IADR Abstract Archives
Recruitment of Gingival Fibroblasts by Periodontal Ligament Cells
Objectives: Mesenchymal stem cell (MSC) implantation may contribute to tissue regeneration directly, but may also act indirectly by recruitment of resident cells to undergo differentiation. The aim of this study was to investigate if periodontal ligament MSCs can induce periodontal differentiation of gingival fibroblast MSCs in vitro.
Methods: Fluorescent labelled ( Cell Tracker) human primary PDL MSCs were co cultured with CD90+ve primary gingival fibroblasts (GF). After 3 days co-culture cells were sorted into gingival and PDL cells by fluorescence activated cell sorting and tested for expression of Asporin (APN), Periostin (PSN) and Alkaline phosphatase (ALP) by quantitative RT-PCR. GF and PDL monocultures were used as controls. In further experiments GF were co-cultured with PDL cells in transwell cultures and analysed as before. We also tested the effects of conditioned medium derived from PDL cultures on GF. Finally we tested the effects of addition of the BMP inhibitor noggin and the wnt signalling inhibitor dkk on PDL conditioned medium induction of GF.
Results: GF directly exposed to PDL MSC in co-cultures showed up regulation of Asporin, Periostin and ALP gene expression ( x6 fold, x5 fold and x3 fold upregulation respectively P <0.01) . In transwell cultures, asporin, periostin and ALP were upregulated in GF exposed to PDL cells, compared to GF controls by x 59.4 fold, x6.3 fold and x12 fold respectively ( P < 0.01). PDL conditioned medium showed qualitatively similar results and also induced ALP protein upregulation. Addition of dkk, but not noggin reduced the ALP induction by conditioned medium by up to 60% in a dose dependent manner.
Conclusions: The results suggest that PDL MSC can recruit GF MSCs to differentiate towards a PDL phenotype by secretion of soluble factors. Preliminary blocking experiments suggest that this is at least partly mediated by wnt signalling.
Methods: Fluorescent labelled ( Cell Tracker) human primary PDL MSCs were co cultured with CD90+ve primary gingival fibroblasts (GF). After 3 days co-culture cells were sorted into gingival and PDL cells by fluorescence activated cell sorting and tested for expression of Asporin (APN), Periostin (PSN) and Alkaline phosphatase (ALP) by quantitative RT-PCR. GF and PDL monocultures were used as controls. In further experiments GF were co-cultured with PDL cells in transwell cultures and analysed as before. We also tested the effects of conditioned medium derived from PDL cultures on GF. Finally we tested the effects of addition of the BMP inhibitor noggin and the wnt signalling inhibitor dkk on PDL conditioned medium induction of GF.
Results: GF directly exposed to PDL MSC in co-cultures showed up regulation of Asporin, Periostin and ALP gene expression ( x6 fold, x5 fold and x3 fold upregulation respectively P <0.01) . In transwell cultures, asporin, periostin and ALP were upregulated in GF exposed to PDL cells, compared to GF controls by x 59.4 fold, x6.3 fold and x12 fold respectively ( P < 0.01). PDL conditioned medium showed qualitatively similar results and also induced ALP protein upregulation. Addition of dkk, but not noggin reduced the ALP induction by conditioned medium by up to 60% in a dose dependent manner.
Conclusions: The results suggest that PDL MSC can recruit GF MSCs to differentiate towards a PDL phenotype by secretion of soluble factors. Preliminary blocking experiments suggest that this is at least partly mediated by wnt signalling.