Chen Min, Wang Jian, Yao Shi-Fei, Zhao Yi, Liu Lu, Li Lian-Wen, Xu Ting, Gan Liu-Gen, Xiao Chun-Lan, Shan Zhi-Ling, Zhong Liang, Liu Bei-Zhong
Central Laboratory of Yong-chuan Hospital, Chongqing Medical University, Chongqing, 402160, China.
Key Laboratory of Laboratory Medical Diagnostics, Ministry of Education, Department of Laboratory Medicine, Chongqing Medical University, Chongqing, 400016, China.
Int J Med Sci. 2017 Jul 20;14(9):902-910. doi: 10.7150/ijms.19965. eCollection 2017.
Yes-associated protein (YAP), the nuclear effector of the Hippo pathway, is a candidate oncoprotein and participates in the progression of various malignancies. However, few reports have examined the effect of YAP inhibition in human leukemia HL-60 cells. We examined the effects of YAP knockdown or inhibition using short hairpin RNA (shRNA) or verteporfin (VP), respectively. Western blot assays were used to determine the expression levels of YAP, Survivin, cyclinD1, PARP, Bcl-2, and Bax. Cell proliferation was assessed using the cell counting kit (CCK-8) assay. Cell cycle progression and apoptosis were evaluated by flow cytometry, and apoptotic cell morphology was observed by Hoechst 33342 staining. Knockdown or inhibition of YAP led to cell cycle arrest at the G0/G1 phase and increased apoptosis, inhibited cell proliferation, increased levels of Bax and cleaved PARP, and decreased levels of PARP, Bcl-2, Survivin, and cyclinD1. Moreover, Hoechst 33342 staining revealed increased cell nuclear fragmentation. Collectively, these results show that inhibition of YAP inhibits proliferation and induces apoptosis in HL-60 cells. Therefore, a novel treatment regime involving genetic or pharmacological inhibition of YAP could be established for acute promyelocytic leukemia.
Yes相关蛋白（YAP）是Hippo信号通路的核效应因子，是一种候选癌蛋白，参与多种恶性肿瘤的进展。然而，很少有报道研究YAP抑制对人白血病HL-60细胞的影响。我们分别使用短发夹RNA（shRNA）或维替泊芬（VP）检测YAP敲低或抑制的效果。采用蛋白质免疫印迹法检测YAP、生存素、细胞周期蛋白D1、聚（ADP-核糖）聚合酶（PARP）、Bcl-2和Bax的表达水平。使用细胞计数试剂盒（CCK-8）检测法评估细胞增殖。通过流式细胞术评估细胞周期进程和凋亡情况，并用Hoechst 33342染色观察凋亡细胞形态。YAP的敲低或抑制导致细胞周期停滞在G0/G1期，凋亡增加，细胞增殖受到抑制，Bax和裂解的PARP水平升高，PARP、Bcl-2、生存素和细胞周期蛋白D1水平降低。此外，Hoechst 33342染色显示细胞核碎片化增加。总的来说，这些结果表明抑制YAP可抑制HL-60细胞的增殖并诱导其凋亡。因此，可为急性早幼粒细胞白血病建立一种涉及基因或药物抑制YAP的新型治疗方案。
