Practical Research Division, Honam National Institute of Biological Resources, Mokpo, Jeollanam 58762, Republic of Korea.
Department of Biochemistry, Jeonbuk National University Medical School, Jeonju, Jeollabuk 54907, Republic of Korea.
Mol Med Rep. 2024 Oct;30(4). doi: 10.3892/mmr.2024.13312. Epub 2024 Sep 2.
NADPH oxidases (NOXs) are a family of membrane proteins responsible for intracellular reactive oxygen species (ROS) generation by facilitating electron transfer across biological membranes. Despite the established activation of NOXs by protein kinase C (PKC), the precise mechanism through which PKC triggers NOX activation during breast cancer invasion remains unclear. The present study aimed to investigate the role of NOX1 and NOX5 in the invasion of MCF‑7 human breast cancer cells. The expression and activity of NOXs and matrix metalloprotease (MMP)‑9 were assessed by reverse transcription‑quantitative PCR and western blotting, and the activity of MMP‑9 was monitored using zymography. Cellular invasion was assessed using the Matrigel invasion assay, whereas ROS levels were quantified using a FACSCalibur flow cytometer. The findings suggested that NOX1 and NOX5 serve crucial roles in 12‑O‑tetradecanoylphorbol‑13‑acetate (TPA)‑induced MMP‑9 expression and invasion of MCF‑7 cells. Furthermore, a connection was established between PKC and the NOX1 and 5/ROS signaling pathways in mediating TPA‑induced MMP‑9 expression and cellular invasion. Notably, NOX inhibitors (diphenyleneiodonium chloride and apocynin) significantly attenuated TPA‑induced MMP‑9 expression and invasion in MCF‑7 cells. NOX1‑ and NOX5‑specific small interfering RNAs attenuated TPA‑induced MMP‑9 expression and cellular invasion. In addition, knockdown of NOX1 and NOX5 suppressed TPA‑induced ROS levels. Furthermore, a PKC inhibitor (GF109203X) suppressed TPA‑induced intracellular ROS levels, MMP‑9 expression and NOX activity in MCF‑7 cells. Therefore, NOX1 and NOX5 may serve crucial roles in TPA‑induced MMP‑9 expression and invasion of MCF‑7 breast cancer cells. Furthermore, the present study indicated that TPA‑induced MMP‑9 expression and cellular invasion were mediated through PKC, thus linking the NOX1 and 5/ROS signaling pathways. These findings offer novel insights into the potential mechanisms underlying their anti‑invasive effects in breast cancer.
NADPH 氧化酶（NOXs）是一类膜蛋白，通过促进生物膜内电子转移，负责细胞内活性氧（ROS）的产生。尽管已经证实蛋白激酶 C（PKC）可激活 NOXs，但 PKC 在乳腺癌浸润过程中触发 NOX 激活的确切机制尚不清楚。本研究旨在探讨 NOX1 和 NOX5 在 MCF-7 人乳腺癌细胞浸润中的作用。通过逆转录定量 PCR 和 Western blot 评估 NOXs 和基质金属蛋白酶（MMP）-9 的表达和活性，并通过明胶酶谱法监测 MMP-9 的活性。通过 Matrigel 侵袭测定评估细胞侵袭，使用 FACSCalibur 流式细胞仪定量 ROS 水平。结果表明，NOX1 和 NOX5 在 12-O-十四烷酰佛波醇-13-乙酸酯（TPA）诱导的 MMP-9 表达和 MCF-7 细胞侵袭中发挥重要作用。此外，还发现 PKC 与 NOX1 和 5/ROS 信号通路之间存在联系，可介导 TPA 诱导的 MMP-9 表达和细胞侵袭。值得注意的是，NOX 抑制剂（二苯基碘鎓氯化物和 apocynin）可显著减弱 TPA 诱导的 MCF-7 细胞中 MMP-9 的表达和侵袭。NOX1 和 NOX5 的特异性小干扰 RNA 减弱了 TPA 诱导的 MMP-9 表达和细胞侵袭。此外，敲低 NOX1 和 NOX5 抑制了 TPA 诱导的 ROS 水平。此外，PKC 抑制剂（GF109203X）抑制了 TPA 诱导的 MCF-7 细胞内 ROS 水平、MMP-9 表达和 NOX 活性。因此，NOX1 和 NOX5 可能在 TPA 诱导的 MMP-9 表达和 MCF-7 乳腺癌细胞浸润中发挥重要作用。此外，本研究表明，TPA 诱导的 MMP-9 表达和细胞侵袭是通过 PKC 介导的，从而将 NOX1 和 5/ROS 信号通路联系起来。这些发现为其在乳腺癌中的抗侵袭作用提供了潜在机制的新见解。
