Implant dentistry has for years focused on studying the interface between the bone and implant, searching for a suitable seal to prevent the advance of perimplant disease. Long-term prognosis of dental implants, however, depends not only on osseointegration, but also on the quality of the seal between the mucosa and the implant abutment .
The seal provided by the contact between mucosal tissue and the prosthetic abutment seems to be the safest route to implant longevity and long-term tissue stability. Currently, prosthetic abutments have their transmucosal collars manufactured with machined and polished surfaces. In the search for a titanium surface that favors cellular events, this study aimed to analyze if titanium surfaces treated with acid solution for different lengths of time could alter the behavior of fibroblasts, seeking a more stable union and promoting a better seal for perimplant health than otherwise polished surfaces.
The results of the present study showed that, in general, different acid treatment times did not increase the cellular events evaluated, namely cell proliferation, viability, and type I collagen secretion.
Some studies have reported that gingival fibroblasts seeded onto treated surfaces show greater proliferation and cellular viability when compared to machined surfaces [8, 9], indicating an advantage to treating the prosthetic abutment or the cervical portion of the implant to enhance a biological seal. It is important to highlight from the literature that cell type and cell cycle phase as well as surface type and chemical composition of the material may differ, which may in turn generate different results.
In spite of the numerous current options for prosthetic abutment materials given the high esthetic demand, titanium was used in the present study, which is a widely cited material in the literature as the gold standard when used as a prosthetic abutment, due to its high biocompatibility [14, 15]. Thus, the effect of applying an acid treatment protocol, which is widely used for dental implant surface treatment, on the proliferation, synthesis, and secretion of collagen by gingival fibroblasts was studied. The results obtained from the acid treatments allowed for smooth and minimally rough surfaces, according to the criteria adopted by Albrektsson and Wennergerg , respectively, for the acid treatments after 20 min and 60 min. This indicates that, despite acid treatment, 20 min was not sufficient to create surface roughness on the titanium surface with the acid solutions used.
The results showed that, regardless of the treatment used, there was no difference in the values of cell proliferation in the different times of analysis. Similarly, cell viability values did not differ, especially after 24 h and 72 h. Corroborating the results of this study, Baltriukiene et al.  used grade 2 titanium disks with different surface treatments and demonstrated that the modified titanium surface did not affect human gingival fibroblast viability. Interestingly, some studies show that smooth surfaces increase the proliferation and adhesion of fibroblasts [18, 19], which raises the question of whether the recommended acid treatment of titanium surfaces indeed has an advantage in terms of promoting greater biological sealing. In addition, increased surface roughness promotes bacterial colonization and increased retention [20, 21], contributing to increased biofilm formation.
In the present study, human gingival fibroblasts were used in a similar strategy as per other reports in the literature [10, 15, 22,23,24,25]. In spite of in vivo studies representing the ideal environment to investigate tissue responses, in vitro studies facilitate the investigation of a specific and isolated factor of the tissue response . In addition to being widely used in cell biocompatibility studies, considering the various topographic surface features, fibroblasts are responsible for the biosynthesis of collagen and other extracellular matrix molecules.
To establish the amount of protein secreted by fibroblasts, we chose type I collagen, as it is the most abundant protein in the extracellular matrix of gingival connective tissue, representing almost 90% of the total organic matrix in mature bone. The results revealed differences between groups only at 48 h of culture, where the lowest titers were observed when surfaces were treated with acid solutions. At 24 h and 72 h, no significant difference was observed regarding collagen secretion, though the highest quantification occurred at 72 h for the surfaces treated for 60 min. These results are in agreement with the findings by Ramaglia et al.  and Velasco-Ortega et al. , in which they suggested that roughened surfaces may improve the biological behavior of fibroblasts as well as the process of perimplant soft tissue healing and osseointegration compared to machined surfaces.
The connective tissue seal around the abutment is crucial for perimplant health. Several efforts have previously been made to optimize abutment surfaces, though without consensus on the ideal surface features. Blasquez et al.  have shown in their systematic review that different types of surface modifications for implant abutments may provide benefit to the surrounding connective tissue, which generally corroborate the results reported herein. It is noteworthy that different methodologies and cell types, together with different types of acid solutions, might lead to different outcomes compared to the present study.
Cao et al.  with the aim of investigating the effects of different decontamination treatments on the microstructure of titanium surfaces as well as the proliferation and adhesion of human gingival fibroblasts showed that proliferation and adhesive strength were higher in the machined surfaces than on treated surfaces. These results led the group to conclude that proliferation and adhesion increases as surface roughness decreases.