Mechanical regulation of cytoskeleton guides beta-catenin effect on MSC fate

细胞骨架的机械调节引导β-连环蛋白对MSC命运的影响

• 批准号: 9042946
• 负责人: Janet E Rubin
• 金额: $ 33.44万
• 依托单位: UNIV OF NORTH CAROLINA CHAPEL HILL
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-04-01 至 2020-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9042946
• 关键词: Actins; Adhesions; Adipocytes; Adipose tissue; Adult; Bone Formation Stimulation; Bone Marrow; Cell Lineage; Cell Nucleus; Cells; Cytoskeletal Modeling; Cytoskeleton; Data; Exercise; F-Actin; Fatty acid glycerol esters; Focal Adhesions; Goals; Grant; Health; Histologic; In Vitro; Knock-out; Knockout Mice; Lamins; Marrow; Measures; Mechanics; Mesenchymal Stem Cells; Methods; Mus; Nuclear; Obesity; Osteogenesis; Pathway interactions; Phosphotransferases; Physicians; Physiological; Population; Proteins; Regulation; Regulatory Pathway; Resistance; Role; Running; Scientist; Signal Pathway; Signal Transduction; Stem cells; Stress Fibers; Structure; Tamoxifen; Testing; Time; Work; base; beta catenin; bone; bone health; bone strength; feeding; improved; in vivo; innovation; lipid biosynthesis; mouse model; osteogenic; osteoprogenitor cell; prevent; response; sedentary; skeletal; synergism; trafficking

 DESCRIPTION (provided by applicant): Exercise builds skeletal strength by inducing new bone formation, which is accompanied by decreased marrow adipose tissue (MAT). A lack of exercise enhances MSC adipogenesis, and increased MAT has been shown to be associated with bone fragility. Understanding mechanisms by which exercise regulates mdMSC lineage allocation to increase osteogenesis and decrease adipogenesis is thus significant for managing skeletal health in an increasingly sedentary population. We have shown that mechanical activation and nuclear trafficking of ßcatenin prevents adipogenesis and promotes MSC entry into osteoprogenitor lineage. ßcatenin signaling is initiated by recruitment of Fyn kinase to foca adhesions during strain, followed by Fyn activation of the mTORC2->Akt->GSK3ß inhibition cascade leading to increased ßcatenin. ßcatenin effects depend on cellular context: in mdMSC, increased cytoskeletal structure improves osteogenesis and cytoskeletal deficiency promotes adipogenesis. The Fyn/mTORC2 proximal signal also activates RhoA to build complexity and connectivity of the cytoskeleton. We propose that mechanically induced cytoskeletal reorganization promotes nuclear entry of ßcatenin, underwriting its anti-adipogenic, pro-osteogenic effect. Importantly, the role of focal adhesion-based Fyn in initiating ßcatenin and RhoA effectors may be unique to the mdMSC, potentially providing a specific pharmacologic target to enhance MSC response to exercise. To test this, we will delineate the synergistic contributions of Fyn/mTORC2's two downstream effectors, RhoA and ßcatenin, to MSC lineage fate allocation in vitro and in vivo. We will find if RhoA generated cytoskeletal reorganization is necessary or sufficient to alter mdMSC cell fate and define how mechanical load activates RhoA (SA1). In SA2, we ask if activation of ßcatenin can alter cell fate in the face of cytoskeletal deficiency, inquiring whether the cytoskeleton drives nuclear/cytoplasmic partitioning of ßcatenin and considering if the actin-nesprin-LINC-nuclear tether might guide ßcatenin into the nucleus. In SA3, using an innovative method to quantify and localize MAT, combined with measures of bone formation, we will investigate mechanical control of mdMSC lineage in vivo, asking if the Fyn/mTORC2 signaling unique to mdMSC is critical during exercise stimulation of bone formation; for this we will generate a Tamoxifen driven Prx1-Cre knock out of Fyn.

 描述(由申请人提供)：运动通过诱导新骨形成来增强骨骼力量，并伴随着骨髓脂肪组织(MAT)的减少。缺乏运动可以促进MSC的脂肪生成，MAT的增加已被证明与骨脆性有关。因此，了解运动调节mdMSC谱系分配以增加成骨和减少脂肪生成的机制，对于在日益久坐的人群中管理骨骼健康具有重要意义。我们已经证明，连接蛋白的机械激活和核运输阻止脂肪形成，并促进MSC进入骨祖细胞谱系。在应变过程中，Fyn激酶被重新招募到FocA粘连上，随后Fyn激活mTORC2-&gt；Akt-&gt；GSK3？抑制级联，导致？catenin增加。？连环蛋白效应取决于细胞环境：在mdMSC中，细胞骨架结构增加促进成骨，细胞骨架缺乏促进脂肪生成。Fyn/mTORC2近端信号也激活RhoA，建立细胞骨架的复杂性和连接性。我们认为，机械诱导的细胞骨架重组促进了çcatenin的核进入，从而保证了其抗脂肪生成和促进成骨的作用。重要的是，基于粘着斑点的Fyn在启动？连环蛋白和RhoA效应器中的作用可能是mdMSC所独有的，可能为增强MSC对运动的反应提供了一个特定的药理靶点。为了验证这一点，我们将描绘Fyn/mTORC2‘S两个下游效应因子RhoA和çcatenin在体外和体内MSC谱系命运分配中的协同作用。我们将发现RhoA产生的细胞骨架重组是否 改变mdMSC细胞命运和确定机械负荷如何激活RhoA(SA1)所必需的或足够的。在SA2中，我们询问在细胞骨架缺乏的情况下，？连接素的激活是否可以改变细胞的命运，询问细胞骨架是否驱动？连接素的核/胞质分配 并考虑肌动蛋白-奈斯普林-LINC-核绳是否可以引导？连环蛋白进入细胞核。在SA3中，使用一种创新的方法来量化和定位MAT，结合骨形成的测量，我们将研究体内mdMSC谱系的机械控制，询问mdMSC特有的Fyn/mTORC2信号是否在运动刺激成骨过程中起关键作用；为此，我们将产生三苯氧胺驱动的PRX1-Cre敲除Fyn。