探討DHA對人類乳癌細胞雌激素接受器α分佈之影響

Abstract

乳癌對女性的健康造成極大的威脅。根據美國癌症協會的統計資料指出，婦女罹患乳癌的機率已攀升至31%；且致死率更升高為26%。而超過百分之六十的乳癌病患，歸類於雌激素-依賴性（estrogen -dependent）的乳癌。此類乳癌細胞中雌激素接受器α（Estrogen receptor α；ERα）有過量表現的情形，而且ERα在雌激素（17β-Estradiol；E2）的刺激下，會透過訊息傳遞以及ERα轉錄活性的增加，而促進乳癌細胞的增生與惡化。流行病學研究發現，富含多元不飽和脂肪酸DHA (docosahexaenoic acid, C22:6n-3) 的深海魚油，可降低乳癌的發生率，但可能發生機轉仍不詳。因此本論文的目的，為探討DHA是否藉由調控ERα在乳癌細胞中的表現與分佈及其訊息傳遞，而降低乳癌細胞的生長。 研究利用人類乳癌細胞株，MCF-7 cells，為estrogen-dependent且表現ERα的乳癌細胞。細胞培養在含有1% Dextran-charcoal 處理過的胎牛血清且phenol-red free DMEM培養液中，並添加不同濃度的DHA處理。從實驗的結果發現：當添加不同濃度的DHA( 0~100μM )處理24小時後，MCF-7中 DHA 含量占所有脂肪酸重量百分比亦隨之增加；從0.3、3.4、4.7、5.2、8.2、10.5、12.3 增加至14.4 wt %。而從細胞的存活率分析可知DHA的添加會抑制MCF-7的生長，使存活率下降14 % 至40 %。而對於ERα 的影響，則發現MCF-7添加DHA 10或60 μM處理24小時之後，再給予10 nM E2 作用4小時，DHA可以使細胞質中的ERα表現量分別下降32及29 %，但並不影響ERα在細胞核內的表現量。而60 μM DHA處理48小時後，除了使細胞核中ERα表現量降低26.2 %，亦降低25 % ER α在細胞中的總表現量。在與ER α有關的訊息傳遞反應方面，發現在E2的刺激下，其DHA的添加明顯降低MCF-7內mitogen activated protein kinases（MAPK）的磷酸化表現，使其下降56%。且從免疫螢光的影像顯示，添加DHA會減少ERα在細胞質及細胞膜上的分佈。 本篇論文利用人類乳癌細胞MCF-7 cells，探討DHA 對乳癌細胞內ERα的表現及其分佈調控，發現DHA可進入MCF-7，增加其細胞內DHA的含量；並可抑制細胞的生長。且DHA的添加可影響ERα在MCF-7內的表現及在細胞質與細胞核中的分佈；DHA的添加明顯降低ERα在MCF-7細胞質中的分佈，而且可降低由E2刺激所引起的MAPK磷酸化。總而言之，DHA可以調控estrogen-dependent乳癌細胞內ER α的表現與分佈及MAPK的活性，進而降低E2對乳癌細胞的作用。本論文將有助於臨床上estrogen-dependent乳癌治療，且期許有所貢獻。

Many studies showed that docosahexaenoic acid (DHA, 22:6n-3)-rich fish oil decrease breast cancer incidence. However, the mechanism is still not clear. The aim of this study is to investigate DHA may regulate estrogen receptor α (ERα) distribution, expression and its signal transduction by changing the DHA level studied in MCF-7, a hormone dependent human breast cancer cell lines. MCF-7 were incubated in phenol red free-DMEM with 1% Dextran-Charcoal treated FBS supplemented with DHA followed by 10 nM E2 stimulation. It was found that the DHA level was increased from 0.3, 3.4, 4.7, 5.2, 8.2, 10.5, 12.3 and 14.4 % of total fatty acids with the increasing 0、5、10、20、40、 60、 80、100 μM DHA supplementation for 24 hrs, respectively. The viability of MCF-7 was decreased 14 ~ 40% of the controls with the increasing DHA supplementation for 24 hrs.ERα expression in cytosol but not in nucleus was reduced 32 % or 29 % with DHA 10 or 60 uM supplementation for 24 hrs, respectively. ERα expression was significant down regulation not only in cytosol, but also in nucleus and whole cells supplemented with 60 μM DHA for 48 hrs. And the phosphorylated p44/42 MAPK expression was declined 56 % of controls with DHA supplementation for 24 hrs followed by 10 nM E2 stimulating for 15 minutes. The cytosolic ERα staining was diminished in MCF-7 with DHA supplementation. In conclusion, DHA supplementation did increase DHA level, reduce its viability, decrease cytosolic ERα distribution, and phosphorylated p44/42 MAPK expression in MCF-7. It was suggested that DHA may down regulate the E2 actions on estrogen dependent breast cancer cells via modulate ERα distribution.