The mechanisms of wound healing and vascularization are crucial steps of the complex morphological process of tissue reconstruction. In addition to epithelial cells, fibroblasts play an important role in this process. They are characterized by dynamic proliferation and they form the stroma for epithelial cells. In this study, we have used primary cultures of oral fibroblasts, obtained from porcine buccal mucosa. Cells were maintained long-term in in vitro conditions, in order to investigate the expression profile of the molecular markers involved in wound healing and vascularization. Based on the Affymetrix assays, we have observed three ontological groups of markers as wound healing group, response to wounding group and vascularization group, represented by different genes characterized by their expression profile during long-term primary in vitro culture (IVC) of porcine oral fibroblasts. Following the analysis of gene expression in three previously identified groups of genes, we have identified that transforming growth factor beta 1 (TGFB1), ITGB3, PDPN, and ETS1 are involved in all three processes, suggesting that these genes could be recognized as markers of repair specific for oral fibroblasts within the porcine mucosal tissue.

Department of Anatomy and Histology University of Zielona Gora 65 046 Zielona Góra Poland

Department of Anatomy Poznan University of Medical Science 60 781 Poznań Poland

Department of Basic and Preclinical Sciences Institute of Veterinary Medicine Faculty of Biological and Veterinary Sciences Nicolaus Copernicus University in Torun 87 100 Toruń Poland

Department of Biomaterials and Experimental Dentistry Poznan University of Medical Sciences 61 701 Poznań Poland

Department of Craniofacial Development and Stem Cell Biology King's College University of London London WC2R 2LS UK

Department of Diagnostics and Clinical Sciences Nicolaus Copernicus University in Torun 87 100 Toruń Poland

Department of Histology and Embryology Poznan University of Medical Science 60 781 Poznań Poland

Department of Obstetrics and Gynecology University Hospital and Masaryk University 601 77 Brno Czech Republic

Department of Periodontology and Oral Implantology Dental Research Division University of Guarulhos Guarulhos SP 07030 010 Brazil

Department of Veterinary Surgery Nicolaus Copernicus University in Torun 87 100 Toruń Poland

Physiology Graduate Program North Carolina State University Raleigh NC 27695 USA

Zobrazit více v PubMed

Politis C., Schoenaers J., Jacobs R., Agbaje J.O. Wound Healing Problems in the Mouth. Front. Physiol. 2016;7:507. doi: 10.3389/fphys.2016.00507. PubMed DOI PMC

Enoch S., Peake M.A., Wall I., Davies L., Farrier J., Giles P., Kipling D., Price P., Moseley R., Thomas D., et al. ‘Young’ oral fibroblasts are geno/phenotypically distinct. J. Dent. Res. 2010;89:1407–1413. doi: 10.1177/0022034510377796. PubMed DOI

Glim J.E., van Egmond M., Niessen F.B., Everts V., Beelen R.H.J. Detrimental dermal wound healing: What can we learn from the oral mucosa? Wound Repair Regen. 2013;21:648–660. doi: 10.1111/wrr.12072. PubMed DOI

Jones K.B., Klein O.D. Oral epithelial stem cells in tissue maintenance and disease: The first steps in a long journey. Int. J. Oral Sci. 2013;5:121–129. doi: 10.1038/ijos.2013.46. PubMed DOI PMC

Kondo M., Yamato M., Takagi R., Murakami D., Namiki H., Okano T. Significantly different proliferative potential of oral mucosal epithelial cells between six animal species. J. Biomed. Mater. Res. A. 2014;102:1829–1837. doi: 10.1002/jbm.a.34849. PubMed DOI

Winning T.A., Townsend G.C. Oral mucosal embryology and histology. Clin. Dermatol. 2000;18:499–511. doi: 10.1016/S0738-081X(00)00140-1. PubMed DOI

Bryja A., Dyszkiewicz-Konwińska M., Jankowski M., Celichowski P., Stefańska K., Chamier-Gliszczyńska A., Borowiec B., Mehr K., Bukowska D., Antosik P., et al. Cation homeostasis and transport related gene markers are differentially expressed in porcine buccal pouch mucosal cells during long-term cells primary culture in vitro. Med. J. Cell Biol. 2018;6:83–90. doi: 10.2478/acb-2018-0014. DOI

Bryja A., Dyszkiewicz-Konwińska M., Jankowski M., Celichowski P., Stefańska K., Chamier-Gliszczyńska A., Popis M., Mehr K., Bukowska D., Antosik P., et al. Ion homeostasis and transport are regulated by genes differentially expressed in porcine buccal pouch mucosal cells during long-term culture in vitro—A microarray approach. Med. J. Cell Biol. 2018;6:75–82. doi: 10.2478/acb-2018-0013. DOI

Ferguson M.W.J., O’Kane S. Scar-free healing: From embryonic mechanisms to adult therapeutic intervention. Philos. Trans. R. Soc. Lond. B Biol. Sci. 2004;359:839–850. doi: 10.1098/rstb.2004.1475. PubMed DOI PMC

Coolen N.A., Schouten K.C.W.M., Boekema B.K.H.L., Middelkoop E., Ulrich M.M.W. Wound healing in a fetal, adult, and scar tissue model: A comparative study. Wound Repair Regen. 2010;18:291–301. doi: 10.1111/j.1524-475X.2010.00585.x. PubMed DOI

Coolen N.A., Schouten K.C.W.M., Middelkoop E., Ulrich M.M.W. Comparison between human fetal and adult skin. Arch. Dermatol. Res. 2010;302:47–55. doi: 10.1007/s00403-009-0989-8. PubMed DOI PMC

Namazi M.R., Fallahzadeh M.K., Schwartz R.A. Strategies for prevention of scars: What can we learn from fetal skin? Int. J. Dermatol. 2011;50:85–93. doi: 10.1111/j.1365-4632.2010.04678.x. PubMed DOI

Kathju S., Gallo P.H., Satish L. Scarless integumentary wound healing in the mammalian fetus: Molecular basis and therapeutic implications. Birth Defects Res. C Embryo Today. 2012;96:223–236. doi: 10.1002/bdrc.21015. PubMed DOI

Penn J.W., Grobbelaar A.O., Rolfe K.J. The role of the TGF-beta family in wound healing, burns and scarring: A review. Int. J. Burns Trauma. 2012;2:18–28. PubMed PMC

Chen L., Arbieva Z.H., Guo S., Marucha P.T., Mustoe T.A., DiPietro L.A. Positional differences in the wound transcriptome of skin and oral mucosa. BMC Genomics. 2010;11:471. doi: 10.1186/1471-2164-11-471. PubMed DOI PMC

Bryja A., Dyszkiewicz-Konwinska M., Budna J., Ciesiółka S., Kranc W., Borys S., Jeseta M., Urbaniak O., Bukowska D., Antosik P., et al. Expression of cell mitotic progression proteins and keratinocyte markers in porcine buccal pouch mucosal cells during short-term, real-time primary culture. J. Biol. Regul. Homeost. Agents. 2017;31:297–309. PubMed

Bryja A., Dyszkiewicz-Konwinska M., Budna J., Kranc W., Chachula A., Borys S., Ciesiółka S., Sokalski J., Prylinski M., Bukowska D., et al. The biomedical aspects of oral mucosal epithelial cell culture in mammals. J. Biol. Regul. Homeost. Agents. 2017;31:81–85. PubMed

Takaichi S., Muramatsu T., Lee J.-M., Jung H.-S., Shinozaki N., Katakura A., Yamane G.-Y. Re-epithelialization of the buccal mucosa after alkaline chemical injury. Acta Histochem. Cytochem. 2014;47:195–201. doi: 10.1267/ahc.14015. PubMed DOI PMC

Lee H.-G., Eun H.C. Differences between fibroblasts cultured from oral mucosa and normal skin: Implication to wound healing. J. Dermatol. Sci. 1999;21:176–182. doi: 10.1016/S0923-1811(99)00037-7. PubMed DOI

Wong J.W., Gallant-Behm C., Wiebe C., Mak K., Hart D.A., Larjava H., Häkkinen L. Wound healing in oral mucosa results in reduced scar formation as compared with skin: Evidence from the red Duroc pig model and humans. Wound Repair Regen. 2009;17:717–729. doi: 10.1111/j.1524-475X.2009.00531.x. PubMed DOI

Smith P.C., Martínez C., Martínez J., McCulloch C.A. Role of Fibroblast Populations in Periodontal Wound Healing and Tissue Remodeling. Front. Physiol. 2019;10:270. doi: 10.3389/fphys.2019.00270. PubMed DOI PMC

Rahimov C.H., Gasimov E., Guliyev T., Rzayev F., Farzaliyev I. Comparative behavior of wound healing on within different sites of oral mucosa and skin. Experimental study on pigs by electron microscope. Int. J. Oral Maxillofac. Surg. 2015;44:e278. doi: 10.1016/j.ijom.2015.08.289. DOI

Hyun S.-Y., Mun S., Kang K.-J., Lim J.-C., Kim S.-Y., Han K., Jang Y.-J. Amelogenic transcriptome profiling in ameloblast-like cells derived from adult gingival epithelial cells. Sci. Rep. 2019;9:3736. doi: 10.1038/s41598-019-40091-x. PubMed DOI PMC

Khan M., Gasser S. Generating Primary Fibroblast Cultures from Mouse Ear and Tail Tissues. J. Vis. Exp. 2016;107:53565. doi: 10.3791/53565. PubMed DOI PMC

Chomczynski P., Sacchi N. Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. Anal. Biochem. 1987;162:156–159. doi: 10.1016/0003-2697(87)90021-2. PubMed DOI

Huang D.W., Sherman B.T., Tan Q., Collins J.R., Alvord W.G., Roayaei J., Stephens R., Baseler M.W., Lane H.C., Lempicki R.A. The DAVID Gene Functional Classification Tool: A novel biological module-centric algorithm to functionally analyze large gene lists. Genome Biol. 2007;8:R183. doi: 10.1186/gb-2007-8-9-r183. PubMed DOI PMC

Walter W., Sanchez-Cabo F., Ricote M. GOplot: An R package for visually combining expression data with functional analysis. Bioinformatics. 2015;31:2912–2914. doi: 10.1093/bioinformatics/btv300. PubMed DOI

Von Mering C., Jensen L.J., Snel B., Hooper S.D., Krupp M., Foglierini M., Jouffre N., Huynen M.A., Bork P. STRING: Known and predicted protein-protein associations, integrated and transferred across organisms. Nucleic Acids Res. 2005;33:D433–D437. doi: 10.1093/nar/gki005. PubMed DOI PMC

Livak K.J., Schmittgen T.D. Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method. Methods. 2001;25:402–408. doi: 10.1006/meth.2001.1262. PubMed DOI

Mak K., Manji A., Gallant-Behm C., Wiebe C., Hart D.A., Larjava H., Hakkinen L. Scarless healing of oral mucosa is characterized by faster resolution of inflammation and control of myofibroblast action compared to skin wounds in the red Duroc pig model. J. Dermatol. Sci. 2009;56:168–180. doi: 10.1016/j.jdermsci.2009.09.005. PubMed DOI

Gemenetzidis E., Elena-Costea D., Parkinson E.K., Waseem A., Wan H., Teh M.T. Induction of human epithelial stem/progenitor expansion by FOXM1. Cancer Res. 2010;70:9515–9526. doi: 10.1158/0008-5472.CAN-10-2173. PubMed DOI PMC

Nakamura T., Endo K.-I., Kinoshita S. Identification of human oral keratinocyte stem/progenitor cells by neurotrophin receptor p75 and the role of neurotrophin/p75 signaling. Stem Cells. 2007;25:628–638. doi: 10.1634/stemcells.2006-0494. PubMed DOI

Izumi K., Tobita T., Feinberg S.E. Isolation of human oral keratinocyte progenitor/stem cells. J. Dent. Res. 2007;86:341–346. doi: 10.1177/154405910708600408. PubMed DOI

Stephens P., Genever P. Non-epithelial oral mucosal progenitor cell populations. Oral Dis. 2007;13:1–10. doi: 10.1111/j.1601-0825.2006.01314.x. PubMed DOI

Ravikanth M., Soujanya P., Manjunath K., Saraswathi T.R., Ramachandran C.R. Heterogenecity of fibroblasts. J. Oral Maxillofac. Pathol. 2011;15:247–250. doi: 10.4103/0973-029X.84516. PubMed DOI PMC

Higa K., Satake Y., Shimazaki J. The characterization of human oral mucosal fibroblasts and their use as feeder cells in cultivated epithelial sheets. Future Sci. OA. 2017;3:FSO243. doi: 10.4155/fsoa-2017-0074. PubMed DOI PMC

Van Wyk C.W., Olivier A., Hoal-van Helden E.G., Grobler-Rabie A.F. Growth of oral and skin fibroblasts from patients with oral submucous fibrosis. J. Oral Pathol. Med. 1995;24:349–353. doi: 10.1111/j.1600-0714.1995.tb01198.x. PubMed DOI

Mah W., Jiang G., Olver D., Gallant-Behm C., Wiebe C., Hart D.A., Koivisto L., Larjava H., Häkkinen L. Elevated CD26 Expression by Skin Fibroblasts Distinguishes a Profibrotic Phenotype Involved in Scar Formation Compared to Gingival Fibroblasts. Am. J. Pathol. 2017;187:1717–1735. doi: 10.1016/j.ajpath.2017.04.017. PubMed DOI

Gurtner G.C., Werner S., Barrandon Y., Longaker M.T. Wound repair and regeneration. Nature. 2008;453:314–321. doi: 10.1038/nature07039. PubMed DOI

Murphy-Ullrich J.E., Poczatek M. Activation of latent TGF-beta by thrombospondin-1: Mechanisms and physiology. Cytokine Growth Factor Rev. 2000;11:59–69. doi: 10.1016/S1359-6101(99)00029-5. PubMed DOI

Szpaderska A.M., Walsh C.G., Steinberg M.J., DiPietro L.A. Distinct patterns of angiogenesis in oral and skin wounds. J. Dent. Res. 2005;84:309–314. doi: 10.1177/154405910508400403. PubMed DOI

Szpaderska A.M., Zuckerman J.D., DiPietro L.A. Differential injury responses in oral mucosal and cutaneous wounds. J. Dent. Res. 2003;82:621–626. doi: 10.1177/154405910308200810. PubMed DOI

Lu S.-L., Reh D., Li A.G., Woods J., Corless C.L., Kulesz-Martin M., Wang X.-J. Overexpression of transforming growth factor beta1 in head and neck epithelia results in inflammation, angiogenesis, and epithelial hyperproliferation. Cancer Res. 2004;64:4405–4410. doi: 10.1158/0008-5472.CAN-04-1032. PubMed DOI

Januszyk M., Rennert R.C., Sorkin M., Maan Z.N., Wong L.K., Whittam A.J., Whitmore A., Duscher D., Gurtner G.C. Evaluating the Effect of Cell Culture on Gene Expression in Primary Tissue Samples Using Microfluidic-Based Single Cell Transcriptional Analysis. Microarrays. 2015;4:540–550. doi: 10.3390/microarrays4040540. PubMed DOI PMC

Miyoshi K., Horiguchi T., Tanimura A., Hagita H., Noma T. Gene Signature of Human Oral Mucosa Fibroblasts: Comparison with Dermal Fibroblasts and Induced Pluripotent Stem Cells. BioMed Res. Int. 2015;2015:121575. doi: 10.1155/2015/121575. PubMed DOI PMC

Larjava H., Wiebe C., Gallant-Behm C., Hart D.A., Heino J., Hakkinen L. Exploring scarless healing of oral soft tissues. J. Can. Dent. Assoc. 2011;77:b18. PubMed

Wang R., Ghahary A., Shen Q., Scott P.G., Roy K., Tredget E.E. Hypertrophic scar tissues and fibroblasts produce more transforming growth factor-beta1 mRNA and protein than normal skin and cells. Wound Repair Regen. 2000;8:128–137. doi: 10.1046/j.1524-475x.2000.00128.x. PubMed DOI

Czuwara-Ladykowska J., Sementchenko V.I., Watson D.K., Trojanowska M. Ets1 is an effector of the transforming growth factor beta (TGF-beta ) signaling pathway and an antagonist of the profibrotic effects of TGF-beta. J. Biol. Chem. 2002;277:20399–20408. doi: 10.1074/jbc.M200206200. PubMed DOI

Ebisawa K., Kato R., Okada M., Sugimura T., Latif M.A., Hori Y., Narita Y., Ueda M., Honda H., Kagami H. Gingival and dermal fibroblasts: Their similarities and differences revealed from gene expression. J. Biosci. Bioeng. 2011;111:255–258. doi: 10.1016/j.jbiosc.2010.11.014. PubMed DOI