J Adv Prosthodont.  2015 Apr;7(2):138-145. 10.4047/jap.2015.7.2.138.

Effect of laser-dimpled titanium surfaces on attachment of epithelial-like cells and fibroblasts

Affiliations
  • 1Department of Periodontology, Veterans Health Service Medical Center, Seoul, Republic of Korea.
  • 2Department of Dentistry, Graduate School, Korea University, Seoul, Republic of Korea.
  • 3Nano-Convergence Mechanical System Research Division, Korea Institute of Machinery and Materials, Daejeon, Republic of Korea.
  • 4Department of Biochemistry and Molecular Biology, College of Medicine, Korea University, Seoul, Republic of Korea.
  • 5Division of Periodontology, Ostrow School of Dentistry, University of Southern California, Los Angeles, California, USA.
  • 6Division of Biomedical Sciences, Ostrow School of Dentistry, University of Southern California, Los Angeles, California, USA.
  • 7Department of Periodontology, College of Dentistry, Yonsei University, Seoul, Republic of Korea.
  • 8Department of Prosthodontics, College of Medicine, Korea University, Seoul, Republic of Korea. koprosth@unitel.co.kr

Abstract

PURPOSE
The objective of this study was to conduct an in vitro comparative evaluation of polished and laserdimpled titanium (Ti) surfaces to determine whether either surface has an advantage in promoting the attachment of epithelial-like cells and fibroblast to Ti.
MATERIALS AND METHODS
Forty-eight coin-shaped samples of commercially pure, grade 4 Ti plates were used in this study. These discs were cleaned to a surface roughness (Ra: roughness centerline average) of 180 nm by polishing and were divided into three groups: SM (n=16) had no dimples and served as the control, SM15 (n=16) had 5-microm dimples at 10-microm intervals, and SM30 (n=16) had 5-microm dimples at 25-microm intervals in a 2 x 4 mm2 area at the center of the disc. Human gingival squamous cell carcinoma cells (YD-38) and human lung fibroblasts (MRC-5) were cultured and used in cell proliferation assays, adhesion assays, immunofluorescent staining of adhesion proteins, and morphological analysis by SEM. The data were analyzed statistically to determine the significance of differences.
RESULTS
The adhesion strength of epithelial cells was higher on Ti surfaces with 5-microm laser dimples than on polished Ti surfaces, while the adhesion of fibroblasts was not significantly changed by laser treatment of implant surfaces. However, epithelial cells and fibroblasts around the laser dimples appeared larger and showed increased expression of adhesion proteins.
CONCLUSION
These findings demonstrate that laser dimpling may contribute to improving the periimplant soft tissue barrier. This study provided helpful information for developing the transmucosal surface of the abutment.

Keyword

Laser; Topography; Attachment; Soft tissue; Dental implant; Epithelial cells

MeSH Terms

Carcinoma, Squamous Cell
Cell Proliferation
Dental Implants
Epithelial Cells
Fibroblasts*
Humans
Lung
Titanium*
Dental Implants
Titanium

Figure

  • Fig. 1 High magnification (5000×) scanning electron microscopy (SEM) image of a laser-dimpled titanium surface.

  • Fig. 2 The 5-µm laser-dimpled surface with an area of 2 × 4 mm2 at the center of a polished titanium disc.

  • Fig. 3 Light microscopy images of each group (40× magnification). A, SM15: 5-µm dimple and 15-µm center distance. B, SM30: 5-µm dimple and 30-µm center distance.

  • Fig. 4 Proliferation of epithelial cells (YD-38) cultured on titanium discs for 3 days. SM: Smooth surface, served as a control, SM15: 5-µm dimples and 15-µm center distance, and SM30: 5-µm dimples and 30-µm center distance. The value was assumed 100 in control disc at day 1. All values are expressed as percentage.

  • Fig. 5 Proliferation of fibroblast cells (MRC-5) cultured on titanium discs for 3 days. SM: Smooth Surface, served as a control, SM15: 5-µm dimples and 15-µm center distance, and SM30: 5-µm dimples and 30-µm center distance. The value was assumed 100 in control disc at day 1. All values are expressed as percentage.

  • Fig. 6 (A) Adherence of epithelial cells (YD-38) cultured on titanium discs for 1 and 3 days. No significant difference was observed in the adhesion strength between SM, SM15, and SM30 discs on day 1. SM: Smooth surface, served as a control, SM15: 5-µm dimples and 15-µm center distance, and SM30: 5-µm dimples and 30-µm center distance. (B) Cell adherence of YD-38 cells cultured on titanium discs for 3 days. **P<.01, SM vs. SM15, SM vs. SM30. The value was assumed 100 in control disc at day 1. All values are expressed as percentage.

  • Fig. 7 Adherence of fibroblast cells (MRC-5) cultured on titanium discs for 1 and 3 days. SM: Smooth surface, served as a control, SM15: 5-µm dimples and 15-µm center distance, and SM30: 5-µm dimples and 30-µm center distance. The value was assumed 100 in control disc at day 1. All values are expressed as percentage.

  • Fig. 8 High magnification (250×, 1000×) scanning electron microscopy (SEM) images of epithelial cells (YD-38) cultured on SM, SM15, and SM30 discs for 3 days. The dotted line represents the border between the dimpled and non-dimpled areas. Two circles among the visible dimples textured on the disc were marked, and imply the dimple zone. SM: Smooth surface, served as a control, SM15: 5-µm dimples and 15-µm center distance, and SM30: 5-µm dimples and 30-µm center distance.

  • Fig. 9 Immunofluorescence images (400× magnification) showing actin filaments and integrin-β4 in epithelial cells (YD-38) cultured on SM and SM30 discs for 3 days. White circles indicate the laser dimples. SM: Smooth surface, served as a control and SM30: 5-µm dimples and 30-µm center distance on a polished titanium surface.

  • Fig. 10 Immunofluorescence images (400× magnification) showing actin filaments and vinculin in fibroblast cells (MRC-5) cultured on SM and SM30 discs for 3 days. White circles indicate the laser dimples. SM: Smooth surface, served as a control and SM30: 5-µm dimples and 30-µm center distance.


Reference

1. Abrahamsson I, Berglundh T, Wennström J, Lindhe J. The peri-implant hard and soft tissues at different implant systems. A comparative study in the dog. Clin Oral Implants Res. 1996; 7:212–219.
2. Brånemark PI, Adell R, Albrektsson T, Lekholm U, Lundkvist S, Rockler B. Osseointegrated titanium fixtures in the treatment of edentulousness. Biomaterials. 1983; 4:25–28.
3. Furuhashi A, Ayukawa Y, Atsuta I, Okawachi H, Koyano K. The difference of fibroblast behavior on titanium substrata with different surface characteristics. Odontology. 2012; 100:199–205.
4. Chehroudi B, Gould TR, Brunette DM. The role of connective tissue in inhibiting epithelial downgrowth on titaniumcoated percutaneous implants. J Biomed Mater Res. 1992; 26:493–515.
5. Geurs NC, Vassilopoulos PJ, Reddy MS. Soft tissue considerations in implant site development. Oral Maxillofac Surg Clin North Am. 2010; 22:387–405.
6. Linkevicius T, Apse P. Biologic width around implants. An evidence-based review. Stomatologija. 2008; 10:27–35.
7. Meffert RM. How to treat ailing and failing implants. Implant Dent. 1992; 1:25–33.
8. Berglundh T, Lindhe J. Dimension of the periimplant mucosa. Biological width revisited. J Clin Periodontol. 1996; 23:971–973.
9. Hansson S. The implant neck: smooth or provided with retention elements. A biomechanical approach. Clin Oral Implants Res. 1999; 10:394–405.
10. Ikeda H, Yamaza T, Yoshinari M, Ohsaki Y, Ayukawa Y, Kido MA, Inoue T, Shimono M, Koyano K, Tanaka T. Ultrastructural and immunoelectron microscopic studies of the peri-implant epithelium-implant (Ti-6Al-4V) interface of rat maxilla. J Periodontol. 2000; 71:961–973.
11. Rompen E, Domken O, Degidi M, Pontes AE, Piattelli A. The effect of material characteristics, of surface topography and of implant components and connections on soft tissue integration: a literature review. Clin Oral Implants Res. 2006; 17:55–67.
12. Abrahamsson I, Zitzmann NU, Berglundh T, Linder E, Wennerberg A, Lindhe J. The mucosal attachment to titanium implants with different surface characteristics: an experimental study in dogs. J Clin Periodontol. 2002; 29:448–455.
13. Berry CC, Campbell G, Spadiccino A, Robertson M, Curtis AS. The influence of microscale topography on fibroblast attachment and motility. Biomaterials. 2004; 25:5781–5788.
14. Huh JB, Rheu GB, Kim YS, Jeong CM, Lee JY, Shin SW. Influence of Implant transmucosal design on early peri-implant tissue responses in beagle dogs. Clin Oral Implants Res. 2014; 25:962–968.
15. Xing R, Salou L, Taxt-Lamolle S, Reseland JE, Lyngstadaas SP, Haugen HJ. Surface hydride on titanium by cathodic polarization promotes human gingival fibroblast growth. J Biomed Mater Res A. 2014; 102:1389–1398.
16. Baharloo B, Textor M, Brunette DM. Substratum roughness alters the growth, area, and focal adhesions of epithelial cells, and their proximity to titanium surfaces. J Biomed Mater Res A. 2005; 74:12–22.
17. Könönen M, Hormia M, Kivilahti J, Hautaniemi J, Thesleff I. Effect of surface processing on the attachment, orientation, and proliferation of human gingival fibroblasts on titanium. J Biomed Mater Res. 1992; 26:1325–1341.
18. Nevins M, Kim DM, Jun SH, Guze K, Schupbach P, Nevins ML. Histologic evidence of a connective tissue attachment to laser microgrooved abutments: a canine study. Int J Periodontics Restorative Dent. 2010; 30:245–255.
19. Okawachi H, Ayukawa Y, Atsuta I, Furuhashi A, Sakaguchi M, Yamane K, Koyano K. Effect of titanium surface calcium and magnesium on adhesive activity of epithelial-like cells and fibroblasts. Biointerphases. 2012; 7:27.
20. Goodwin AE, Pauli BU. A new adhesion assay using buoyancy to remove non-adherent cells. J Immunol Methods. 1995; 187:213–219.
21. Eisenbarth E, Velten D, Schenk-Meuser K, Linez P, Biehl V, Duschner H, Breme J, Hildebrand H. Interactions between cells and titanium surfaces. Biomol Eng. 2002; 19:243–249.
22. An N, Rausch-fan X, Wieland M, Matejka M, Andrukhov O, Schedle A. Initial attachment, subsequent cell proliferation/viability and gene expression of epithelial cells related to attachment and wound healing in response to different titanium surfaces. Dent Mater. 2012; 28:1207–1214.
23. Qu Z, Rausch-Fan X, Wieland M, Matejka M, Schedle A. The initial attachment and subsequent behavior regulation of osteoblasts by dental implant surface modification. J Biomed Mater Res A. 2007; 82:658–668.
24. Zhao G, Schwartz Z, Wieland M, Rupp F, Geis-Gerstorfer J, Cochran DL, Boyan BD. High surface energy enhances cell response to titanium substrate microstructure. J Biomed Mater Res A. 2005; 74:49–58.
25. Drake DR, Paul J, Keller JC. Primary bacterial colonization of implant surfaces. Int J Oral Maxillofac Implants. 1999; 14:226–232.
26. Listgarten MA. Soft and hard tissue response to endosseous dental implants. Anat Rec. 1996; 245:410–425.
27. Atsuta I, Yamaza T, Yoshinari M, Goto T, Kido MA, Kagiya T, Mino S, Shimono M, Tanaka T. Ultrastructural localization of laminin-5 (gamma2 chain) in the rat peri-implant oral mucosa around a titanium-dental implant by immuno-electron microscopy. Biomaterials. 2005; 26:6280–6287.
28. Kim S, Myung WC, Lee JS, Cha JK, Jung UW, Yang HC, Lee IS, Choi SH. The effect of fibronectin-coated implant on canine osseointegration. J Periodontal Implant Sci. 2011; 41:242–247.
29. Teng FY, Ko CL, Kuo HN, Hu JJ, Lin JH, Lou CW, Hung CC, Wang YL, Cheng CY, Chen WC. A comparison of epithelial cells, fibroblasts, and osteoblasts in dental implant titanium topographies. Bioinorg Chem Appl. 2012; 2012:687291.
30. Atsuta I, Ayukawa Y, Furuhashi A, Ogino Y, Moriyama Y, Tsukiyama Y, Koyano K. In vivo and in vitro studies of epithelial cell behavior around titanium implants with machined and rough surfaces. Clin Implant Dent Relat Res. 2014; 16:772–781.
31. Lee JH, Ogawa T. The biological aging of titanium implants. Implant Dent. 2012; 21:415–421.
32. Walboomers XF, Croes HJ, Ginsel LA, Jansen JA. Growth behavior of fibroblasts on microgrooved polystyrene. Biomaterials. 1998; 19:1861–1868.
33. Walboomers XF, Croes HJ, Ginsel LA, Jansen JA. Contact guidance of rat fibroblasts on various implant materials. J Biomed Mater Res. 1999; 47:204–212.
34. Goldmann WH, Schindl M, Cardozo TJ, Ezzell RM. Motility of vinculin-deficient F9 embryonic carcinoma cells analyzed by video, laser confocal, and reflection interference contrast microscopy. Exp Cell Res. 1995; 221:311–319.
35. Mierke CT, Kollmannsberger P, Zitterbart DP, Diez G, Koch TM, Marg S, Ziegler WH, Goldmann WH, Fabry B. Vinculin facilitates cell invasion into three-dimensional collagen matrices. J Biol Chem. 2010; 285:13121–13130.
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