Mitsuhiro Ohshima1, Yoko Yamaguchi2, Kimiharu Ambe3, Masafumi Horie4, Akira Saito4, Takahide Nagase4, Keisuke Nakashima5, Hidero Ohki6, Toshihisa Kawai7, Yoshimitsu Abiko8, Patrick Micke9, Kai Kappert10. 1. Department of Biochemistry, Ohu University School of Pharmaceutical Sciences, Koriyama, Fukushima, Japan. 2. Department of Biochemistry, Nihon University School of Dentistry, Tokyo, Japan. 3. Department of Morphological Biology, Ohu University School of Dentistry, Koriyama, Fukushima, Japan. 4. Department of Respiratory Medicine, Graduate School of Medicine and Faculty of Medicine, University of Tokyo, Tokyo, Japan. 5. Division of Periodontology, Department of Oral Function, Kyushu Dental University, Fukuoka, Japan. 6. First Department of Oral Surgery, Nihon University School of Dentistry, Tokyo, Japan. 7. Department of Immunology, The Forsyth Institute, Cambridge, MA, USA. 8. Department of Molecular Biology and Biochemistry, Nihon University School of Dentistry at Matsudo, Chiba, Japan. 9. Department of Immunology, Genetics and Pathology, Uppsala University, Hospital, Uppsala, Sweden. 10. Institute of Laboratory Medicine, Clinical Chemistry and Pathobiochemistry, Center for Cardiovascular Research (CCR), Charité-University Medicine Berlin, Berlin, Germany.
Abstract
AIM: Degradation of extracellular matrices is an integral part in periodontitis. For antagonizing this pathophysiological mechanism, we aimed at identifying gene expression profiles in disease progression contributing periodontitis-associated fibroblasts (PAFs) versus normal gingival fibroblasts to determine their molecular repertoire, and exploit it for therapeutic intervention. MATERIALS AND METHODS: Applying an exploratory analysis using a small number of microarrays in combination with a three dimensional (3D) in vitro culture model that incorporates some aspects of periodontitis, PAFs were initially characterized by gene-expression analyses, followed by targeted gene down-regulation and pharmacological intervention in vitro. Further, immunohistochemistry was applied for phosphorylation analyses in tissue specimens. RESULTS: PAFs were characterized by 42 genes being commonly up-regulated >1.5-fold, and by five genes that were concordantly down-regulated (<0.7-fold). Expression of vascular endothelial growth factor (VEGF)-receptor 1 (Flt-1) was highly enhanced, and was thus further explored in in vitro culture models of periodontal fibroblasts without accounting for the microbiome. Phosphorylation of the VEGF-receptor 1 was enhanced in PAFs. Receptor inhibition by a specific VEGF-receptor inhibitor or intrinsic down-regulation by RNAi of the VEGF-receptor kinase in 3D gel cultures resulted in significant reduction in collagen degradation associated with increased tissue inhibitor of metalloproteinase expression, suggesting that Flt-1 may contribute to periodontitis. CONCLUSION: Based on the finding that VEGF-receptor kinase inhibition impaired collagen degradation pathways, Flt-1 may represent a candidate for therapeutic approaches in periodontitis.
AIM: Degradation of extracellular matrices is an integral part in periodontitis. For antagonizing this pathophysiological mechanism, we aimed at identifying gene expression profiles in disease progression contributing periodontitis-associated fibroblasts (PAFs) versus normal gingival fibroblasts to determine their molecular repertoire, and exploit it for therapeutic intervention. MATERIALS AND METHODS: Applying an exploratory analysis using a small number of microarrays in combination with a three dimensional (3D) in vitro culture model that incorporates some aspects of periodontitis, PAFs were initially characterized by gene-expression analyses, followed by targeted gene down-regulation and pharmacological intervention in vitro. Further, immunohistochemistry was applied for phosphorylation analyses in tissue specimens. RESULTS: PAFs were characterized by 42 genes being commonly up-regulated >1.5-fold, and by five genes that were concordantly down-regulated (<0.7-fold). Expression of vascular endothelial growth factor (VEGF)-receptor 1 (Flt-1) was highly enhanced, and was thus further explored in in vitro culture models of periodontal fibroblasts without accounting for the microbiome. Phosphorylation of the VEGF-receptor 1 was enhanced in PAFs. Receptor inhibition by a specific VEGF-receptor inhibitor or intrinsic down-regulation by RNAi of the VEGF-receptor kinase in 3D gel cultures resulted in significant reduction in collagen degradation associated with increased tissue inhibitor of metalloproteinase expression, suggesting that Flt-1 may contribute to periodontitis. CONCLUSION: Based on the finding that VEGF-receptor kinase inhibition impaired collagen degradation pathways, Flt-1 may represent a candidate for therapeutic approaches in periodontitis.
Authors: Olga Kuten-Pella; Andrea De Luna; Karina Kramer; Markus Neubauer; Stefan Nehrer; Zsombor Lacza Journal: Curr Issues Mol Biol Date: 2021-07-11 Impact factor: 2.976