Department of Biochemistry, College of Medicine, The Catholic University of Korea, Seoul 137-701, Republic of Korea.
Oncol Rep. 2012 Dec;28(6):2049-56. doi: 10.3892/or.2012.2034. Epub 2012 Sep 17.
Anthocyanins (ATCs) have been reported to induce apoptosis in various types of cancer cells, stimulating the development of ATCs as a cancer chemotherapeutic or chemopreventive agent. It was recently reported that ATCs can induce autophagy, however, the mechanism for this remains unclear. In the present report, we carried out mechanistic studies of the mechanism involved in ATC-induced autophagy using ATCs extracted from black soybeans (cv. Cheongja 3, Glycine max L.). ATCs clearly induced hallmarks of autophagy, including LC3 puncta formation and the conversion of LC3-I to LC3-II in U2OS human osteosarcoma cells. The induction of autophagy was accompanied by the phosphorylation of multiple protein kinases including extracellular signal-regulated kinase (ERK)1/2, p38 mitogen-activated protein kinase (MAPK), c-Jun N-terminal kinase (JNK), protein kinase B (AKT) and adenosyl mono-phosphate-dependent protein kinase (AMPK). While chemical inhibitors against ERK1/2, p38 MAPK, JNK and AKT failed to inhibit ATC-induced autophagy, the suppression of AMPK by compound C (CC) as well as siRNA against AMPK reduced ATC-induced autophagy. The treatment of ATCs resulted in a decrease in intracellular ATP contents and the activation of AMPK by AICAR treatment also induced autophagy. It is noteworthy that the reduction of autophagy via the inhibition of AMPK resulted in enhanced apoptosis in ATC-treated cells. In addition, siRNA against forkhead box O3A (FOXO3a), a downstream target of AMPK, suppressed ATC-induced autophagy and p27KIP1 siRNA increased apoptosis in ATC-treated cells. Collectively, it can be concluded that ATCs induce autophagy in U2OS cells via activation of the AMPK-FOXO3a pathway and protect cells from ATC-induced apoptosis via the AMPK-p27KIP1 pathway. These results also suggest that autophagy-modulating agents could contribute to the efficient development of ATCs as anticancer therapy.
花色苷（ATCs）已被报道能诱导各种类型的癌细胞凋亡，这刺激了 ATCs 作为癌症化疗或化学预防剂的发展。最近有报道称，ATCs 可以诱导自噬，然而，其机制尚不清楚。在本报告中，我们使用从黑豆（cv. 青荚 3，Glycine max L.）中提取的 ATCs 进行了 ATC 诱导自噬的机制研究。ATCs 明显诱导了自噬的标志性特征，包括 LC3 斑点形成和 LC3-I 向 LC3-II 的转化，在 U2OS 人骨肉瘤细胞中。自噬的诱导伴随着多种蛋白激酶的磷酸化，包括细胞外信号调节激酶（ERK）1/2、p38 丝裂原活化蛋白激酶（p38 MAPK）、c-Jun N 端激酶（JNK）、蛋白激酶 B（AKT）和腺苷单磷酸依赖性蛋白激酶（AMPK）。虽然针对 ERK1/2、p38 MAPK、JNK 和 AKT 的化学抑制剂未能抑制 ATC 诱导的自噬，但用化合物 C（CC）抑制 AMPK 以及针对 AMPK 的 siRNA 减少了 ATC 诱导的自噬。ATCs 的处理导致细胞内 ATP 含量减少，而 AICAR 处理激活 AMPK 也诱导了自噬。值得注意的是，通过抑制 AMPK 减少自噬导致 ATC 处理的细胞中凋亡增加。此外，针对 AMPK 的下游靶标叉头框 O3A（FOXO3a）的 siRNA 抑制了 ATC 诱导的自噬，而 p27KIP1 siRNA 增加了 ATC 处理的细胞中的凋亡。总之，可以得出结论，ATCs 通过激活 AMPK-FOXO3a 途径诱导 U2OS 细胞中的自噬，并通过 AMPK-p27KIP1 途径保护细胞免受 ATC 诱导的凋亡。这些结果还表明，自噬调节剂可能有助于高效开发 ATCs 作为抗癌疗法。
