Basic fibroblast growth factor regulates phosphate/pyrophosphate regulatory genes in stem cells isolated from human exfoliated deciduous teeth

Abstract

BackgroundBasic fibroblast growth factor (bFGF) regulates maintenance of stemness and modulation of osteo/odontogenic differentiation and mineralization in stem cells from human exfoliated deciduous teeth (SHEDs). Mineralization in the bones and teeth is in part controlled by pericellular levels of inorganic phosphate (Pi), a component of hydroxyapatite, and inorganic pyrophosphate (PPi), an inhibitor of mineralization. The progressive ankylosis protein (gene ANKH; protein ANKH) and ectonucleotide pyrophosphatase phosphodiesterase 1 (ENPP1/ENPP1) increase PPi and inhibit mineralization, while tissue-nonspecific alkaline phosphatase (ALPL; TNAP) is a critical pro-mineralization enzyme that hydrolyzes PPi. We hypothesized that regulation by bFGF of mineralization in SHEDs occurs by modulation of Pi/PPi-associated genes.MethodsCells were isolated from human exfoliated deciduous teeth and characterized for mesenchymal stem cell characteristics. Cells were treated with bFGF, and the osteogenic differentiation ability was determined. The mRNA expression was evaluated using real-time polymerase chain reaction. The mineralization was examined using alizarin red S staining.ResultsCells isolated from primary teeth expressed mesenchymal stem cell markers, CD44, CD90, and CD105, and were able to differentiate into osteo/odontogenic and adipogenic lineages. Addition of 10 ng/ml bFGF to SHEDs during in vitro osteo/odontogenic differentiation decreased ALPL mRNA expression and ALP enzyme activity, increased ANKH mRNA, and decreased both Pi/PPi ratio and mineral deposition. Effects of bFGF on ALPL and ANKH expression were detected within 24 h. Addition of 20 mM fibroblast growth factor receptor (FGFR) inhibitor SU5402 revealed the necessity of FGFR-mediated signaling, and inclusion of 1 μg/ml cyclohexamide (CHX) implicated the necessity of protein synthesis for effects on ALPL and ANKH. Addition of exogenous 10 μm PPi inhibited mineralization and increased ANKH, collagen type 1a1 (COL1A1), and osteopontin (SPP1) mRNA, while addition of exogenous Pi increased mineralization and osterix (OSX), ANKH, SPP1, and dentin matrix protein 1 (DMP1) mRNA. The effects of PPi and Pi on mineralization could be replicated by short-term 3- and 7-day treatments, suggesting signaling effects in addition to physicochemical regulation of mineral deposition.ConclusionThis study reveals for the first time the effects of bFGF on Pi/PPi regulators in SHEDs and implicates these factors in how bFGF directs osteo/odontogenic differentiation and mineralization by these cells.Electronic supplementary materialThe online version of this article (10.1186/s13287-018-1093-9) contains supplementary material, which is available to authorized users.

Activation of nuclear factor-kappa B by TNF promotes nucleus pulposus mineralization through inhibition of ANKH and ENPP1

“…We also observed that either TNF treatment or enforced TNF expression further enhanced the mineral deposition in TNAP-overexpressing bNPs in vitro. Our gene expression analysis showed that TNF decreased ANKH mRNA by approximately 50%, in agreement with reports showing that TNF promotes ectopic mineralization in hASMCs by suppressing ANKH mRNA expression by half 21 , and other authors in several tissues showing induced matrix mineralization and 50% reduction of ANKH mRNA expression in target cells as a result of TGF-β1 57 , ePi/forskolin 58 , or IL-1β 24 , 25 treatment, whereas basic fibroblast growth factor (bFGF) 59 and IL-6 60 treatment increased ANKH mRNA by twofold while still driving mineralization. Regarding ENPP1 expression modification by cytokines, it has been reported that IL-1β decreases ENPP in hMSCs 23 and in bNPs 25 , mRNA for ENPP was also reduced in rat ASMCs by ePi/forskolin treatment 58 , and ENPP is decreased in IDD patients with advanced degeneration and mineralization 56 .…”

“…Regarding ENPP1 expression modification by cytokines, it has been reported that IL-1β decreases ENPP in hMSCs 23 and in bNPs 25 , mRNA for ENPP was also reduced in rat ASMCs by ePi/forskolin treatment 58 , and ENPP is decreased in IDD patients with advanced degeneration and mineralization 56 . Treatment of human deciduous teeth stem cells with IL-6 had little effect on ENPP1 mRNA expression, whereas bFGF resulted in nearly a doubling of baseline levels 59 , 60 , and rat endplate chondrocytes increase ENPP mRNA following TGF-β1 treatment 61 . There appears to be a complicated tissue- and factor-specific influence on ANKH and ENPP expression modulation.…”

Effect of fibronectin, FGF-2, and BMP4 in the stemness maintenance of BMSCs and the metabolic and proteomic cues involved

“…Multiple levels of cell-autonomous and extrinsic signals tightly control fate determination of MSCs [ 11 , 16 – 19 ]. It is possible that extrinsic factors derived from the local niche and systemic environment shape the epigenetic landscape of stem cells, which influences the gene expressions of stemness markers to dictate cell fate [ 20 – 26 ]. Previous studies revealed that designing artificial niches through adding extracellular matrix (ECM) proteins or growth factors in the ex vivo culture condition proved to be an effective way to maintain the potential capability of BMSCs [ 7 , 26 – 28 ].…”

Promoting Effect of Basic Fibroblast Growth Factor in Synovial Mesenchymal Stem Cell-Based Cartilage Regeneration

“…This may be due to the timing of evaluation, because SOX9 gene expression was assessed at 28 days after bFGF stimulation, which may be too late to detect SOX9 upregulation by bFGF. On the other hand, bFGF controlled the maintenance of stemness in MSCs [ 29 , 30 ], and suppressed the cellular senescence of MSCs by downregulating transforming growth factor β2 (TGF-β2) [ 31 ]. Cellular senescence downregulates the therapeutic potential of human MSCs by reducing their migratory, homing, and immunoregulatory abilities in vivo [ 32 , 33 ].…”