Biological response of epithelial and connective tissue cells to titanium surfaces with different ranges of roughness: An in-vitro study

This study aimed to investigate how titanium (Ti) surface with different range roughness created by industrial machining influence the biological response of primary human gingival fibroblasts (HGFB) and keratinocytes (HGKC) in terms of cell proliferation and cytotoxicity. Four Ti surfaces of different roughness ranges were investigated: smooth (S: 0.08–0.1 µm), minimally rough (MM: 0.3–0.5 µm), moderately rough (MR: 1.2–1.4 µm) and rough (R: 3.3–3.7 µm). Discs topography and surface roughness were evaluated by scanning electron microscopy (SEM) and non-contact profilometer. Both cell lines were cultured, expanded, and maintained according to their supplier’s protocols. Cell proliferation and cytotoxicity were evaluated at days 1, 3, 5, and 10 using cell viability and cytotoxicity colorimetric assays. Data were analysed via two-way ANOVA, one-way ANOVA and Tukey’s post hoc test (p = 0.05 for all tests). Both cell lines showed comparable initial proliferation activity of 70–86% for all the investigated roughnesses. HGKC showed better and higher proliferation % with S surface at all time points than all the other investigated surfaces which was significantly higher than MM at day 3 and higher than all the other investigated surfaces at day 5 and 10. On the other hand, HGFB exhibited the best proliferation with both MM and R surfaces with no significant differences from the other two surfaces (S and MR). Different surface roughnesses and exposure times showed significant effect on cell proliferation in both cell lines. Cytotoxicity for both cell lines was generally the highest on day 3, with the following order from highest to lowest: S (19.86%)> R> MR> MM for HGKC and MM (39.48%)> MR> S> R for HGFB. Different exposure times showed a significant effect on cell cytotoxicity in both cell lines and a significant effect of surface roughness in HGFB. All investigated roughness levels were sufficiently biologically compatible with cells representative of the major population of the soft tissue surrounding dental implants. However, the S surface was most cytotoxic to HGKC, while the MM surface was most cytotoxic to HGFB cells.