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GIUSEPPE CALVARUSO

WIN modulates osteosarcoma MG63 cell migration by inhibiting MMPs activity and adjusting intra- and extra-cellular SPARC differential expression

  • Autori: Notaro, A.; Sabella, S.; DI FIORE, R.; Calvaruso, G.; Giuliano, M.
  • Anno di pubblicazione: 2014
  • Tipologia: Proceedings (TIPOLOGIA NON ATTIVA)
  • OA Link: http://hdl.handle.net/10447/96218

Abstract

Invasion of cancer cells into surrounding tissue is an initial step in tumor metastasis. This event, which requires migration of cancer cells and attachment to extracellular matrix (ECM), is regulated by elements of the local microenvironment, including ECM architecture. After having demonstrated the ability of the synthetic cannabinoid WIN55,512 to induce osteosarcoma MG63 cell death (1), we studied the effects of WIN on MG63 cell migration. Wound healing assay was performed to measure the ability of cells to migrate and fill the gap obtained by physical disruption of cell monolayer (2). We observed a significant delay in wound closure in 5 M WIN treated cells compared to untreated cells that almost completed the healing in 24 hours (20% vs 87% of wound closure). The addition of conditioned medium obtained by confluent control cells to WIN treated cultures largely reverted the delaying WIN action. To evaluate the influence of matrix metalloproteinases (MMPs) in the migratory ability, we analyzed MMP-2 and MMP-9 activities by zymography in WIN-treated culture medium. MMP-9 zymographic activity was strongly lower in WIN-treated culture medium in comparison with medium from control cells, suggesting that WIN inhibits MMP-9 activity. Since it is known that cell/ECM interactions are mediated by SPARC (Secreted Protein Acidic and Rich in Cysteine) that also modulates MMPs activity (3-4), we evaluated intra- and extracellular levels of SPARC in our experimental conditions. RT-PCR and western blotting analysis showed in WIN-treated cells an increase both in mRNA and protein expression of intracellular SPARC, while a decrease in extracellular protein level was observed. Studies are in progress to study the possible involvement of SPARC in MMPs activation and MG63 cell migration.