ABSTRACT: Fibroblasts are widely used cells for regenerative medicine in clinics, such as gingival or facial skin treatment. In fact, fibroblasts are considered as a mixture of various types of cells with "spindle shape" and there is no available clear marker. Gingival and dermal fibroblasts are similar in their morphology and function; however it is considered that the cultured cells retain their original characteristics depending on the origin, which may contribute to the differential therapeutic effects. For example, gingival wounds are known to heal relatively quickly with less scar formation compared with skin, which may imply their higher capability for regeneration as a therapeutic effect. The reason for this phenomenon may be partly due to characteristic differences between gingival and dermal fibroblasts including the expression of migration stimulating factor and matrix formation but these differences remain largely unknown. Recently, the characteristics of dermal fibroblasts have been reported to be different depending on body sites, such as face, trunk and plamoplantar skin. Although the expression of fibronectin and their splicing variants were known to be different between trunk and oral mucosal fibroblasts, there is still no detailed report on the functional differences between gingival and dermal fibroblasts. In this study, we investigated differential gene expression in normal gingival and dermal fibroblasts using DNA microarray to investigate the difference between the vague fibroblast-type cells from different tissue origin to achieve higher therapeutic effect in cell therapy. Gingival and dermal tissues were collected from healthy patients. After successive stages primary culture and RNA extraction and hybridization on Affymetrix microarrays (Genome Focus Array). Dermal and gingival tissues were obtained from healthy volunteers (8 cases for dermal tissue; 6 females, 2 males , average age 48 and 8 cases for buccal gingival tissue; 6 females, 2 males, average age 43) whose informed consent was obtained according to a protocol approved by the ethics committee of Nagoya University Hospital. After enzymatic digestion, tissues were cultured in Dulbecco's modified Eagle's medium (DMEM) containing 10% fetal bovine serum at 37°C in the presence of 5% CO2 for about 4 weeks as reported previously. Total mRNAs were extracted from cells between passages 4-5 by Trizol reagent (Invitrogen, Carlsbad, CA, USA) and were applied to Human Focus Arrays (Affymetrix, Santa Clara, CA, USA) for microarray analysis according to the manufacturer's protocol (http://www.affymetrix.com/support/technical/manuals.affx). The gene expression data were analyzed by Arrayassist (Stratagene, La Jolla, CA, USA). Briefly, 8,500 probes on the array, normalization and scaling (MAS5), flag-positive gene selection, unpaired t-test, and CV selection (<20) resulted in 5,284 genes to analyze. GO (Gene ontology) analysis was performed using the software default settings to find the gene group related to the same category of biological function by searching common key terms that were reported for each gene.
