碱性成纤维细胞因子在人牙髓损伤修复过程中的表达

Expression of basic fibroblast growth factor in the progress of repairing pulp injury

目的研究不同状态人牙髓组织中碱性成纤维细胞因子(b FGF)的表达特点,以及大肠杆菌脂多糖(LPS)刺激后人牙髓细胞(h DPC)中b FGF的表达水平,探讨b FGF在牙髓损伤修复过程中的可能作用。方法采用实时荧光定量聚合酶链反应(PCR)和免疫蛋白印迹方法(Western blot)分别检测正常、深龋及牙髓炎牙髓组织中b FGF m RNA和蛋白表达情况。实时荧光定量PCR测定1 mg/L LPS刺激h DPC 6、12、24、48 h后b FGF和热休克蛋白70(HSP70)表达水平的变化;Western blot和细胞免疫荧光染色检测LPS刺激h DPC后b FGF蛋白表达变化。结果实时荧光定量PCR和Western blot结果表明,深龋牙髓组织中b FGF水平显著上调,而正常和牙髓炎牙髓组织中b FGF表达无差异。实时荧光定量PCR检测到LPS刺激h DPC后,b FGF和HSP70 m RNA水平同步上调,在12 h达峰值;Western blot显示,LPS刺激h DPCs 12、24、48 h后b FGF蛋白表达水平均高于正常h DPC;细胞免疫荧光染色证实,LPS刺激12 h后h DPC中b FGF呈强阳性表达,而正常h DPC中b FGF呈弱阳性表达。结论 b FGF在深龋牙髓组织中高表达,且LPS刺激早期可上调h DPC内b FGF表达,推测b FGF可能参与牙髓组织防御修复反应,可能是细胞抗损伤的重要调节机制之一。

Objective To investigate the expression of basic fibroblast growth factor（b FGF） in human dental pulp tissues of different state and the b FGF level in human dental pulp cells（h DPCs）stimulated by lipopolysaccharide（LPS）, and explore the potential role of b FGF in the progress of repairing pulp injury. Methods The messenger RNA and protein level of b FGF in normal and inflammatory pulp were detected by fluorescence quantitative-polymerase chain reaction（q PCR） and western blot. HDPCs were collected after stimulated by 1 mg / L LPS for 6, 12, 24 and 48 h. The expression of b FGF and HSP70 in h DPCs was evaluated by q PCR. In addition, the protein level of b FGF in in normal and LPS treated h DPCs were detected by western blot and immunofluorescence staining. Results The m RNA and protein level of b FGF was significantly up-regulated in the pulp with deep caries compared with healthy pulps, while inflamed dental pulp did not show significant difference.QPCR showed that b FGF and HSP70 m RNA were concomitantly increased from 0 h to 12 h after stimulated by LPS. Similarly, it was shown that b FGF was increased in LPS treated h DPCs compared with normal h DPCs by western blot. Moreover, immunofluorescence staining results demonstrated that b FGF was strongly positive stained in LPS treated h DPCs, while it was weakly expressed in normal h DPCs. Conclusion BFGF is up-regulated in pulp with deep caries and h DPCs induced by LPS,indicating that b FGF may participates in the progress of repairing pulp injury.