【摘要】 目的:基于泪管-耳-齿-指(LADD)综合征的临床线索，探讨成纤维细胞生长因子受体(FGFR)3对人牙髓干细胞(hDPSCs)生物学行为及牙根发育的调控作用。方法:选取C57BL/6小鼠随机分为注射DMSO的对照组和泛FGFR抑制剂NVP-BGJ398的实验组，通过Micro-CT观察牙根发育表型。利用siRNA技术特异性沉默hDPSCs的FGFR3基因，通过CCK-8法及克隆形成实验评估其增殖/自我更新能力，利用Western blot检测其干性标志物表达，通过实时荧光定量PCR检测其成牙/成骨相关基因的动态变化，通过碱性磷酸酶(ALP)染色及茜素红S染色(ARS)评估其体外矿化功能的完整性，通过Western blot检测其细胞外信号调节激酶(ERK)信号通路的变化。结果:体内实验显示，阻断FGFR信号后，小鼠磨牙牙根长度短于对照组(P<0.05)。体外实验显示，特异性沉默FGFR3后，hDPSCs增殖活性受抑，干性标志物表达下调；si-FGFR3组成骨诱导早期ALP mRNA水平代偿性升高(P<0.05),但Runt相关转录因子2(RUNX2)、骨桥蛋白(OPN)、牙本质涎磷蛋白(DSPP)未见明显上调(P>0.05);成骨诱导后期，ALP活性降低(P<0.05),ARS矿化结节形成显著减少(P<0.05),细胞内磷酸化ERK(p-ERK)水平降低(P<0.05)。结论:泛FGFR信号阻断可导致小鼠牙根短小。FGFR3沉默与hDPSCs增殖和干性维持受损、成牙/成骨分化受损以及ERK通路活性下降相关，提示FGFR3可能通过调控hDPSCs的自我更新与分化过程参与牙根发育。更多还原

【Abstract】 Objective: Based on clinical clues from lacrimo-auriculo-dento-digital(LADD) syndrome, this study aimed to investigate the role of fibroblast growth factor receptor 3(FGFR3) in regulating the biological behavior of human dental pulp stem cells(hDPSCs) and tooth root development. Methods: C57BL/6 mice were locally injected with the pan-FGFR inhibitor NVP-BGJ398 in the mandible to establish an FGFR-blockade model, and root phenotypes were evaluated by Micro-CT. FGFR3 in hDPSCs was specifically silenced by siRNA. Cell proliferation and self-renewal were assessed by CCK-8 and colony-formation assays. Stemness markers were examined by Western blot. Dynamic changes in odontogenic/osteogenic marker genes were examined by real-time fluorescent quantitative PCR; alkaline phosphatase(ALP) staining and alizarin red S(ARS) staining were performed to evaluate the integrity of in vitro mineralization capacity. ERK signaling was analyzed by Western blot. Results: In vivo, NVP-BGJ398-treated mice showed shorter molar roots than controls(P<0.05). FGFR3 knockdown inhibited hDPSC proliferation and reduced stemness marker expression. In the si-FGFR3 group, ALP mRNA showed an early compensatory increase(P<0.05), but no significant upregulation was observed for Runt-related transcription factor 2(RUNX2), osteopontin(OPN), and dentin sialophosphoprotein(DSPP)(P>0.05). In the late stage of osteogenic induction, ALP activity decreased(P<0.05), ARS mineralized nodule formation was significantly reduced(P<0.05), and intracellular phosphorylated ERK(p-ERK) levels were decreased(P<0.05). Conclusions: Pan-FGFR signaling blockade can lead to short tooth roots in mice. FGFR3 silencing is associated with impaired proliferation and stemness maintenance, compromised odontogenic/osteogenic differentiation, and reduced ERK pathway activity in hDPSCs, suggesting that FGFR3 may participate in tooth root development by regulating hDPSC self-renewal and differentiation.更多还原