Connexin 43在SOST基因调控假体周围骨溶解与骨重建中的机制研究

批准号:
81874008
项目类别:
面上项目
资助金额:
55.0 万元
负责人:
徐炜
依托单位:
学科分类:
H0605.骨、关节、软组织移植与重建
结题年份:
2022
批准年份:
2018
项目状态:
已结题
项目参与者:
佘昶、方菁嶷、李光飞、谢晔、林苏滨、郏新宇、张载航
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中文摘要
我们研究发现骨细胞和SOST基因参与了假体周围骨溶解过程,抑制SOST表达,减少了骨溶解。同时发现缝隙连接蛋白connexin43的表达随SOST表达同步增高,但机制不明确。connexin43是骨细胞向成骨和破骨细胞传递信息的蛋白通道,我们猜想connexin43同时作为调控蛋白参与磨损颗粒诱导的骨溶解,并影响SOST信号的传递。拟通过:1)构建connexin43敲除的小鼠骨溶解模型,观察SOST表达、骨形成和骨吸收的变化。2)磨损颗粒孵育骨细胞,离体观察靶向干预connexin43对SOST表达的影响,以及干预SOST引起connexin43的变化。3)骨细胞分别与成骨和破骨细胞共培养,connexin43阻断前后,检测骨细胞SOST基因对成骨、破骨细胞功能和OPG/RANKL信号通路影响。以此探索connexin43参与SOST调控骨溶解的机制,为假体松动的靶向干预提供理论依据。
英文摘要
Our latest study showed that osteocyte and SOST, its characteristic gene, were associated with periprosthetic osteolysis. Depression of SOST expression reduced the severity of Ti particle-induced osteolysis. And our research also found the expression of connexin 43 went up with increase of SOST expression. But the mechanism is still unknown. We know that gap junctions are intercellular channels which permit the direct cell-to-cell communication of signals and connexin 43 is the most abundant gap junction in bone. Some research showed osteocyte coupling osteoblast and osteoclast is mediated by connexin 43. Based on our previous study and the latest results, we have a hypothesis that connexin 43 plays an important role in the process of periprosthetic osteolysis and bone remodeling, and affects SOST signal transmission as a regulator protein of signal pathway. To test the hypothesis, we will plan to build the osteolysis model of connexin 43 cKO mouse and detect the SOST expression. Bone resorption and bone formation will be assessed by using histomorphology, Micro-CT, molecular biotechology, mechanical pull-out test and so on. Osteocytes with connexin 43 down-regulation or up-regulation through the virus transfection will be cultivated with different concentration of particulates titanium alloy. cell proliferation, apoptosis, gene and protein expression of the characteristic marker, OPG/RANKL and inflammatory cytokine will be evaluated or measured by Brdu assay, flow cytometry, real-time PCR, Western Blot and Elisa kit respectively. Furthermore, osteocyte with knockdown of connexin 43 will be co-cultured with osteoblast and osteoclast respectively. Then the characteristic marker will be detected to evaluate the anabolic effect of osteoblast and the catabolic effect of osteoclast. The effect of connexin 43 on OPG/RANKL signaling pathways also will be examined. At last, after the protein products of SOST being added, we will evaluate the effects of osteocyte with the depression of connexin 43 on osteoblast and osteoclast again..Through a series of experiments, we want to confirm our hypothesis and explict the role of connexin 43 on osteocyte regulating osteoblast and osteoclast in the process of periprosthetic osteolysis. Therefore the research may provide a scientific evident for connexin 43 as a target for prevention and cure of periprosthetic osteolysis and joint loosening.
本研究通过小鼠骨溶解模型的构建,钛颗粒(Ti)干预骨细胞,骨细胞-成骨细胞和骨细胞-破骨细胞共培养等体外模型的构建,以及基因敲除等方式,研究了Ti颗粒干预下,骨细胞Cx43的表达变化以及其对成骨,破骨分化的影响,探索骨细胞及connexin43(Cx43)在假体周围骨溶解中的作用机制。首先在骨溶解模型中,颅骨和股骨内的骨细胞Cx43在Ti颗粒的干预下表达显著增高;Ti颗粒干预骨细胞株MLO-Y4,Cx43蛋白表达增加,β-catenin表达减少,wnt信号通路被抑制,证实了骨溶解过程中骨细胞Cx43的变化规律。进而通过构建Cx43条件性基因敲除鼠和Cx43过表达和低表达的骨细胞株进一步研究。成骨方面,Cx43基因敲除使骨溶解部分缓解,成骨标记物表达水平增高。Ti颗粒干预下,将骨细胞与成骨细胞共培养。发现Ti颗粒引起骨细胞Cx43表达增加,与之接触的成骨细胞Cx43表达增加,β-catenin被限制在成骨细胞膜下,wnt信号通路靶蛋白Runx-2减少,抑制了成骨细胞的分化。Cx43敲低后,β-catenin表达增加,成骨细胞成骨标记物表达水平增高,成骨细胞受Ti颗粒的抑制作用部分被缓解。破骨方面, Cx43敲除后,小鼠外周血中RANKL/OPG,CTX的含量较野生鼠明显下降;来源于股骨的破骨标志物表达下降。43低表达的骨细胞,在钛颗粒干预下所获得培养上清,刺激破骨细胞的分化,培养基中的RANKL/OPG下降,破骨细胞的分化指标下降,Cx43低表达后缓解Ti颗粒对破骨的活化。通过一系列实验证明骨细胞及其Cx43参与了假体周围骨溶解过程,通过负向调节wnt信号通路抑制骨形成和加速骨吸收,为假体周围骨溶解机制的研究提供新的思路,为将Cx43作为防治人工关节无菌松动新的靶点提供科学依据。
期刊论文列表
专著列表
科研奖励列表
会议论文列表
专利列表
0.5-Gy X-ray irradiation induces reorganization of cytoskeleton and differentiation of osteoblasts
0.5Gy X射线照射诱导细胞骨架重组和成骨细胞分化
DOI:10.3892/mmr.2021.12018
发表时间:2021-05-01
期刊:MOLECULAR MEDICINE REPORTS
影响因子:3.4
作者:Huang,Qun;Chai,Hao;Xu,Wei
通讯作者:Xu,Wei
Mid-term study on the effects of arthroscopic discoid lateral meniscus plasty on patellofemoral joint: An observational study.
关节镜下盘状外侧半月板成形术对髌股关节影响的中期观察性研究
DOI:10.1097/md.0000000000031760
发表时间:2022-11-11
期刊:Medicine
影响因子:1.6
作者:
通讯作者:
First dorsal metacarpal artery flap with dorsal digital nerve with or without dorsal branch of the proper digital nerve produces comparable short-term sensory outcomes.
第一掌背动脉 fap 与指背神经(带或不带适当指神经的背支)产生可比较的短期感觉结果
DOI:10.1186/s13018-021-02838-z
发表时间:2021-11-18
期刊:Journal of orthopaedic surgery and research
影响因子:2.6
作者:Feng SM;Zhao JJ;Migliorini F;Maffulli N;Xu W
通讯作者:Xu W
The Study of Biomechanics and Clinical Anatomy on a Novel Plate Designed for Posterolateral Tibial Plateau Fractures via Anterolateral Approach.
新型前外侧入路胫骨平台后外侧骨折钢板的生物力学和临床解剖学研究
DOI:10.3389/fbioe.2022.818610
发表时间:2022
期刊:Frontiers in bioengineering and biotechnology
影响因子:5.7
作者:Ren W;Zhang W;Jiang S;Peng J;She C;Li L;Mao Y;Zhou H;Xu W
通讯作者:Xu W
Reduction of SOST gene promotes bone formation through the Wnt/β‐catenin signalling pathway and compensates particle‐induced osteolysis
SOST 基因的减少通过 Wnt/β-catenin 信号通路促进骨形成并补偿颗粒诱导的骨溶解
DOI:10.1111/jcmm.15084
发表时间:2020-03
期刊:Journal of Cellular and Molecular Medicine
影响因子:5.3
作者:张载航;郏新宇;方菁嶷;柴灏;黄群;佘昶;贾鹏;耿德春;徐炜
通讯作者:徐炜
SOST/sclerostin在骨细胞参与假体周围骨溶解过程中的作用和机制研究
- 批准号:81472105
- 项目类别:面上项目
- 资助金额:72.0万元
- 批准年份:2014
- 负责人:徐炜
- 依托单位:
国内基金
海外基金
