細胞自噬及粒線體在爵床素A誘導人類膀胱癌細胞之細胞毒性的角色

膀胱癌在台灣癌症死因排名第十四位，全球癌症死因排名第九位。目前美國食品藥品監督管理局（US FDA）認可的膀胱癌第一線臨床用藥cisplatin及gemcitabine合併使用，對人體具有副作用及高復發的問題，因此需要尋找新藥物。爵床素A（JA）由傳統中草藥爵床全草萃取的純化物，本實驗室先前已發現，JA對人類正常周邊血單細胞毒性非常低，但對許多人類癌細胞株具毒殺能力、影響粒線體膜電位改變、引起細胞凋亡、誘導細胞自噬，JA也可以抑制腫瘤生長。因此，本研究將探討JA對於膀胱癌細胞T24及TSGH8301是否具毒殺能力、影響粒線體膜電位改變、誘導細胞自噬，及誘發選擇性粒線體細胞自噬（mitophagy），並結合臨床用藥，觀察是否能提高臨床用藥敏感性。以SRB assay檢測毒殺能力，發現毒殺能力與劑量呈正相關。利用acridine orange（AO）染劑及西方墨點法檢測，發現可以誘導細胞自噬的產生。利用染劑rhodamine123染劑檢測粒線體膜電位，發現可以誘導T24粒線體膜電位改變，JA並可誘發PINK1-Parkin及Bnip3，產生mitophagy。經NAO及MitoTracker deep red染劑及西方墨點法發現，T24的粒線體會有生合成及膨脹的現象。JA與cisplatin及gemcitabine合併使用，在T24細胞呈現加成效果。整體而言，本研究結果顯示，JA可誘發膀胱癌細胞T24產生毒殺及提升藥物敏感性，可能與JA誘導細胞自噬及mitophagy現象有關。

關鍵字

爵床素A ；人類膀胱癌細胞；細胞自噬；選擇性粒線體自噬； cisplatin ； gemcitabine

並列摘要

Bladder cancer is the 14th death of cancer in Taiwan, and is the 9th of death of cancer in the world. The US FDA-approved first line clinical drugs, cisplatin combined with gemcitabine, exhibit side effects and relapse problems. Thus, to develop new drugs for bladder cancer is urgent. Justicidine A (JA) is a purified compound isolated from whole Justicia procumbens. Our previous published reports indicate that JA showed low cytotoxicity to human normal peripheral blood mononuclear cells, whereas JA induced cytotoxicity, disrupted mitochondrial membrane potential, and induced apoptosis and autophagy in many human cancer cell lines. JA also suppressed tumor growth. The present study is proposed to investigate if JA can exhibit cytotoxicity, disrupt mitochondrial membrane potential, induce autophagy and mitophagy, as well as enhance the sensitivity of clinical drugs against human bladder cancer cell lines T24 and TSGH8301. Here we show that JA inhibited the population growth of both cells using SRB assay. Flow cytometric and Western analysis indicate that JA induced autophagy in both cell lines. JA also disrupted mitochondrial membrane potential in T24 cells after staining with rhodamin123. The JA-induced mitophagy was via PINK1-Parkin and Bnip3 pathways in T24 cells. By staining with NAO and MitoTracker deep red, JA promoted mitochondrial biogenesis and swelling phenomena. Synergistic effect was found when T24 cells were treated with JA in combination with cisplatin and gemcitabine. Taken together, our data suggest that JA-induced cytotoxicity and enhanced sensitivity of clinical drugs may be associated with the induction of autophagy and mitophagy in T24 cells.

並列關鍵字

Justicidin A ； human bladder cancer ； autophagy ； mitophagy ； cisplatin ； gemcitabine

參考文獻

Akita M, Suzuki-Karasaki M, Fujiwara K, Nakagawa C, Soma M,

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Yoshida Y, Ochiai T, Tokuhashi Y, Suzuki-Karasaki Y. Mitochondrial division inhibitor-1 induces mitochondrial hyperfusion and sensitizes human cancer cells to TRAIL-induced apoptosis. (2014). Int J Oncol. 45(5):1901-12. doi: 10.3892/ijo.2014.2608.