中文摘要 砷化物是廣泛存在於自然環境中的毒性物質，與人類皮膚癌、肺癌及肝癌的發生息息相關。近年來許多學者的研究指出，砷化物所造成的毒性與砷化物會產生自由基有關。在細胞中為了抵抗氧化壓力，細胞自己便會發展許多防禦的機制。Thioredoxin peroxidase II（TPxII）是一種受氧化性壓力（oxidative stress）誘導的酵素，也是一新發現的抗氧化蛋白質，其功能涵蓋細胞的分裂及分化、保護其他蛋白質免於受到氧化傷害、參與細胞中訊息的傳遞。 在本論文中，我們研究不同砷化物對各種哺乳類細胞中TPx II表現的影響及調控機制。由結果發現，在SA7N細胞中，亞砷酸鈉會誘發TPx II大量表現。而對其他細胞株例如：H1299、H460、HFW、IRC、KB、HaCaT等，亞砷酸鈉則無法誘導其TPx II大量表現。此外，也發現在亞砷酸鈉處理下，SA7N細胞內過氧化氫（H2O2）的累積量有增加的現象，顯示亞砷酸鈉誘導TPx II的表現，可能與其透過對細胞產生氧化性壓迫有關。 我們進一步以SA7N細胞研究亞砷酸鈉誘導TPx II表現的訊號傳遞途徑。結果顯示蛋白質激酶C（PKC）的抑制劑包括Rottlerin，Staurosprine可以抑制亞砷酸鈉所誘導TPx II的表現，而且在鈣離子 螯合劑EGTA和BAPTA/AM的處理下，亞砷酸鈉所誘導的TPx II大量表現也都沒有被抑制。進一步研究發現，對PKCδ有高專一性的抑制劑G06983、Calphostin C也可以有效地抑制亞砷酸鈉誘導TPx II的表現。而蛋白質激酶A（PKA）抑制劑（H89）、ERK抑制劑（PD98059）、p38抑制劑（SB203580）、酪胺酸激酶抑制劑（Genistein）則都無法抑制TPx II的表現。此一結果顯示，蛋白質激酶Cδ可能參與在SA7N細胞中亞砷酸鈉誘導TPx II的表現。 Abstract Arsenic is a common environmental toxicant associated with human skin, lung and liver cancers. Recently, numerous studies suggested that arsenic-induced toxicity is associated with the generation of free radicals. To cope with the oxidative stress, the cell has developed various defense mechanism. Thioredoxin peroxidase II (TPx II) is an oxidative stress-inducible enzyme. TPx II functions in cell proliferation and differentiation, and protects other proteins from oxidative damage. In this study, we investigate the effect of arsenic compounds on the expression of TPx II. The results showed that arsenite induces TPx II protein expression in SA7N cell (a revertant of arsenic resistant Chinese hamster ovary cells), but not in other cell lines such as H1299、H460、HFW、IRC、KB、HaCaT. Moreover, the induction of TPx II also accompanied with H2O2 accumulation in SA7N cells. We therefore further explore the signaling pathway involved in TPx II induction by arsenite in SA7N cells. The results showed that pre-treatment of SA7N cells with inhibitors of protein kinase A (PKA), extracellular-regulated kinase (ERK), p38 and tyrosine kinase had no effect on the induction of TPx-II expression by arsenite. Calcium chelators, such as EGTA and BAPTA/AM, also showed no effect on TPx-II induction by arsenite. However, arsenite-induced TPx-II expression was dose-dependently reduced by two broad–based protein kinase C (PKC) inhibitors, Rottlerin and Staurosprine. Furthermore, two highly specific inhibitors of PKCδ, Go6983 and Calphostin C , also decreased arsenite-induced TPx II expression. The present results suggested that PKCδ might be involved in arsenite-induced TPx II expression in SA7N cells.
