Modulation of the cross-talk between macrophages and osteoblasts by titanium-based particles Gema Valle ´s a , Enrique Gil-Garay b,c , Luis Munuera c,d , Nuria Vilaboa a,c, * a Unidad de Investigacio ´n, Hospital Universitario La Paz, Paseo de la Castellana 261, 28046 Madrid, Spain b Departamento de Traumatologı ´a y Cirugı ´a Ortope ´dica, Hospital Universitario La Paz, Paseo de la Castellana 261, 28046 Madrid, Spain c Centro de Investigacio ´n Biome ´dica en Red de Bioingenierı ´a, Biomateriales y Nanomedicina, CIBER-BBN, Madrid, Spain d Departmento de Cirugı ´a, Universidad Auto ´noma de Madrid, Arzobispo Morcillo 2, 28029 Madrid, Spain Received 24 November 2007; accepted 13 February 2008 Available online 3 March 2008 Abstract Titanium (Ti) and its alloys have widespread uses as implant materials for orthopaedic and dental applications. To improve their surface characteristics, modifications that give rise to an outer ceramic layer of rutile have been developed. It is expected that after a long period of service, rutile particles will arise from these modified surfaces. Rutile particles have recently been proposed as reinforcement agents of substrates designed for bone tissue engineering applications. In this study, the ability of Ti and rutile particles to modulate secretion of soluble factors involved in bone turnover has been assayed in an in vitro co-culture system of macrophages and human osteoblasts that allows the exchange of soluble factors between both cell types without direct cell contact. Exposure of co-cultured macrophages to sub-cytotoxic doses of Ti or rutile particles did not modify the osteoblastic expression of surface RANKL or the secretion of OPG into the media. Both IL-6 and PGE 2 levels increased to a similar extent after treatment with rutile or Ti particles. M-CSF and GM-CSF levels were lower after treatment with rutile particles than with Ti. Experiments employing neutralising antibodies indicate that exposure of co-cultured macrophages to both Ti-based particles induces the release of M-CSF, GM-CSF, IL-6 and PGE 2 through up-regulation of IL-1b and TNF-a. We comparatively examined the response of co-cultured macrophages, osteoblasts or both types of cells after exposure to particles. The results indicate that interactions of osteoblasts with particles can modulate the extent of the response initiated by macrophages. Maximal levels of secretions of all tested factors were reached after exposure of co-cultured cells to Ti particles, which is suggestive of the lower bioreactivity of rutile particles. Ó 2008 Elsevier Ltd. All rights reserved. Keywords: Macrophage; Osteoblast; Cytokine; Titanium; Titanium oxide; Osteolysis 1. Introduction Total joint arthroplasty (TJA) is the most frequent and successful procedure in orthopaedic surgery. Periprosthetic osteolysis, a common long-term complication of TJA, is induced by a combination of events that includes the genera- tion of wear particles, corrosion products and reactive oxygen species that ultimately induce an inflammatory reaction. Over time, it progresses to aseptic loosening and implant failure [1]. Upon stimulation by particles, macrophages and various other types of cells, including osteoblasts, release cytokines, chemo- kines and other soluble factors into the periprosthetic milieu [2]. These cellular mediators act in concert in paracrine and autocrine fashions, inducing resorption directly by enhancing proliferation and/or activity of cells along the osteoclast line- age as well as indirectly by acting on stromal or osteoblastic cells which ultimately results in loss of bone stock. Media from monocytes incubated with metallic particles induce the differentiation of osteoclast precursors co-cultured with mesenchymal support cells [3], and the resorptive activity of culture media from macrophages exposed to diverse types of particles has been well documented [4,5]. Conditioned media * Corresponding author. Edificio IþD, Hospital La Paz, Paseo de La Castel- lana 261, 28046 Madrid, Spain. Tel./fax: þ34 912071512. E-mail address: nvilaboa.hulp@salud.madrid.org (N. Vilaboa). 0142-9612/$ - see front matter Ó 2008 Elsevier Ltd. All rights reserved. doi:10.1016/j.biomaterials.2008.02.011 Available online at www.sciencedirect.com Biomaterials 29 (2008) 2326e2335 www.elsevier.com/locate/biomaterials