Significance of the neuronal guidance molecule Semaphorin 3A for bone remodelling during orthodontic tooth movement

点击翻译按钮获取中文摘要

Malocclusions and jaw misalignments appear with high prevalence in school children and in adults. Besides various cytokines and growth factors neuronal guidance molecules (NGM) gained attention for their roles in bone homeostasis and thus potential roles during tooth movement. Recently we have shown that Ephrin ligands and Eph receptors are involved in bone remodeling during tooth movement. Among the NGMs which have recently been implicated in the regulation of bone remodeling Semaphorin 3A (Sema3A) is the most interesting as it concurrently induces osteogenic differentiation in osteoblasts and disturbs osteoclast differentiation. To continue my analysis of neuronal guidance molecules in orthodontic tooth movement and to elucidate the underlying mechanisms the objectives of this project are (i.) to investigate if different mechanical forces modulate the expression of Sema3A and its receptors Neuropilin1 and PlexinA1 in periodontal fibroblasts and osteoblasts of the alveolar bone, (ii.) to study which biochemical signals are involved in the mechanical modulation of Sema3A (iii.) to investigate the effects of Sema3A stimulation on alveolar bone osteoblasts differentiation and (iv.) to elucidate the molecular mechanisms underlying the Sema3A dependent osteoblast differentiation. To achieve this, I will first stimulate periodontal fibroblasts and alveolar bone osteoblasts mechanically by applying compressive and tension forces. Culture of both cell types, as well as the mechanical stimulation is well established in our laboratory. The expression of Sema3A and its receptors will be analyzed by means of quantitative PCR and western blot. Second, I will investigate the potential mechanoinductive pathway leading to Sema3A alterations. To this end, different MAPKinases are tested for phosphorylation and the involvement of the putative Sema3A transcription factor Osterix will be studied by qPCR and western blot. Third, alveolar osteoblasts will be stimulated by recombinant human Sema3A and its impact on osteogenic differentiation will be analyzed by monitoring the expression of different osteoblast differentiation markers by quantitative PCR. Fourth, the mechanism underlying the Sema3A mediated osteoblast differentiation will be investigated by studying the involvement of the Rac1 GTPase and the beta-catenin transcription factor using pull down assays and immunofluorescent staining and western blot, respectively. Own research on members of the Ephrin/Eph family and preliminary data on Sema3A suggest that the role of neuronal guidance molecules in the regulation of orthodontic tooth movement might have been underestimated. Thus, this project might illuminate additional cellular and molecular events which might be useful as targets for pharmacological manipulations aiming for a side effect free and accelerated orthodontic tooth movement.