Role of NOX1 and NOX5 in protein kinase C/reactive oxygen species‑mediated MMP‑9 activation and invasion in MCF‑7 breast cancer cells

Song,Hyun-Kyung; Kim,Jeong-Mi; Noh,Eun-Mi; Youn,Hyun Jo; Lee,Young-Rae

doi:10.3892/mmr.2024.13312

Role of NOX1 and NOX5 in protein kinase C/reactive oxygen species‑mediated MMP‑9 activation and invasion in MCF‑7 breast cancer cells

Authors:
- Hyun-Kyung Song
- Jeong-Mi Kim
- Eun-Mi Noh
- Hyun Jo Youn
- Young-Rae Lee
View Affiliations

Affiliations: 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, Department of Oral Biochemistry, School of Dentistry, Wonkwang University, Iksan, Jeollabuk 54538, Republic of Korea, Department of Surgery, Research Institute of Clinical Medicine, Jeonbuk National University Hospital, Jeonbuk National University and Biomedical Research Institute, Jeonju, Jeollabuk 54907, Republic of Korea
Published online on: August 20, 2024 https://doi.org/10.3892/mmr.2024.13312
Article Number: 188
Copyright: © Song et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

Metrics: Total Views: 0 (Spandidos Publications: | PMC Statistics: )

Metrics: Total PDF Downloads: 0 (Spandidos Publications: | PMC Statistics: )

Cited By (CrossRef): 0 citations Loading Articles...

Abstract

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.

View Figures

View References

1	Jemal A, Murray T, Ward E, Samuels A, Tiwari RC, Ghafoor A, Feuer EJ and Thun MJ: Cancer statistics, 2005. CA Cancer J Clin. 55:10–30. 2005. View Article : Google Scholar : PubMed/NCBI
2	Anne N, Sulger E and Pallapothu R: Primary squamous cell carcinoma of the breast: A case report and review of the literature. J Surg Case Rep. 2019:rjz1822019. View Article : Google Scholar : PubMed/NCBI
3	Zagelbaum NK, Ward MF II, Okby N and Karpoff H: Invasive ductal carcinoma of the breast with osteoclast-like giant cells and clear cell features: A case report of a novel finding and review of the literature. World J Surg Oncol. 14:2272016. View Article : Google Scholar : PubMed/NCBI
4	Berman AT, Thukral AD, Hwang WT, Solin LJ and Vapiwala N: Incidence and patterns of distant metastases for patients with early-stage breast cancer after breast conservation treatment. Clin Breast Cancer. 13:88–94. 2013. View Article : Google Scholar : PubMed/NCBI
5	Chambers AF and Matrisian LM: Changing views of the role of matrix metalloproteinases in metastasis. J Natl Cancer Inst. 89:1260–1270. 1997. View Article : Google Scholar : PubMed/NCBI
6	Woessner JF Jr: Matrix metalloproteinases and their inhibitors in connective tissue remodeling. FASEB J. 5:2145–2154. 1991. View Article : Google Scholar : PubMed/NCBI
7	Mizutani K, Kofuji K and Shirouzu K: The significance of MMP-1 and MMP-2 in peritoneal disseminated metastasis of gastric cancer. Surg Today. 30:614–621. 2000. View Article : Google Scholar : PubMed/NCBI
8	Zeigler ME, Chi Y, Schmidt T and Varani J: Role of ERK and JNK pathways in regulating cell motility and matrix metalloproteinase 9 production in growth factor-stimulated human epidermal keratinocytes. J Cell Physiol. 180:271–284. 1999. View Article : Google Scholar : PubMed/NCBI
9	Hozumi A, Nishimura Y, Nishiuma T, Kotani Y and Yokoyama M: Induction of MMP-9 in normal human bronchial epithelial cells by TNF-alpha via NF-kappa B-mediated pathway. Am J Physiol Lung Cell Mol Physiol. 281:L1444–L1452. 2001. View Article : Google Scholar : PubMed/NCBI
10	Weng CJ, Chau CF, Hsieh YS, Yang SF and Yen GC: Lucidenic acid inhibits PMA-induced invasion of human hepatoma cells through inactivating MAPK/ERK signal transduction pathway and reducing binding activities of NF-kappaB and AP-1. Carcinogenesis. 29:147–156. 2008. View Article : Google Scholar : PubMed/NCBI
11	Lee HB, Yu MR, Song JS and Ha H: Reactive oxygen species amplify protein kinase C signaling in high glucose-induced fibronectin expression by human peritoneal mesothelial cells. Kidney Int. 65:1170–1179. 2004. View Article : Google Scholar : PubMed/NCBI
12	Wu WS, Tsai RK, Chang CH, Wang S, Wu JR and Chang YX: Reactive oxygen species mediated sustained activation of protein kinase C alpha and extracellular signal-regulated kinase for migration of human hepatoma cell Hepg2. Mol Cancer Res. 4:747–758. 2006. View Article : Google Scholar : PubMed/NCBI
13	Traore K, Sharma RB, Burek CL and Trush MA: Role of ROS and MAPK in TPA-induced ICAM-1 expression in the myeloid ML-1 cell line. J Cell Biochem. 100:1010–1021. 2007. View Article : Google Scholar : PubMed/NCBI
14	McCubrey JA, Lahair MM and Franklin RA: Reactive oxygen species-induced activation of the MAP kinase signaling pathways. Antioxid Redox Signal. 8:1775–1789. 2006. View Article : Google Scholar : PubMed/NCBI
15	Huang L, Lin H, Chen Q, Yu L and Bai D: MPPa-PDT suppresses breast tumor migration/invasion by inhibiting Akt-NF-κB-dependent MMP-9 expression via ROS. BMC Cancer. 19:11592019. View Article : Google Scholar : PubMed/NCBI
16	Lee GH, Jin SW, Kim SJ, Pham TH, Choi JH and Jeong HG: Tetrabromobisphenol A induces MMP-9 expression via NADPH Oxidase and the activation of ROS, MAPK, and Akt pathways in human breast cancer MCF-7 Cells. Toxicol Res. 35:93–101. 2019. View Article : Google Scholar : PubMed/NCBI
17	Hsu TC, Young MR, Cmarik J and Colburn NH: Activator protein 1 (AP-1)- and nuclear factor kappaB (NF-kappaB)-dependent transcriptional events in carcinogenesis. Free Radic Biol Med. 28:1338–1348. 2000. View Article : Google Scholar : PubMed/NCBI
18	Huang Q, Shen HM and Ong CN: Inhibitory effect of emodin on tumor invasion through suppression of activator protein-1 and nuclear factor-kappaB. Biochem Pharmacol. 68:361–371. 2004. View Article : Google Scholar : PubMed/NCBI
19	Savaraj N, Wei Y, Unate H, Liu PM, Wu CJ, Wangpaichitr M, Xia D, Xu HJ, Hu SX and Tien Kuo M: Redox regulation of matrix metalloproteinase gene family in small cell lung cancer cells. Free Radic Res. 39:373–381. 2005. View Article : Google Scholar : PubMed/NCBI
20	Karin M: The regulation of AP-1 activity by mitogen-activated protein kinases. J Biol Chem. 270:16483–16486. 1995. View Article : Google Scholar : PubMed/NCBI
21	Madrid LV, Mayo MW, Reuther JY and Baldwin AS Jr: Akt stimulates the transactivation potential of the RelA/p65 Subunit of NF-kappa B through utilization of the Ikappa B kinase and activation of the mitogen-activated protein kinase p38. J Biol Chem. 276:18934–18940. 2001. View Article : Google Scholar : PubMed/NCBI
22	Brandes RP, Weissmann N and Schroder K: Nox family NADPH oxidases: Molecular mechanisms of activation. Free Radic Biol Med. 76:208–226. 2014. View Article : Google Scholar : PubMed/NCBI
23	Schmittgen TD and Livak KJ: Analyzing real-time PCR data by the comparative C(T) method. Nat Protoc. 3:1101–1108. 2008. View Article : Google Scholar : PubMed/NCBI
24	Kim JM, Park J, Noh EM, Song HK, Kang SY, Jung SH, Kim JS, Park BH, Lee YR and Youn HJ: Bruton's agammaglobulinemia tyrosine kinase (Btk) regulates TPA-induced breast cancer cell invasion via PLCγ2/PKCβ/NF-κB/AP-1-dependent matrix metalloproteinase-9 activation. Oncol Rep. 45:562021. View Article : Google Scholar : PubMed/NCBI
25	Noh EM, Park YJ, Kim JM, Kim MS, Kim HR, Song HK, Hong OY, So HS, Yang SH, Kim JS, et al: Fisetin regulates TPA-induced breast cell invasion by suppressing matrix metalloproteinase-9 activation via the PKC/ROS/MAPK pathways. Eur J Pharmacol. 764:79–86. 2015. View Article : Google Scholar : PubMed/NCBI
26	Piszczatowska K, Przybylska D, Sikora E and Mosieniak G: Inhibition of NADPH oxidases activity by diphenyleneiodonium chloride as a mechanism of senescence induction in human cancer cells. Antioxidants (Basel). 9:12482020. View Article : Google Scholar : PubMed/NCBI
27	Ren G, Luo W, Sun W, Niu Y, Ma DL, Leung CH, Wang Y, Lu JJ and Chen X: Psoralidin induced reactive oxygen species (ROS)-dependent DNA damage and protective autophagy mediated by NOX4 in breast cancer cells. Phytomedicine. 23:939–947. 2016. View Article : Google Scholar : PubMed/NCBI
28	Nasimian A, Farzaneh P, Tamanoi F and Bathaie SZ: Cytosolic and mitochondrial ROS production resulted in apoptosis induction in breast cancer cells treated with Crocin: The role of FOXO3a, PTEN and AKT signaling. Biochem Pharmacol. 177:1139992020. View Article : Google Scholar : PubMed/NCBI
29	D'Anneo A, Carlisi D, Emanuele S, Buttitta G, Di Fiore R, Vento R, Tesoriere G and Lauricella M: Parthenolide induces superoxide anion production by stimulating EGF receptor in MDA-MB-231 breast cancer cells. Int J Oncol. 43:1895–1900. 2013. View Article : Google Scholar : PubMed/NCBI
30	Kaikai S, Lu F, Xie J, Wu M, Cai B, Liu Y, Zhang H, Tan H, Pan Y and Xu H: Cambogin exerts anti-proliferative and pro-apoptotic effects on breast adenocarcinoma through the induction of NADPH oxidase 1 and the alteration of mitochondrial morphology and dynamics. Oncotarget. 7:50596–50611. 2016. View Article : Google Scholar : PubMed/NCBI
31	Jacobson PB, Kuchera SL, Metz A, Schächtele C, Imre K and Schrier DJ: Anti-inflammatory properties of Gö 6850: A selective inhibitor of protein kinase C. J Pharmacol Exp Ther. 275:995–1002. 1995.PubMed/NCBI
32	Augsburger F, Filippova A, Rasti D, Seredenina T, Lam M, Maghzal G, Mahiout Z, Jansen-Dürr P, Knaus UG, Doroshow J, et al: Pharmacological characterization of the seven human NOX isoforms and their inhibitors. Redox Biol. 26:1012722019. View Article : Google Scholar : PubMed/NCBI
33	Siegel R, Ma J, Zou Z and Jemal A: Cancer statistics, 2014. CA Cancer J Clin. 64:9–29. 2014. View Article : Google Scholar : PubMed/NCBI
34	Redig AJ and McAllister SS: Breast cancer as a systemic disease: A view of metastasis. J Intern Med. 274:113–126. 2013. View Article : Google Scholar : PubMed/NCBI
35	Jiang WG, Sanders AJ, Katoh M, Ungefroren H, Gieseler F, Prince M, Thompson SK, Zollo M, Spano D, Dhawan P, et al: Tissue invasion and metastasis: Molecular, biological and clinical perspectives. Semin Cancer Biol. 35 (Suppl):S244–S275. 2015. View Article : Google Scholar : PubMed/NCBI
36	Duffy MJ, Maguire TM, Hill A, McDermott E and O'Higgins N: Metalloproteinases: Role in breast carcinogenesis, invasion and metastasis. Breast Cancer Res. 2:252–257. 2000. View Article : Google Scholar : PubMed/NCBI
37	Scorilas A, Karameris A, Arnogiannaki N, Bassilopoulos P, Trangas T and Talieri M: Overexpression of matrix-metalloproteinase-9 in human breast cancer: A potential favourable indicator in node-negative patients. Br J Cancer. 84:1488–1496. 2001. View Article : Google Scholar : PubMed/NCBI
38	Yan C and Boyd DD: Regulation of matrix metalloproteinase gene expression. J Cell Physiol. 211:19–26. 2007. View Article : Google Scholar : PubMed/NCBI
39	Landry WD and Cotter TG: ROS signalling, NADPH oxidases and cancer. Biochem Soc Trans. 42:934–938. 2014. View Article : Google Scholar : PubMed/NCBI
40	Jiang F, Zhang Y and Dusting GJ: NADPH oxidase-mediated redox signaling: Roles in cellular stress response, stress tolerance, and tissue repair. Pharmacol Rev. 63:218–242. 2011. View Article : Google Scholar : PubMed/NCBI
41	Bedard K and Krause KH: The NOX family of ROS-generating NADPH oxidases: Physiology and pathophysiology. Physiol Rev. 87:245–313. 2007. View Article : Google Scholar : PubMed/NCBI
42	Lambeth JD: NOX enzymes and the biology of reactive oxygen. Nat Rev Immunol. 4:181–189. 2004. View Article : Google Scholar : PubMed/NCBI
43	Kamata T: Roles of Nox1 and other Nox isoforms in cancer development. Cancer Sci. 100:1382–1388. 2009. View Article : Google Scholar : PubMed/NCBI
44	Block K and Gorin Y: Aiding and abetting roles of NOX oxidases in cellular transformation. Nat Rev Cancer. 12:627–637. 2012. View Article : Google Scholar : PubMed/NCBI
45	Deep G, Kumar R, Jain AK, Dhar D, Panigrahi GK, Hussain A, Agarwal C, El-Elimat T, Sica VP, Oberlies NH and Agarwal R: Graviola inhibits hypoxia-induced NADPH oxidase activity in prostate cancer cells reducing their proliferation and clonogenicity. Sci Rep. 6:231352016. View Article : Google Scholar : PubMed/NCBI
46	Shinohara M, Adachi Y, Mitsushita J, Kuwabara M, Nagasawa A, Harada S, Furuta S, Zhang Y, Seheli K, Miyazaki H and Kamata T: Reactive oxygen generated by NADPH oxidase 1 (Nox1) contributes to cell invasion by regulating matrix metalloprotease-9 production and cell migration. J Biol Chem. 285:4481–4488. 2010. View Article : Google Scholar : PubMed/NCBI
47	Juhasz A, Ge Y, Markel S, Chiu A, Matsumoto L, van Balgooy J, Roy K and Doroshow JH: Expression of NADPH oxidase homologues and accessory genes in human cancer cell lines, tumours and adjacent normal tissues. Free Radic Res. 43:523–532. 2009. View Article : Google Scholar : PubMed/NCBI
48	Dho SH, Kim JY, Lee KP, Kwon ES, Lim JC, Kim CJ, Jeong D and Kwon KS: STAT5A-mediated NOX5-L expression promotes the proliferation and metastasis of breast cancer cells. Exp Cell Res. 351:51–58. 2017. View Article : Google Scholar : PubMed/NCBI
49	Lim PS, Sutton CR and Rao S: Protein kinase C in the immune system: From signalling to chromatin regulation. Immunology. 146:508–522. 2015. View Article : Google Scholar : PubMed/NCBI
50	Parekh DB, Ziegler W and Parker PJ: Multiple pathways control protein kinase C phosphorylation. EMBO J. 19:496–503. 2000. View Article : Google Scholar : PubMed/NCBI
51	Kim JM, Park J, Noh EM, Song HK, Kang SY, Jung SH, Kim JS, Youn HJ and Lee YR: Downregulation of matriptase suppresses the PAR-2/PLCγ2/PKC-mediated invasion and migration abilities of MCF-7 breast cancer cells. Oncol Rep. 46:2472021. View Article : Google Scholar : PubMed/NCBI
52	Wu WS: The signaling mechanism of ROS in tumor progression. Cancer Metastasis Rev. 25:695–705. 2006. View Article : Google Scholar : PubMed/NCBI
53	Eid BG, Abu-Sharib AT, El-Bassossy HM, Balamash K and Smirnov SV: Enhanced calcium entry via activation of NOX/PKC underlies increased vasoconstriction induced by methylglyoxal. Biochem Biophys Res Commun. 506:1013–1018. 2018. View Article : Google Scholar : PubMed/NCBI
54	Cui X, Li X, He Y, Yu J, Fu J, Song B and Zhao RC: Combined NOX/ROS/PKC signaling pathway and metabolomic analysis reveals the mechanism of TRAM34-Induced endothelial progenitor cell senescence. Stem Cells Dev. 30:671–682. 2021. View Article : Google Scholar : PubMed/NCBI
55	Chen F, Yu Y, Haigh S, Johnson J, Lucas R, Stepp DW and Fulton DJ: Regulation of NADPH oxidase 5 by protein kinase C isoforms. PLoS One. 9:e884052014. View Article : Google Scholar : PubMed/NCBI
56	Lee TH, Chen JL, Liu PS, Tsai MM, Wang SJ and Hsieh HL: Rottlerin, a natural polyphenol compound, inhibits upregulation of matrix metalloproteinase-9 and brain astrocytic migration by reducing PKC-delta-dependent ROS signal. J Neuroinflammation. 17:1772020. View Article : Google Scholar : PubMed/NCBI
57	Brandes RP and Schroder K: NOXious phosphorylation: Smooth muscle reactive oxygen species production is facilitated by direct activation of the NADPH oxidase Nox1. Circ Res. 115:898–900. 2014. View Article : Google Scholar : PubMed/NCBI