The Effect And Mechanism Of BFGF Affect HDPSCs Mineralization And Its Signaling

The stem cells in human oral tissue are mainly comprised of dental pulpstem cells（hDPSCs）, Periodontal LigamentCells and Stem cells from the apicalpapilla. hDPSCs are characterized by highly proliferation and multipledifferentiation proficiency. Nowadays, hDPSCs is a hot spot in the field of toothregeneration research. It not only can differentiate into odontoblast, neurocyte,vascular endothelial cell and adipocyte, but also can form pulp-dentin complex.Therefore, the culturing of hDPSCs in vitro can be used as an model to researchon dental pulp regeneration. And it has great significance in tooth regenerationresearch.Many research shows that Basic Fibroblast Growth Factor (bFGF) canimprove bone regeneration in vivo research. However, there are many debateson how it affects stem cell bone differentiation in vitro. Similar debates alsoexist in hDPSCs. Some research shows that bFGF can improve the activity ofalkalinephosphatase (ALP). Some shows that bFGF can inhibit the odontoblastdifferentiation of hDPSCs. AS we all known, the formation ofreparative dentin is an important mechanism in pulp-dentin complex repair.Finding out how bFGF affect odontoblastdifferentiation of hDPSCs is verysignificant in tooth regeneration research.Our research use hDPSCs as the seed cell, refine the culture condition andbFGF stimulation in order to figure out how how bFGF affectodontoblastdifferentiation of hDPSCs in vitro. We choose MAPKs and researchon how the signal mechanism of MAPKs regulates bFGF stimulation onhDPSCs. Our research will provide evidence to the mechanism of bFGF affectodontoblastdifferentiation of hDPSCs s and bring out new thoughts and methodson tooth regeneration research.The results are as follows:1. Separation, culture and identification of hDPSCsWe use enzymic digestion to separate hDPSCs from dental pulp tissue. Wepick up single clone by limiting dilution. Flow cytometry andimmunocytochemistry are used to identify the Surface markers of hDPSCs. Andour cells are identified as hDPSCs. We induce our cells to differentiate intoodontoblast and adipocyte which prove our cells have multiple differentiatepotential.2. How bFGF affect mineralization ability of hDPSCs in vitroWe refine the culture condition and bFGF stimulation of hDPSCs in order tofind out how bFGF affect hDPSCs mineralization ability. Our result shows thatbFGF can inhibit the mineralization ability of hDPSCs in the mineralizationperiod. In the proliferation period, how long the stimulation last determines theeffect of bFGF. One week stimulation of bFGF will improve mineralizationability of hDPSCs, however two weeks stimulation of bFGF will inhibit mineralization ability of hDPSCs. And the effect of bFGF stimulation can passto the next generation by subcultivition.3. How MAPK pathway regulate bFGF stimulation on hDPSCsbFGF stimulation can rapidly activate MAPKs pathway, especially ERKpathway. After we inhibit ERK, JNK and P38, bFGF can not activate JNK andP38, but ERK can still be activated, however the phosphorylationlevel of ERKis lower than the group without inhibition.bFGF stimulation can improve theexpression of stemness gene: Nanog, Oct4, Sox2.After we inhibit JNK and P38,the expression of Nanog, Oct4and Sox2are down regulated. However, after weinhibit ERK, the expression of Nanog, Oct4are down regulated, but theexpression of Sox2is up regulated. The results indicate that other pathwayinvolved in the regulation of bFGF stimulation besides MAPKs pathway.In conclusion, bFGF has essential effect on the mineralization of hDPSCs.In different period of hDPSCs, the bFGF stimulation will have different effect.bFGF can activate MAPK pathway, but there are other pathway regulate bFGFstimulation besides MAPK pathway. Our results provide evidence to theresearch on hDPSCs differentiationand bring out new thoughts and methods ontooth regeneration research.