Anti-Migratory Effect of Dipotassium Glycyrrhizinate on Glioblastoma Cell Lines: Microarray Data for the Identification of Key MicroRNA Signatures

Author(s)

Bonafe, Gabriel Alves ; dos Santos, Jessica Silva ; Fernandes, Anna Maria Alves de Piloto ; Ziegler, Jussara Vaz ; Marson, Fernando Augusto Lima ; Rocha, Thalita ; Carvalho, Patricia de Oliveira ; Ortega, Manoela Marques

• Document type: Journal article
• Source: FRONTIERS IN ONCOLOGY; v. 12, p. 10-pg., 2022-08-03.

Abstract

The nuclear factor kappa B (NF-kappa B) pathway has been reported to be responsible for the aggressive disease phenomenon observed in glioblastoma (GBM). Dipotassium glycyrrhizinate (DPG), a dipotassium salt of glycyrrhizic acid isolated from licorice, has recently demonstrated an anti-tumoral effect on GBM cell lines U87MG and T98G through NF-kappa B suppression by IRAK2- and TRAF6-mediating microRNA (miR)-16 and miR-146a, respectively. Thus, the present study aimed to evaluate the expression profiles of miRNAs related to NF-kappa B suppression in T98G GBM cell line after DPG exposure using miRNA microarray (Affymetrix Human miRNA 4.0A), considering only predicted miRNAs as NF-kappa B regulator genes. Additional assays using U251 and U138MG cells were performed to validate the array results. DPG cytotoxicity was determined by (4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay, and cellular apoptosis was quantified by DNA fragmentation and terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) assay. The anti-proliferative effect was observed by cell proliferation and wound-healing assays, and the sphere formation assay examined whether DPG reduced stem cell subpopulation formation. The most over-expressed miRNAs were miR-4443 and miR-3620. The cytotoxic effect of DPG in U251 and U138MG was observed with an IC50 of 32 and 20 mM for 48 h, respectively. The IC50 of each cell line was used in all further assays. DPG treatment-induced apoptosis is observed by DNA fragmentation and increased TUNEL-positive cells. Cell proliferation and wound-healing assays showed an anti-proliferative and anti-migratory effect by DPG on the evaluated cell lines. In addition, DPG treatment led to a 100% reduction in sphere formation. The qPCR results in U251 and U138MG cells showed that DPG increased miR-4443 (2.44 vs. 1.11, p-value = 0.11; 8.27 vs. 1.25, p-value = 0.04) and miR-3620 expression (1.66 vs. 1.00, p-value = 0.03; 8.47 vs. 1.01, p-value = 0.03) and decreased CD209 (0.44 vs. 1.10, p-value = 0.03; 0.49 vs. 1.07, p-value = 0.04) and TNC (0.20 vs. 1.03, p-value = 0.001; 0.39 vs. 1.06, p-value = 0.01) mRNA levels compared to controls. Our results suggest that DPG inhibits cell viability by activating apoptosis and inhibiting cell proliferation and stem cell subpopulation formation through miR-4443 and miR-3620 upregulation. Both miRNAs are responsible for the post-transcriptional inhibition of NF-kappa B by CD209 and TNC modulation. (AU)

• FAPESP's process: 18/05930-0 - Identification and characterization of microRNAs involved in inhibition of the NF-kB signaling pathway by the action of glycyrrhizinate dipotassium in glioblastoma multiforme cell lines
• Grantee: Gabriel Alves Bonafé
• Support Opportunities: Scholarships in Brazil - Master
• FAPESP's process: 15/03870-1 - Identification of single-nucleotide polymorphisms in microRNAs involved with the glioblastoma susceptibility
• Grantee: Manoela Marques Ortega
• Support Opportunities: Regular Research Grants