初级纤毛在关节软骨中的生物力学效应

生物力学因素对于关节软骨的稳态维持具有至关重要的作用。初级纤毛是一种可同时感受力学信号和化学信号的细胞器,并于软骨细胞膜表面也存在有初级纤毛分布。其与多个信号转导通路相关,共同参与软骨细胞表型维持和物质代谢的过程。同时,初级纤毛的异常也关联到多种人类的骨关节类疾病。主要论述初级纤毛在软骨细胞力学微环境中的作用,以及与其他信号通路的交互作用机制,探讨其与骨关节疾病的联系,以期为骨科临床和基础科研提供一定的科学依据。     

Shh和初级纤毛在腭部发育中的作用

Sonic hedgehog （ Shh）通路在脊椎动物组织和器官发育中扮演重要角色,并且有证据表明其牵涉到包括腭裂等发育缺陷。近年关于Shh及相关因子在发育中腭部的表达模式以及条件性基因敲除小鼠的相关研究着重突出了上皮和间充质细胞间的互动在腭发育中的作用。此外, Hedgehog通路还因其独特的初级纤毛依赖性备受瞩目。文章以Shh在腭部发育中的作用,以及其与初级纤毛的关系为重点来介绍近年来相关发现。     

初级纤毛在肿瘤发生发展中的研究进展

初级纤毛是一种由微管构成的不具有运动功能的细胞器,它参与了细胞的生长发育、内环境稳态、信号转导以及肿瘤的发生和发展。该文就初级纤毛的基本结构、组装与解体,初级纤毛相关的主要信号通路,初级纤毛在肿瘤中的研究进展进行系统性的综述。     

内皮细胞中力学信号的传递与传导

力学作用对血管内皮细胞的生理功能以及相关疾病病机理有着十分重要的影响。因此，力学信号在内皮细胞中的传播方式日益受到关注，并不断有新的有意义的研究结果公诸于世。本文正是基于这些研究结果，从传递和传导两个过程既相互独立、又相互联系的角度予邓评述。     

力学刺激在骨愈合中的作用及机制研究进展

骨组织是一种完全可再生组织,骨愈合的过程可以认为是骨痂中骨组织在生物力学环境下增殖、适应改建的过程.近年来,随着创伤骨科治疗水平的提高与生物力学的不断发展,新型内固定材料的不断发明与进步,骨折愈合机制的分子基础研究不断明确,骨科医生也从早期骨折手术治疗的绝对稳定固定理念转变为符合生物力学理念下的相对稳定固定.而理念的转...     

内皮细胞中力学信号的传递与传导

力学作用对血管内皮细胞的生理功能以及相关疾病的发病机理有着十分重要的影响。因此，力学信号在内皮细胞中的传播方式日益受到关注，并不断有新的有意义的研究结果公诸于世。本文正是基于这些研究结果，从传递和传导两个过程既相互独立、又相互联系的角度予以评述。     

初级纤毛/鞭毛转运系统介导力学反应性信号通路促骨髓基质干细胞成骨分化

背景：目前已证实力学刺激可以促进骨髓基质干细胞成骨分化,但其机制未完全明了。初级纤毛是重要的力学感受器并调控TGF-β1/BMP-2/SMAD等多种信号通路,很可能是骨髓基质干细胞力学调控的重要靶点。目的：探讨流体剪切力对骨髓基质干细胞成骨分化的影响及机制。方法：将大鼠骨髓基质干细胞分为对照组、力学刺激组(通过摇床施加流体剪切力学干预)、力学刺激+IFT88沉默组(力学刺激+使用siRNA沉默IFT88表达),干预24 h后,采用qRT-PCR检测转化生长因子β1、骨形成蛋白2的表达、Western blot检测磷酸化SMAD2/3蛋白的表达,初级纤毛免疫荧光染色及形态学分析。结果与结论：剪切力刺激可促进骨髓基质干细胞的初级纤毛表达,转化生长因子β1及骨形成蛋白2基因转录激活,提高磷酸化SMAD2/3蛋白表达。siRNA干扰初级纤毛生成后,这一力学反应效应明显减低。骨髓基质干细胞的初级纤毛面积改变比值与转化生长因子β1及骨形成蛋白2基因转录增高比例具有Spearman相关性。结果表明：初级纤毛/鞭毛转运系统介导了流体剪切力反应性的TGF-β1/BMP-2/SMAD信号通路激活,促进骨髓...     

基于数字图像处理的初级纤毛定量分析

初级纤毛是一种由微管构成并突起于细胞表面的细胞器。初级纤毛的结构在某些疾病中会发生改变,特别对肿瘤的发生发展具有重要的影响。基于显微影响,分析初级纤毛的形态结构,有助于相关疾病的研究和诊断。传统分析方法主要采用Photoshop软件画线量取其长度指标,因此存在准确度低、重复性差、分析效率低等问题。本研究结合数字图像处理技术,应用色彩分离技术,将细胞与初级纤毛有效分离;并对比分析中值滤波、均值滤波、Sobel、Roberts、Prewitt、Log算子多种算法对初级纤毛形态边缘的保持效果,选取Prewitt、Log算子,改进并提升边界提取效果;再利用最大类间方差法去除杂质;最后通过连通域标记获得每个初级纤毛的周长与面积参数。与传统方法对比,本研究方法测量初级纤毛结构参数的时间由50~80 min减少为2 s,相对误差ω提高了19.08%以上,达到了快速精确测量的目的。     

初级纤毛在白藜芦醇诱导大鼠骨髓基质细胞分化为神经元样细胞中的作用

目的:研究初级纤毛在白藜芦醇诱导大鼠骨髓基质细胞(MSCs)分化为神经元样细胞中的作用.方法:全骨髓贴壁法分离、培养、纯化MSCs.含15 μmol/L白藜芦醇的无血清DMEM/F12诱导MSCs分化.倒置显微镜下观察细胞形态,免疫印迹检测神经元特异性烯醇化酶(NSE)、微管相关蛋白-2(MAP-2)的表达.免疫荧光法检测MSCs增殖情况.扫描电子显微镜检测MSCs表面初级纤毛,免疫荧光法检测初级纤毛蛋白(Ac-Tu)的表达.结果:白藜芦醇诱导后60％的细胞胞体收缩,立体感增强,类似神经元.免疫印迹显示MSCs及对照组细胞有NSE蛋白的轻度表达,诱导后NSE、MAP-2蛋白表达阳性,随着诱导时间延长,NSE、MAP-2的表达渐增强.经过24 h饥饿后,90％的MSCs处于生长静止状态.扫描电子显微镜及免疫荧光显示,处于生长静止期的MSCs具有初级纤毛,且白藜芦醇可诱导具有初级纤毛的MSCs分化为神经元样细胞.对照组则无明显变化.结论:白藜芦醇能诱导MSCs分化为神经元样细胞.在此过程中,初级纤毛可能发挥着重要作用.     

血流切应力对血管内皮细胞PCP信号通路与初级纤毛发生的影响

目的研究不同血流切应力(fluid shear stress, FSS)对内皮细胞平面细胞极性通路(planar cell polarity, PCP)的调节作用,进一步探讨FSS、PCP信号通路以及纤毛发生之间的关系。方法建立可调控FSS的流体动力学细胞培养模型,qPCR以及免疫荧光检测不同FSS作用下PCP信号通路核心蛋白Dvl2及纤毛装配蛋白IFT88的mRNA表达与细胞定位以及两者的共定位,Western bolt (WB)检测不同FSS作用18 h时Dvl2蛋白表达。结果 qPCR结果显示,与1.5 Pa比较,Dvl2 mRNA的表达量在0.1 Pa FSS作用6 h和18 h时均升高(P0.05)、12 h时显著升高(P0.01);IFT88 mRNA表达量在0.1 Pa FSS作用18 h时显著升高(P0.01)。WB结果显示,与0 h比较,Dvl2蛋白的表达在0.1 Pa FSS作用18 h时升高(P0.05),在1.5 Pa FSS作用18 h时显著降低(P0.05);与1.5 Pa FSS比较,Dvl2蛋白的表达在0.1 Pa FSS作用18 h时升高(P0.05)。免疫荧光结果显示,Dvl2蛋白阳性表达随FSS作用时间的增加增多,且逐渐聚集于细胞核周边一点;IFT88蛋白阳性表达在0.1 Pa FSS作用下逐渐由细胞核向细胞质转移并聚集为一点,1.5 Pa FSS作用下逐渐减少且解聚;蛋白Dvl2、IFT88在0.1 Pa FSS作用下均定位于细胞的同一位置,在1.5 Pa FSS作用18 h内均定位于细胞的同一位置,18 h后由于蛋白IFT88发生解聚,未观察到共定位。结论层流FSS作用能够抑制PCP信号通路的转导并阻碍纤毛发生,低FSS促进其转导,且PCP信号通路可能通过Dvl2调控FSS诱导的初级纤毛发生。     

Microfluidic devices for studies of primary cilium mediated cellular response to dynamic flow conditions

We present the first microfabricated microfluidic devices designed specifically for studies of primary cilium mediated cellular response to dynamic flow. The primary cilium functions as a mechano-sensor in renal tubular epithelium, sensing the extracellular fluid flow. Malfunction of cilia has been implicated in e.g. polycystic kidney disease and other pathological conditions. Bending of the primary cilium by fluid flow has been shown to give rise to an intracellular calcium signal, however little is known about the sensitivity to flow duration, magnitude and direction. This paper presents a novel method for studying cilia forming cells in asymmetric microfluidic environments. The microfluidic devices presented here were designed for a dynamic control of the local fluid flow on a cellular level, and thus, enables studies of cellular responses to an amplitude, frequency and direction controlled cilium movement.     

Primary cilium and brain aging: role in neural stem cells,neurodegenerative diseases and glioblastoma

Brain aging is characterized by a progressive loss of tissue integrity and function as a consequence of impaired homeostasis and regeneration capacities. The primary cilium is a highly conserved organelle that projects from the cell surface in a single copy in virtually all mammalian cell types including neural stem/progenitors cells and neurons. Increasing evidence in the last decade points out that primary cilium could be a relevant mediator of neural stem cell activity, neurogenesis, neuronal maturation and maintenance, and brain tumorigenesis. In this review, we summarize the current knowledge about primary cilia roles in these processes. There is currently sufficient background to propose that defective primary cilia contribute to age-related cognitive decline and brain tumor development due to their critical roles in cell cycle control and signaling transduction. This might have potential applications on therapy against age-associated brain diseases.     

力学刺激促进成骨和骨再生

肌骨系统的结构与功能、疾病的防治与康复均与力学环境和干预存在着密切的关系。伴随细胞分子生物学的发展,骨科生物力学研究从器官、组织水平深入到细胞、分子、基因水平。医用生物力学的发展促进了基于力学刺激原理的理疗设备的研发,使力学刺激促进成骨和骨再生成为目前骨科基础和应用研究的热点。《医用生物力学》杂志近年来刊登了一系列力学刺激对肉骨系统的基础和应用研究,这次选登了几项力学刺激促进成骨和骨再生,尤其在具有挑战性的骨折疏松骨折愈合促进方面的基础和应用基础成果,借此进一步加强相关领域中紧密结合临床需求的科学研究的开展。     

Role of subcellular shear-stress distributions in endothelial cell mechanotransduction

The endothelium of blood vessels presents a wavy surface to the flowing blood. The subcellular distribution of shear stress depends on the shape and orientation of the cells and on their spatial arrangement within the monolayer. By studying details of the distribution of stress at this scale and the morphological responses that serve to modify the distribution, we can gain insight into the physical mechanisms by which the cell senses its fluid mechanical environment. The rapidly growing body of evidence indicates that endothelial cells discriminate between subtle variations in the exact loading conditions including differences in temporal and spatial gradients of shear stress, steady and pulsatile laminar flow, and laminar and turbulent flows. While in a few studies the effects of these individual flow characteristics have been carefully isolated, it is difficult to assess the relative importance of any one parameter. To interpret the relationships between isolated flow characteristics or the integrated effects of combined loading conditions and the biochemical signaling events that mediate the cell response, a full stress analysis of the cell is needed. The microscopic distribution of shear stress acting upon the cell surface provides the boundary condition for such an analysis. Experimental and analytical tools are being developed to assess the stress distribution throughout the cellular structures that might be involved in mechanotransduction. © 2002 Biomedical Engineering Society. PAC2002: 8716Xa, 8719Uv, 8719Xx     

The Fuzzy planar cell polarity protein (FUZ), necessary for primary cilium formation,is essential for pituitary development

The primary cilium is an essential organelle that is important for normal cell signalling during development and homeostasis but its role in pituitary development has not been reported. The primary cilium facilitates signal transduction for multiple pathways, the best-characterised being the SHH pathway, which is known to be necessary for correct pituitary gland development. FUZ is a planar cell polarity (PCP) effector that is essential for normal ciliogenesis, where the primary cilia of Fuz^−/−mutants are shorter or non-functional. FUZ is part of a group of proteins required for recruiting retrograde intraflagellar transport proteins to the base of the organelle. Previous work has reported ciliopathy phenotypes in Fuz^−/− homozygous null mouse mutants, including neural tube defects, craniofacial abnormalities, and polydactyly, alongside PCP defects including kinked/curly tails and heart defects. Interestingly, the pituitary gland was reported to be missing in Fuz^−/− mutants at 14.5 dpc but the mechanisms underlying this phenotype were not investigated. Here, we have analysed the pituitary development of Fuz^−/− mutants. Histological analyses reveal that Rathke's pouch (RP) is initially induced normally but is not specified and fails to express LHX3, resulting in hypoplasia and apoptosis. Characterisation of SHH signalling reveals reduced pathway activation in Fuz^−/− mutant relative to control embryos, leading to deficient specification of anterior pituitary fate. Analyses of the key developmental signals FGF8 and BMP4, which are influenced by SHH, reveal abnormal patterning in the ventral diencephalon, contributing further to abnormal RP development. Taken together, our analyses suggest that primary cilia are required for normal pituitary specification through SHH signalling.     

雌激素受体在骨生长、代谢及其力学响应中的作用和机制

力学信号在骨组织的生长、重建和疾病治疗中发挥着重要的作用。近年来的研究发现雌激素受体(estrogenreceptor,ER)能介导雌激素效应调节骨组织细胞的增殖、凋亡以及功能活性,从而影响骨形成和骨吸收,在骨组织生长、骨重建中发挥重要作用;同时参与骨组织及其细胞对力学刺激的响应过程。骨组织响应力学刺激的效应要受到ER数量和(或)功能活性的影响。这些研究提示了力学刺激和雌激素可能通过一些共同的信号通路来调节骨组织细胞的功能活性。本文着重关注ER在骨组织及其力学响应中的作用和机制。     

流体剪切力在骨生长、重建中的重要作用

骨组织具有最优化结构,是一个典型的结构-功能受生物力学控制的例子。力学因素在骨的生长、重建和成形中起着十分重要的作用,但力学刺激的本质形式至今无法确认。骨组织存在多孔结构,力学负荷引起的变形会促使流体流动对骨细胞产生作用,离体实验也证实骨细胞能对流体产生响应,因此流体剪切力是研究骨组织受力学调控时考虑的一个重要方面。现有的研究主要体现在流体的有效作用形式,细胞的生物学反应,以及流体剪切力在细胞中的力学转导;各个方面的研究都提示,流体剪切力至少部分的参与了骨组织内的力转导。作者就这方面的研究进展作一综述。     

N‐terminal dual lipidation‐coupled molecular targeting into the primary cilium

The primary cilium functions as an “antenna” for cell signaling, studded with characteristic transmembrane receptors and soluble protein factors, raised above the cell surface. In contrast to the transmembrane proteins, targeting mechanisms of nontransmembrane ciliary proteins are poorly understood. We focused on a pathogenic mutation that abolishes ciliary localization of retinitis pigmentosa 2 protein and revealed a dual acylation‐dependent ciliary targeting pathway. Short N‐terminal sequences which contain myristoylation and palmitoylation sites are sufficient to target a marker protein into the cilium in a palmitoylation‐dependent manner. A Golgi‐localized palmitoyltransferase DHHC‐21 was identified as the key enzyme controlling this targeting pathway. Rapid turnover of the targeted protein was ensured by cholesterol‐dependent membrane fluidity, which balances highly and less‐mobile populations of the molecules within the cilium. This targeting signal was found in a set of signal transduction molecules, suggesting a general role of this pathway in proper ciliary organization, and dysfunction in ciliary disorders.     

Mechanisms linking mitochondrial mechanotransduction and chondrocyte biology in the pathogenesis of osteoarthritis

Mechanical loading is essential for chondrocyte health. Chondrocytes can sense and respond to various extracellular mechanical signals through an integrated set of mechanisms. Recently, it has been found that mitochondria, acting as critical mechanotransducers, are at the intersection between extracellular mechanical signals and chondrocyte biology. Much attention has been focused on identifying how mechanical loading-induced mitochondrial dysfunction contributes to the pathogenesis of osteoarthritis. In contrast, little is known regarding the mechanisms underlying functional alterations in mitochondria induced by mechanical stimulation. In this review, we describe how chondrocytes perceive environmental mechanical signals. We discuss how mechanical load induces mitochondrial functional alterations and highlight the major unanswered questions in this field. We speculate that AMP-activated protein kinase (AMPK), a master regulator of energy homeostasis, may play an important role in coupling force transmission to mitochondrial health and intracellular biological responses.     

力传导中亚细胞水平的分子FRET成像

体内细胞受到含有化学和力学因素的生理和病理生理的刺激,故研究这些因素在细胞和器官水平如何调节功能就尤为重要。有关细胞和器官对化学因素的反应已开展诸多研究,而力学因素的影响却鲜有报道。近年来,荧光蛋白和显微镜技术的发展已成为阐明力传导过程的有用工具,先进的信号活细胞成像技术促进了力学生物学中分子机制的时空因素研究。本文综述荧光蛋白的基本知识以及其在生物学研究中的应用,特别讨论了以荧光共振能量迁移(fluorescence proteins and microscopy,FRET)技术为基础的生物传感器的发展和特征。基因编码的FRET生物传感器能够实现分子时空活动的成像和定量,使得活细胞中生物化学信号在力学刺激下的反应和传导可视化。同时,本文重点阐述分子水平力学刺激下的活细胞信号传导。