Src and ROCK Kinases Differentially Regulate Mineralization of Human Osteosarcoma Saos-2 Cells

Agnieszka Strzelecka-Kiliszek, Marta Romiszewska, Lukasz Bozycki, Saida Mebarek, Joanna Bandorowicz-Pikula, Rene Buchet, Slawomir Pikula
2019 International Journal of Molecular Sciences  
Osteoblasts initiate bone mineralization by releasing matrix vesicles (MVs) into the extracellular matrix (ECM). MVs promote the nucleation process of apatite formation from Ca2+ and Pi in their lumen and bud from the microvilli of osteoblasts during bone development. Tissue non-specific alkaline phosphatase (TNAP) as well as annexins (among them, AnxA6) are abundant proteins in MVs that are engaged in mineralization. In addition, sarcoma proto-oncogene tyrosine-protein (Src) kinase and
more » ... ciated coiled-coil (ROCK) kinases, which are involved in vesicular transport, may also regulate the mineralization process. Upon stimulation in osteogenic medium containing 50 μg/mL of ascorbic acid (AA) and 7.5 mM of β-glycerophosphate (β-GP), human osteosarcoma Saos-2 cells initiated mineralization, as evidenced by Alizarin Red-S (AR-S) staining, TNAP activity, and the partial translocation of AnxA6 from cytoplasm to the plasma membrane. The addition of 4-amino-5-(4-chlorophenyl)-7-(t-butyl)pyrazolo [3,4-d] pyrimidine (PP2), which is an inhibitor of Src kinase, significantly inhibited the mineralization process when evaluated by the above criteria. In contrast, the addition of (R)-(+)-trans-4-(1-aminoethyl)-N-(4-pyridyl) cyclohexane carboxamide hydrochloride (Y-27632), which is an inhibitor of ROCK kinase, did not affect significantly the mineralization induced in stimulated Saos-2 cells as denoted by AR-S and TNAP activity. In conclusion, mineralization by human osteosarcoma Saos-2 cells seems to be differently regulated by Src and ROCK kinases.
doi:10.3390/ijms20122872 pmid:31212828 pmcid:PMC6628028 fatcat:mfmnz64i45h57fiyhcvzw2xtou