Zhou Guang-Zhou, Zhang Shuai-Na, Zhang Lu, Sun Gang-Chun, Chen Xiao-Bing
College of Bioengineering .
Pharm Biol. 2014 Jan;52(1):111-6. doi: 10.3109/13880209.2013.816971. Epub 2013 Sep 18.
Curcumin exhibits growth-suppressive activity against a variety of cancer cells, but low bioavailability restricts its application in chemotherapeutic trials. Nowadays, a growing number of curcumin derivatives or analogs are known, hoping to replace curcumin and circumvent this problem. Hydrazinobenzoylcurcumin (HBC) has been synthesized and identified as a potent inhibitor of cell proliferation in previous reports.
This study presents a novel mechanism of cell autophagy induced by HBC in the human non-small lung epithelial carcinoma (A549) cells.
Cells were cultured and treated with HBC at different concentrations (10-80 μM) and at different time periods (1-24 h). Microscopic analysis was used to detect the morphology changes and autophagolysosomes of A549 cells. An acridine orange staining assay was conducted to evaluate the autophagolysosomes and autophagic vacuoles was analyzed by monodansylcadaverine (MDC) and GFP-LC3 transfection analysis. Western blotting was used to assess the conversion of microtubule-associated protein light chain 3 (LC3).
HBC could induce A549 cells autophagolysosomes formation in a dose and time-dependent manner and the inhibitory rate of HBC (80 μM) on the viability of A549 cells reached 76.68 ± 5.81% after 24 h of treatment. Autophagic vacuoles increased in a concentration-dependent manner in HBC-treated cell. Furthermore, conversion of LC3-I to LC3-II, accumulation of GFP-tagged LC3 positive intracellular vacuoles and increased fusion of autophagosomes with lysosomes suggested the occurrence of autophagy.
Our data indicate that HBC induced A549 cell autophagy, which is a novel cell death mechanism induced by curcumin derivatives.
姜黄素对多种癌细胞具有生长抑制活性，但低生物利用度限制了其在化疗试验中的应用。如今，已知越来越多的姜黄素衍生物或类似物，希望能取代姜黄素并解决这一问题。肼基苯甲酰姜黄素（HBC）已被合成，并在先前的报道中被鉴定为一种有效的细胞增殖抑制剂。
本研究提出了HBC诱导人非小肺上皮癌（A549）细胞自噬的新机制。
将细胞培养并用不同浓度（10 - 80μM）的HBC在不同时间段（1 - 24小时）进行处理。采用显微镜分析检测A549细胞的形态变化和自噬溶酶体。进行吖啶橙染色试验以评估自噬溶酶体，并用单丹磺酰尸胺（MDC）和绿色荧光蛋白-微管相关蛋白轻链3（GFP-LC3）转染分析自噬空泡。采用蛋白质免疫印迹法评估微管相关蛋白轻链3（LC3）的转化。
HBC可呈剂量和时间依赖性诱导A549细胞自噬溶酶体形成，处理24小时后，HBC（80μM）对A549细胞活力的抑制率达到76.68±5.81%。在HBC处理的细胞中，自噬空泡以浓度依赖性方式增加。此外，LC3-I向LC3-II的转化、绿色荧光蛋白标记的LC3阳性细胞内空泡的积累以及自噬体与溶酶体融合增加提示自噬的发生。
我们的数据表明，HBC诱导A549细胞自噬，这是一种由姜黄素衍生物诱导的新的细胞死亡机制。
