Retinol/inflammation affect stemness and differentiation potential of gingival stem/progenitor cells via Wnt/β‐catenin

Abstract

Background and objective Inflammatory cytokines impact the course of periodontal disease, repair, and regeneration. Vitamin A and its metabolites are inflammation‐modulatory biomolecules, affecting cellular pluripotency. The aim of this study was to investigate the effect of retinol and periodontal inflammatory cytokines (IL‐1β/TNF‐α/IFN‐γ) on pluripotency and proliferative properties of gingival mesenchymal stem/progenitor cells (G‐MSCs), for the first time. Material and methods Human G‐MSCs (n = 5) were STRO‐1 immuno‐magnetically sorted, characterized and expanded 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 group), in basic medium with retinol (20 μmol/L) (retinol group) and with retinol added to the inflammatory group (inflammatory/retinol group). β‐catenin levels at 1 hour, cellular proliferation over 14 days, and colony‐forming units (CFUs) at 14 days were investigated. Pluripotency gene expressions were examined at 1, 3, and 5 days via reverse transcription‐polymerase chain reaction (RT‐PCR). Multilineage differentiation potential was evaluated, following 5 days priming, using qualitative and quantitative histochemistry and RT‐PCR. Results G‐MSCs were CD14−, CD34−, CD45−, CD73+, CD90+, CD105+, and showed mesenchymal stem/progenitor cells’ hallmarks, CFUs, and multilineage differentiation potential. Intracellular β‐catenin significantly declined in the stimulated groups (P < 0.001, Friedman test). Cellular proliferation at 72 hours was most prominent in the control and inflammatory group [Median cell numbers (Q25/Q75); 6806 (4983/7312) and 5414 (4457/7230), respectively], followed by an upsurge in the retinol group. At 14 days, the retinol group exhibited the highest CFUs [Median CFUs (Q25/Q75); 35 (20/58), P = 0.043, Wilcoxon signed‐rank]. Nanog was most expressed in the inflammatory and retinol group [Median gene expression/PGK1 (Q25/Q75); 0.0006 (0.0002/0.0014) and 0.0005 (0.0003/0.0008)]. Inflammation significantly upregulated Sox2 expression [0.0002 (0.0008/0.0005)], while its expression was diminished in the retinol and inflammatory/retinol group (P < 0.001, Friedman test). Inflammatory/retinol group exhibited the highest multilineage differentiation potential. Conclusion Controlled short‐term inflammatory/retinol stimuli activate the Wnt/β‐catenin pathway, affecting G‐MSCs’ pluripotency, proliferation, and differentiation. The present findings provide further insights into the inflammatory‐regenerative interactions and their modulation potential for G‐MSCs‐mediated periodontal repair/regeneration.

Ascorbic Acid, Inflammatory Cytokines (IL-1β/TNF-α/IFN-γ), or Their Combination’s Effect on Stemness, Proliferation, and Differentiation of Gingival Mesenchymal Stem/Progenitor Cells

“…The current study examined the effect of the inflammatory cytokines mixture together and in combination with AA on the pluripotency markers' expression, namely Nanog, Oct4A, and Sox2 at 24, 72, and 120 hours. In line with earlier studies, in the present one Nanog, Oct4A, and Sox2 expressions were amplified in the AA- [3,20,48] and inflammation-challenged [19,49] G-MSCs. The AAinduced increase in the pluripotency markers Oct4A and Nanog could be ascribed primarily to the capability of AA, similar to Retinol [19], to activate the ten-eleventranslocation (TET) demethylases, eliminating methylation of the DNA and thereby inciting intracellular epigenetic reprogramming actions, encompassing pluripotency amplification [19,50,51] in G-MSCs.…”

“…Reparative/regenerative approaches in periodontology are based principally on recapitulating the chief periodontal developmental events, encompassing stem/progenitor cells' proliferation, migration, homing, differentiation, and finally maturation [28]. Clinically, these healing stages primarily take place in an initially inflamed periodontal microenvironment, with inflammatory cytokines orchestrating the course of the inflammatory periodontal disease progression [29], possible healing/regeneration [30], as well as periodontal stem/progenitor cells' attributes [18,19]. Apart from its important roles in periodontal wound healing, tissue regeneration [31], and collagen synthesis of bone, teeth, and gingiva, AA demonstrates potent cellular protective antioxidative properties in response to periodontitis-induced oxidative inflammatory reactions [32,33].…”

Maresin-1 and Resolvin E1 Promote Regenerative Properties of Periodontal Ligament Stem Cells Under Inflammatory Conditions

“…Nevertheless, TNF-α may also induce stem cell death and inhibit osteogenic differentiation of hPDLSCs (28,31). A combination of TNF-α and IL-1β creates an even greater inflammatory stimulus and up-regulates Sox2 expression on stem cells in tissues such as the gingiva (38). In turn, double Oct4 + Sox2 + cells have shown immunomodulatory activity by reducing the expression of both cytokines (39), suggesting that the inflammatory environment provides a two-way interaction where the synthesis of pro-inflammatory mediators can also activate resolution and a return to homeostasis.…”

Dental Stem Cell-Derived Secretome/Conditioned Medium: The Future for Regenerative Therapeutic Applications

“…Dental MSCs are unique adult MSCs, derived from the ectomesenchyme's neural cells [32,33]. They include dental pulp mesenchymal stem/progenitor cells (dental pulp MSCs) isolated from dental pulpal tissues of permanent teeth [34], stem/progenitor cells extracted from pulpal tissues of human shed deciduous teeth (SHED) [35,36], periodontal ligament mesenchymal stem/progenitor cells (periodontal ligament MSCs) isolated from the periodontal ligament [37,38], dental follicle mesenchymal stem/progenitor cells (dental follicle MSCs), usually isolated from the dental follicle surrounding the third molar [39], alveolar bone proper-derived mesenchymal stem/progenitor cells (alveolar bone MSCs) [40][41][42], mesenchymal stem/progenitor cells isolated from the apical dental papilla (MSCs from apical papilla) at the apices of the immature permanent teeth [38,43], tooth germ progenitor cells, isolated from late bell stage third molar's tooth germs [44], and gingival mesenchymal stem/progenitor cells (gingival MSCs), isolated from gingival tissues [45][46][47][48][49].…”