RESUMEN
During therapeutic application, mesenchymal stem cells (MSCs) may interact with their environment via their expressed toll-like-receptors (TLRs) leading to pro- or anti-inflammatory immune responses. The present study aimed to describe the gingival margin-derived stem/progenitor cells' (G-MSCs) TLR-induced immune regulatory response to specific TLR agonists. Gingival cells were obtained, immunomagnetically sorted via anti-STRO-1 antibodies and seeded out to achieve colony forming units (CFUs). G-MSCs were investigated for stem cell characteristics and TLR expression. Specific TLR agonists were applied and m-RNA expression of pro- and anti-inflammatory factors was analyzed via real-time polymerase chain reaction. G-MSCs showed all characteristics of stem/progenitor cells. All TLR agonists induced pro-inflammatory cytokines, except for the TLR3 agonist, which significantly promoted the anti-inflammatory response. (p⩽0.05, Wilcoxon-Signed-Ranks-Test). TLR-induced immunomodulation by G-MSCs could impact their therapeutic potential in vivo. Two distinctive pro-inflammatory and an anti-inflammatory TLR-induced phenotypes of G-MSCs become noticeable in this study.
Asunto(s)
Citocinas/inmunología , Inmunomodulación/inmunología , Células Madre Mesenquimatosas/inmunología , Receptores Toll-Like/inmunología , Diferenciación Celular/efectos de los fármacos , Diferenciación Celular/genética , Diferenciación Celular/inmunología , Células Cultivadas , Citocinas/genética , Citocinas/metabolismo , Expresión Génica/efectos de los fármacos , Expresión Génica/inmunología , Encía/citología , Humanos , Lipopéptidos/farmacología , Células Madre Mesenquimatosas/efectos de los fármacos , Células Madre Mesenquimatosas/metabolismo , Poli I-C/farmacología , Receptor Toll-Like 3/agonistas , Receptor Toll-Like 3/inmunología , Receptor Toll-Like 3/metabolismo , Receptores Toll-Like/agonistas , Receptores Toll-Like/metabolismoRESUMEN
AIM: This study investigated the periodontal regenerative potential of gingival margin-derived stem/progenitor cells (G-MSCs) in conjunction with IL-1ra-releasing hyaluronic acid synthetic extracellular matrix (HA-sECM). MATERIALS AND METHODS: Periodontal defects were induced at four sites in eight miniature pigs in the premolar/molar area (-4 weeks). Autologus G-MSCs were isolated from the free gingival margin and magnetically sorted, using anti-STRO-1 antibodies. Colony formation and multilineage differentiation potential were tested. The G-MSCs were expanded and incorporated into IL-1ra-loaded/unloaded HA-sECM. Within every miniature pig, four periodontal defects were randomly treated with IL-1ra/G-MSCs/HA-sECM (test group), G-MSCs/HA-sECM (positive-control), scaling and root planing (SRP; negative control-1) or left untreated (no-treatment group; negative control 2). Differences in clinical attachment level (ΔCAL), probing depth (ΔPD), gingival recession (ΔGR), radiographic defect volume (ΔRDV), and changes in bleeding on probing (BOP) between baseline and 16 weeks post-transplantation, as well as periodontal attachment level (PAL), junctional epithelium length (JE), connective tissue adhesion (CTA), cementum regeneration (CR) and bone regeneration (BR) at 16 weeks post-transplantation were evaluated. RESULTS: Isolated G-MSCs showed stem/progenitor cell characteristics. IL-1ra loaded and unloaded G-MSCs/HA-sECM showed higher ΔCAL, ΔPD, ΔGR, PAL, CR and BR as well as a lower JE compared to their negative controls and improved BOP. CONCLUSION: G-MSCs in conjunction with IL-1ra-loaded/unloaded HA-sECM show a significant periodontal regenerative potential.
Asunto(s)
Encía/citología , Regeneración Tisular Guiada Periodontal/métodos , Ácido Hialurónico/química , Hidrogeles/química , Proteína Antagonista del Receptor de Interleucina 1/uso terapéutico , Trasplante de Células Madre/métodos , Andamios del Tejido/química , Pérdida de Hueso Alveolar/terapia , Animales , Regeneración Ósea/fisiología , Diferenciación Celular/fisiología , Cementogénesis/fisiología , Tejido Conectivo/patología , Raspado Dental/métodos , Inserción Epitelial/patología , Femenino , Recesión Gingival/terapia , Masculino , Pérdida de la Inserción Periodontal/terapia , Índice Periodontal , Bolsa Periodontal/terapia , Periodontitis/terapia , Distribución Aleatoria , Aplanamiento de la Raíz/métodos , Células Madre/fisiología , Porcinos , Porcinos EnanosRESUMEN
Cytokines play major roles in tissue destruction/repair. The present study investigates proliferative and osteogenic differentiation potentials of gingival mesenchymal stem/progenitor cells (G-MSCs), influenced by IL-1/TNF-α inflammatory/anti-inflammatory conditions. Human G-MSCs were isolated, characterized, and cultured in basic medium (control group, M1), in basic medium with IL-1ß, TNF-α, and IFN-γ (inflammatory group, M2) and with IL-1ra/TNF-αi added to M2 (anti-inflammatory group, M3). MTT tests at days 1, 3, and 7 and CFU assay at day 12 were conducted. Osteogenic differentiation was analyzed by bone-specific transcription factors (RUNX2), alkaline phosphatase (ALP), type I collagen (Col-I), osteopontin (OPN), and osteonectin (ON) expression at days 1, 3, 7, and 14 and Alizarin red staining at day 14. At day 3, the control group showed the highest cell numbers. At day 7, cell numbers in inflammatory and anti-inflammatory group outnumbered the control group. At day 12, CFUs decreased in the inflammatory and anti-inflammatory groups, with altered cellular morphology. The anti-inflammatory group demonstrated elevated bone-specific transcription factors at 14 days. After 14 days of osteogenic induction, calcified nodules in the anti-inflammatory group were higher compared to control and inflammatory groups. For regeneration, initial inflammatory stimuli appear essential for G-MSCs' proliferation. With inflammatory persistence, this positive effect perishes and is followed by a short-term stimulatory one on osteogenesis. At this stage, selective anti-inflammatory intervention could boost G-MSCs' differentiation.