Osteopontin expression in co-cultures of human squamous cell carcinoma-derived cells and osteoblastic cells and its effects on the neoplastic cell phenotype and osteoclastic activation
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This study evaluated the temporal expression of osteopontin (OPN) in co-cultures of human osteoblastic cells (SAOS-2) and oral squamous cell carcinoma (OSCC)-derived cells (SCC9) and examined the effects of osteoblast-derived OPN on the neoplastic cell phenotype. Additionally, the effects of these co-cultures on subsequent osteoclastic activity were explored. SCC9 cells were plated on Transwell® membranes that were either coated or not coated with Matrigel and were then co-cultured with SAOS-2 cells during the peak of OPN expression. SCC9 cells exposed to OPN-silenced SAOS-2 cultures and SCC9 cells cultured alone served as controls. SCC9 cells were quantitatively evaluated for cell adhesion, proliferation, migration, and invasion into Matrigel. The impact of co-culturing SAOS-2 and SCC9 cells on the resorptive capacity of U-937-derived osteoclastic cells was also investigated. Furthermore, a reciprocal induction of SAOS-2 and SCC9 cells in terms of OPN expression over the co-culture interval was identified. SAOS-2-secreted OPN altered the SCC9 cell phenotype, leading to enhanced cell adhesion and proliferation and higher Matrigel invasion. This invasion was also enhanced, albeit to a lesser degree, by co-culture with OPN-silenced SAOS-2 cells. Cell migration was not affected. Co-culture with SAOS-2 cells—mainly during the period of peak OPN expression—promoted over-expression of IL-6 and IL-8 by SCC9 cells and enhanced the resorptive capacity of osteoclastic cells. Taken together, these results suggest that osteoblast-derived OPN affects the interactions among OSCC-derived epithelial cells, osteoblasts, and osteoclasts, which could contribute to the process of bone destruction during bone invasion by OSCC.
KeywordsOsteoblasts Osteopontin Oral squamous cell carcinoma Co-culture Bone invasion
The authors would like to thank the State of Sao Paulo Research Foundation (FAPESP, grants #2012/07531-9, #2012/20863-0, and #2012/08605-6) and the National Council of Scientific and Technological Development (CNPq, grant #308200/2012-8) for financial support. Roger Rodrigo Fernandes and Fabíola Singaretti de Oliveira are acknowledged for technical assistance. The mouse monoclonal anti-rat osteopontin (MPIIIB10-1) antibody, developed by Michael Solursh and Ahnders Franzen, was obtained from the Developmental Studies Hybridoma Bank, created by the NICHD of the NIH and maintained at The University of Iowa, Department of Biology, Iowa City, IA 52242.
Compliance with ethical standards
The State of Sao Paulo Research Foundation (FAPESP, Brazil), grants #2012/07531-9, #2012/20863-0, and #2012/08605-6, and the National Council of Scientific and Technological Development (CNPq, Brazil), grant #308200/2012-8.
Conflicts of interest
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