ERK5信号通路介导流体剪切力对成骨细胞OPG、RANKL表达影响的研究

发布时间：2018-03-23 18:02

本文选题：流体剪切力　切入点：MC3T3-E1成骨细胞　出处：《兰州大学》2014年硕士论文

【摘要】：目的：随着中国逐步进入老龄化社会,老年人的健康问题成为全社会的一大焦点,骨质疏松由于其发病率高而成为一个公共卫生问题,目前主要是通过药物以及运动完成骨质疏松的治疗,但由于目前对于骨骼感受应力的机制不是很明确,导致临床医生无法正确地指导患者进行合理的运动促进康复,因此本研究应用自制的体外流体剪切力(Fluid shear stress, FSS)加载装置用生理强度的流体剪切力加载成骨细胞(MC3T3-E1),明确ERK5信号通路的传导机制,以及对成骨有很重要作用的两个因子骨保护素(osteoprotegerin, OPG)和破骨细胞分化因子(Osteoclast differentiation factor, ODF),又称细胞核因子κB受体活化因子配体(receptor activator of NF-κB ligand, RANKL)的表达情况。以期能够为临床骨质减少性疾病患者正确合理的运动提供理论依据。方法：将ERK5特异性阻断剂BIX02188配制成不同浓度,并干预MC3T3-E1成骨细胞,后用MTT法检测490nm OD值,观察成骨细胞的增殖状态,并运用荧光定量PCR检测ERK5mRNA的表达情况。用生理强度为12dyne/cm2的流体剪切力加载成骨细胞,后检测OPG、RANKL mRNA的表达情况。先用之前实验得出的浓度BIX02188干预成骨细胞,之后再给成骨细胞加载生理强度为12dyne/cm的流体剪切力,检测OPG、RANKL mRNA的表达情况。结果：流体剪切力促进成骨细胞的增殖；浓度为15μM的ERK5特异阻断剂BIX02188能够有效的抑制ERK5mRNA的表达；MTT结果证实：ERK5特异性阻断剂阻断ERK5活化后,成骨细胞增殖受到明显抑制；强度为12dyne/cm的流体剪切力能够促进成骨细胞OPG mRNA表达(P0.05),降低RANKL mRNA表达(P0.05)；当BIX02188干预成骨细胞后,流体剪切力对成骨细胞OPG、RANKL mRNA表达的影响明显减弱(P0.05)。结论：生理强度为12dyne/cm2的流体剪切力能促进成骨细胞增殖,对OPG mRNA的表达有促进作用,对RANKL mRNA的表达有抑制作用；ERK5能够有效介导流体剪切力对成骨细胞OPG. RANKL mRNA的表达。
[Abstract]:Objective: as China has gradually entered an aging society, the health of the elderly has become a major focus in the whole society, and osteoporosis has become a public health problem due to its high incidence. At present, the treatment of osteoporosis is mainly done through medicine and exercise. However, because the mechanism of bone stress is not very clear at present, the clinicians are unable to correctly guide the patients to carry out reasonable exercise to promote rehabilitation. Therefore, in order to clarify the transduction mechanism of ERK5 signaling pathway, a self-made in vitro fluid shear stress (FSS) loading device was used to load osteoblasts MC3T3-E1 with physiological strength of fluid shear force. And the expression of osteoprotegerin (OPG) and osteoclast differentiation factor (Osteoclast differentiation factor), also known as nuclear factor 魏 B receptor activating factor receptor activator of NF- 魏 B ligand (RANKL). The correct and reasonable exercise in patients with osteopenia disease provides theoretical basis. Methods: BIX02188, a specific inhibitor of ERK5, was prepared into different concentrations, and MC3T3-E1 osteoblasts were interfered with. The OD value of 490nm was detected by MTT method, and the proliferation of osteoblasts was observed. Fluorescence quantitative PCR was used to detect the expression of ERK5mRNA. Osteoblasts were loaded with fluid shear force with physiological strength of 12dyne/cm2, and then the expression of RANKL mRNA was detected. Firstly, the concentration of BIX02188 was used to interfere with osteoblasts. Then the osteoblasts were subjected to fluid shear stress with physiological strength of 12dyne/cm to detect the expression of OPGG RANKL mRNA. Results: the proliferation of osteoblasts was promoted by fluid shear stress, and the proliferation of osteoblasts was inhibited by BIX02188, a specific inhibitor of ERK5 at concentration of 15 渭 M, which could effectively inhibit the expression of ERK5mRNA. The results showed that the proliferation of osteoblasts was significantly inhibited after the activation of ERK5 was blocked by the specific blocker of 1: ERK5. The fluid shear stress with 12dyne/cm strength could promote the expression of OPG mRNA and decrease the expression of RANKL mRNA in osteoblasts, but the effect of fluid shear stress on the expression of mRNA in osteoblasts was significantly weakened after BIX02188 intervention. Conclusion: fluid shear stress with physiological strength of 12dyne/cm2 can promote the proliferation of osteoblasts, promote the expression of OPG mRNA, and inhibit the expression of RANKL mRNA. ERK5 can effectively mediate the expression of osteoblast OPG. RANKL mRNA by fluid shear stress.
【学位授予单位】：兰州大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R318.01

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