Mechanotransduction is a key determinant of tissue homeostasis and tumor progression. It is driven by intercellular adhesions, cell contractility, and forces generated within the microenvironment ...and is dependent on extracellular matrix composition, organization, and compliance. We show that caveolin-1 (Cav1) favors cell elongation in three-dimensional cultures and promotes Rho- and force-dependent contraction, matrix alignment, and microenvironment stiffening through regulation of p190RhoGAP. In turn, microenvironment remodeling by Cav1 fibroblasts forces cell elongation. Cav1-deficient mice have disorganized stromal tissue architecture. Stroma associated with human carcinomas and melanoma metastases is enriched in Cav1-expressing carcinoma-associated fibroblasts (CAFs). Cav1 expression in breast CAFs correlates with low survival, and Cav1 depletion in CAFs decreases CAF contractility. Consistently, fibroblast expression of Cav1, through p190RhoGAP regulation, favors directional migration and invasiveness of carcinoma cells in vitro. In vivo, stromal Cav1 remodels peri- and intratumoral microenvironments to facilitate tumor invasion, correlating with increased metastatic potency. Thus, Cav1 modulates tissue responses through force-dependent architectural regulation of the microenvironment.
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► Cav1 controls cell contractility and matrix remodeling through p190RhoGAP regulation ► Biomechanical remodeling by stromal Cav1 regulates tissue architecture ► Carcinoma-associated stroma is enriched in Cav1 ► Stromal Cav1 promotes local tumor invasion and metastasis
The most dangerous aspect of cancer lies in metastatic progression. Tumor cells will successfully form life-threatening metastases when they undergo sequential steps along a journey from the primary ...tumor to distant organs. From a biomechanics standpoint, growth, invasion, intravasation, circulation, arrest/adhesion, and extravasation of tumor cells demand particular cell-mechanical properties in order to survive and complete the metastatic cascade. With metastatic cells usually being softer than their non-malignant counterparts, high deformability for both the cell and its nucleus is thought to offer a significant advantage for metastatic potential. However, it is still unclear whether there is a finely tuned but fixed mechanical state that accommodates all mechanical features required for survival throughout the cascade or whether tumor cells need to dynamically refine their properties and intracellular components at each new step encountered. Here, we review the various mechanical requirements successful cancer cells might need to fulfill along their journey and speculate on the possibility that they dynamically adapt their properties accordingly. The mechanical signature of a successful cancer cell might actually be its ability to adapt to the successive microenvironmental constraints along the different steps of the journey.
Gensbittel et al. discuss the mechanical properties of cancer cells and how these contribute to their metastatic ability. This perspective discusses how deformability and adaptation drive metastatic capability.
Extracellular vesicles (EVs) are released by most cell types but providing evidence for their physiological relevance remains challenging due to a lack of appropriate model organisms. Here, we ...developed an in vivo model to study EV function by expressing CD63-pHluorin in zebrafish embryos. A combination of imaging methods and proteomic analysis allowed us to study biogenesis, composition, transfer, uptake, and fate of individual endogenous EVs. We identified a subpopulation of EVs with exosome features, released in a syntenin-dependent manner from the yolk syncytial layer into the blood circulation. These exosomes are captured, endocytosed, and degraded by patrolling macrophages and endothelial cells in the caudal vein plexus (CVP) in a scavenger receptor- and dynamin-dependent manner. Interference with exosome biogenesis affected CVP growth, suggesting a role in trophic support. Altogether, our work represents a system for studying endogenous EV function in vivo with high spatiotemporal accuracy, demonstrating functional inter-organ communication by exosomes.
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•Single endogenous EVs can be live-visualized in the zebrafish embryo with CD63-pHluorin•Syntenin in the YSL regulates exosome release into the blood for further dissemination•YSL exosomes are taken up by macrophages and endothelial cells in the tail of the embryo•Uptake is scavenger receptor- and dynamin-dependent and provides trophic support
Verweij et al. develop an in vivo model using zebrafish embryos to live-track the production, journey, and fate of individual exosomes. Using a combination of imaging methods and proteomic analysis, they investigate the composition of endogenous exosomes and the molecular mechanisms controlling their biogenesis, fates, and functions in receiving cells.
Metastasis is a dynamic succession of events involving the dissemination of tumour cells to distant sites within the body, ultimately reducing the survival of patients with cancer. To colonize ...distant organs and, therefore, systemically disseminate within the organism, cancer cells and associated factors exploit several bodily fluid systems, which provide a natural transportation route. Indeed, the flow mechanics of the blood and lymphatic circulatory systems can be co-opted to improve the efficiency of cancer cell transit from the primary tumour, extravasation and metastatic seeding. Flow rates, vessel size and shear stress can all influence the survival of cancer cells in the circulation and control organotropic seeding patterns. Thus, in addition to using these fluids as a means to travel throughout the body, cancer cells exploit the underlying physical forces within these fluids to successfully seed distant metastases. In this Review, we describe how circulating tumour cells and tumour-associated factors leverage bodily fluids, their underlying forces and imposed stresses during metastasis. As the contribution of bodily fluids and their mechanics raises interesting questions about the biology of the metastatic cascade, an improved understanding of this process might provide a new avenue for targeting cancer cells in transit.
Metastases go with the flow Goetz, Jacky G
Science (American Association for the Advancement of Science),
2018-Nov-30, Letnik:
362, Številka:
6418
Journal Article
Tumor extracellular vesicles (EVs) mediate the communication between tumor and stromal cells mostly to the benefit of tumor progression. Notably, tumor EVs travel in the bloodstream, reach distant ...organs, and locally modify the microenvironment. However, visualizing these events in vivo still faces major hurdles. Here, we describe an approach for tracking circulating tumor EVs in a living organism: we combine chemical and genetically encoded probes with the zebrafish embryo as an animal model. We provide a first description of tumor EVs’ hemodynamic behavior and document their intravascular arrest. We show that circulating tumor EVs are rapidly taken up by endothelial cells and blood patrolling macrophages and subsequently stored in degradative compartments. Finally, we demonstrate that tumor EVs activate macrophages and promote metastatic outgrowth. Overall, our study proves the usefulness and prospects of zebrafish embryo to track tumor EVs and dissect their role in metastatic niches formation in vivo.
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•MemBright allows for bright and specific staining of EVs•The zebrafish embryo allows tracking of tumor EVs at high spatiotemporal resolution•Circulating tumor EVs are mostly taken up by endothelial cells and patrolling macrophages•Zebrafish melanoma EVs favor metastatic outgrowth in zebrafish embryos
Tumor extracellular vesicles (EVs) promote tumor progression. However, their behavior in body fluids remains mysterious. Hyenne et al. show that the zebrafish embryo can be used to track and assess the function of circulating tumor EVs in vivo and provide a high-resolution description of their dissemination and uptake.
The pattern of blood flow has long been thought to play a significant role in vascular morphogenesis, yet the flow-sensing mechanism that is involved at early embryonic stages, when flow forces are ...low, remains unclear. It has been proposed that endothelial cells use primary cilia to sense flow, but this has never been tested in vivo. Here we show, by noninvasive, high-resolution imaging of live zebrafish embryos, that endothelial cilia progressively deflect at the onset of blood flow and that the deflection angle correlates with calcium levels in endothelial cells. We demonstrate that alterations in shear stress, ciliogenesis, or expression of the calcium channel PKD2 impair the endothelial calcium level and both increase and perturb vascular morphogenesis. Altogether, these results demonstrate that endothelial cilia constitute a highly sensitive structure that permits the detection of low shear forces during vascular morphogenesis.
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•Endothelial cilia are present during angiogenesis and deflected by low flow forces•Cilia deflection leads to endothelial calcium increase as flow forces increase•Endothelial cilia ultrastructure is unique, which makes it highly flexible•Cilia, flow, and PKD2 are necessary for early angiogenesis
Blood flow is a major mechanical stimulus that controls vascular development, yet the mechanodetection mechanisms that drive angiogenesis are poorly understood. Vermot and colleagues now show that primary cilia located at the endothelial cell surface regulate early angiogenesis. The authors demonstrate that cilia-mediated mechanodetection detects low flow forces through PKD2-mediated calcium fluxes.
Studying key biological events within complex model systems relies on dynamic and functional imaging at optimum spatial and temporal resolutions. Intravital correlative light and electron microscopy ...(intravital CLEM) combines imaging living multicellular model systems with electron microscopy, and offers full ultrastructural details of dynamic or transient events in vivo . However, routine use of intravital CLEM is hindered by multiple technological challenges faced when targeting a micron-size object (e.g., single cells or organelles) in a complex living organism. Recently, various approaches have been developed to overcome these limitations. In this review we outline the current methods and present the power of intravital CLEM in different fields of research. Finally, we describe approaches that will make intravital CLEM a routine, quantitative method for high-resolution cell biology in vivo.
Fluorescent polymer nanoparticles for long‐term labeling and tracking of living cells with any desired color code are developed. They are built from biodegradable poly(lactic‐co‐glycolic acid) ...polymer loaded with cyanine dyes (DiO, DiI, and DiD) with the help of bulky fluorinated counterions, which minimize aggregation‐caused quenching. At the single particle level, these particles are ≈20‐fold brighter than quantum dots of similar color. Due to their identical 40 nm size and surface properties, these nanoparticles are endocytosed equally well by living cells. Mixing nanoparticles of three colors in different proportions generates a homogeneous RGB (red, green, and blue) barcode in cells, which is transmitted through many cell generations. Cell barcoding is validated on 7 cell lines (HeLa, KB, embryonic kidney (293T), Chinese hamster ovary, rat basophilic leucemia, U97, and D2A1), 13 color codes, and it enables simultaneous tracking of co‐cultured barcoded cell populations for >2 weeks. It is also applied to studying competition among drug‐treated cell populations. This technology enabled six‐color imaging in vivo for (1) tracking xenografted cancer cells and (2) monitoring morphogenesis after microinjection in zebrafish embryos. In addition to a robust method of multicolor cell labeling and tracking, this work suggests that multiple functions can be co‐localized inside cells by combining structurally close nanoparticles carrying different functions.
Structurally close but spectrally distinct fluorescent polymer nanoparticles are prepared through loading of cyanine dyes with the help of bulky hydrophobic counterions, preventing aggregation‐caused quenching. Due to identical internalization, these nanoparticles can be combined together for long‐term labeling and tracking of cells in vitro and in vivo with at least 13 color codes.
Exosomes are secreted vesicles arising from the fusion of multivesicular bodies (MVBs) with the plasma membrane. Despite their importance in various processes, the molecular mechanisms controlling ...their formation and release remain unclear. Using nematodes and mammary tumor cells, we show that Ral GTPases are involved in exosome biogenesis. In Caenorhabditis elegans, RAL-1 localizes at the surface of secretory MVBs. A quantitative electron microscopy analysis of RAL-1-deficient animals revealed that RAL-1 is involved in both MVB formation and their fusion with the plasma membrane. These functions do not involve the exocyst complex, a common Ral guanosine triphosphatase (GTPase) effector. Furthermore, we show that the target membrane SNARE protein SYX-5 colocalizes with a constitutively active form of RAL-1 at the plasma membrane, and MVBs accumulate under the plasma membrane when SYX-5 is absent. In mammals, RalA and RalB are both required for the secretion of exosome-like vesicles in cultured cells. Therefore, Ral GTPases represent new regulators of MVB formation and exosome release.
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