Ascorbic Acid/Retinol and/or Inflammatory Stimuli’s Effect on Proliferation/Differentiation Properties and Transcriptomics of Gingival Stem/Progenitor Cells
Karim M. Fawzy El‐Sayed, Amira Bittner, Kristina Schlicht, Mohamed Mekhemar, Kim Enthammer, Marc P. Höppner, M. Es‐Souni, Juliane Schulz, Matthias Laudes, Christian Graetz, Christof E. Dörfer, Dominik M. Schulte
Abstract
The present study explored the effects of ascorbic-acid (AA)/retinol and timed inflammation on the stemness, the regenerative potential, and the transcriptomics profile of gingival mesenchymal stem/progenitor cells’ (G-MSCs). STRO-1 (mesenchymal stem cell marker) immuno-magnetically sorted G-MSCs were cultured in basic medium (control group), in basic medium with IL-1β (1 ng/mL), TNF-α (10 ng/mL) and IFN-γ (100 ng/mL, inflammatory-medium), in basic medium with AA (250 µmol/L) and retinol (20 µmol/L) (AA/retinol group) or in inflammatory medium with AA/retinol (inflammatory/AA/retinol group; n = 5/group). The intracellular levels of phosphorylated and total β-Catenin at 1 h, the expression of stemness genes over 7 days, the number of colony-forming units (CFUs) as well as the cellular proliferation aptitude over 14 days, and the G-MSCs’ multilineage differentiation potential were assessed. Next-generation sequencing was undertaken to elaborate on up-/downregulated genes and altered intracellular pathways. G-MSCs demonstrated all mesenchymal stem/progenitor cells characteristics. Controlled inflammation with AA/retinol significantly elevated NANOG (p < 0.05). The AA/retinol-mediated reduction in intracellular phosphorylated β-Catenin was restored through the effect of controlled inflammation (p < 0.05). Cellular proliferation was highest in the AA/retinol group (p < 0.05). AA/retinol counteracted the inflammation-mediated reduction in G-MSCs’ clonogenic ability and CFUs. Amplified chondrogenic differentiation was observed in the inflammatory/AA/retinol group. At 1 and 3 days, the differentially expressed genes were associated with development, proliferation, and migration (FOS, EGR1, SGK1, CXCL5, SIPA1L2, TFPI2, KRATP1-5), survival (EGR1, SGK1, TMEM132A), differentiation and mineral absorption (FOS, EGR1, MT1E, KRTAP1-5, ASNS, PSAT1), inflammation and MHC-II antigen processing (PER1, CTSS, CD74) and intracellular pathway activation (FKBP5, ZNF404). Less as well as more genes were activated the longer the G-MSCs remained in the inflammatory medium or AA/retinol, respectively. Combined, current results point at possibly interesting interactions between controlled inflammation or AA/retinol affecting stemness, proliferation, and differentiation attributes of G-MSCs.