PRASE, DSST, Lebanese University, Beirut, Lebanon; Biology Department, Faculty of Sciences-I, Lebanese University, Beirut, Lebanon; Laboratory of Cancer Immunology and Cell Signaling (LCICS), Lebanese University, Faculty of Sciences, Beirut, Lebanon.
College of Medicine, Department of Basic Medical Sciences, Qatar university, Qatar.
Cell Signal. 2018 Oct;50:37-47. doi: 10.1016/j.cellsig.2018.06.009. Epub 2018 Jun 20.
PACAP has opposing roles ranging from activation to inhibition of tumor growth and PACAP agonists/antagonists could be used in tumor therapy. In this study, the effect of PACAP stimulation on signaling pathways was investigated in MCF-7 human adenocarcinoma breast cancer cells. Results showed that MCF-7 cells express VPAC1 and VPAC2, but not PAC1, receptors. In addition, PACAP increased the phosphorylation levels of STAT1, Src and Raf within seconds, confirming their involvement in early stages of PACAP signaling whereas maximal phosphorylation of AKT, ERK and p38 was reached 10 to 20 min later. Moreover, selective inhibition of Src or PI3K resulted in a significant decrease in the phosphorylation of ERK and AKT, but not p38, demonstrating that PACAP signaling follows Src/Raf/ERK and PI3K/AKT pathways. On the other hand, selective inhibition of PLC or PKA resulted in a significant decrease in the phosphorylation of p38, but not AKT or ERK, indicating that PACAP signaling also follows the PLC and PKA/cAMP pathways. Furthermore, PACAP induced ROS through H₂O₂ production whereas pretreatment with NAC inhibitor decreased AKT and ERK phosphorylation, but not p38. Selective NOX2 inhibition affected Src/Raf/Erk and PI3K/Akt pathways, without affecting the p38/PLC/PKA pathway whereas other inhibitors (ML171, VAS2870) had no effect on PACAP induced ROS generation. On the other hand, PACAP induced calcium release, which was decreased by pretreatment with PLC inhibitor. Finally, PACAP stimulation promoted apoptosis by increasing Bax and decreasing Bcl2 expression. In conclusion, we demonstrated that PACAP signaling in MCF-7 cells follows the Src/Raf/ERK and PI3K/AKT pathways and is VPAC1 dependent in a ROS dependent manner, whereas it follows PLC and PKA/cAMP pathways and is VPAC2 dependent through p38 MAP kinase activation involving calcium.
PACAP 对肿瘤生长具有促进或抑制的双重作用，PACAP 激动剂/拮抗剂可用于肿瘤治疗。本研究旨在探讨 PACAP 刺激对 MCF-7 人腺癌细胞信号通路的影响。结果显示 MCF-7 细胞表达 VPAC1 和 VPAC2 受体，但不表达 PAC1 受体。此外，PACAP 在数秒内即可增加 STAT1、Src 和 Raf 的磷酸化水平，证实它们参与了 PACAP 信号的早期阶段，而 AKT、ERK 和 p38 的最大磷酸化则在 10-20min 后达到。此外，Src 或 PI3K 的选择性抑制可显著降低 ERK 和 AKT 的磷酸化，但不影响 p38 的磷酸化，表明 PACAP 信号转导遵循 Src/Raf/ERK 和 PI3K/AKT 途径。另一方面，PLC 或 PKA 的选择性抑制可显著降低 p38 的磷酸化，但不影响 AKT 或 ERK 的磷酸化，表明 PACAP 信号转导也遵循 PLC 和 PKA/cAMP 途径。此外，PACAP 通过 H₂O₂的产生诱导 ROS，而 NAC 抑制剂预处理可降低 AKT 和 ERK 的磷酸化，但不影响 p38 的磷酸化。选择性 NOX2 抑制剂影响 Src/Raf/Erk 和 PI3K/Akt 途径，而不影响 p38/PLC/PKA 途径，而其他抑制剂（ML171、VAS2870）对 PACAP 诱导的 ROS 生成无影响。另一方面，PACAP 刺激可通过增加 Bax 表达和降低 Bcl2 表达促进细胞凋亡。综上所述，我们证实了 MCF-7 细胞中的 PACAP 信号转导通过 Src/Raf/ERK 和 PI3K/AKT 途径，并且依赖于 ROS 和 VPAC1，而通过 PLC 和 PKA/cAMP 途径，依赖于 p38 MAP 激酶激活和钙来激活 VPAC2。
