The BMP and FGF pathways reciprocally regulate odontoblast differentiation

Previous studies demonstrated that the exposure of primary dental pulp (DP) cultures to fibroblast growth factor 2 (FGF2) between days 3–7 exerted significant and long-lasting stimulatory effects on odontoblast differentiation and <i>Dspp</i> expression. These effects involved the increased expression of components of bone morphogenetic protein (BMP) signaling and were reverted by a BMP inhibitor noggin. FGF2 also transiently stimulated osteoblast differentiation and the expression of <i>Ibsp</i> and <i>Dmp1</i>. The present study aimed to further explore interactions between BMP and FGF signaling during odontoblast and osteoblast differentiation in DP cultures. Cultures were established using DP tissue isolated from non-transgenic and fluorescent reporter (DSPP-Cerulean, BSP-GFP, and DMP1-mCherry) transgenic mice and exposed to BMP2, FGF2, SU5402 (an FGF receptor inhibitor), and noggin between days 3–7. Mineralization, gene expression, fluorescent protein expression, and odontoblast formation were examined using xylenol orange, quantitative PCR, fluorometric analysis, and immunocytochemistry, respectively. BMP2 activated SMAD1/5/8 but not ERK1/2 signaling, whereas FGF2 exerted opposite effects. BMP2 did not affect mineralization, the expression of <i>Ibsp</i> and <i>Dmp1</i>, and the percentage of DSPP-Cerulean+ odontoblasts but significantly increased <i>Dspp</i> and DSPP-Cerulean. In cultures exposed to BMP2 and FGF2, respectively, both SU5402 and noggin led to long-lasting decreases in <i>Dspp</i> and DSPP-Cerulean and transient decreases in <i>Dmp1</i> and DMP1-mCherry without affecting <i>Ibsp</i> and BSP-GFP. BMP2 and FGF2 exerted reciprocal stimulatory effects on odontoblast differentiation, whereas their effects on osteoblast differentiation were mediated independently. These data will further elucidate the perspectives of using BMP2 and FGF2 for dentin regeneration/repair.

既往研究表明，在第3至7天期间将原代牙髓（dental pulp, DP）培养物暴露于成纤维细胞生长因子2（fibroblast growth factor 2, FGF2），可对成牙本质细胞分化及<i>Dspp</i>的表达产生显著且持久的促进作用。该效应涉及骨形态发生蛋白（bone morphogenetic protein, BMP）信号通路组分的表达上调，且可被BMP抑制剂头蛋白（noggin）逆转。FGF2还可短暂促进成骨细胞分化以及<i>Ibsp</i>和<i>Dmp1</i>的表达。本研究旨在进一步探究DP培养物中，成牙本质细胞与成骨细胞分化过程中BMP与FGF信号通路的相互作用。本研究通过从非转基因及携带荧光报告基因（DSPP-Cerulean、BSP-GFP与DMP1-mCherry）的转基因小鼠中分离的DP组织构建培养体系，并在第3至7天期间将其暴露于BMP2、FGF2、SU5402（一种FGF受体抑制剂）及头蛋白。分别通过二甲酚橙染色法、定量PCR、荧光光度分析及免疫细胞化学技术检测矿化水平、基因表达、荧光蛋白表达与成牙本质细胞形成情况。结果显示，BMP2可激活SMAD1/5/8信号通路，但无法激活ERK1/2信号通路；而FGF2则呈现相反的调控效应。BMP2不会影响矿化水平、<i>Ibsp</i>与<i>Dmp1</i>的表达以及DSPP-Cerulean阳性成牙本质细胞的占比，但可显著上调<i>Dspp</i>的表达及DSPP-Cerulean的荧光信号。在分别暴露于BMP2与FGF2的培养体系中，SU5402与头蛋白均可使<i>Dspp</i>的表达及DSPP-Cerulean信号产生持久下调，同时使<i>Dmp1</i>的表达及DMP1-mCherry信号出现短暂下调，但不会影响<i>Ibsp</i>的表达与BSP-GFP的荧光信号。BMP2与FGF2对成牙本质细胞分化存在协同促进效应，而二者对成骨细胞分化的调控则相互独立。本研究结果将进一步阐明利用BMP2与FGF2进行牙本质再生/修复的应用前景。