探討甲型-硫辛酸對PC-3和DU-145人類前列腺癌細胞的氧化還原狀態、細胞增殖及遷移之影響

本研究使用具轉移特性之人類前列腺癌細胞株PC-3、DU-145，探討甲型-硫辛酸 (α-lipoic acid) 對前列腺癌細胞影響。本研究以含不同濃度α-lipoic acid (0、50、100、500、750、1000μM) 之細胞培養液處理前列腺癌細胞，以trypan blue assay計算細胞數目變化，並測試細胞移動變化及基質金屬蛋白酶 (matrix metalloproteinase；MMPs)活性。結果顯示，PC-3在處理72小時後，α-lipoic acid濃度50、100μM增加其細胞增生，500μM以上組別細胞增殖程度則顯著受抑制 (p＜0.05)；DU-145在處理72小時後，細胞增殖程度隨濃度增加而顯著受抑制 (p<0.05)。α-lipoic acid隨濃度促進PC-3細胞移動距離；於低濃度 (50、100μM) 促進DU-145細胞移動，高濃度 (500、750、1000μM)則抑制細胞移動距離，進一步使用明膠蛋白酵素電泳法探究α-lipoic acid是否藉由基質金屬蛋白酶活性而增加或抑制細胞轉移，結果顯示，PC-3經α-lipoic acid處理72小時，其MMP-2、MMP-9活性顯著增加 (p<0.01)；DU-145經處理72小時，其低濃度α-lipoic acid (50μM以上) MMP-2、MMP-9活性顯著增加 (p<0.01)，於750、1000μM，MMP-2、MMP-9活性略微降低。並測試細胞內多種抗氧化指標，包含GSH、GSSG、GPx、GR、GST、SOD、CAT，發現α-lipoic acid可改變細胞內氧化還原平衡，使細胞內榖胱甘肽(GSH)濃度隨α-lipoic acid處理有增加趨勢，並增加GSH/GSSG比值，而GSH/GSSG比值與細胞增殖 (proliferation)、細胞凋亡 (apotosis) 有關。利用西方墨點法偵測發現細胞凋亡相關蛋白質：caspase 3、caspase 9均受α-lipoic acid影響，表現量上升，促進細胞凋亡。本篇結果顯示，甲型-硫辛酸可能藉由改變細胞氧化/還原平衡、活化caspase 3、caspase 9，達到促進前列腺癌細胞凋亡等效果，但可能促進前列腺癌細胞轉移。

We examined the effects of α-lipoic acid (ALA) on cell behaviors in human prostate cancer cell lines with metastatic characteristic, PC-3 and DU-145. Cells were treated with concentrations of α-lipoic acid (0, 50, 100, 500, 750, 1000 μM) for studying the growth of prostate cancer cells by trypan blue assay and its migration by wound healing test and the underlying mechanisms evaluated by matrix metalloproteinase (MMPs) activity. The results show that the growth of PC-3 cell increases following the treatment of moderate concentrations (50, 100μM) of α-lipoic acid after 72 hour, while high concentration of ALA (500μM) significantly inhibited the proliferation (p＜0.05). Differential pattern of proliferation was observed in DU-145 cell, in which increasing concentrations significantly inhibited cell growth after 72 hours treatment (p＜0.05). ALA promoted distance of migration of PC-3 cells along with the increasing concentrations. While in DU-145 cells, migration distance was promoted only in the low concentrations (50, 100μM), but inhibited in the higher concentration (500, 750, 1000μM). Zymographic analysis of matrix metalloproteinase (MMPs) activity showed that, ALA treatment significantly increased the MMP-2 and MMP-9 activity in PC-3 cell after 72 hours (p＜0.01). In DU-145, the MMP-2 and MMP-9 activity significantly increased in low concentration of α-lipoic acid (50μM) for 72 hours (p＜0.01), but slightly reduced in 750, 1000μM groups. Test of a series of intracellular antioxidant indices, including GSH, GSSG, GPx, GR, GST, SOD, CAT, found that α-lipoic acid treatment may alter intracellular redox balance as the followings: intracellular glutathione (GSH) concentration and the GSH / GSSG ratio tended to increase, which may indirectly linked to either subsequent cell proliferation or apoptosis. Western blot analysis of apoptosis-related proteins revealed that proteins Caspase 3, and Caspase 9 increased by α-lipoic acid, and subsequently promote apoptosis. Our results indicate that, α-lipoic acid may enhance human prostate cancer cell apoptpsis but simultaneously promote metastasis through the alteration of GSH redox ratio, and caspase 3, caspase 9 protein expression.