12‐O‐tetradecanoylphorbol‐13‐acetate‐induced invasion/migration of glioblastoma cells through activating PKCα/ERK/NF‐κB‐dependent MMP‐9 expression

Abstract

An increase in MMP-9 gene expression and enzyme activity with stimulating the migration of GBM8401 glioma cells via wound healing assay by 12-O-tetradecanoylphorbol-13-acetate (TPA) was detected in glioblastoma cells GBM8401. TPA-induced translocation of protein kinase C (PKC)alpha from the cytosol to membranes, and migration of GBM8401 elicited by TPA was suppressed by adding the PKCalpha inhibitors, GF109203X and H7. Activation of extracellular signal-regulated kinase (ERK) and c-Jun-N-terminal kinase (JNK) by TPA was identified, and TPA-induced migration and MMP-9 activity was significantly blocked by ERK inhibitor PD98059 and U0126, but not JNK inhibitor SP600125. Activation of NF-kappaB protein p65 nuclear translocation and IkappaBalpha protein phosphorylation with increased NF-kappaB-directed luciferase activity by TPA were observed, and these were blocked by the PD98059 and IkB inhibitor BAY117082 accompanied by reducing migration and MMP-9 activity induced by TPA in GBM8401 cells. Transfection of GBM8401 cells with PKCalpha siRNA specifically reduced PKCalpha protein expression with blocking TPA-induced MMP-9 activation and migration. Additionally, suppression of TPA-induced PKCalpha/ERK/NK-kappaB activation, migration, and MMP-9 activation by flavonoids including kaempferol (Kae; 3,5,7,4'-tetrahydroxyflavone), luteolin (Lut; 5,7,3'4'-tetrahydroxyflavone), and wogonin (Wog; 5,7-dihydroxy-8-methoxyflavone) was demonstrated, and structure-activity relationship (SAR) studies showed that hydroxyl (OH) groups at C4' and C8 are critical for flavonoids' action against MMP-9 enzyme activation and migration/invasion of glioblastoma cells elicited by TPA. Application of flavonoids to prevent the migration/invasion of glioblastoma cells through blocking PKCalpha/ERK/NF-kappaB activation is first demonstrated herein.

Silibinin inhibits TPA-induced cell migration and MMP-9 expression in thyroid and breast cancer cells

“…TPA was found to significantly enhance cell migration abilities of human cancer cells including hepatoma and breast cancer cells (18,19). In addition, TPA-induced migration and invasion of glioblastoma cells were prevented by blocking PKCα-dependent pathways (20). Consistent with these reports, we found that TPA increased cell migration in thyroid and breast cancer cells.…”

Galangin and Kaempferol Suppress Phorbol-12-Myristate-13-Acetate-Induced Matrix Metalloproteinase-9 Expression in Human Fibrosarcoma HT-1080 Cells

“…Although suppression of MMP-9 expression by kaempferol, quercetin, and morin was reported in different conditions Lee et al, 2013;Lin et al, 2010;Sivaramakrishnan and Niranjali Devaraj, 2009;Yi et al, 2012), our study showed that MMP-9 secretion from HT-1080 cells was severely decreased by galangin as well as kaempferol, and moderately decreased by fisetin. Interestingly, quercetin weakly affected and morin did not affect MMP-9 secretion.…”

“…In addition MMP inhibition, it has recently been shown that flavonols can regulate MMP function through transcriptional suppression. For example, kaempferol suppresses PMA-induced MMP-9 expression in glioblastoma GBM8401 cells (Lin et al, 2010), MMP-2 expression in tongue squamous cell carcinoma SCC4 cells , and MMP-2 and MMP-9 expression in human osteosarcoma U-2 OS cells . Quercetin downregulates oxidized LDL-induced expression of MMP-1, MMP-2, and MMP-9 in human umbilical vein endothelial EA.hy926 cells (Yi et Cells were starved for 24 h, preincubated for 30 min with 30 M of galangin, kaempferol, or gossypetin (as a negative control), and stimulated for 30 min with 30 ng/ml PMA.…”

Growth factors and kinases in glioblastoma growth

“…Once triggered, these proteins can activate and interact with other cascades such as MAPK and PI3K, and regulate processes such as proliferation, differentiation, apoptosis, and angiogenesis [110,111] . These events make PKC an important target in cancer, particularly in glioma where its importance has been thoroughly discussed [112][113][114] as PKC shows increased expression and activity in this kind of tumors [115] . There are reports indicating that PKC is important for progesterone-receptor-mediated proliferation in glioma cells, adding to the possible mechanisms of PKC function [114] .…”

“…The importance of PKC in therapy resistance has been highlighted, particularly PKC's implication in cell cycle regulation [116] . PKCα activates MAPK and MMP, thus increasing infiltration ability [112] ; PKCδ has been linked to radiation-induced apoptosis as well as MMP activation [119,120] ; and PKCε induces proliferation and apoptosis evasion [121] . Moreover, our group has reported that PKCα and PKCδ, through progesterone receptor phosphorylation, induce proliferation and migration [114] .…”