NMDA型谷氨酸受体在骨祖细胞和成骨细胞表达并介导力学信号转导

Identification of NMDA Glutamate Receptors Expressed in Bone Precursor Cell and Osteoblastic Cells and it's Involvement in Mechanotransduction

目的:鉴定骨髓间充质干细胞D1和成骨细胞MC3T3-E1、MC3T3-E1亚克隆14是否表达NMDA型谷氨酸受体(NMDAR),并初步探讨NMDAR是否参与力学信号转导。方法:采用RT-PCR技术、免疫荧光染色技术、蛋白免疫杂交技术和体外细胞循环拉伸装置,鉴定了上述3个细胞系中NMDAR的表达情况,并初步探讨了在MC3T3-E1亚克隆14细胞系中,NMDAR在力学信号转导中的可能作用。结果:3个细胞系均表达NMDA受体亚基1(NR1)和不同的亚基2(NR2A-NR2D)。细胞微丝骨架的破坏并没有阻断力学应变诱导的c-jun、c-fos的表达上调;而阻断NMDAR后,却抑制了力学应变诱导的c-jun、c-fos和成骨细胞特异的转录因子Cbfa1/Runx2的表达上调。结论:NMDAR在骨中表达并发挥功能,参与了骨的力学信号转导过程。

Objective： To identify the transcription of particular subunits of ionotropie NMDA glutamate reeeptors（NMDAR） in mouse bone marrow stromal precursor cell line D1, osteoblastie cell MC3T3-E1 and MC3T3-E1 subelone 14, investigate the involved glutamate signaling underlying the potential mechanisms of meehanotransduetion. Methods： Immunofluorescence and western blotting were employed to identify the NMDAR subunit 1 in above three cell lines, RT-PCR was applied to discern the NMDAR subunits 2 and discriminate the pattern of AP-1 components and Cbfal/Runx2 response to cyclic strain under different conditions. Results： The NMDAR subunit 1 and different types of subunit 2 were identified in these cell lines, the upregnlation of AP-1 components c-jun and e-los induced by cyclic strain was not blocked by the application of the eytoskeleton disrupter eytochalasin D, while the upregnlation of e-jun, e-los and bone specific transcription factor Cbfal/Runx2 induced by cyclic strain were inhibited by the NMDAR noncompetitive blocker MK801. Conclusion： These results demonstrated the expression of NMDAR in these three cells, suggested that the NMDAR may be functional in these cells that as in vitro models to investigate the involved glutamate-signalling underlying the potential meehanisms of mechanotransduction.