Glutathione peroxidase 4 maintains a stemness phenotype, oxidative homeostasis and regulates biological processes in Panc‑1 cancer stem‑like cells

Abstract

Reactive oxygen species (ROS) have been widely accepted as critical molecules playing regulatory roles in various biological processes, including proliferation, differentiation and apoptotic/ferroptotic/necrotic cell death. Emerging evidence suggests that ROS may be involved in the induction of epithelial-to-mesenchymal transition (EMT), which has been reported to promote cancer stem-like cell (CSC) generation. Recent data indicate that altered accumulation of ROS is associated with CSC generation, EMT and hypoxia exposure, but the underlying mechanisms are poorly understood. In the present study, we derived CSCs from Panc-1 human pancreatic cancer cells and characterized them using serial replating assays and western blot analysis. Functional identification of viable cells was performed using the CCK-8 assay and colony formation assays. The expression of various antioxidant enzymes, including superoxide dismutase (SOD) and glutathione peroxidase (GPX), was measured by western blot analysis in Panc-1 CSCs. The role of GPX4 in regulating biological processes of Panc-1 CSCs was assessed by proliferation, sphere formation and invasion assays with or without oxidative stress. Manipulation of GPX4 expression by siRNA knockdown or an overexpression vector was performed to assess functions including proliferation, colony formation and invasion. EMT hallmark genes were detected after GPX4 alteration by RT-qPCR and western blot analysis. Panc-1 CSCs displayed more resistance to hypoxia exposure. Compared with the parental Panc-1 cells, Panc-1 CSCs expressed an obviously higher endogenous GPX4 level, indicating their role in maintaining homeostasis. During GPX4 knockdown, ROS accumulation was promoted following oxidative stress exposure to either H 2 O 2 or erastin. Additionally, overexpression of GPX4 eliminated ROS induction by oxidative stress exposure and thus, exerted protective effects on physiological processes in the Panc-1 CSCs. Knockdown of GPX4 arrested cell cycle progression at the G 1 /G 0 phase; inhibited cell proliferation, colony formation, invasion and the stemness phenotype in the Panc-1 CSCs; and decreased the EMT phenotype. Collectively, GPX4 plays a critical role in maintaining oxidative homeostasis and regulates several biological processes, including stemness and EMT, in Panc-1 CSCs.

The NRF2, Thioredoxin, and Glutathione System in Tumorigenesis and Anticancer Therapies

“…[242] GPX4 deficiency induces G1/G0 cell cycle arrest and inhibits tumorigenesis in pancreatic cancer stem-like cells. [278] GPX2 overexpression correlates with the activation of epithelial to mesenchymal transition, the activation of β-catenin-WNT signaling, and the increased proliferation and metastasis of cervical cancer cells.…”

“…GSTP1 is also downregulated by the methylation of the CpG island [277], while the recruitment of early B cell factor 1 to its promoter upregulates GSTP1 [255]. The overexpression of GSTP1 can induce cell cycle arrest [254], as well as GPX4 deficiency [278], and can thus be used as potential targets in anticancer therapy. GSTPs catalytic activity varies depending on the target.…”

mTOR-Mediated Antioxidant Activation in Solid Tumor Radioresistance

“…Non-small-cell lung cancer (NSCLC) cells overexpressing GPX1 were resistant to cisplatin-induced ROS through PI3K/AKT pathway activation [85]. In pancreatic cancer cells, GPX4 was shown to be essential for the maintenance of stemness, and PRX1 was required for p38-mediated invasion [86]. A high level of PRX2 in colorectal cancer patients was associated with tumor progression and poor diagnosis [87].…”

Deconvolution of autoencoders to learn biological regulatory modules from single cell mRNA sequencing data

“…The direction of change in hidden units that could signify stemness or progenitor states are up in CMP, i.e. WNT-{beta}catenin-signaling, described as key stemness factor [30], and DNA repair and hypoxia, both associated with stemness [31,32]. Relative to the control, the CMPs show less activity in pathways that could be associated with differentiation, division and maturation, in terms like mitotic spindle, Apical changes and Hedgehog signaling [33].…”