Long term oncostatin M treatment induces an osteocyte-like differentiation on osteosarcoma and calvaria cells Bénédicte Brounais a,b , Emmanuelle David a,b , Céline Chipoy a,b, 1 , Valérie Trichet a,b , Virginie Ferré c,d , Céline Charrier a,b , Laurence Duplomb a,b,c , Martine Berreur a,b , Françoise Rédini a,b , Dominique Heymann a,b,c , Frédéric Blanchard a,b, ⁎ a INSERM, ERI 7, Nantes, F-44035, France b Université de Nantes, Nantes Atlantique Universités, Laboratoire de Physiopathologie de la Résorption Osseuse et Thérapie des Tumeurs Osseuses Primitives, EA3822, Nantes, F-44035, France c CHU, Hôtel Dieu, Nantes, France d Université de Nantes, Nantes Atlantique Universités, Laboratoire d'Immunovirologie et Polymorphisme Génétique, EA4271, Nantes, F-44035, France abstract article info Article history: Received 24 July 2008 Revised 15 December 2008 Accepted 18 December 2008 Available online 3 January 2009 Edited by: J. Aubin Keywords: Oncostatin M Lentivirus Osteosarcoma Osteoblast Osteocyte Previous in vitro studies on primary osteoblastic and osteosarcoma cells (normal and transformed osteoblasts) have shown that oncostatin M (OSM), a member of the interleukin-6 family, possesses cytostatic and pro-apoptotic effects in association with complex and poorly understood activities on osteoblast differentiation. In this study, we use rat osteosarcoma cells transduced with lentiviral particles encoding OSM (lvOSM) to stably produce this cytokine. We show that after several weeks of culture, transduced OSRGA and ROS 17/2.8 cells are growth inhibited and sensitized to apoptosis induced by the kinase inhibitor Staurosporine (Sts). Moreover, this long term OSM treatment induces (i) a decrease in osteoblastic markers, (ii) morphological changes leading to an elongated and/or stellate shape and (iii) an increase in osteocytic markers (sclerostin and/or E11), suggesting an osteocyte-like differentiation. We also show that non transformed rat calvaria cells transduced with lvOSM differentiate into stellate shaped cells expressing sclerostin, E11, Phex and functional hemichannels. Together, these results indicate that osteosarcoma cells stably producing OSM do not develop resistance to this cytokine and thus could be a valuable new tool to study the anti-cancer effect of OSM in vivo. Moreover, OSM-over-expressing osteoblastic cells differentiate into osteocyte-like cells, the major cellular contingent in bone, providing new culture conditions for this cell type which is difficult to obtain in vitro. © 2009 Elsevier Inc. All rights reserved. Introduction Oncostatin M (OSM) is a multifunctional cytokine belonging to the Interleukin (IL)-6 family [1] which also includes IL-11, IL-27, IL-31, leukemia inhibitory factor (LIF), ciliary neurotrophic factor (CNTF), cardiotrophin-1 (CT-1), cardiotrophin-like cytokine (CLC) and neuro- poietin (NP). OSM binds to a specific receptor complex composed of gp130 and OSMRβ and activates two main signaling pathways: the Janus Kinase (JAK)/Signal Transducer and Activator of Transcription 3 (STAT3) and the Mitogen-Activated Protein Kinase (MAPK) signaling pathways [2,3]. In comparison to other IL-6-type cytokines, OSM is more active to induce renal inflammatory response [4], differentiation of fetal hepatocytes [5], chronic joint inflammation and destruction [6] and growth inhibition of various solid tumor cell lines derived from melanoma [7], breast and lung cancer [8], hepatoma [9], glioblastoma [10] or osteosarcoma [11]. Osteosarcomas are rare primitive bone tumors derived from osteoblasts. Patient survival largely depends on the response of tumor cells to chemotherapy, reaching 60–70% in the best responsive patient groups and only 30% when pulmonary metastases are detected at the time of diagnosis. Thus, a better understanding of osteosarcoma and osteoblast biology is necessary in order to improve this treatment. On osteosarcoma cell lines and normal proliferating osteoblasts, we previously showed that OSM inhibits the proliferation and sensitizes to apoptosis driven by various death inducers such as the kinase inhibitor Staurosporine [12,13]. Moreover, in a rat osteosarcoma model, OSM in combination with Midostaurin, a derivate of Staurosporine, reduced the progression of the primary bone tumor, in association with an increased tumor necrosis and tissue repair [14]. The inhibitory effect of OSM on cancer cell proliferation is tightly coupled to induced differentiation. OSM, through activation of the Bone 44 (2009) 830–839 ⁎ Corresponding author. Laboratoire de Physiopathologie de la Résorption Osseuse et Thérapie des Tumeurs Osseuses Primitives, Faculté de Médecine, 1 rue Gaston Veil, F-44035 Nantes cedex 1, France. E-mail address: frederic.blanchard@univ-nantes.fr (F. Blanchard). 1 Present address: Molecular Oncology Group, McGill University, Montreal, Canada. 8756-3282/$ – see front matter © 2009 Elsevier Inc. All rights reserved. doi:10.1016/j.bone.2008.12.021 Contents lists available at ScienceDirect Bone journal homepage: www.elsevier.com/locate/bone