Purpose: The purpose of this study was to evaluate the effect of different titanium surface characteristics on the behavior and function of oral fibroblasts as well as the deposition pattern of collagen within the extracellular matrix. Materials and Methods: Titanium surfaces created by machining, acid etching with sulfuric acid (AE1), or acid etching with hydrofluoric acid (AE2) were analyzed using scanning electron microscopy (SEM) and energy-dispersive x-ray spectroscopy. Rat oral fibroblasts were cultured on different surfaces. Cell spread and morphology of extracellular matrix were evaluated using SEM. Attachment and proliferation of cells were examined by comparing the numbers of attached to detached cells and cell count, respectively. Gene expression was analyzed via reverse transcriptase polymerase chain reaction. Collagen production and deposition were examined via a Sirius red–based stain assay and confocal laser scanning microscopy. Results: The machined surface showed a flat profile with isotropic grooves, the AE1 surface showed a uniformly microscale roughened surface, and the AE2 surface had a grooved profile with intermediate surface roughness. The AE2 surface contained fluoride atoms (2.45% ± 0.44% as F/Ti atomic ratio). Cell attachment was significantly weaker on the machined surface than on the AE1 and AE2 surfaces, whereas no differences were observed between the AE1 and AE2 surfaces. The cell counts on the machined and AE2 surfaces were higher, with a parallel orientation, whereas the cell count was lower and randomly distributed on the AE1 surface. The expression level of fibroblastic genes was similar among surfaces for all time points tested. Collagen production was highest on the machined surface, followed by AE2 and AE1 surfaces. Collagen deposition displayed a parallel pattern on the machined surface, while it was multidirectional on the AE1 and AE2 surfaces. Conclusion: The surface characteristics of titanium affect attachment, spread, and proliferative activity of oral fibroblasts as well as the deposition pattern of collagen in vitro. Int J Oral Maxillofac Implants 2009;24:419–431