Influence of microgroove dimension on cell behavior of human gingival fibroblasts cultured on titanium substrata

Abstract: Objective: The purpose of this study was to determine the dimension of surface microgrooves on titanium (Ti) substrata that shows the greatest positive influence on characterizing specific cell behavior of cultured human gingival fibroblasts. Material and methods: Commercially pure Ti disks with surface microgrooves of monotonous 3.5 μm in depth and respective 15, 30, and 60 μm in width were fabricated using photolithography and used as the culture substrata in the three experimental groups in this study (TiD15, TiD30, and TiD60 groups), whereas the smooth Ti disk was used as the control substrata (smooth Ti group). Human gingival fibroblasts were cultured on the four groups of Ti substrata on successive timelines. Cell behaviors, such as adhesion, morphology, viability and proliferation, and gene expression were analyzed and compared between all groups using crystal violet stain, scanning electron microscopy (SEM), XTT assay, and reverse transcriptase-polymerase chain reaction, respectively. Results: SEM demonstrated that cells were able to readily descend into the microgrooves of TiD30 at the early phase of culture. Cells on the ridge edges or in groove corners were spindle shaped with abundant filopodia formation toward the acid-etched surface inside the microgrooves, thus mimicked the cell shape in three-dimensional (3D) nanoenvironment. TiD15 significantly increased the cell viability and proliferation compared with the smooth Ti substrata after 72 h of culture. Up-regulation of fibronectin (FN) and α5 integrin genes was noted in cells cultured on TiD15 and TiD30. Gene expression pattern specific to the cells in 3D-matrix culture, such as down-regulation of α-smooth muscle actin gene along with up-regulation of FN and p21 genes, was pronounced in cells cultured on TiD30. Conclusion: This study indicates that surface microgrooves of both 15 and 30 μm in width and a monotonous 3.5 μm in depth on Ti substrata increase various cell behaviors of cultured human gingival fibroblasts.