uptake in adipocytes. However, none of the ERK/MAPK inhibitors, such as PD98059 and U0126, significantly blocked the anti-ET-1 effects of EGCG on adipocyte glucose uptake. These data suggest that PI3K activation is necessary for the effects of EGCG on the acutely ET-1-stimulated glucose uptake in adipocytes. Interestingly, we observed that pretreatment with the JAK2/STAT3 inhibitors, such as AG490(50 ?M ), increased the basal glucose uptake and the ET-1-dependent glucose uptake, suggesting the JAK2/STAT3-dependent effect of EGCG. The EGCG receptor [also known as the 67-kDa laminin receptor (67LR)] was identified in fat cells. Pretreatment of adipocytes with 67LR antibody, but not normal rabbit immunoglobulin, prevented the effects of EGCG on ET-1-induced glucose uptake in adipocytes. These data suggest that EGCG exerts its anti-ET-1 actions on adipocyte glucose uptake via the 67LR. However, NAC(an antioxidant) acutely increased the basal glucose uptake and ET-1 stimulated glucose uptake, indicating that NAC possibly affected glucose uptake by suppressing not only the ET-1-dependent ROS production but also the ET-1-independent ROS production. Pretreatment with the AMPK inhibitor, such as compound C, blocked the anti-ET-1 effects of EGCG on adipocyte glucose uptake, and prevented the effects of EGCG on the chronically ET-1-stimulated GLUT-1 protein expression. This suggests that EGCG inhibits the chronically ET-1 action on adipocyte glucose uptake through the activation of AMPK pathway that results in alterations of GLUT-1 protein expression. Results of this study may help understand the mechanism of how green tea EGCG modulates hormone-mediated obesity.
