Effects of short-term inflammatory and/or hypoxic pretreatments on periodontal ligament stem cells: in vitro and in vivo studies

Abstract

In this study, we extensively screened the in vitro and in vivo effects of PDLSCs following short-term inflammatory and/or hypoxic pretreatments. We found that the 24-h hypoxic pretreatment of PDLSCs significantly enhanced cell migration and improved cell surface CXCR4 expression. In addition, hypoxia-pretreated PDLSCs exhibited improved cell colony formation and proliferation. Cells that were dually stimulated also formed more colonies compared to untreated cells but their proliferation did not increase. Importantly, the hypoxic pretreatment of PDLSCs enhanced cell differentiation as determined by elevated RUNX-2 and ALP protein expression. In this context, the inflammatory stimulus impaired cell OCN protein expression, while dual stimuli led to decreased RUNX-2 and OCN mRNA levels. Although preconditioning PDLSCs with inflammatory and/or hypoxic pretreatments resulted in no differences in the production of matrix proteins, hypoxic pretreatment led to the generation of thicker cell sheets; the inflammatory stimulus weakened the ability of cells to form sheets. All the resultant cell sheets exhibited clear bone regeneration following ectopic transplantation as well as in periodontal defect models; the amount of new bone formed by hypoxia-preconditioned cells was significantly greater than that formed by inflammatory stimulus- or dual-stimuli-treated cells or by nonpreconditioned cells. The regeneration of new cementum and periodontal ligaments was only identified in the hypoxia-stimulus and no-stimulus cell groups. Our findings suggest that PDLSCs that undergo short-term hypoxic pretreatment show improved cellular behavior in vitro and enhanced regenerative potential in vivo. The preconditioning of PDLSCs via combined treatments or an inflammatory stimulus requires further investigation.

The periodontal stem/progenitor cell inflammatory‐regenerative cross talk: A new perspective

“…PDLSCs cultured for 24 hours in the IL‐1β (5 ng/mL)/TNFα (10 ng/mL) inflammatory micro‐environment upregulated their expression of CXCR4, to attach to distant SDF‐1/CXCL12, while downregulated it in their resident tissues (see above), thereby increasing their cell migration/homing ability. However, no notable increase in their CFUs ability was observed . Again, it appeared that the two stages of proliferation and homing could not be activated at the same time in the periodontal stem/progenitor cells and need to be timely synchronized.…”

“…Periodontal ligament stem/progenitor cells cultured for 24 hours in IL‐1β (5 ng/mL)/TNFα (10 ng/mL) inflammatory micro‐environment slightly downregulated their expression of RUNX2, ALP, and OC. This effect was even more pronounced, when the inflammatory micro‐environment was combined with a hypoxic state . PDLSCs cultured in osteogenic medium supplemented by monocyte‐derived TNFα exhibited a reduced osteogenic potential.…”

Hypoxia and low‐dose inflammatory stimulus synergistically enhance bone marrow mesenchymal stem cell migration

“…To detect the expression levels of osteogenic (RUNX‐2, ALP and OCN) and adipogenic (PPAR‐γ) genes and proteins, total RNA and protein were collected at day 7 and then subjected to analysis via real‐time PCR and Western blotting, respectively. The sequences of the primers used for real‐time PCR analysis were described previously . Furthermore, the osteogenic differentiation of the cells in each group was assessed through Alizarin Red staining, while the adipogenic potential was analysed using Oil Red O staining.…”

“…The effects of different pretreatments on the expression of osteo- previously. 18 Furthermore, the osteogenic differentiation of the cells in each group was assessed through Alizarin Red staining, while the adipogenic potential was analysed using Oil Red O staining. In parallel, the cells in all groups were incubated with osteogenic medium and adipogenic medium.…”

Administration of signalling molecules dictates stem cell homing for in situ regeneration

“…Additionally, short‐term stimulation of BMMSCs with combinations of various cytokines demonstrated up‐regulated cell surface and intracellular CXCR4 expression, leading to improved cell trafficking potential in vitro and in vivo . Although numerous investigations have revealed that either a hypoxic or an inflammatory stimulus may increase the capacity for cell migration, dual stimuli combined with hypoxia and inflammation do not necessarily lead to a synergistic effect . Further strategies to increase and control cell migration in vivo and to control this process may bring us closer to reaching the original goals of stem cell therapy.…”