中国科技论文统计源期刊 中文核心期刊
美国《化学文摘》《国际药学文摘》
《乌利希期刊指南》
WHO《西太平洋地区医学索引》来源期刊
日本科学技术振兴机构数据库（JST）
第七届湖北十大名刊提名奖
美国《化学文摘》《国际药学文摘》
《乌利希期刊指南》
WHO《西太平洋地区医学索引》来源期刊
日本科学技术振兴机构数据库（JST）
第七届湖北十大名刊提名奖
, 杨祥良
, YANG Xiangliang
随着纳米技术快速发展,新型纳米探针从组织、细胞乃至分子水平深入揭示了肿瘤力学微环境演化,而一些新型纳米操作子通过对肿瘤组织及肿瘤细胞进行力学干预实现杀伤肿瘤或提高肿瘤杀伤效果,提示纳米技术应用于肿瘤力学生物学已形成了一个新的研究热点——生物力纳米肿瘤学。该文试图定义生物力纳米肿瘤学,并总结新型生物力纳米探针与生物力纳米操作子在肿瘤力学生物学的最新研究进展,探讨当前所面临的挑战和可能的研究思路。
With the rapid development of nanotechnology,novel nanoprobes have revealed the evolution of tumor mechano-microenvironment from tissue,cell and molecule levels; and some new nanooperators have achieved tumor killing or improved the efficacy of cancer therapy by intervening tumor mechano-microenvironment or cancer cell mechanics.The application of nanotechnology in tumor mechanobiology has fostered a new research hotspot,namely mechano-nanooncology.In this review, the authors attempt to define the term mechano-nanooncology and summarize the latest advances in leveraging mechano-nanoprobes and mechano-nanooperators for the study of tumor mechanobiology.Finally,the authors discuss current challenges of mechano-nanooncology and look forward to its future development.
开放科学(资源服务)标识码(OSID)
近年来,随着纳米技术的快速发展[1],结合多模纳米成像技术、原子力显微镜表征技术、微/纳尺度光刻加工技术所发展的纳米探针从组织、细胞乃至分子水平深入揭示肿瘤力学微环境演化;另一方面,新型纳米操作子通过对肿瘤组织或细胞进行力学干预实现肿瘤杀伤。纳米技术应用于肿瘤力学生物学所取得的突破提示生物力纳米肿瘤学作为一个新的学术方向逐渐形成。生物力纳米肿瘤学旨在运用纳米技术最新进展,结合力学和工程学的原理和方法,从基因、蛋白质到细胞、组织、器官乃至整体多层面研究肿瘤力学微环境及其调控的相关机制,为肿瘤的早期诊断、精准治疗提供新的原理和方法。肿瘤组织与肿瘤细胞涉及的生物力一般介于皮牛(pN)至纳牛(nN)级别,作用范围在纳米(nm)至微米(μm)尺度,常规检测技术很难实现精准测量,而纳米技术在肿瘤力学生物学上具备其他技术无法比拟的优势。本文拟总结生物力纳米探针与生物力纳米操作子最新研究进展,讨论生物力纳米肿瘤学所面临的挑战,并展望其未来发展方向。
生物力纳米探针指能测定肿瘤组织或肿瘤细胞力学性能的纳米探针,生物力纳米探针分为两类:①可主动对肿瘤组织或细胞施加机械力,通过检测肿瘤组织或细胞受力后的反应测得肿瘤组织或细胞的力学性能;②不能主动对肿瘤组织或细胞施加机械力,但可以在肿瘤组织或细胞产生的机械力作用下产生变形,进而反推出肿瘤组织或细胞力。
肿瘤组织异常的力学性能主要体现在“三高”:高肿瘤组织硬度、高间质液体压力和高固体应力(solid stress)[2]。肿瘤组织硬度表征的是肿瘤组织对弹性变形的阻力,液体压力主要来源于肿瘤间质液体施加的压力,而固体应力主要来源于肿瘤组织内肿瘤细胞、间质细胞以及细胞外基质(extracellular matrix,ECM)等固体成分[3]。检测肿瘤组织固体应力有以下3种方法:①将肿瘤块对半切开,肿瘤切面在残余固体应力驱使下变形:向外凸出意味着局部压缩力,而向内凹陷意味着局部张力;②将肿瘤切成薄片,残余的固体应力使肿瘤片在三维空间变形,此方法尤其适用固体应力小的转移灶;③利用芯针活检释放固体应力,该方法能原位检测肿瘤内部固体应力,同时离体测量癌旁组织对肿瘤组织施加的固体应力[4]。上述方法原理相同:先以可控的方式人为释放肿瘤内固体应力,然后利用超声成像技术重构切面形变并结合有限元理论计算出肿瘤组织固体应力[5]。然而,这种侵入式肿瘤固体应力测量会对机体造成损伤,借助临床现有检测仪器和技术,如磁共振弹性成像、超声弹性成像或超声横波弹性成像,新型纳米探针有望以无损的方式测定恶性实体瘤固体应力。发展能够活体、原位、实时、无损检测肿瘤固体应力的纳米探针对于阐明肿瘤转移、复发及药物治疗中肿瘤力学微环境的演化过程具有重要意义。
组织硬化是乳腺癌、肝癌等恶性实体瘤临床触诊的理论依据。不同于肿瘤固体应力的测量,生物力纳米探针在肿瘤组织硬度测量已经有重要应用。利用压电材料锆钛酸铅制备的两种生物力纳米探针提高了肿瘤针刺活检取样成功率[6]。第一种可独立工作直接用于针刺活检,第二种则可以加载到临床使用的穿刺针针头。二者均能直接对接触对象施加一定应力(
肿瘤细胞力学性能异常主要体现在三方面:低细胞硬度、高牵引力和低力学性能感知。利用原子力显微镜检测细胞杨氏模量已有较多报道[7,11],相关研究进展已经有系统性总结[12]。此外,细胞机械性能的检测方法也已有介绍[13]。当前新型生物力纳米探针在肿瘤细胞力学性能检测方面的研究进展如下。
1993年,WANG等[14]发明磁力扭曲仪,其作用原理是通过4 μm磁球对细胞施加一定应力,同时检测在应力作用下细胞产生的应变,最终由应力和应变的比值计算出细胞弹性模量[15]。利用磁力扭曲仪测得经力学筛选方法得到的肿瘤干细胞弹性模量为50 Pa,而相应的肿瘤细胞弹性模量为200 Pa[16]。此外,多电极磁镊已被用来检测人膀胱癌T24细胞的杨氏模量[17]。由于700 nm磁球可通过细胞摄取进入细胞内部,多电极磁镊能够精准测定胞内细胞质杨氏模量。利用多电极磁镊测得的肿瘤细胞长轴杨氏模量为1.5 kPa,而短轴仅为1.1 kPa,两者之间存在显著性差异。磁力扭曲仪和磁镊技术都可以通过改变磁场强度调控磁球对细胞施加的机械力,包括力的大小、方向以及频率。新型生物力纳米探针可以揭示传统检测手段无法探测的细胞内部机械性能。上述细胞力学性能检测手段均依赖于外源性纳米探针。尽管布里渊显微镜可不依赖外源性纳米探针直接测量肿瘤细胞和多细胞球体弹性模量[18,19],但是该方法在活体肿瘤组织中的应用面临挑战。
牛顿第三定律指出,对于一个作用力,都存在一个大小相等、方向相反的反作用力,遵循这一定律,肿瘤细胞将产生作用力,简称细胞力(cell generated force),以响应周围力学微环境的改变并维持力学稳态[20]。当前生物力纳米探针主要通过直接或间接地测定细胞力作用导致的形变,从而实现对细胞力的测量。牵引力显微镜是目前应用最广泛的测定方法[21],其通过测定细胞力作用后的基质形变反算出细胞力[22]。该方法中基质形变的测定取决于荧光纳米粒,荧光纳米粒扮演生物力纳米探针的角色。TAN等[23]改进了牵引力显微镜,制备了由聚二甲基硅氧烷(polydimethylsiloxane,PDMS)组成的弹性微柱生物力纳米探针。PDMS柱在细胞力作用下会逐渐弯曲变形。通过测量微柱顶面在细胞力作用下的位移量,可以计算出单个微柱所受力的方向和大小,进而得到细胞力。然而,PDMS微柱之间相互独立,细胞与柱表面不能完全接触,并不能较好地模拟细胞原位微环境。利用DNA分子张力探针[24],LIU 等[25]发现T细胞受体(T cell receptor,TCR)与抗原结合时需要12~19 pN机械力以激活下游信号通路。该生物力纳米探针主要由修饰在金纳米粒、荧光分子以及淬灭基团标记的DNA发夹组成[26]。当探针与T细胞结合力不足以打开DNA发夹时,荧光共振能量转移以及纳米金属表面能转移双重淬灭机制会使荧光分子的荧光淬灭,探针不发出荧光[27]。当探针与TCR的结合力足以拉伸DNA发夹结构使荧光分子与淬灭基团及金纳米颗粒分开时,探针的荧光会得到恢复并能反映TCR与抗原结合时所产生的机械力。分子张力纳米探针时空分辨率高,能够在单分子水平上测量细胞力,但该探针目前仅适用二维平面培养的细胞,难以在活体组织中应用。
肿瘤细胞的力学感知能力在增殖、迁移和侵袭等过程中起着重要作用。实体瘤内的细胞受多重机械力刺激,包括静液压力、剪切应力、压应力、张力等[28]。肿瘤细胞通过细胞膜上的蛋白和细胞骨架网络将感知的机械力信号转换成生化信号,并进一步传递到细胞核内激活基因的表达。由于传统表征手段无法测量肿瘤细胞力学感知能力,基于弹性基底材料的生物力纳米探针在肿瘤细胞力学感知能力的研究上具有不可替代的作用。借助弹性基底生物力纳米探针发现,乳腺肌上皮细胞对基底刚度的感知由不同类型整合素的键动力学决定[29]。α5β1整合素在细胞表面组成型表达(constitutively expressed),αvβ6整合素在癌症的发生和发展中选择性表达,研究提示整合素αvβ6可作为一种新的乳腺癌治疗靶标。此外,肿瘤细胞和正常细胞的刚度感知能力存在显著差异[30]。在不同软硬的PDMS柱上培养肿瘤细胞,并对细胞的刚度感知单位(contractile units,CUs)进行检测发现,由于细胞骨架蛋白水平的改变,肿瘤细胞在不同刚度PDMS柱上无法产生足够数量的CUs感知刚度刺激,并产生异常的生长信号。细胞骨架蛋白恢复正常水平,可以重新使肿瘤细胞获得刚度感知能力,进而抑制肿瘤形成。研究肿瘤细胞的机械感知能力对于揭示肿瘤细胞与细胞之间、细胞与基质之间的相互作用机制具有重要意义。
生物力纳米操作子为能够对肿瘤组织或肿瘤细胞进行力学调控的纳米颗粒或器件。生物力纳米操作子分为两类,一类通过调控肿瘤组织力学微环境中的组成成分从而改善肿瘤力学微环境,该类生物力纳米操作子不具备主动施加机械力的能力;另一类可以对肿瘤组织或肿瘤细胞施加机械力,激活力学信号通路,最终决定肿瘤组织或细胞存亡。
实体瘤异常力学微环境是纳米药物递送过程的主要屏障之一,其导致纳米药物在肿瘤部位分布不均—靠近肿瘤血管部位纳米药物蓄积量高而远离肿瘤血管部位纳米药物蓄积量低。化疗不足的危害在临床已多有报道。本课题组在细胞、斑马鱼以及小鼠中证实非均质分布纳米药物导致肿瘤细胞上调转化生长因子β分泌量,加速肿瘤细胞上皮间质转化进程,促进肿瘤细胞侵袭与转移[31]。在此,笔者阐述生物力纳米操作子在调控肿瘤力学微环境,提高药物递送效率及肿瘤杀伤效果中的应用。粒径220 nm负载靶向赖氨酸氧化酶抗体的聚乳酸-羟基乙酸共聚物纳米粒在4T1小鼠乳腺癌模型中证实,该生物力纳米操纵子可以显著降低肿瘤细胞外基质的量并破坏其致密结构,最终抑制肿瘤的生长[32]。粒径19 nm铁磁性纳米粒在交变磁场的作用下可以使肿瘤温度提高至42 ℃,从而破坏胶原纤维。这使得纳米药物更容易穿透肿瘤组织,取得良好的抑瘤效果[33]。粒径100 nm包载胶原酶的脂质体可保护胶原酶在血液循环过程不会被蛋白酶降解。富集在肿瘤部位的胶原酶通过降解胰腺癌组织中的胶原纤维,提高包载紫杉醇脂质体肿瘤穿透能力及抗肿瘤疗效[34]。尽管上述生物力纳米操纵子可以显著减少胶原纤维,但是肿瘤组织的力学性能未能得到测定。研究肿瘤力学性能与肿瘤细胞外基质以及药物递送之间的关系对发展新型生物力纳米操作子具有重要价值[35]。此外,肿瘤缺氧微环境与肿瘤组织中纤维沉积密切相关,本课题组证实高压氧治疗提高肿瘤组织中氧含量,抑制HIF-1α/CTGF/ Collagen-1通路的激活,降低肿瘤组织中胶原纤维沉积量,最终提高纳米药物脂质体阿霉素肿瘤富集量与抗肿瘤疗效[36]。
近十年来,研究人员已经在单细胞层面报道了一些能够对肿瘤细胞施加机械力的生物力纳米操作子,并对其工作原理及生物医学应用进行了总结[37,38]。根据生物力纳米操纵子对细胞施加机械力后所产生的效果,可以将其细分为两类:第一类仅激活细胞力学信号通路;第二类可以杀死肿瘤细胞。磁力扭曲仪中的微米磁球可作为一种激活细胞力学信号通路的生物力纳米操作子[16]。结合三维细胞磁力扭曲仪和受激发射损耗纳米显微镜交互超高分辨三维细胞磁力扭曲系统,研究人员首次在细胞内报道机械力可以直接拉伸染色质并上调基因转录[39]。近红外光响应生物力纳米操作子由金纳米棒构成核层,聚异丙基丙酰胺水凝胶组成壳层,水凝胶外周修饰有可以和细胞表面的整合素结合的精氨酸-甘氨酸-天冬氨酸(Arg-Gly-Asp,RGD)肽[40]。常温时,聚异丙基丙酰胺水凝胶处于舒张状态。在近红外激光照射下,金纳米棒将光能转化为热能,使聚异丙基丙酰胺高分子链受热收缩,并同时拉动RGD以及细胞表面整合素,最终对细胞施加机械力。利用DNA生物力纳米探针进一步测得每个RGD分子在收缩过程中对细胞产生了13~50 pN的机械力。微米磁球生物力纳米操作子施加机械力的大小、方向及频率可控,并且能够进行高通量细胞操控与测量[41]。但是磁球自身的重力会影响细胞表面理化性质以及力学信号通路。近红外光响应生物力纳米操作子虽然有很高的时/空间精度,但是其施加的机械力大小、方向及频率可控性有待进一步提高。
第二类生物力纳米操作子主要通过施加机械力直接杀死肿瘤细胞。表面修饰靶向死亡受体4抗体的粒径15 nm Zn0.4Fe2.6O4铁磁性纳米粒在磁场诱导下与死亡受体4结合并且聚集,从而激活下游凋亡通路,引起肿瘤细胞凋亡[42]。不施加磁场时,纳米粒虽然与细胞表面死亡受体4结合但不聚集,因此不会引发下游凋亡通路。ZHANG等[43]制备出的生物力纳米操作子和溶酶体结合后会在动态磁场发生器中旋转运动,破坏溶酶体结构完整性,导致细胞凋亡。利用相似的工作原理,研究人员利用磁场诱导铁磁性生物力纳米操作子产生机械力杀死脑癌细胞[44,45]。铁磁性纳米粒在磁场作用下组装为纳米棒,并产生超过100 pN的机械力破坏肿瘤细胞细胞膜和溶酶体膜,造成肿瘤细胞程序性死亡和坏死[46]。此类生物力纳米操作子主要通过施加磁力破坏细胞器或细胞膜的完整性,最终杀死肿瘤细胞,但其临床有效性需要进一步验证。MITCHELL等[47]将粒径100 nm PLGA纳米粒或粒径500 nm聚苯乙烯纳米粒通过化学键偶联到肿瘤细胞表面,制备了不依赖于磁力的生物力纳米操作子。该生物力纳米操作子能增强细胞膜局部受到的剪切力,进而显著提高TRAIL蛋白对肿瘤细胞的杀伤效率。相比于磁场诱导产生的磁力,生物体内剪切力的可控性更差,因此,此类生物力纳米操作子的临床转化面临更大的挑战。
肿瘤力学微环境与肿瘤发生发展、转移复发以及治疗息息相关,肿瘤力学生物学已经成为肿瘤生物学以及肿瘤临床转化医学的研究热点。传统研究方法面临诸多挑战。近年来,纳米技术的迅猛发展为肿瘤力学生物学的研究提供了可靠的研究手段与检测方法。笔者提出生物力纳米肿瘤学这一新的研究方向,并给出其明确定义,进一步凝练出生物力纳米探针以及生物力纳米操作子。通过实例展示生物力纳米探针在检测肿瘤组织力学性能和肿瘤细胞力学性能的具体应用。结合最新研究进展展示生物力纳米操作子在调控肿瘤力学微环境和细胞力学性能两方面的具体应用。笔者相信,对生物力纳米探针与生物力纳米操作子的深入研究将有助于揭示肿瘤异常力学微环境及其对肿瘤的发生、发展、转移、复发及药物治疗的影响规律及机制,有望为恶性实体瘤早期诊断和精准治疗带来颠覆性理论和变革性技术。
谨以此文感谢2020年1月11日在华中科技大学参加首届“生物力纳米肿瘤学研讨会”的13位学者,特别是从美国及中国香港、北京、天津、上海、常州赴武汉参会的朋友们。
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| [2] |
Physical forces have a crucial role in tumor progression and cancer treatment. The application of principles of engineering and physical sciences to oncology has provided powerful insights into the mechanisms by which these forces affect tumor progression and confer resistance to delivery and efficacy of molecular, nano-, cellular, and immuno-medicines. Here, we discuss the mechanics of the solid and fluid components of a tumor, with a focus on how they impede the transport of therapeutic agents and create an abnormal tumor microenvironment (TME) that fuels tumor progression and treatment resistance. We also present strategies to reengineer the TME by normalizing the tumor vasculature and the extracellular matrix (ECM) to improve cancer treatment. Finally, we summarize various mathematical models that have provided insights into the physical barriers to cancer treatment and revealed new strategies to overcome these barriers.
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| [3] |
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| [4] |
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| [5] |
Solid stress, distinct from both tissue stiffness and fluid pressure, is a mechanical stress that is often elevated in both murine and human tumors. The importance of solid stress in tumor biology has been recognized in initial studies: solid stress promotes tumor progression and lowers the efficacy of anticancer therapies by compressing blood vessels and contributing to hypoxia. However, robust, reproducible, and objective methods that go beyond demonstration and bulk measurements have not yet been established. We have developed three new techniques to rigorously measure and map solid stress in both human and murine tumors that are able to account for heterogeneity in the tumor microenvironment. We describe here these methods and their independent advantages: 2D spatial mapping of solid stress (planar-cut method), sensitive estimation of solid stress in small tumors (slicing method), and in situ solid-stress quantification (needle-biopsy method). Furthermore, the preservation of tissue morphology and structure allows for subsequent histological analyses in matched tumor sections, facilitating quantitative correlations between solid stress and markers of interest. The three procedures each require approximately 2 h of experimental time per tumor. The required skill sets include basic experience in tumor resection and/or biopsy (in mice or humans), as well as in intravital imaging (e.g., ultrasonography).
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| [6] |
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| [7] |
Cancer initiation and progression follow complex molecular and structural changes in the extracellular matrix and cellular architecture of living tissue. However, it remains poorly understood how the transformation from health to malignancy alters the mechanical properties of cells within the tumour microenvironment. Here, we show using an indentation-type atomic force microscope (IT-AFM) that unadulterated human breast biopsies display distinct stiffness profiles. Correlative stiffness maps obtained on normal and benign tissues show uniform stiffness profiles that are characterized by a single distinct peak. In contrast, malignant tissues have a broad distribution resulting from tissue heterogeneity, with a prominent low-stiffness peak representative of cancer cells. Similar findings are seen in specific stages of breast cancer in MMTV-PyMT transgenic mice. Further evidence obtained from the lungs of mice with late-stage tumours shows that migration and metastatic spreading is correlated to the low stiffness of hypoxia-associated cancer cells. Overall, nanomechanical profiling by IT-AFM provides quantitative indicators in the clinical diagnostics of breast cancer with translational significance.
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| [8] |
SummaryTumors are stiffer than normal tissue, and tumors have altered integrins. Because integrins are mechanotransducers that regulate cell fate, we asked whether tissue stiffness could promote malignant behavior by modulating integrins. We found that tumors are rigid because they have a stiff stroma and elevated Rho-dependent cytoskeletal tension that drives focal adhesions, disrupts adherens junctions, perturbs tissue polarity, enhances growth, and hinders lumen formation. Matrix stiffness perturbs epithelial morphogenesis by clustering integrins to enhance ERK activation and increase ROCK-generated contractility and focal adhesions. Contractile, EGF-transformed epithelia with elevated ERK and Rho activity could be phenotypically reverted to tissues lacking focal adhesions if Rho-generated contractility or ERK activity was decreased. Thus, ERK and Rho constitute part of an integrated mechanoregulatory circuit linking matrix stiffness to cytoskeletal tension through integrins to regulate tissue phenotype. |
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Tissue mechanics regulate development and homeostasis and are consistently modified in tumor progression. Nevertheless, the fundamental molecular mechanisms through which altered mechanics regulate tissue behavior and the clinical relevance of these changes remain unclear. We demonstrate that increased matrix stiffness modulates microRNA expression to drive tumor progression through integrin activation of beta-catenin and MYC. Specifically, in human and mouse tissue, increased matrix stiffness induced miR-18a to reduce levels of the tumor suppressor phosphatase and tensin homolog (PTEN), both directly and indirectly by decreasing levels of homeobox A9 (HOXA9). Clinically, extracellular matrix stiffness correlated directly and significantly with miR-18a expression in human breast tumor biopsies. miR-18a expression was highest in basal-like breast cancers in which PTEN and HOXA9 levels were lowest, and high miR-18a expression predicted poor prognosis in patients with luminal breast cancers. Our findings identify a mechanically regulated microRNA circuit that can promote malignancy and suggest potential prognostic roles for HOXA9 and miR-18a levels in stratifying patients with luminal breast cancers.
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| [10] |
Biomechanical and biochemical cues within a tissue collaborate across length scales to direct cell fate during development and are critical for the maintenance of tissue homeostasis. Loss of tensional homeostasis in a tissue not only accompanies malignancy but may also contribute to oncogenic transformation. High mechanical stress in solid tumors can impede drug delivery and may additionally drive tumor progression and promote metastasis. Mechanistically, biomechanical forces can drive tumor aggression by inducing a mesenchymal-like switch in transformed cells so that they attain tumor-initiating or stem-like cell properties. Given that cancer stem cells have been linked to metastasis and treatment resistance, this raises the intriguing possibility that the elevated tissue mechanics in tumors could promote their aggression by programming their phenotype toward that exhibited by a stem-like cell.Significance: Recent findings argue that mechanical stress and elevated mechanosignaling foster malignant transformation and metastasis. Prolonged corruption of tissue tension may drive tumor aggression by altering cell fate specification. Thus, strategies that could reduce tumor mechanics might comprise effective approaches to prevent the emergence of treatment-resilient metastatic cancers. Cancer Discov; 7(11); 1224-37. (c)2017 AACR.
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| [11] |
Change in cell stiffness is a new characteristic of cancer cells that affects the way they spread. Despite several studies on architectural changes in cultured cell lines, no ex vivo mechanical analyses of cancer cells obtained from patients have been reported. Using atomic force microscopy, we report the stiffness of live metastatic cancer cells taken from the body (pleural) fluids of patients with suspected lung, breast and pancreas cancer. Within the same sample, we find that the cell stiffness of metastatic cancer cells is more than 70% softer, with a standard deviation over five times narrower, than the benign cells that line the body cavity. Different cancer types were found to display a common stiffness. Our work shows that mechanical analysis can distinguish cancerous cells from normal ones even when they show similar shapes. These results show that nanomechanical analysis correlates well with immunohistochemical testing currently used for detecting cancer.
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| [12] |
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| [13] |
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| [14] |
Mechanical stresses were applied directly to cell surface receptors with a magnetic twisting device. The extracellular matrix receptor, integrin beta 1, induced focal adhesion formation and supported a force-dependent stiffening response, whereas nonadhesion receptors did not. The cytoskeletal stiffness (ratio of stress to strain) increased in direct proportion to the applied stress and required intact microtubules and intermediate filaments as well as microfilaments. Tensegrity models that incorporate mechanically interdependent struts and strings that reorient globally in response to a localized stress mimicked this response. These results suggest that integrins act as mechanoreceptors and transmit mechanical signals to the cytoskeleton. Mechanotransduction, in turn, may be mediated simultaneously at multiple locations inside the cell through force-induced rearrangements within a tensionally integrated cytoskeleton.
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| [15] |
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| [16] |
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| [17] |
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| [18] |
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| [19] |
The role and importance of mechanical properties of cells and tissues in cellular function, development and disease has widely been acknowledged, however standard techniques currently used to assess them exhibit intrinsic limitations. Recently, Brillouin microscopy, a type of optical elastography, has emerged as a non-destructive, label- and contact-free method that can probe the viscoelastic properties of biological samples with diffraction-limited resolution in 3D. This led to increased attention amongst the biological and medical research communities, but it also sparked debates about the interpretation and relevance of the measured physical quantities. Here, we review this emerging technology by describing the underlying biophysical principles and discussing the interpretation of Brillouin spectra arising from heterogeneous biological matter. We further elaborate on the technique's limitations, as well as its potential for gaining insights in biology, in order to guide interested researchers from various fields.
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Cells exert, sense, and respond to physical forces through an astounding diversity of mechanisms. Here we review recently developed tools to quantify the forces generated by cells. We first review technologies based on sensors of known or assumed mechanical properties, and discuss their applicability and limitations. We then proceed to draw an analogy between these human-made sensors and force sensing in the cell. As mechanics is increasingly revealed to play a fundamental role in cell function we envisage that tools to quantify physical forces may soon become widely applied in life-sciences laboratories.
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Forces generated by cells are critical regulators of cell adhesion, signaling, and function, and they are also essential drivers in the morphogenetic events of development. Over the past 20 years, several methods have been developed to measure these forces. However, despite recent substantial interest in understanding the contribution of these forces in biology, implementation and adoption of the developed methods by the broader biological community remain challenging because of the inherently multidisciplinary expertise required to conduct and interpret the measurements. In this review, we introduce the established methods and highlight the technical challenges associated with implementing each technique in a biological laboratory.
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| [22] |
Responses of cells to mechanical properties of the adhesion substrate were examined by culturing normal rat kidney epithelial and 3T3 fibroblastic cells on a collagen-coated polyacrylamide substrate that allows the flexibility to be varied while maintaining a constant chemical environment. Compared with cells on rigid substrates, those on flexible substrates showed reduced spreading and increased rates of motility or lamellipodial activity. Microinjection of fluorescent vinculin indicated that focal adhesions on flexible substrates were irregularly shaped and highly dynamic whereas those on firm substrates had a normal morphology and were much more stable. Cells on flexible substrates also contained a reduced amount of phosphotyrosine at adhesion sites. Treatment of these cells with phenylarsine oxide, a tyrosine phosphatase inhibitor, induced the formation of normal, stable focal adhesions similar to those on firm substrates. Conversely, treatment of cells on firm substrates with myosin inhibitors 2,3-butanedione monoxime or KT5926 caused the reduction of both vinculin and phosphotyrosine at adhesion sites. These results demonstrate the ability of cells to survey the mechanical properties of their surrounding environment and suggest the possible involvement of both protein tyrosine phosphorylation and myosin-generated cortical forces in this process. Such response to physical parameters likely represents an important mechanism of cellular interaction with the surrounding environment within a complex organism.
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| [23] |
We describe an approach to manipulate and measure mechanical interactions between cells and their underlying substrates by using microfabricated arrays of elastomeric, microneedle-like posts. By controlling the geometry of the posts, we varied the compliance of the substrate while holding other surface properties constant. Cells attached to, spread across, and deflected multiple posts. The deflections of the posts occurred independently of neighboring posts and, therefore, directly reported the subcellular distribution of traction forces. We report two classes of force-supporting adhesions that exhibit distinct force-size relationships. Force increased with size of adhesions for adhesions larger than 1 microm(2), whereas no such correlation existed for smaller adhesions. By controlling cell adhesion on these micromechanical sensors, we showed that cell morphology regulates the magnitude of traction force generated by cells. Cells that were prevented from spreading and flattening against the substrate did not contract in response to stimulation by serum or lysophosphatidic acid, whereas spread cells did. Contractility in the unspread cells was rescued by expression of constitutively active RhoA. Together, these findings demonstrate a coordination of biochemical and mechanical signals to regulate cell adhesion and mechanics, and they introduce the use of arrays of mechanically isolated sensors to manipulate and measure the mechanical interactions of cells.
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| [24] |
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| [25] |
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| [26] |
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| [27] |
Studying chemomechanical coupling at interfaces is important for fields ranging from lubrication and tribology to microfluidics and cell biology. Several polymeric macro- and microscopic systems and cantilevers have been developed to image forces at interfaces, but few materials are amenable for molecular tension sensing. To address this issue, we have developed a gold nanoparticle sensor for molecular tension-based fluorescence microscopy. As a proof of concept, we imaged the tension exerted by integrin receptors at the interface between living cells and a substrate with high spatial (<1 mum) resolution, at 100 ms acquisition times and with molecular specificity. We report integrin tension values ranging from 1 to 15 pN and a mean of ~1 pN within focal adhesions. Through the use of a conventional fluorescence microscope, this method demonstrates a force sensitivity that is 3 orders of magnitude greater than is achievable by traction force microscopy or polydimethylsiloxane micropost arrays, which are the standard in cellular biomechanics.
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Over the past decade, researchers have highlighted the importance of mechanical cues of the metastatic niche such as matrix stiffness, topography, mechanical stresses, and deformation on cells in influencing tumor growth and proliferation. Understanding the cellular and molecular basis and fine-tuning the mechano-response of cancer cells to this niche could lead to new and novel therapeutic interventions. In this review, we discuss the importance of mechanical cues surrounding tumor microenvironment that govern the growth and progression of cancer. We also highlight some emergent principles underlying the mechanosensing and mechanotransduction mechanisms that link cellular responses such as gene expression to phenotypic changes arising from such external cues. Recent technological advancements to visualize, quantify, model, and test these crucial steps with great precision will further advance our understanding of this phenomenon. We will conclude by showcasing potential applications of mechanobiology in controlling cancer growth as alternative cancer treatment regimes.
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| [29] |
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| [30] |
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| [31] |
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| [32] |
A cancer nanotherapeutic has been developed that targets the extracellular matrix (ECM)-modifying enzyme lysyl oxidase (LOX) and alters the ECM structure. Poly(d,l-lactide-co-glycolide) nanoparticles ( approximately 220 nm) coated with a LOX inhibitory antibody bind to ECM and suppress mammary cancer cell growth and invasion in vitro as well as tumor expansion in vivo, with greater efficiency than soluble anti-LOX antibody. This nanomaterials approach opens a new path for treating cancer with higher efficacy and decreased side effects.
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Several studies propose nanoparticles for tumor treatment, yet little is known about the fate of nanoparticles and intimate interactions with the heterogeneous and ever-evolving tumor environment. The latter, rich in extracellular matrix, is responsible for poor penetration of therapeutics and represents a paramount issue in cancer therapy. Hence new strategies start aiming to modulate the neoplastic stroma. From this perspective, we assessed the efficacy of 19 nm PEG-coated iron oxide nanocubes with optimized magnetic properties to mediate mild tumor magnetic hyperthermia treatment. After injection of a low dose of nanocubes (700 mug of iron) into epidermoid carcinoma xenografts in mice, we monitored the effect of heating nanocubes on tumor environment. In comparison with the long-term fate after intravenous administration, we investigated spatiotemporal patterns of nanocube distribution, evaluated the evolution of cubes magnetic properties, and examined nanoparticle clearance and degradation processes. While inside tumors nanocubes retained their magnetic properties and heating capacity throughout the treatment due to a mainly interstitial extracellular location, the particles became inefficient heaters after cell internalization and transfer to spleen and liver. Our multiscale analysis reveals that collagen-rich tumor extracellular matrix confines the majority of nanocubes. However, nanocube-mediated hyperthermia has the potential to
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Overexpressed extracellular matrix (ECM) in pancreatic ductal adenocarcinoma (PDAC) limits drug penetration into the tumor and is associated with poor prognosis. Here, we demonstrate that a pretreatment based on a proteolytic-enzyme nanoparticle system disassembles the dense PDAC collagen stroma and increases drug penetration into the pancreatic tumor. More specifically, the collagozome, a 100 nm liposome encapsulating collagenase, was rationally designed to protect the collagenase from premature deactivation and prolonged its release rate at the target site. Collagen is the main component of the PDAC stroma, reaching 12.8 +/- 2.3% vol in diseased mice pancreases, compared to 1.4 +/- 0.4% in healthy mice. Upon intravenous injection of the collagozome, approximately 1% of the injected dose reached the pancreas over 8 h, reducing the level of fibrotic tissue to 5.6 +/- 0.8%. The collagozome pretreatment allowed increased drug penetration into the pancreas and improved PDAC treatment. PDAC tumors, pretreated with the collagozome followed by paclitaxel micelles, were 87% smaller than tumors pretreated with empty liposomes followed by paclitaxel micelles. Interestingly, degrading the ECM did not increase the number of circulating tumor cells or metastasis. This strategy holds promise for degrading the extracellular stroma in other diseases as well, such as liver fibrosis, enhancing tissue permeability before drug administration.
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| [35] |
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| [36] |
DOI:10.1002/advs.v5.8
URL
[本文引用:1]
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| [37] |
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| [38] |
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| [39] |
Mechanical forces play critical roles in the function of living cells. However, the underlying mechanisms of how forces influence nuclear events remain elusive. Here, we show that chromatin deformation as well as force-induced transcription of a green fluorescent protein (GFP)-tagged bacterial-chromosome dihydrofolate reductase (DHFR) transgene can be visualized in a living cell by using three-dimensional magnetic twisting cytometry to apply local stresses on the cell surface via an Arg-Gly-Asp-coated magnetic bead. Chromatin stretching depended on loading direction. DHFR transcription upregulation was sensitive to load direction and proportional to the magnitude of chromatin stretching. Disrupting filamentous actin or inhibiting actomyosin contraction abrogated or attenuated force-induced DHFR transcription, whereas activating endogenous contraction upregulated force-induced DHFR transcription. Our findings suggest that local stresses applied to integrins propagate from the tensed actin cytoskeleton to the LINC complex and then through lamina-chromatin interactions to directly stretch chromatin and upregulate transcription.
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To control receptor tension optically at the cell surface, we developed an approach involving optomechanical actuator nanoparticles that are controlled with near-infrared light. Illumination leads to particle collapse, delivering piconewton forces to specific cell surface receptors with high spatial and temporal resolution. We demonstrate optomechanical actuation by controlling integrin-based focal adhesion formation, cell protrusion and migration, and T cell receptor activation.
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The mechanical properties of cells influence their cellular and subcellular functions, including cell adhesion, migration, polarization, and differentiation, as well as organelle organization and trafficking inside the cytoplasm. Yet reported values of cell stiffness and viscosity vary substantially, which suggests differences in how the results of different methods are obtained or analyzed by different groups. To address this issue and illustrate the complementarity of certain approaches, here we present, analyze, and critically compare measurements obtained by means of some of the most widely used methods for cell mechanics: atomic force microscopy, magnetic twisting cytometry, particle-tracking microrheology, parallel-plate rheometry, cell monolayer rheology, and optical stretching. These measurements highlight how elastic and viscous moduli of MCF-7 breast cancer cells can vary 1,000-fold and 100-fold, respectively. We discuss the sources of these variations, including the level of applied mechanical stress, the rate of deformation, the geometry of the probe, the location probed in the cell, and the extracellular microenvironment.
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| [42] |
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The ability to control the movement of nanoparticles remotely and with high precision would have far-reaching implications in many areas of nanotechnology. We have designed a unique dynamic magnetic field (DMF) generator that can induce rotational movements of superparamagnetic iron oxide nanoparticles (SPIONs). We examined whether the rotational nanoparticle movement could be used for remote induction of cell death by injuring lysosomal membrane structures. We further hypothesized that the shear forces created by the generation of oscillatory torques (incomplete rotation) of SPIONs bound to lysosomal membranes would cause membrane permeabilization, lead to extravasation of lysosomal contents into the cytoplasm, and induce apoptosis. To this end, we covalently conjugated SPIONs with antibodies targeting the lysosomal protein marker LAMP1 (LAMP1-SPION). Remote activation of slow rotation of LAMP1-SPIONs significantly improved the efficacy of cellular internalization of the nanoparticles. LAMP1-SPIONs then preferentially accumulated along the membrane in lysosomes in both rat insulinoma tumor cells and human pancreatic beta cells due to binding of LAMP1-SPIONs to endogenous LAMP1. Further activation of torques by the LAMP1-SPIONs bound to lysosomes resulted in rapid decrease in size and number of lysosomes, attributable to tearing of the lysosomal membrane by the shear force of the rotationally activated LAMP1-SPIONs. This remote activation resulted in an increased expression of early and late apoptotic markers and impaired cell growth. Our findings suggest that DMF treatment of lysosome-targeted nanoparticles offers a noninvasive tool to induce apoptosis remotely and could serve as an important platform technology for a wide range of biomedical applications.
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| [44] |
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| [45] |
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| [46] |
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| [47] |
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