ARTICLE IN PRESS 1.0 0.9 2.0 15 80.1 0.0 6 6 Days d 5.0 4.0 2.5 .0 6 15 10 321 9是5E 09876543 510 目 broblasts (a, c e)an d centrifuged in order to eliminate residual particles Supernatant were used for human type I expressed as mean+SEM, n=9 for culture in the presence of particles and n= 12 for controls: *p< 0.05. *p<0.01(compared to control cultures). At higher magnification(high power field: 40x ) we clearly order to avoid confusion between the visualization of particles and observed the presence of particles in the synovial membrane for all blood deposition, the presence of particles was confirmed with two animals, which were injected with alumina or zirconia(Fig. 7). In different colorations (Goldner trichrome and toluidin blue). As lease cite this article in press as: Roualdes O, et al, In vitro and in vivo evaluation of an., Biomaterials(2009), doi: 10.1016/ j biomaterials 2009. 11.107At higher magnification (high power field: 40), we clearly observed the presence of particles in the synovial membrane for all animals, which were injected with alumina or zirconia (Fig. 7). In order to avoid confusion between the visualization of particles and blood deposition, the presence of particles was confirmed with two different colorations (Goldner trichrome and toluidin blue). As 3 6 10 15 0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 a Days 3 6 10 15 0.0 0.5 1.0 1.5 2.0 2.5 3.0 3.5 4.0 b Days 3 6 10 15 0.0 0.5 1.0 1.5 2.0 2.5 3.0 3.5 4.0 + c Days 3 6 10 15 0.0 0.5 1.0 1.5 2.0 2.5 3.0 3.5 4.0 4.5 d 5.0 Days Days 3 6 10 15 0 5 10 15 20 25 30 35 ++ e Days 3 6 10 15 0 1 2 3 4 5 6 7 8 9 10 11 12 13 f Cell proliferation (optical density) Cell proliferation (optical density) Human type I collagen (µg/mL) Human type I collagen (µg/mL) Human fibronectin (µg/mL) Human fibronectin (µg/mL) Fig. 4. Fibroblasts (a, c, e) and osteoblasts (b, d, f) were cultured for 3, 6, 10 and 15 days in the presence of particles (100 mg/mL) of alumina ( ), zirconia ( ) or without particles ( ) under standard cell culture conditions in 24-well culture plates. At each time of culture, cell proliferation was assessed using the MTT assay (a, b) and culture media were harvested and centrifuged in order to eliminate residual particles. Supernatant were used for human type I collagen (c, d) and human fibronectin (e, f) synthesis quantification. Values are expressed as mean  SEM, n ¼ 9 for culture in the presence of particles and n ¼ 12 for controls; þp < 0.05, þþp < 0.01 (compared to control cultures). 6 O. Roualdes et al. / Biomaterials xxx (2009) 1–12 ARTICLE IN PRESS Please cite this article in press as: Roualdes O, et al., In vitro and in vivo evaluation of an..., Biomaterials (2009), doi:10.1016/ j.biomaterials.2009.11.107