J Korean Acad Prosthodont JKAP The Journal of Korean Academy of Prosthodontics 0301-2875 2005-3789 The Korean Academy of Prosthodontics 10.4047/jkap.2017.55.4.361 Original Article Regulation of human gingival fibroblast gene expression on microgrooves: A DNA microarray study https://orcid.org/0000-0002-7546-0525 Lee Kyungho 1 https://orcid.org/0000-0002-8381-723X Leesungbok Richard 2 https://orcid.org/0000-0003-2128-1561 Ahn Su-Jin 2 https://orcid.org/0000-0002-4111-2231 Park Su-Jung 2 https://orcid.org/0000-0003-2726-3567 Lee Suk Won 2 Department of Dentistry, Graduate School, Kyung Hee University, Seoul, Republic of Korea. Department of Biomaterials & Prosthodontics, Kyung Hee University Hospital at Gangdong, School of Dentistry, Kyung Hee University, Seoul, Republic of Korea. Corresponding Author: Suk Won Lee. Department of Biomaterials & Prosthodontics, Kyung Hee University Hospital at Gangdong, Institute of Oral Biology, School of Dentistry, Kyung Hee University, 892 Dongnam-ro, Gangdong-gu, Seoul 05278, Republic of Korea. +82 (0)2 440 7519: ysprosth@hanmail.net 10 2017 26 10 2017 55 4 361 371 08 06 2017 11 09 2017 14 09 2017 © 2017 The Korean Academy of Prosthodontics 2017 The Korean Academy of Prosthodontics This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited. Purpose

We aimed to investigate the gene expression of human gingival fibroblasts on microgroove surface using DNA microarray.

 Materials and methods

Microgrooves were applied on grade II titanium discs to have 0/0 µm (NE0, control group), 60/10 µm (E60/10, experimental group) of respective width/depth by photolithography. The entire surface of the microgrooved Ti substrata was further acid etched and used as the two experimental groups in this study. Human gingival fibroblasts were cultured in the experimental group and the control group, and total RNA was extracted. The oligonucleotide microarray was performed to confirm the changes of various gene expression levels between experimental group and control group. Changes of gene expression level were determined at the pathway level by mapping the expression results of DNA chips, using the KEGG (Kyoto Encyclopedia of Genes and Genomes) pathway analysis.

 Results

Gene expression levels on E60/10 and NE0 were analyzed, there were 123 genes showing significant differences in expression more than 1.5 times on E60/10 microgrooved surface compared to NE0 surface, and 19 genes showing significant differences in expression more than 2 times. The KEGG pathway analysis confirmed the changes in gene expression levels under experimental conditions. Cell signaling, proliferation, and activity among the various gene expression results were identified.

 Conclusion

Microgrooved surfaces induce gene expression changes and related cell signaling. According to the results of this study, microgrooves can be used as the surface of various biomaterials which need to improve cell activity through gene expression changes and activation of cell signaling.

 Titanium Microgrooves Human gingival fibroblasts Gene expression changes National Research Foundation of Korea http://dx.doi.org/10.13039/501100003725 NRF-2015R1D1A1A01057956

This research was supported by the Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Education, Republic of Korea (NRF-2015R1D1A1A01057956).

 Kasemo B Lausmaa J Surface science aspects on inorganic biomaterials CRC critical reviews in biocompatibility Boca Raton FL CRC Press 1986 335 380 Albrektsson T Wennerberg A Oral implant surfaces: Part 1-review focusing on topographic and chemical properties of different surfaces and in vivo responses to them Int J Prosthodont 2004 17 536 543 Morra M Cassinelli C Bruzzone G Carpi A Di Santi G Giardino R Fini M Surface chemistry effects of topographic modification of titanium dental implant surfaces: 1. Surface analysis Int J Oral Maxillofac Implants 2003 18 40 45 Zhao L Mei S Chu PK Zhang Y Wu Z The influence of hierarchical hybrid micro/nano-textured titanium surface with titania nanotubes on osteoblast functions Biomaterials 2010 31 5072 5082 Lee SW Kim SY Rhyu IC Chung WY Leesungbok R Lee KW Influence of microgroove dimension on cell behavior of human gingival fibroblasts cultured on titanium substrata Clin Oral Implants Res 2009 20 56 66 Chou L Firth JD Uitto VJ Brunette DM Substratum surface topography alters cell shape and regulates fibronectin mRNA level, mRNA stability, secretion and assembly in human fibroblasts J Cell Sci 1995 108 1563 1573 Jockusch BM Bubeck P Giehl K Kroemker M Moschner J Rothkegel M Rüdiger M Schlüter K Stanke G Winkler J The molecular architecture of focal adhesions Annu Rev Cell Dev Biol 1995 11 379 416 Pankov R Yamada KM Fibronectin at a glance J Cell Sci 2002 115 3861 3863 Park SJ Leesungbok R Ahn SJ Im BJ Lee DY Jee YJ Yoon JH Cui T Lee SC Lee SW Effect of microgrooves and fibronectin conjugation on the osteoblast marker gene expression and differentiation J Adv Prosthodont 2015 7 496 505 Lai EC Notch signaling: control of cell communication and cell fate Development 2004 131 965 973 Halper J Kjaer M Basic components of connective tissues and extracellular matrix: elastin, fibrillin, fibulins, fibrinogen, fibronectin, laminin, tenascins and thrombospondins Progress in Heritable Soft Connective Tissue Diseases Springer 2014 31 47 Yun-Feng W Matsuo N Sumiyoshi H Yoshioka H Sp7/Osterix up-regulates the mouse pro-alpha3(V) collagen gene (Col5a3) during the osteoblast differentiation Biochem Biophys Res Commun 2010 394 503 508 Kadler KE Hill A Canty-Laird EG Collagen fibrillogenesis: fibronectin, integrins, and minor collagens as organizers and nucleators Curr Opin Cell Biol 2008 20 495 501 Mitra SK Hanson DA Schlaepfer DD Focal adhesion kinase: in command and control of cell motility Nat Rev Mol Cell Biol 2005 6 56 68 Zhang W Liu HT MAPK signal pathways in the regulation of cell proliferation in mammalian cells Cell Res 2002 12 9 18 Yang P Roy SK A novel mechanism of FSH regulation of DNA synthesis in the granulosa cells of hamster preantral follicles: involvement of a protein kinase C-mediated MAP kinase 3/1 self-activation loop Biol Reprod 2006 75 149 157 Balko JM Schwarz LJ Bhola NE Kurupi R Owens P Miller TW Go′mez H Cook RS Arteaga CL Activation of MAPK pathways due to DUSP4 loss promotes cancer stem cell-like phenotypes in basal-like breast cancer Cancer Res 2013 73 6346 6358 Leonard WJ Lin JX Cytokine receptor signaling pathways J Allergy Clin Immunol 2000 105 877 888 Baumann H Morella KK White DW Dembski M Bailon PS Kim H Lai CF Tartaglia LA The full-length leptin receptor has signaling capabilities of interleukin 6-type cytokine receptors Proc Natl Acad Sci USA 1996 93 8374 8378 Peschon JJ Morrissey PJ Grabstein KH Ramsdell FJ Maraskovsky E Gliniak BC Park LS Ziegler SF Williams DE Ware CB Meyer JD Davison BL Early lymphocyte expansion is severely impaired in interleukin 7 receptor-deficient mice J Exp Med 1994 180 1955 1960 Mi S Lee X Hu Y Ji B Shao Z Yang W Huang G Walus L Rhodes K Gong BJ Miller RH Pepinsky RB Death receptor 6 negatively regulates oligodendrocyte survival, maturation and myelination Nat Med 2011 17 816 821 Versteeg HH Spek CA Peppelenbosch MP Richel DJ Tissue factor and cancer metastasis: the role of intracellular and extracellular signaling pathways Mol Med 2004 10 6 11 Rawlings JS Rosler KM Harrison DA The JAK/STAT signaling pathway J Cell Sci 2004 117 1281 1283 Fabrication of Ti substrata using photolithography. Field emission scanning electron microscopic images of (A) NE0 (×250), (B) E60/10 (×250) by Fe-SEM. A. The difference in the degree of gene expression was examined by hierarchical clustering analysis. Datas were obtained using median-center, average linkage. B is a cluster of 123 genes showing 1.5 fold changes and C is a cluster of 19 genes showing 2 fold changes. Changes in gene expression level in cell communication. COMP, COL5A3 were up-regulated and KRT18, COL11A1, KRTHA4, KRT7 and KRT19 were downregulated. Changes in gene expression level in MAPK signaling pathway. PLA2G4A, DUSP4 were up-regulated. Changes in gene expression level in Focal adhesion. COMP, COL5A3 were up-regulated and COL11A1, MYLK were down-regulated. Changes in gene expression level in Cytokine-cytokine receptor interaction. LEPR, IL7R, TNFRSF21 were down-regulated. Changes in gene expression level in Calcium signaling pathway. OXTR, ADRA1B, MYLK were down-regulated. Changes in gene expression level in JAK-STAT signaling pathway. LEPR, IL7R were down-regulated. Changes in gene expression level in ECM-receptor interaction. COL5A3 were up-regulated and COL11A1 were down-regulated. Genes showing significant differences in expression more than 2 times Changes of gene expression level were determined at the pathway level using the KEGG (Kyoto Encyclopedia of Genes and Genomes) pathway analysis