IADR Abstract Archives
MMP20 Silencing Inhibits Key Tumorigenic Hallmarks of Oral Cancer
Objectives: We determined recently that MMP20 whose expression hitherto was thought to be tooth-specific is expressed in human oral squamous cell carcinoma (OSCC) with evidence that it also directly interacts with dentin sialophosphoprotein (DSPP), a member of the small integrin-binding ligand N-linked glycoproteins (SIBLINGs). MMP20 therefore represents the cognate MMP partner for DSPP. Our earlier studies also showed that DSPP is highly upregulated in OSCC where upregulation is associated with tumor aggressiveness. As preliminary step towards determining the function/mechanisms of MMP20-DSPP interaction in OSCC, the current study investigated the effects of MMP20-silencing on the tumorigenic profiles of the oral cancer cell line, OSC2.
Methods: Multiple regions of MMP20 transcript were targeted for shRNA interference using hMMP20-shRNA lentiviral particles designed to silence MMP20 gene. Control shRNA plasmid encoding a scrambled sequence incapable of degrading any known cellular mRNA was used for negative control. Following puromycin selection of stable lines of MMP20-silenced OSC2 cells, phenotypic hallmarks of oral carcinogenesis were assayed by western blot, RT-PCR, MTT (cell-viability), colony-formation, modified Boyden-Chamber (migration and invasion), and flow cytometry (cell-cycle and apoptosis) analyses.
Results: MMP20-silenced OSC2 cells showed altered cell morphology, reduced viability, decreased colony-formation ability, decreased migration and invasion, G0/G1 cell-cycle arrest, and increased tumor cell sensitivity to cisplatin-induced apoptosis. Furthermore, MMP-2, MMP-3, MMP-9, VEGF, Ki-67, EGFR, and DSPP were down-regulated. There was a direct correlation between the degree of MMP20-silencing and DSPP suppression, as indicated by least squares regression analysis.
Conclusions: MMP20-silencing in OSC2 cell decreased salient hallmarks of oral tumorigenesis and provides the first functional evidence of a potential key role for MMP20 in oral cancer biology. The down-regulation of MMP2, MMP3, MMP9, EGFR, and VEGF in DSPP-silenced OSC2 cells provides a significant functional/molecular framework for deciphering the mechanisms of MMP20/DSPP activities in oral cancer.
Methods: Multiple regions of MMP20 transcript were targeted for shRNA interference using hMMP20-shRNA lentiviral particles designed to silence MMP20 gene. Control shRNA plasmid encoding a scrambled sequence incapable of degrading any known cellular mRNA was used for negative control. Following puromycin selection of stable lines of MMP20-silenced OSC2 cells, phenotypic hallmarks of oral carcinogenesis were assayed by western blot, RT-PCR, MTT (cell-viability), colony-formation, modified Boyden-Chamber (migration and invasion), and flow cytometry (cell-cycle and apoptosis) analyses.
Results: MMP20-silenced OSC2 cells showed altered cell morphology, reduced viability, decreased colony-formation ability, decreased migration and invasion, G0/G1 cell-cycle arrest, and increased tumor cell sensitivity to cisplatin-induced apoptosis. Furthermore, MMP-2, MMP-3, MMP-9, VEGF, Ki-67, EGFR, and DSPP were down-regulated. There was a direct correlation between the degree of MMP20-silencing and DSPP suppression, as indicated by least squares regression analysis.
Conclusions: MMP20-silencing in OSC2 cell decreased salient hallmarks of oral tumorigenesis and provides the first functional evidence of a potential key role for MMP20 in oral cancer biology. The down-regulation of MMP2, MMP3, MMP9, EGFR, and VEGF in DSPP-silenced OSC2 cells provides a significant functional/molecular framework for deciphering the mechanisms of MMP20/DSPP activities in oral cancer.