ArticleLiterature Review

Specialized Intercellular Communications via Cytonemes and Nanotubes

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Abstract

In recent years, thin membrane protrusions such as cytonemes and tunneling nanotubes have emerged as a novel mechanism of intercellular communication. Protrusion-based cellular interactions allow for specific communication between participating cells and have a distinct spectrum of advantages compared to secretion- and diffusion-based intercellular communication. Identification of protrusion-based signaling in diverse systems suggests that this mechanism is a ubiquitous and prevailing means of communication employed by many cell types. Moreover, accumulating evidence indicates that protrusion-based intercellular communication is often involved in pathogenesis, including cancers and infections. Here we review our current understanding of protrusion-based intercellular communication. Expected final online publication date for the Annual Review of Cell and Developmental Biology Volume 34 is October 6, 2018. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.

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... Communication between cells as well as cells with their microenvironment is fundamental to preserving tissue homeostasis and restoring normal function upon damage or deterioration. Logically, a high degree of specificity is required for accurate transfer; it is vital that a particular initiating cell communicate the appropriate message to the correct recipient (Yamashita et al., 2018). While numerous processes have been described that mediate cell-cell communication such as the diffusion of secreted soluble molecules or the release of microvesicles or cargo-loaded exosomes, these lack the strict specificity or reliability required in some cases (Wolpert, 2016;Caviglia and Ober, 2018). ...
... Indeed, the microenvironment of damaged tissues is highly toxic and proteolytic, yet transplanted mesenchymal stem cells are able to transfer healthy organelles and vesicles to rescue damaged cells, suggesting a more 'protected' means of communication must exist. Alternatively, protrusion-based cellular interactions such as filopodia (Gardel et al., 2010;Jacquemet et al., 2015), cytonemes (Kornberg and Roy, 2014;Stanganello and Scholpp, 2016) and although less well studied, the tunneling nanotubes (TNTs) (Onfelt et al., 2004;Rustom et al., 2004;Baker, 2017;Yamashita et al., 2018;Hanna et al., 2019;Genna et al., 2023) would provide specificity over soluble-al., 2013;Reglero-Real et al., 2016;Caporali et al., 2017;Johnson et al., 2017). These combined observations support a mechanism whereby protrusion-based structures enable highly specific, long-range intercellular communication and may explain the propagation of signals through complex and potentially lethal environments to influence cell fate in vivo. ...
... Despite their initial description nearly 20 years ago (Rustom et al., 2004), the extracellular and intracellular signals and specific molecules that either trigger or specifically inhibit the induction, formation and function of these protrusions are largely unknown (Belian et al., 2023). TNTs appear to form in response to metabolic stress such as deprivation of serum proteins and high glucose conditions, but few specific molecules have been identified that induce TNT formation (Yamashita et al., 2018;Belian et al., 2023). Logically, both the membrane and the cytoskeleton must play a role in protrusion formation. ...
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Transmembrane CD13 assembles protein complexes at the plasma membrane to enable diverse cellular processes such as cell-cell adhesion, focal adhesion turnover, endocytosis and recycling of cell surface proteins. In this study, we demonstrate a novel CD13-dependent assembly platform that regulates phosphoinositide (PI) signal transduction during the formation of Tunneling Nanotubes (TNTs). TNTs are actin-based, membrane-delimited bridges that facilitate intercellular communication by connecting distant cells to physically transfer subcellular cargoes. TNTs form between various cell types under stress conditions, but few molecular TNT-inducers exist. Human Kaposi sarcoma-derived endothelial cells (KSECs) readily form stress-induced TNTs capable of transferring calcium ion and membrane molecules between cells, with clear accumulation of CD13 and actin at the base of the protrusions. Alternatively, CD13-null KSECs form fewer TNTs and calcium ion transfer is markedly reduced. Mechanistically, CD13-mediated TNT formation requires activation of CD13, Src, FAK and Cdc42 to allow tethering of the IQGAP1 and ARF6 complex at the membrane to activate the phosphatidylinositol-4-phosphate-5-kinase PI5K. This increases local phosphatidylinositol 4,5-bisphosphate levels to promote the actin-polymerization and membrane protrusion necessary for TNT formation. Therefore, CD13 is a novel molecular PIP regulator and TNT trigger that will facilitate the dissection of downstream pathways and mechanisms regulating TNT formation.
... Some of this communication occurs by diffusing molecules (Hu et al., 2010;Govern and ten Wolde, 2012;Bialek and Setayeshgar, 2008;Endres and Wingreen, 2009) like morphogens. However, recently, an alternative cell-cell communication mechanism has been revealed to be long, thin cellular protrusions extending tens to hundreds of micrometers (Eom, 2020;Yamashita et al., 2018;Caviglia and Ober, 2018;Sanders et al., 2013;Bressloff and Kim, 2019;Inaba et al., 2015). These include cytonemes (Kornberg and Roy, 2014), tunneling nanotubes (Zurzolo, 2021), tenocyte projections (Subramanian et al., 2018), and airinemes in zebrafish (Volkening and Sandstede, 2018;Volkening, 2020;Eom and Parichy, 2017;Eom et al., 2015), shown in Figure 1A. ...
... These include cytonemes (Kornberg and Roy, 2014), tunneling nanotubes (Zurzolo, 2021), tenocyte projections (Subramanian et al., 2018), and airinemes in zebrafish (Volkening and Sandstede, 2018;Volkening, 2020;Eom and Parichy, 2017;Eom et al., 2015), shown in Figure 1A. One of the difficulties delaying their discovery and characterization is their thin, suboptical width, and the fact that they only form at specific stages of development (Eom, 2020;Yamashita et al., 2018;Caviglia and Ober, 2018). ...
... The growing catalog of non-canonical cellular protrusions (Eom, 2020;Yamashita et al., 2018;Caviglia and Ober, 2018;Sanders et al., 2013;Bressloff and Kim, 2019;Inaba et al., 2015;Kornberg and Roy, 2014;Parker et al., 2017;Subramanian et al., 2018;Wang and Gerdes, 2015) includes strikingly different shapes. For example, some tunneling nanotubes in cancer cells (Parker et al., 2017) are straight compared to airinemes. ...
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In addition to diffusive signals, cells in tissue also communicate via long, thin cellular protrusions, such as airinemes in zebrafish. Before establishing communication, cellular protrusions must find their target cell. Here we demonstrate that the shapes of airinemes in zebrafish are consistent with a finite persistent random walk model. The probability of contacting the target cell is maximized for a balance between ballistic search (straight) and diffusive search (highly curved, random). We find that the curvature of airinemes in zebrafish, extracted from live cell microscopy, is approximately the same value as the optimum in the simple persistent random walk model. We also explore the ability of the target cell to infer direction of the airineme's source, finding that there is a theoretical trade-off between search optimality and directional information. This provides a framework to characterize the shape, and performance objectives, of non-canonical cellular protrusions in general.
... (v) Intercellular communication: Long nanotubes known as cytonemes [45,46] are known to serve as conduits for communication between two cells through exchange of matter and information. (vi) Collecting cues and signals: Many protrusive structures, like filopodia and stereocilia, are used for exploring the environment in search of different biochemical cues and mechanical signals [35]. ...
... Protrusion: (i) Pollen tube is an example of such protrusion which doesn't cease growing till the tip establishes contact with the ovary for the delivery of sperm [209]. (ii) Cytonemes and tunnelling nanotubes keep growing till they hit the target cell with which the host cell has to establish connection with [45]. (iv) Axon [72]. ...
... Certain transient protrusions like neurites transform into more permanent structures like dendrites and maintain this new identity throughout their lifetime [74]. In certain cases, filopodia get converted to cytoskeletal bridges [45]. ...
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A living cell uses long tubular appendages for locomotion and sensory purposes. Hence, assembling and maintaining a protrusion of correct length is crucial for its survival and overall performance. Usually the protrusions lack the machinery for the synthesis of building blocks and imports them from the cell body. What are the unique features of the transport logistics which facilitate the exchange of these building blocks between the cell and the protrusion? What kind of `rulers' and `timers' does the cell use for constructing its appendages of correct length on time? How do the multiple appendages coordinate and communicate among themselves during different stages of their existence? How frequently do the fluctuations drive the length of these dynamic protrusions out of the acceptable bounds? These questions are addressed from a broad perspective in this review which is organized in three parts. In part-I the list of all known cell protrusions is followed by a comprehensive list of the mechanisms of length control of cell protrusions reported in the literature. We review not only the dynamics of the genesis of the protrusions, but also their resorption and regrowth as well as regeneration after amputation. As a case study in part-II, the specific cell protrusion that has been discussed in detail is eukaryotic flagellum (also known as cilium); this choice was dictated by the fact that flagellar length control mechanisms have been studied most extensively over more than half a century in cells with two or more flagella. Although limited in scope, brief discussions on a few non-flagellar cell protrusions in part-III of this review is intended to provide a glimpse of the uncharted territories and challenging frontiers of research on subcellular length control phenomena that awaits vigorous investigations.
... Cytonemes can reach lengths of several hundred microns (most mammalian cells have a diameter of 10 to 100 microns). The length of the filopodia is consistent with the radius of distribution of signal molecules secreted [51,61,62]. These protrusions can deliver signals in both ways: from the sender to the recipient and back [48,54,60,[63][64][65][66][67]. ...
... The protrusions are heterogeneous [57] and can form, for example, TNTs or tumor microtubes (MTs), which consist of actin and function as intercellular bridges connecting a variety of cell types. TMs are longer and have a larger diameter compared with TNTs observed in vitro [58,60,62,68,69]. TNTs play an important role in signaling through various types of immune cells [70]. ...
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52 years have passed since President Nixon launched the "War on Cancer". The goals outlined by the President were not achieved because cancer treatment applied such as chemotherapy, radiotherapy, and targeted therapy have not fully met expectations. We suggest a new chemotherapeutic strategy: disrupting the communication between cancer cells and their microenvironment by chemical means. Immunological synapses that form between cancer cells and immune or other stromal cells provide an attractive target for this approach. Synapses form ligand-receptor clusters within interface of the interacting cells. Despite their differences, synapses share common properties: intercellular protein clusters; the proximity of these proteins; and their cooperative interaction making a synapse an unified functional unit. Synapses provide the limited space for the focused intercellular exchange of signaling molecules and particles. Therefore, destruction of synapses is expected to cause collapse of various tumor types. Additionally, the clustered arrangement of synapse components offers opportunities to increase treatment safety by reducing the concentration of cell impermeable agents used, to enhance its specificity applying cross-linking reagents, thus restricting modifications of surface-exposed molecules. By attaching a cleavable cell permeable toxic agent to a crosslinker should further enhance a killing potential in treating cancer. The proposed approach promises to be simple, universal, and less expensive than existing cancer therapy methods.
... The length of the filopodia is consistent with the radius of distribution of signal molecules secreted [39,64,65]. These protrusions can deliver signals in both ways: from the sender to the recipient and back [38,43,55,58,61,62,66,67]. The protrusions are heterogeneous [46] and can form, for example, TNTs or tumor microtubes (TMs), which consist of actin and function as intercellular bridges connecting a variety of cell types. ...
... These protrusions can deliver signals in both ways: from the sender to the recipient and back [38,43,55,58,61,62,66,67]. The protrusions are heterogeneous [46] and can form, for example, TNTs or tumor microtubes (TMs), which consist of actin and function as intercellular bridges connecting a variety of cell types. TMs are longer and have a larger diameter compared with TNTs observed in vitro [49,55,57,65,68]. Synapses are specific intercellular interfaces that have a unique membrane organization and provide relevant signal transmission between contacting cells. ...
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Fifty-two years have passed since President Nixon launched the “War on Cancer”. Despite unparalleled efforts and funds allocated worldwide, the outlined goals were not achieved because cancer treatment approaches such as chemotherapy, radiation therapy, hormonal and targeted therapies have not fully met the expectations. Based on the recent literature, a new direction in cancer therapy can be proposed which targets connections between cancer cells and their microenvironment by chemical means. Cancer–stromal synapses such as immunological synapses between cancer and immune cells provide an attractive target for this approach. Such synapses form ligand–receptor clusters on the interface of the interacting cells. They share a common property of involving intercellular clusters of spatially proximate and cooperatively acting proteins. Synapses provide the space for the focused intercellular signaling molecules exchange. Thus, the disassembly of cancer–stromal synapses may potentially cause the collapse of various tumors. Additionally, the clustered arrangement of synapse components offers opportunities to enhance treatment safety and precision by using targeted crosslinking chemical agents which may inactivate cancer synapses even in reduced concentrations. Furthermore, attaching a cleavable cell-permeable toxic agent(s) to a crosslinker may further enhance the anti-cancer effect of such therapeutics. The highlighted approach promises to be universal, relatively simple and cost-efficient. We also hope that, unlike chemotherapeutic and immune drugs that interact with a single target, by using supramolecular large clusters that include many different components as a target, the emergence of a resistance characteristic of chemo- and immunotherapy is extremely unlikely.
... However, various organizations of TNT-like structures can be observed, where membrane fusion and cytoplasmic connection may not necessarily occur. This depends on the cell type and the cellular context [30]. This variety of similar structures within the category of membrane protrusions has led to the use of several acceptable terms: thin membrane protrusions, membrane extensions, TNT-like protrusions, cellular bridges, specialized filopodia, and signaling filopodia [30]. ...
... This depends on the cell type and the cellular context [30]. This variety of similar structures within the category of membrane protrusions has led to the use of several acceptable terms: thin membrane protrusions, membrane extensions, TNT-like protrusions, cellular bridges, specialized filopodia, and signaling filopodia [30]. In this context, we include the TNT-like structures observed in G. duodenalis. ...
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Giardia doudenalis (lamblia, intestinalis) is a protozoan parasite that inhabits the lumen of the upper small intestine of vertebrates, causing chronic abdominal pains and severe diarrhea, symptoms of giardiasis, a persistent and recurrent infection. This characteristic is mainly due to the presence of membrane variant-specific surface proteins (VSPs) that give this parasite the ability to successively infect the host through antigenic variation. Using high-resolution scanning microscopy (HR-SM), we observed the presence, formation, and extension of tunneling-nanotube-like surface structures in Giardia, especially following parasite challenges with VSP antibodies. They were seen all over the parasite surface, both in vitro and in vivo, showing that G. duodenalis nanotube formation occurs in complex environments such as the gut. In addition, we also observed that some of these nanotubes displayed a periodic strangulation that produces 100 nm vesicles that seemed to be released in a process similar to that previously observed in Trypanosoma brucei. The presence of nanotube-like structures in G. duodenalis highlights yet another strategy of cellular communication utilized by these parasites, whether between themselves or with the host cell.
... In rat hippocampal neurons and astrocytes, p53 leads to caspase-3 activation, subsequently leading to the cleavage of the calcium-binding protein S100A4 in TNT-initiating cells. This consequently results in an extracellular gradient of S100A4 which was shown to direct TNT formation towards other cell [62] . It is yet the only known mechanism of guidance of TNTs via chemotactic cues, but it leads us to think that the general directionality of TNT growth might be regulated through similar processes. ...
... A major drawback of such a method is the increase in false negatives. This analysis allows the identification of a specific subtype of long, non-adherent TNTs, yet does not consider structures too close to the substrate or smaller than 10 μm [55][56][57][58][59][60][61][62][63][64][65][66][67][68] . Finally, the field of TNTs faces the same challenge as studies on EVs, or even filopodia. ...
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Over the years, the influence of secretory mechanisms on intercellular communication has been extensively studied. In the central nervous system (CNS), both trans-synaptic (neurotransmitter-based) and long-distance (extracellular vesicles-based) communications regulate activities and homeostasis. In less than a couple of decades, however, there has been a major paradigm shift in our understanding of intercellular communication. Increasing evidence suggests that besides secretory mechanisms (via extracellular vesicles), several cells are capable of establishing long-distance communication routes referred to as Tunneling Nanotubes (TNTs). TNTs are membranous bridges classically supported by F-Actin filaments, allowing for the exchange of different types of intracellular components between the connected cells, ranging from ions and organelles to pathogens and toxic protein aggregates. The roles of TNTs in pathological spreading of several neurodegenerative conditions such as Prion diseases, Alzheimer’s disease (AD), Parkinson’s disease (PD), and Huntington’s disease (HD) have been well established. However, the fragile nature of these structures and lack of specific biomarkers raised some skepticism regarding their existence. In this review, we will first place TNTs within the spectrum of intercellular communication mechanisms before discussing their known and hypothesized biological relevance in vitro and in vivo in physiological and neurodegenerative contexts. Finally, we discuss the challenges and promising prospects in the field of TNT studies.
... TNTs containing mitochondria have also been shown to be found between liver macrophages, cardiomyocytes, and cardiofibroblasts [105]. There is evidence of the presence of TNTs between cell types of different origins in vitro, namely, epithelial, endothelial, mesenchymal, neuronal, muscular, and immune cells [106]. ...
Article
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A number of our studies have shown that low-frequency (LF) oscillations in the functional parameters of the oxygen transport system are stable and synchronized with one another. The literature presents a large number of examples of LF oscillations in various functional parameters that are directly or indirectly related to energy metabolism. In parallel, artificially induced damped and constant spontaneous oscillations related to energy produced by the mitochondria over a range of LF frequencies have been studied for more than 40 years. A parameter study is therefore needed to find the connection between the oscillation amplitudes and the physical characteristics both of the oxygen transport system and mitochondria that operate on common LF range (0.003–0.03 Hz). We believe the nature of all these oscillation amplitudes to be affected by the periodic dynamics of energy dissipation in mitochondria that form an interconnected network. The process of creating these oscillations occurs in two phases. In the 1st phase, the amount of Са2+ entering the mitochondria exceeds the amount of Са2+ released by mitochondria thereby promoting an increase in oxidative phosphorylation efficiency. In the 2nd phase, Са2+ efflux from mitochondria prevails over Са2+ influx and is accompanied by inhibition of oxidative phosphorylation. The oscillations remain stable and spontaneous and arise from an “autocatalytic” interaction based on feedback mechanisms. The inertia of the processes of a full cycle (1st and 2nd phases) that lasts 1–3 minutes may be due to the capacity of the phosphate buffer of mitochondria. The structural basis for synchronizing oscillations at the tissue level may be mitochondrial networks of excitable tissues. Synchronization at the organism level between mitochondrial oscillations and fluctuations in parameters associated with energy metabolism can be achieved through a system of tunnel nanotubes.
... Importantly, macrophages can also affect the function of beta cells through other cytokines or direct interactions, such as the formation of tunneling nanotubes and the phagocytosis of insulin vesicles. 16,48 Our study did not investigate whether NL-SDT plays a role in other interactions between macrophages and beta cells. Furthermore, the mechanisms underlying DRP1 activation during NL-SDT are not fully understood, and further exploration is needed. ...
Article
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Obesity plays a crucial role in the development and progression of type 2 diabetes mellitus (T2DM) by causing excessive release of free fatty acid from adipose tissue, which in turn leads to systemic infiltration of macrophages. In individuals with T2DM, the infiltration of macrophages into pancreatic islets results in islet inflammation that impairs beta cell function, as evidenced by increased apoptosis and decreased glucose‐stimulated insulin secretion. The present study aimed to investigate the effects of non‐lethal sonodynamic therapy (NL‐SDT) on bone marrow‐derived macrophages (BMDMs) exposed to high glucose and palmitic acid (HG/PA). These findings indicate that NL‐SDT facilitates the expression of DRP1 through the transient production of mitochondrial ROS, which subsequently promotes mitophagy. This mitophagy was shown to limit the activation of the NLRP3 inflammasome and the secretion of IL‐1β in BMDMs exposed to HG/PA. In co‐culture experiments, beta cells exhibited significant dysfunction when interacting with HG/PA‐treated BMDMs. However, this dysfunction was markedly alleviated when the BMDMs had undergone NL‐SDT treatment. Moreover, NL‐SDT was found to lower blood glucose levels and elevate serum insulin concentrations in db/db mice. Furthermore, NL‐SDT effectively reduced the infiltration of F4/80‐positive macrophages and the expression of CASP1 within islets. These findings provide fundamental insights into the mechanisms through which NL‐SDT may serve as a promising approach for the treatment of T2DM.
... By direct we mean a sense that could (almost) exactly signal the current position of another individual, for example, through directly touching or sighting the neighbour; indirect refers to detection through an intermediary, such as chemical cues or tracks left by the neighbour, that may indicate its recent presence. Whether a sensing is direct or indirect, within heterogeneous groups the interaction ranges of the homotypic and heterotypic interactions could vary considerably: animals range widely with respect to the form and range of their sensory systems-certain species of baleen whales are believed to be able to communicating over 100s of kilometres [9]; different cells can extend a spectrum of cell protrusions from shorter to long range [10]. ...
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In a chase-and-run dynamic, the interaction between two individuals is such that one moves towards the other (the chaser), while the other moves away (the runner). Examples can be found in both interacting cells and animals. Here, we investigate the behaviours that can emerge at a population level, for a heterogeneous group that contains subpopulations of chasers and runners. We show that a wide variety of patterns can form, from stationary patterns to oscillatory and population-level chase-and-run, where the latter describes a synchronized collective movement of the two populations. We investigate the conditions under which different behaviours arise, specifically focusing on the interaction ranges: the distances over which cells or organisms can sense one another’s presence. We find that when the interaction range of the chaser is sufficiently larger than that of the runner—or when the interaction range of the chase is sufficiently larger than that of the run—population-level chase-and-run emerges in a robust manner. We discuss the results in the context of phenomena observed in cellular and ecological systems, with particular attention to the dynamics observed experimentally within populations of neural crest and placode cells.
... Было также показано, что TNT, содержащие митохондрии, обнаружены между макрофагами печени, кардиомиоцитами и кардиофибробластами [105]. Имеются данные о присутствии TNT между типами клеток различного происхождения in vitro, а именно эпителиальными, эндотелиальными, мезенхимальными, нейрональными, мышечными и иммунными [106]. ...
... [9][10][11][12][13] Potential approaches for mitochondrial transfer comprise tunneling nanotubes (TNTs), extracellular vesicles (EVs), gap junctions, various cytoplasmic bridges, and cell fusion. [14][15][16][17][18][19][20][21] Among these, TNTs are considered a major intercellular conduit for mitochondrial exchanges, and relevant mechanisms have been widely reported. By contrast, studies on cell-fusion approaches are relatively scarce, and most cell fusions in these studies refer to partial cell fusion. ...
... TNTs containing mitochondria have also been shown to be found between liver macrophages, cardiomyocytes, and cardiofibroblasts [105]. There is evidence of the presence of TNTs between cell types of different origins in vitro, namely, epithelial, endothelial, mesenchymal, neuronal, muscular, and immune cells [106]. ...
Article
Full-text available
It has been shown in a number of our studies that low-frequency (LF) oscillations in the functional parameters of the oxygen transport system are stable and synchronized with each other. The literature presents a large number of examples of LF oscillations of various functional indicators that are directly or indirectly related to energy metabolism. In parallel, for more than 40 years, artificially induced attenuated and constant spontaneous oscillations in the energization levels of mitochondria in the same LF range have been studied. The aim of this review is to consider a possible relationship between oscillations in the functional parameters of the oxygen transport system and the functional parameters of mitochondria in the very-low-frequency (VLF) range common to them (0.003–0.03 Hz). We believe that a common source for all these oscillations is the periodic dynamics of “energization” in mitochondria united in mitochondrial networks. The process of generating these oscillations proceeds in two phases. In the first phase, the inflow of Ca2+ into the mitochondria exceeds the outflow and enhances the activity of oxidative phosphorylation. In the second phase, the outflow of Ca2+ from the mitochondria prevails over the inflow and is accompanied by the inhibition of oxidative phosphorylation. The oscillations are of a constant spontaneous nature and are based on autocatalytic regulation based on the feedback principle. The inertia of the full cycle processes (first and second phases) lasting 1–3 min may be due to the capacity of the mitochondrial phosphate buffer. The mitochondrial networks of excitable tissues can be the structural basis for synchronizing oscillations at the tissue level. Synchronization at the body level between mitochondrial oscillations and oscillations in indicators related to energy metabolism can be carried out through a system of tunneling nanotubes.
... By direct we mean a sense that could (almost) exactly signal the current position of another individual, for example through directly touching or sighting the neighbour; indirect refers to detection through an intermediary, such as chemical cues or tracks left by the neighbour, that may indicate its recent presence. Whether a sensing is direct or indirect, within heterogeneous groups the interaction ranges of the homotypic and heterotypic interactions could vary considerably: animals range widely with respect to the form and range of their sensory systemscertain species of baleen whales are believed to be possible of communicating over 100s of kilometres [9]; different cells can extend a spectrum of cell protrusions from shorter to long range [10]. ...
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In a chase-and-run dynamic, the interaction between two individuals is such that one moves towards the other (the chaser), while the other moves away (the runner). Examples can be found in both interacting cells and interacting animals. Here we investigate the behaviours that can emerge at a population level, for a heterogeneous group that contains subpopulations of chasers and runners. We show that a wide variety of patterns can form, from stationary patterns to oscillatory and population-level chase-and-run, where the latter describes a synchronised collective movement of the two populations. We investigate the conditions under which different behaviours arise, specifically focusing on the interaction ranges: the distances over which cells or organisms can sense one another's presence. We find that when the interaction range of the chaser is sufficiently larger than that of the runner - or when the interaction range of the chase is sufficiently larger than that of the run -population-level chase-and-run emerges in a robust manner. We discuss the results in the context of phenomena observed in cellular and ecological systems, with particular attention to the dynamics observed experimentally within populations of neural crest and placode cells.
... All of them are extended by either signal-sending or signal-receiving cells or both, establishing physical contact with their specific target cells. Signaling molecules, including major morphogens, are moved along the protrusions, or packaged into the vesicles and delivered (Kornberg and Roy, 2014b;Roy et al., 2011;Yamashita et al., 2018). ...
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Specialized cellular protrusions facilitate local intercellular communications in various species, including mammals. Among these, airinemes play a crucial role in pigment pattern formation in zebrafish by mediating long-distance Notch signaling between pigment cells. Remarkably, airinemes exhibit large vesicle-like structure at their tips, which are pulled by a macrophage subpopulation and delivered to target cells. The interaction between macrophages and Delta-ligand carrying airineme vesicles is essential for initiating airineme-mediated signaling, yet the molecular detail of this interaction remains elusive. Through high-resolution live imaging and genetic in vivo manipulations, we found that adhesive interactions via the extracellular domain of CD44, a class I transmembrane glycoprotein, between macrophages and airineme vesicles are critical for airineme signaling. Mutants lacking the extracellular domain of CD44 lose their adhesiveness, resulting in a significant reduction in airineme extension and pigment pattern defects. Our findings provide valuable insights into the role of adhesive interactions between signal-sending cells and macrophages in long-range intercellular signaling.
... Cargo passes through TNTs bidirectionally either in a native state, as in the case of organelles and large molecules, or encapsulated in vesicles for nucleic acids. Unlike gap junctions, TNTs do not allow passive diffusion so calcium transients do not propagate, and membrane-embedded proteins cannot diffuse laterally (Yamashita et al., 2018). TNTs might therefore serve as a faster, more direct method of intercellular communication than exo-or endocytosis, although it is unknown whether EVs released into the extracellular milieu have different cargo compared to TNT EVs (Driscoll et al., 2022). ...
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Non‐coding RNAs (ncRNAs) are highly plastic RNA molecules that can sequester cellular proteins and other RNAs, serve as transporters of cellular cargo and provide spatiotemporal feedback to the genome. Mounting evidence indicates that ncRNAs are central to biology, and are critical for neuronal development, metabolism and intra‐ and intercellular communication in the brain. Their plasticity arises from state‐dependent dynamic structure states that can be influenced by cell type and subcellular environment, which can subsequently enable the same ncRNA with discrete functions in different contexts. Here, we highlight different classes of brain‐enriched ncRNAs, including microRNA, long non‐coding RNA and other enigmatic ncRNAs, that are functionally important for both learning and memory and adaptive immunity, and describe how they may promote cross‐talk between these two evolutionarily ancient biological systems. image
... CoNPs, TiO 2 NPs and MWCNTs are widely accepted as engineered nanomaterials that can enter the body and reach the central nervous system, raising concerns about their neurotoxicity. The original discovery of TNTs was closely related to homeostasis and pathogenesis [36,37], especially in neurotoxicity induced by oxidative stress [38]. Here, we show, for the first time, that three types of engineered nanomaterials can promote TNTs formation and mitochondrial transfer via the induction of oxidative stress, a common protective strategy in response to nanomaterial exposure that restores ATP production and cell viability. ...
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Background As the demand and application of engineered nanomaterials have increased, their potential toxicity to the central nervous system has drawn increasing attention. Tunneling nanotubes (TNTs) are novel cell–cell communication that plays a crucial role in pathology and physiology. However, the relationship between TNTs and nanomaterials neurotoxicity remains unclear. Here, three types of commonly used engineered nanomaterials, namely cobalt nanoparticles (CoNPs), titanium dioxide nanoparticles (TiO2NPs), and multi-walled carbon nanotubes (MWCNTs), were selected to address this limitation. Results After the complete characterization of the nanomaterials, the induction of TNTs formation with all of the nanomaterials was observed using high-content screening system and confocal microscopy in both primary astrocytes and U251 cells. It was further revealed that TNT formation protected against nanomaterial-induced neurotoxicity due to cell apoptosis and disrupted ATP production. We then determined the mechanism underlying the protective role of TNTs. Since oxidative stress is a common mechanism in nanotoxicity, we first observed a significant increase in total and mitochondrial reactive oxygen species (namely ROS, mtROS), causing mitochondrial damage. Moreover, pretreatment of U251 cells with either the ROS scavenger N-acetylcysteine or the mtROS scavenger mitoquinone attenuated nanomaterial-induced neurotoxicity and TNTs generation, suggesting a central role of ROS in nanomaterials-induced TNTs formation. Furthermore, a vigorous downstream pathway of ROS, the PI3K/AKT/mTOR pathway, was found to be actively involved in nanomaterials-promoted TNTs development, which was abolished by LY294002, Perifosine and Rapamycin, inhibitors of PI3K, AKT, and mTOR, respectively. Finally, western blot analysis demonstrated that ROS and mtROS scavengers suppressed the PI3K/AKT/mTOR pathway, which abrogated TNTs formation. Conclusion Despite their biophysical properties, various types of nanomaterials promote TNTs formation and mitochondrial transfer, preventing cell apoptosis and disrupting ATP production induced by nanomaterials. ROS/mtROS and the activation of the downstream PI3K/AKT/mTOR pathway are common mechanisms to regulate TNTs formation and mitochondrial transfer. Our study reveals that engineered nanomaterials share the same molecular mechanism of TNTs formation and intercellular mitochondrial transfer, and the proposed adverse outcome pathway contributes to a better understanding of the intercellular protection mechanism against nanomaterials-induced neurotoxicity. Graphical abstract
... In the search to identify the causative mechanism for MT, several possibilities have emerged, including cell fusion, endocytosis (Generous et al., 2019), and gap junctions (Valiunas et al., 2005). Recent transplantation studies determined that tunneling nanotubes (TNTs), consisting of actin-rich cytoplasmic bridges connecting neighboring cells (Rustom et al., 2004;Yamashita et al., 2018), mediate donor-to-host photoreceptor MT in photoreceptors (Kalargyrou et al., 2021;Ortin-Martinez et al., 2021). Nanotube-mediated horizontal MT also occurs between endogenous photoreceptors during development (Kalargyrou et al., 2021;Heisterkamp et al., 2022). ...
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Intercellular cytoplasmic material transfer (MT) occurs between transplanted and developing photoreceptors and ambiguates cell origin identification in developmental, transdifferentiation, and transplantation experiments. Whether MT is a photoreceptor-specific phenomenon is unclear. Retinal ganglion cell (RGC) replacement, through transdifferentiation or transplantation, holds potential for restoring vision in optic neuropathies. During careful assessment for MT following human stem cell-derived RGC transplantation into mice, we identified RGC xenografts occasionally giving rise to labeling of donor-derived cytoplasmic, nuclear, and mitochondrial proteins within recipient Muller glia. Critically, nuclear organization is distinct between human and murine retinal neurons, which enables unequivocal discrimination of donor from host cells. MT was greatly facilitated by internal limiting membrane disruption, which also augments retinal engraftment following transplantation. Our findings demonstrate that retinal MT is not unique to photoreceptors and challenge the isolated use of species-specific immunofluorescent markers for xenotransplant identification. Assessment for MT is critical when analyzing neuronal replacement interventions.
... It's reported that CoNPs, TiO 2 NPs and MWCNTs were wildly accepted as engineering nanomaterials, which could enter the body and reach the central nervous system, which arising concern about their neurotoxicity. The original discovery of TNTs was closely related to homeostasis and pathogenesis 31,32 , especially in neurotoxicity induced by oxidative stress 33 . Here we show, for the rst time, that three types of engineering nanomaterials could promote TNTs formation and mitochondrial transfer via the induction of oxidative stress is a common protective strategy in response to nanomaterials exposure, which restores ATP production and cell viability. ...
Preprint
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Background As the demand and application of engineering nanomaterials rise, their potential toxicity in the central nervous system has drawn increasing concerns. As a novel cell-cell communication, tunneling nanotubes (TNTs) plays a crucial role in pathology and physiology. Unfortunately, the relationship between TNTs and nanomaterials neurotoxicity remains scarce. Here, three types of commonly used engineering nanomaterials, Cobalt nanoparticles (CoNPs), titanium dioxide nanoparticles (TiO2NPs), and multi-walled carbon nanotubes (MWCNTs) were selected to address this limitation. Results After complete characterization of the nanomaterials, the induction of TNTs formation by all of the nanomaterials was observed by high-content screening system and confocal microscopy in both primary astrocyte and U251 cells. The positive role of TNTs formation was further unveiled, which protected against the nanomaterials-induced neurotoxicity from cell apoptosis to ATP production dysfunction. We then ought to determine the underline mechanism of TNTs positive role. Since the generation of oxidative stress is a common mechanism in nanotoxicity, we first observed a significant increase in total and mitochondrial reactive oxygen species (namely ROS, mtROS), causing mitochondrial damage. Moreover, the pretreatment of U251 cells with either a ROS scavenger N-acetylcysteine or a mtROS scavenger mitoquinone attenuated the nanomaterial-induced neurotoxicity and TNTs generation, suggesting the central role of ROS in nanomaterials-induced TNTs formation. Furthermore, a vigorous downstream pathway of ROS, i.e., PI3K/AKT/mTOR pathway was found to be actively involved in the nanomaterials-promoted TNTs development, which was abolished by LY294002, Perifosine and Rapamycin, the inhibitors of PI3K, AKT, and mTOR, respectively. Finally, Western blot analysis demonstrated that ROS and mtROS scavenger suppressed PI3K/AKT/mTOR pathway, which then abrogated TNTs formation. Conclusion Our findings indicate that various types of nanomaterials promote TNTs formation through the generation of ROS/mtROS and the activation of downstream PI3K/AKT/mTOR pathway, which could fight against cell apoptosis and ATP production dysfunction induced by nanomaterials. Our study contributes to a better understanding of the intercellular protection mechanism against neurotoxicity induced by various kinds of nanomaterials, and sheds light on potential treatments to activate the cell-cell defense system against environmental toxicants.
... First, the cell bodies can be highly deformable, where frequent protrusions of the membrane -pseudopodia [51] -locally extend parts of the membrane far beyond the average diameter. Second, a diversity of more specialised membrane protrusions have been identified [219,121,180] -variously termed cytonemes, tunnelling nanotubes, microtubes -that in some cases extend the order of 100s of microns. Thus, a contact can be achieved between cells separated by multiple cell diameters, and a non-local description is warranted. ...
Preprint
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The employment of nonlocal PDE models to describe biological aggregation and other phenomena has gained considerable traction in recent years. For cell populations, these methods grant a means of accommodating essential elements such as cell adhesion, critical to the development and structure of tissues. For animals, they can be used to describe how the nearby presence of conspecifics and/or heterospecifics influence movement behaviour. In this review, we will focus on classes of biological movement models in which the advective (or directed) component to motion is governed by an integral term that accounts for how the surrounding distribution(s) of the population(s) impact on a member's movement. We recount the fundamental motivation for these models: the intrinsic capacity of cell populations to self-organise and spatially sort within tissues; the wide-ranging tendency of animals towards spatial structuring, from the formations of herds and swarms to territorial segregation. We examine the derivation of these models from an individual level, illustrating in the process methods that allow models to be connected to data. We explore a growing analytical literature, including methods of stability and bifurcation analysis, and existence results. We conclude with a short section that lays out some future challenges and connections to the modelling of sociological phenomena including opinion dynamics.
... In addition to cytonemes, closed membrane protrusions also include microtubulebased nanotubes (MT-nanotubes) [163], while tunneling nanotubes (TNTs) and membrane nanotubes (cell tubes, bridges or ring channels) are mostly actin-based structures with an open end, although in some cases microtubules are also present in their cytoskeletons [164]. Thanks to tunneling and membrane nanotubes, the exchange and transfer of soluble cytoplasmic components, molecules, vesicles, organelles and pathogens between cells is ensured [165,166]. ...
Article
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In multicellular organisms, interactions between cells and intercellular communications form the very basis of the organism’s survival, the functioning of its systems, the maintenance of homeostasis and adequate response to the environment. The accumulated experimental data point to the particular importance of intercellular communications in determining the fate of cells, as well as their differentiation and plasticity. For a long time, it was believed that the properties and behavior of cells were primarily governed by the interactions of secreted or membrane-bound ligands with corresponding receptors, as well as direct intercellular adhesion contacts. In this review, we describe various types of other, non-classical intercellular interactions and communications that have recently come into the limelight—in particular, the broad repertoire of extracellular vesicles and membrane protrusions. These communications are mediated by large macromolecular structural and functional ensembles, and we explore here the mechanisms underlying their formation and present current data that reveal their roles in multiple biological processes. The effects mediated by these new types of intercellular communications in normal and pathological states, as well as therapeutic applications, are also discussed. The in-depth study of novel intercellular interaction mechanisms is required for the establishment of effective approaches for the control and modification of cell properties both for basic research and the development of radically new therapeutic strategies.
... Ca 2+ , cAMP, ATP) has been proposed to mediate both tissue morphogenesis and homeostasis [39,40] (Fig. 2A). Distant cells can also establish direct contacts through nanotubes and cytonemes, which are thin membranous protrusions that span several cell diameters and facilitate intercellular signalling [41,42] (Fig. 2B). For example, tunneling nanotubes were reported to facilitate Sonic Hedgehog (SHH) signalling between cells during limb bud patterning in chick embryos [43]. ...
Article
The process by which biological systems such as cells, tissues and organisms acquire shape has been named as morphogenesis and it is central to a plethora of biological contexts including embryo development, wound healing, or even cancer. Morphogenesis relies in both self-organising properties of the system and in environmental inputs (biochemical and biophysical). The classical view of morphogenesis is based on the study of external biochemical molecules, such as morphogens. However, recent studies are establishing that the mechanical environment is also used by cells to communicate within tissues, suggesting that this mechanical crosstalk is essential to synchronise morphogenetic transitions and self-organisation. In this article we discuss how tissue interaction drive robust morphogenesis, starting from a classical biochemical view, to finalise with more recent advances on how the biophysical properties of a tissue feedback with their surroundings to allow form acquisition. We also comment on how in silico models aid to integrate and predict changes in cell and tissue behaviour. Finally, considering recent advances from the developmental biomechanics field showing that mechanical inputs work as cues that promote morphogenesis, we invite to revisit the concept of morphogen.
... This may be of particular interest to increase the efficiency of assisted reproductive technologies. More broadly, our results may be valuable for the comprehension of other cellular models relying on distant cell-cell contact communication to function (48). It may notably provide clues as to how cellular development is modulated by exogenous regulation of gene expression through direct intercellular dialogue. ...
Article
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The oocyte must grow and mature before fertilization, thanks to a close dialogue with the somatic cells that surround it. Part of this communication is through filopodia-like protrusions, called transzonal projections (TZPs), sent by the somatic cells to the oocyte membrane. To investigate the contribution of TZPs to oocyte quality, we impaired their structure by generating a full knockout mouse of the TZP structural component myosin-X (MYO10). Using spinning disk and super-resolution microscopy combined with a machine-learning approach to phenotype oocyte morphology, we show that the lack of Myo10 decreases TZP density during oocyte growth. Reduction in TZPs does not prevent oocyte growth but impairs oocyte-matrix integrity. Importantly, we reveal by transcriptomic analysis that gene expression is altered in TZP-deprived oocytes and that oocyte maturation and subsequent early embryonic development are partially affected, effectively reducing mouse fertility. We propose that TZPs play a role in the structural integrity of the germline–somatic complex, which is essential for regulating gene expression in the oocyte and thus its developmental potential.
... This foregrounds the imperatives for shielding or sequestering various internal states from other internal states, which brings us back to MBs; however, these are internal MBs that define an internal morphology or message-passing structure. One might conjecture that much of biological self-organization is concerned with isolation and shielding, as a necessary part of internal autopoiesis (e.g., the role of enzymes and catalysts, gap junctions, and many other highly controllable mechanisms for setting up signaling paths and boundaries [98,99,100,101]. This occurs at all scales, from subcellular organelles that partition biophysical and chemical reactions to nascent organ compartment boundaries, to the dynamics that guide which members of a swarm pass messages to which others [102,103,104,105,106]. ...
Article
Full-text available
We show how any finite physical system with morphological, i.e. three-dimensional embedding or shape, degrees of freedom and locally limited free energy will, under the constraints of the free energy principle, evolve over time towards a neuromorphic morphology that supports hierarchical computations in which each “level” of the hierarchy enacts a coarse-graining of its inputs, and dually, a fine-graining of its outputs. Such hierarchies occur throughout biology, from the architectures of intracellular signal transduction pathways to the large-scale organization of perception and action cycles in the mammalian brain. Formally, the close formal connections between cone-cocone diagrams (CCCD) as models of quantum reference frames on the one hand, and between CCCDs and topological quantum field theories on the other, allow the representation of such computations in the fully-general quantum-computational framework of topological quantum neural networks.
... But how is canonical Wnt signal transduction spatially restricted to four or five layers of dermal fibroblast progenitors between E11.5 and E13.5? The range of diffusion-mediated Wnt signaling varies widely depending on the context and there is emerging evidence that Wnt ligands can be moved from their site of synthesis via membrane extensions, exosomes and microvesicles (Parchure et al., 2018;Stewart et al., 2019;Yamashita et al., 2018). In other contexts, Wnt signaling can be regulated by epigenetic regulators, such as polycomb repressive complex 2 (PRC2) (Nehila et al., 2020). ...
Article
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Skin is largely composed of an epidermis that overlies a supporting dermis. Recent advancements in our understanding of how diverse groups of dermal fibroblasts regulate epidermal and hair follicle growth and differentiation have been fueled by tools capable of resolving molecular heterogeneity at a single-cell level. Fibroblast heterogeneity can be traced back to their developmental origin before their segregation into spatially distinct fibroblast subtypes. The mechanisms that drive this lineage diversification during development are being unraveled, with studies showing that both large- and small-scale positional signals play important roles during dermal development. Here, we first delineate what is known about the origins of the dermis and the central role of Wnt/β-catenin signaling in its specification across anatomical locations. We then discuss how one of the first morphologically recognizable fibroblast subtypes, the hair follicle dermal condensate lineage, emerges. Leveraging the natural variation of skin and its appendages between species and between different anatomical locations, these collective studies have identified shared and divergent factors that contribute to the extraordinary diversity of skin.
... How do cytonemes remain in contact during either loading or unloading of their cargo, and when vesicles are involved in PIT cytonemes, what determines when the transfer process stops? A number of recent reviews cover the variety of systems in which cytonemes play a role [10,23,37,41,49], but much remains to characterize the details of the processes involved. ...
Preprint
Spatial distributions of morphogens provide positional information in developing systems, but how the distributions are established and maintained remains an open problem. Transport by diffusion has been the traditional mechanism, but recent experimental work has shown that cells can also communicate by filopodia-like structures called cytonemes that make direct cell-to-cell contacts. Here we investigate the roles each may play individually in a complex tissue and how they can jointly establish a reliable spatial distribution of a morphogen.
... Rustom et al. (2004), who first described the yet unknown type of ultrafine intercellular connections, referred to as TNTs, which are formed a few hours after cell plating. They are very similar to another kind of membrane bridges, named cytonemes, described as filopodia-like structures connecting cellular membranes [2,3]. However, cytonemes have been reported to serve as tracks for membrane-associated molecules, which moved outside them, while TNTs are able to transfer inside them cytoplasmic materials [2]. ...
Article
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Cellular communication and the transfer of information from one cell to another is crucial for cell viability and homeostasis. During the last decade, tunneling nanotubes (TNTs) have attracted scientific attention, not only as a means of direct intercellular communication, but also as a possible system to transport biological cargo between distant cells. Peculiar TNT characteristics make them both able to increase cellular survival capacities, as well as a potential target of neurodegenerative disease progression. Despite TNT formation having been documented in a number of cell types, the exact mechanisms triggering their formation are still not completely known. In this review, we will summarize and highlight those studies focusing on TNT formation in the nervous system, as well as their role in neurodegenerative diseases. Moreover, we aim to stress some possible mechanisms and important proteins probably involved in TNT formation in the nervous system.
... This foregrounds the imperatives for shielding or sequestering various internal states from other internal states, which brings us back to MBs; however, these are internal MBs that define an internal morphology or message-passing structure. One might conjecture that much of biological self-organization is concerned with isolation and shielding, as a necessary part of internal autopoiesis (e.g. the role of enzymes and catalysts, gap junctions, and many other highly controllable mechanisms for setting up signaling paths and boundaries [98][99][100][101]. This occurs at all scales, from subcellular organelles that partition biophysical and chemical reactions to nascent organ compartment boundaries, to the dynamics that guide which members of a swarm pass messages to which others [102][103][104][105][106]. ...
Preprint
We show how any system with morphological degrees of freedom and locally limited free energy will, under the constraints of the free energy principle, evolve toward a neuromorphic morphology that supports hierarchical computations in which each level of the hierarchy enacts a coarse-graining of its inputs, and dually a fine-graining of its outputs. Such hierarchies occur throughout biology, from the architectures of intracellular signal transduction pathways to the large-scale organization of perception and action cycles in the mammalian brain. Formally, the close formal connections between cone-cocone diagrams (CCCD) as models of quantum reference frames on the one hand, and between CCCDs and topological quantum field theories on the other, allow the representation of such computations in the fully-general quantum-computational framework of topological quantum neural networks.
... Les cellules tumorales augmentent ainsi leur nombre de mitochondries saines et donc leur production d'ATP [49]. Les mitochondries circulent d'une cellule à l'autre en utilisant des ponts cytoplasmiques, par un mécanisme actif dans lequel Miro1 [50] transporte les mitochondries le long des microtubules [51,52]. Dans les cellules tumorales, un des principaux mécanismes impliqués dans ce type de transfert mitochondrial est la constitution de nanotubes formant des tunnels (TNT, pour tunneling nanotubes), des nanostructures composées de filaments d'actine partagés entre les deux cellules [53]. ...
Article
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La reprogrammation métabolique est l’un des marqueurs de la carcinogenèse. Au cœur de cette reprogrammation se trouvent les mitochondries qui produisent l’énergie sous forme de molécules d’ATP. La régulation spatio-temporelle de la production d’ATP, indispensable pour fournir l’énergie au bon endroit et au bon moment, est assurée par le transport intracellulaire des mitochondries. Les complexes Miro/TRAK présents à la surface des mitochondries se lient aux protéines motrices de la cellule (dynéine, kinésine, myosine) pour transporter les mitochondries le long du cytosquelette. Ces acteurs du transport mitochondrial sont souvent dérégulés dans le cancer. Nous présentons dans cette revue les mécanismes par lesquels le transport mitochondrial contribue à la migration, à la division cellulaire et à la réponse au stress des cellules cancéreuses. Décrypter ces mécanismes pourrait ouvrir la voie à de nouvelles approches thérapeutiques en oncologie.
... In addition to the juxtacrine signaling type of contact-dependent cellto-cell communication, the formation of thin membrane protrusions that connect cells located far apart from each other warrants attention. Currently, these thin cellular protrusions are structurally classified into three major groups based on their cytoskeletal components and on their tip shapes (Yamashita et al., 2018). The first group encompasses open-ended and actin-based connections termed tunneling nanotubes (TNTs) that enable the exchange of soluble cytoplasmic components and intracellular vesicles between connecting cells (Gerdes and Carvalho, 2008;Rustom, 2016;Rustom et al., 2004). ...
Article
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Actin-based protrusions called cytonemes are reported to function in cell communication by supporting events such as morphogen gradient establishment and pattern formation. Despite the crucial roles of cytonemes in cell signaling, the molecular mechanism for cytoneme establishment remains elusive. In this study, we showed that the leukocyte common antigen-related (LAR) receptor protein tyrosine phosphatase plays an important role in cytoneme-like protrusion formation. Overexpression of LAR in HEK293T cells induced the formation of actin-based protrusions, some of which exceeded 200 µm in length and displayed a complex morphology with branches. Upon focusing on the regulation of LAR dimerization or clustering and the resulting regulatory effects on LAR phosphatase activity, we found that longer and more branched protrusions were formed when LAR dimerization was artificially induced and when heparan sulfate was applied. Interestingly, although the truncated form of LAR lacking phosphatase-related domains promoted protrusion formation, the phosphatase-inactive forms did not show clear changes, suggesting that LAR dimerization triggers the formation of cytoneme-like protrusions in a phosphatase-independent manner. Our results thus emphasize the importance of LAR and its dimerization in cell signaling. This article has an associated First Person interview with the first author of the paper.
... Moreover, other ways of introducing long-range cell-cell communication are possible: examples include the exchange of molecules through cellular protrusions, such as filopodia or cytonemes [57,58], or other cellular channels, such as epithelial bridges [59] and tunnelling nanotubes [60,61]. Our model could account for this means of intercellular communication, replacing the diffusion in the lateral intercellular space with that through cellular protrusions. ...
Article
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Cells can measure shallow gradients of external signals to initiate and accomplish a migration or a morphogenetic process. Recently, starting from mathematical models like the local-excitation global-inhibition (LEGI) model and with the support of empirical evidence, it has been proposed that cellular communication improves the measurement of an external gradient. However, the mathematical models that have been used have over-simplified geometries (e.g., they are uni-dimensional) or assumptions about cellular communication, which limit the possibility to analyze the gradient sensing ability of more complex cellular systems. Here, we generalize the existing models to study the effects on gradient sensing of cell number, geometry and of long- versus short-range cellular communication in 2D systems representing epithelial tissues. We find that increasing the cell number can be detrimental for gradient sensing when the communication is weak and limited to nearest neighbour cells, while it is beneficial when there is long-range communication. We also find that, with long-range communication, the gradient sensing ability improves for tissues with more disordered geometries; on the other hand, an ordered structure with mostly hexagonal cells is advantageous with nearest neighbour communication. Our results considerably extend the current models of gradient sensing by epithelial tissues, making a step further toward predicting the mechanism of communication and its putative mediator in many biological processes.
Article
Spatial distributions of morphogens provide positional information in developing systems, but how the distributions are established and maintained remains an open problem. Transport by diffusion has been the traditional mechanism, but recent experimental work has shown that cells can also communicate by filopodia-like structures called cytonemes that make direct cell-to-cell contacts. Here we investigate the roles each may play individually in a complex tissue and how they can jointly establish a reliable spatial distribution of a morphogen. To this end, we formulate models that capture fundamental aspects of various cytoneme-based transport mechanisms. In simple cases, exact solutions are attainable, and in more complex cases, we discuss results of numerical simulations.
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Mitophagy is the intracellular recycling system that disposes damaged/inefficient mitochondria and allows biogenesis of new organelles to ensure mitochondrial quality is optimized. Dysfunctional mitophagy has been implicated in human aging and diseases. Multiple evolutionarily selected, redundant mechanisms of mitophagy have been identified, but their specific roles in human health and their potential exploitation as therapeutic targets are unclear. Recently, the characterization of the endosomal−lysosomal system has revealed additional mechanisms of mitophagy and mitochondrial quality control that operate via the production of mitochondria-derived vesicles (MDVs). Circulating MDVs can be isolated and characterized to provide an unprecedented opportunity to study this type of mitochondrial recycling in vivo and to relate it to human physiology and pathology. Defining the role of MDVs in human physiology, pathology, and aging is hampered by the lack of standardized methods to isolate, validate, and characterize these vesicles. Hence, some basic questions about MDVs remain unanswered. While MDVs are generated directly through the extrusion of mitochondrial membranes within the cell, a set of circulating extracellular vesicles leaking from the endosomal−lysosomal system and containing mitochondrial portions have also been identified and warrant investigation. Preliminary research indicates that MDV generation serves multiple biological roles and contributes to restoring cell homeostasis. However, studies have shown that MDVs may also be involved in pathological conditions. Therefore, further research is warranted to establish when/whether MDVs are supporting disease progression and/or are extracting damaged mitochondrial components to alleviate cellular oxidative burden and restore redox homeoastasis. This information will be relevant for exploiting these vesicles for therapeutic purpose. Herein, we provide an overview of preclinical and clinical studies on MDVs in aging and associated conditions and discuss the interplay between MDVs and some of the hallmarks of aging (mitophagy, inflammation, and proteostasis). We also outline open questions on MDV research that should be prioritized by future investigations.
Chapter
Intercellular communication is indispensable across multicellular organisms, and any aberration in this process can give rise to significant anomalies in developmental and homeostatic processes. Thus, a comprehensive understanding of its mechanisms is imperative for addressing human health-related concerns. Recent advances have expanded our understanding of intercellular communication by elucidating additional signaling modalities alongside established mechanisms. Notably, cellular protrusion-mediated long-range communication, characterized by physical contact through thin and elongated cellular protrusions between cells involved in signal transmission and reception, has emerged as a significant intercellular signaling paradigm. This chapter delves into the exploration of a signaling cellular protrusion termed ‘airinemes,’ discovered in the zebrafish skin. It covers their identified signaling roles and the cellular and molecular mechanisms that underpin their functionality.
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From the moment a cell is on the path to malignant transformation, its interaction with other cells from the microenvironment becomes altered. The flow of molecular information is at the heart of the cellular and systemic fate in tumors, and various processes participate in conveying key molecular information from or to certain cancer cells. For instance, the loss of tight junction molecules is part of the signal sent to cancer cells so that they are no longer bound to the primary tumors and are thus free to travel and metastasize. Upon the targeting of a single cell by a therapeutic drug, gap junctions are able to communicate death information to by-standing cells. The discovery of the importance of novel modes of cell–cell communication such as different types of extracellular vesicles or tunneling nanotubes is changing the way scientists look at these processes. However, are they all actively involved in different contexts at the same time or are they recruited to fulfill specific tasks? What does the multiplicity of modes mean for the overall progression of the disease? Here, we extend an open invitation to think about the overall significance of these questions, rather than engage in an elusive attempt at a systematic repertory of the mechanisms at play.
Article
Tunnelling nanotubes (TNTs) connect distant cells and mediate cargo transfer for intercellular communication in physiological and pathological contexts. How cells generate these actin-mediated protrusions to span lengths beyond those attainable by canonical filopodia remains unknown. Through a combination of micropatterning, microscopy, and optical tweezer-based approaches, we demonstrate that TNTs formed through the outward extension of actin achieve distances greater than the mean length of filopodia and that branched Arp2/3-dependent pathways attenuate the extent to which actin polymerizes in nanotubes, thus limiting their occurrence. Proteomic analysis using epidermal growth factor receptor kinase substrate 8 (Eps8) as a positive effector of TNTs showed that, upon Arp2/3 inhibition, proteins enhancing filament turnover and depolymerization were reduced and Eps8 instead exhibited heightened interactions with the inverted Bin/Amphiphysin/Rvs (I-BAR) domain protein IRSp53 that provides a direct connection with linear actin polymerases. Our data reveals how common protrusion players (Eps8 and IRSp53) form tunnelling nanotubes, and that when competing pathways overutilizing such proteins and monomeric actin in Arp2/3 networks are inhibited, processes promoting linear actin growth dominate to favour tunnelling nanotube formation.
Article
The employment of nonlocal PDE models to describe biological aggregation and other phenomena has gained considerable traction in recent years. For cell populations, these methods grant a means of accommodating essential elements such as cell adhesion, critical to the development and structure of tissues. For animals, they can be used to describe how the nearby presence of conspecifics and/or heterospecifics influence movement behaviour. In this review, we will focus on classes of biological movement models in which the advective (or directed) component to motion is governed by an integral term that accounts for how the surrounding distribution(s) of the population(s) impact on a member’s movement. We recount the fundamental motivation for these models: the intrinsic capacity of cell populations to self-organise and spatially sort within tissues; the wide-ranging tendency of animals towards spatial structuring, from the formations of herds and swarms to territorial segregation. We examine the derivation of these models from an individual level, illustrating in the process methods that allow models to be connected to data. We explore a growing analytical literature, including methods of stability and bifurcation analysis, and existence results. We conclude with a short section that lays out some future challenges and connections to the modelling of sociological phenomena including opinion dynamics.
Article
Full-text available
Much attention has been focused on the possibility that cytoplasmic proteins and RNA may be conveyed between cells in extracellular vesicles (EVs) and tunneling nanotube (TNT) structures. Here, we set up two quantitative delivery reporters to study cargo transfer between cells. We found that EVs are internalized by reporter cells but do not efficiently deliver functional Cas9 protein to the nucleus. In contrast, donor and acceptor cells co-cultured to permit cell contact resulted in a highly effective transfer. Among our tested donor and acceptor cell pairs, HEK293T and MDA-MB-231 recorded optimal intercellular transfer. Depolymerization of F-actin greatly decreased Cas9 transfer whereas inhibitors of endocytosis or knock-down of genes implicated in this process had little effect on transfer. Imaging results suggest that intercellular transfer of cargos occurred through open-ended membrane tubular connections. In contrast, cultures consisting only of HEK293T cells form close-ended tubular connections ineffective in cargo transfer. Depletion of human endogenous fusogens, syncytins, especially syncytin-2 in MDA-MB-231 cells, significantly reduced Cas9 transfer. Full-length mouse syncytin, but not truncated mutants, rescued the effect of depletion of human syncytins on Cas9 transfer. Mouse syncytin overexpression in HEK293T cells partially facilitated Cas9 transfer among HEK293T cells. These findings suggest that syncytin may serve as the fusogen responsible for the formation of an open-ended connection between cells.
Thesis
Objectives: Acute Kidney Injury (AKI) is a major global burden with an enormous financial impact on health systems and an extensive impairment of quality of life for the affected patients. The sudden drop in renal function can be caused by a variety of conditions. One of the most common causes for a decrease in renal function is reduced renal perfusion resulting in a lack of oxygen, for example, as a consequence of hypovolemic shock. The pathophysiology of ischemia-reperfusion injury (IRI) is closely related to mitochondrial dysfunction which ultimately leads to a complete disruption of the cellular energy metabolism, resulting in premature cell death. In particular, proximal tubular epithelial cells are dependent on a continuous adenosine triphosphate (ATP) supply in order to maintain the physiological function of the nephron due to their predominantly active reabsorption- and secretion mechanisms. The background of immunological mechanisms for recovery after ischemic tissue injury has been subjected to extensive research. However, it is already known that shortly after tissue injury and as part of the different phases of the immunological response, an invasion of pro-inflammatory M1 macrophages can be seen. At subsequent time points, anti-inflammatory M2 macrophages, which seem to be a contributing factor for tissue regeneration, dominate. Analysis of murine kidney sections with electron microscopy was conducted at the department of Nephropathology, Yale University. It was revealed that macrophages could partially disrupt the tubular basement membrane (TBM) in order to interact with tubular cells via cellular protrusions. Indeed, many experimental studies have suggested that macrophages have the capability to communicate with other cells over long distances via long and thin cytoplasmic protrusions. These processes, termed tunneling nanotubes (TNTs), provide the possibility for the exchange of cell organelles, for example lysosomes or mitochondria, between participating cells. In different organ systems, it has been shown that, after tissue injury, there is a beneficial effect on cell recovery caused by the anterograde transfer of mitochondria in dysfunctional cells originating from macrophages or mesenchymal stem cells. The goal of this in vitro study was to examine, 1) whether macrophages have the capability to transfer mitochondria towards tubular epithelial cells via TNTs and 2) whether this transfer has a positive impact on regeneration after cell injury. The answer for these questions required the establishment of a suitable in vitro model. Design & Methods: The hypothesis was examined with an in vitro co-culture model involving both murine bone marrow derived macrophages as well as primary murine tubular epithelial cells or an immortalized tubular cell line. In order to visualize mitochondria and cell membrane, different fluorescent dyes, transfections with fluorescent constructs and genetically modified mice strains were employed. Live cell imaging was performed with widefield fluorescence and confocal laser scanning microscopy to analyze the presence of TNTs and mitochondrial transfer. Observations & Results: We established a reliable model which provided the possibility to study cell-cell interactions without loss of fluorescence intensity over a course of several days. Moreover, the regions of interest (ROIs) were sometimes misidentified during microscopy, and the sources of these misidentifications and related pitfalls were identified. Surprisingly, the mitochondrial morphology of cells mostly remained intact in the injury condition, even though the weak staining intensity of the fluorescent dye Tetramethylrhodamine methyl ester (TMRM) already indicated a substantial decrease of the mitochondrial membrane potential. Confocal live cell imaging revealed that polarized and unpolarized macrophages were capable of engulfing mitochondrial material from injured and non-injured tubular epithelial cells. To a lesser extent, anterograde mitochondrial transfer from macrophages towards tubular cells was observed. An exchange of organelles between cells of the same cell type was also seen. The transfer seemed to be partially mediated by the formation of TNTs. As an additional finding, we showed that proximal tubular epithelial cells have the capability to phagocytose cell debris. Conclusions: This study confirmed the initial hypothesis that macrophages and tubular have the capability to exchange mitochondria. However, it remained unclear whether transferred mitochondria were functional and provided a benefit for maintenance of cell integrity. Since the fluorescence intensity of mitochondrial material remarkedly decreased after transfer, it is conceivable that mitochondria entered lysosomal degradation as part of mitophagy, a certain kind of autophagy. Further experiments are needed to investigate the functional state of transferred mitochondria and their impact more precisely. Based on the findings of this project, less importance should be attributed as previously thought to the mitochondrial morphology as an indicator for cell integrity. Beyond that, this work pointed out that a reliable quantitative analysis cannot be achieved with confocal microscopy alone due to the range of potential interpretation of 2D image data and its technical limitations. Computational algorithms rendering 2D data in 3D models or alternative methods such as fluorescence activated cell sorting (FACS) should be employed for further quantitative assessment. A more detailed understanding of the triggers of mitochondrial transfer could help to promote therapeutic strategies accelerating the healing process after ischemic kidney injury. Finally, the observation that proximal tubular epithelial cells had the capability to phagocytose dying diphtheria-toxin receptor positive cells during the isolation process when treated with diphtheria toxin, underlines the idea that tubular cells are involved in the removal of apoptotic neighboring cells after tubular necrosis.
Preprint
Full-text available
Much attention has been focused on the possibility that cytoplasmic proteins and RNA may be conveyed between cells in extracellular vesicles (EVs) and tunneling nanotube (TNT) structures. Here, we set up two quantitative delivery reporters to study cargo transfer between cells. We found that EVs are internalized by reporter cells but do not efficiently deliver functional Cas9 protein to the nucleus. In contrast, donor and acceptor cells co-cultured to permit cell contact resulted in a highly effective transfer. Among our tested donor and acceptor cell pairs, HEK293T and MDA MB-231 recorded optimal intercellular transfer. Depolymerization of F-actin greatly decreased Cas9 transfer whereas inhibitors of endocytosis or knock-down of genes implicated in this process had little effect on transfer. Imaging results suggest that intercellular transfer of cargos occurred through open-ended membrane tubular connections. In contrast, cultures consisting only of HEK293T cells form close-ended tubular connections ineffective in cargo transfer. Depletion of human endogenous fusogens, syncytins, especially syncytin-2 in MDA-MB-231 cells, significantly reduced Cas9 transfer. Full-length mouse syncytin, but not truncated mutants, rescued the effect of depletion of human syncytins on Cas9 transfer. Mouse syncytin overexpression in HEK293T cells partially facilitated Cas9 transfer among HEK293T cells. These findings suggest that syncytin may serve as the fusogen responsible for the formation of an open-ended connection between cells.
Article
Living cells use long tubular appendages for locomotion and sensory purposes. Hence, assembling and maintaining a protrusion of correct length is crucial for survival and overall performance. Usually the protrusions lack the machinery for the synthesis of building blocks and imports them from the cell body. What are the unique features of the transport logistics which facilitate the exchange of these building blocks between the cell and the protrusion? What kind of ‘rulers’ and ‘timers’ does the cell use for constructing its appendages of correct length on time? How do the multiple appendages coordinate and communicate among themselves during different stages of their existence? How frequently do the fluctuations drive the length of these dynamic protrusions out of the acceptable bounds? These questions are addressed from a broad perspective in this review which is organized in three parts. In part-I the list of all known cell protrusions is followed by a comprehensive list of the mechanisms of length control of cell protrusions reported in the literature. We review not only the dynamics of the genesis of the protrusions, but also their resorption and regrowth as well as regeneration after amputation. As a case study in part-II, the specific cell protrusion that has been discussed in detail is eukaryotic flagellum (also known as cilium); this choice was dictated by the fact that flagellar length control mechanisms have been studied most extensively over more than half a century in cells with two or more flagella. Although limited in scope, brief discussions on a few non-flagellar cell protrusions in part-III of this review is intended to provide a glimpse of the uncharted territories and challenging frontiers of research on subcellular length control phenomena that awaits rigorous investigations.
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The oocyte must grow and mature before fertilization, thanks to a close dialogue with the somatic cells which surround it. Part of this communication is through filopodia-like protrusions, called transzonal projections (TZPs), sent by the somatic cells to the oocyte membrane. To investigate the contribution of TZPs to oocyte quality, we impaired their structure by generating a full knockout mouse of the TZP structural component Myosin-X (MYO10). Using spinning disk and super-resolution microscopy combined with a machine learning approach to phenotype oocyte morphology, we show that the lack of Myo10 decreases TZP density during oocyte growth. Reduction in TZPs does not prevent oocyte growth but impairs oocyte-matrix integrity. Importantly, we reveal by transcriptomic analysis that gene expression is altered in TZP-deprived oocytes, and that oocyte maturation and subsequent early embryonic development are partially affected, effectively reducing mouse fertility. We propose that TZPs play a role in the structural integrity of the germline-somatic complex, which is essential for regulating gene expression in the oocyte and thus its developmental potential.
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The development of germ cells relies on contact and communication with neighboring somatic cells that provide metabolic support and regulatory signals. In females, contact is achieved through thin cytoplasmic processes that project from follicle cells surrounding the oocyte, extend through an extracellular matrix (ECM) that lies between them, and reach its surface. In mammals, the ECM is termed the zona pellucida and the follicular cell processes are termed transzonal projections (TZPs). TZPs become detectable when the zona pellucida is laid down during early folliculogenesis and subsequently increase in number as oocyte growth progresses. They then rapidly disappear at the time of ovulation, permanently breaking germ‐soma contact. Here we review the life cycle and functions of the TZPs. We begin with an overview of the morphology and cytoskeletal structure of TZPs, in the context of actin‐ and tubulin‐based cytoplasmic processes in other cell types. Next, we review the roles played by TZPs in mediating progression through successive stages of oocyte development. We then discuss two mechanisms that may generate TZPs—stretching at pre‐existing points of granulosa cell‐oocyte contact and elaboration of new processes that push through the zona pellucida—as well as gene products implicated in their formation or function. Finally, we describe the signaling pathways that cause TZPs to be retracted in response to signals that also trigger meiotic maturation and ovulation of the oocyte. The principles and mechanisms that govern TZP behavior may be relevant to understanding communication between physically separated cells in other physiological contexts.
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Microglia are the most prominent immune resident cell population in the central nervous system (CNS). In the healthy CNS, microglia survey their surrounding microenvironment, through recurrent extension and retraction of filopodia-like membrane protrusions, without evident cell body displacement. Microglia undergo dramatic transcriptomic and shape changes upon brain insults or neurodegenerative disease states and adopt a classical immune effector function (producing an extensive array of inflammatory mediators such as cytokines, chemokines, and reactive oxygen species) to re-establish tissue homeostasis. While the biophysical principles underlying microglia morphological changes remain elusive, several recent studies have highlighted the pivotal role of the actin and non-muscle myosin II filamentous cytoskeleton in this process. In this work, we discuss how subcellular topological patterning of the actin and myosin cytoskeleton can control microglial cell shape dynamics and how it can potentially feedback on their functional specialization, which is of great importance to understanding the mechanisms of microglial action in homeostatic conditions and CNS disease states.
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Cellular protrusions generated by the actin cytoskeleton are central to the process of building the body of the embryo. Problems with cellular protrusions underlie human diseases and syndromes, including implantation defects and pregnancy loss, congenital birth defects, and cancer. Cells use protrusive activity together with actin-myosin contractility to create an ordered body shape of the embryo. Here, I review how actin-rich protrusions are used by two major morphological cell types, epithelial and mesenchymal cells, during collective cell migration to sculpt the mouse embryo body. Pre-gastrulation epithelial collective migration of the anterior visceral endoderm is essential for establishing the anterior-posterior body axis. Gastrulation mesenchymal collective migration of the mesoderm wings is crucial for body elongation, and somite and heart formation. Analysis of mouse mutants with disrupted cellular protrusions revealed the key role of protrusions in embryonic morphogenesis and embryo survival. Recent technical approaches have allowed examination of the mechanisms that control cell and tissue movements in vivo in the complex 3D microenvironment of living mouse embryos. Advancing our understanding of protrusion-driven morphogenesis should provide novel insights into human developmental disorders and cancer metastasis.
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Cell-to-cell communication is essential for the organization, coordination, and development of cellular networks and multi-cellular systems. Intercellular communication is mediated by soluble factors (including growth factors, neurotransmitters, and cytokines/chemokines), gap junctions, exosomes and recently described tunneling nanotubes (TNTs). It is unknown whether a combination of these communication mechanisms such as TNTs and gap junctions may be important, but further research is required. TNTs are long cytoplasmic bridges that enable long-range, directed communication between connected cells. The proposed functions of TNTs are diverse and not well understood but have been shown to include the cell-to-cell transfer of vesicles, organelles, electrical stimuli and small molecules. However, the exact role of TNTs and gap junctions for intercellular communication and their impact on disease is still uncertain and thus, the subject of much debate. The combined data from numerous laboratories indicate that some TNT mediate a long-range gap junctional communication to coordinate metabolism and signaling, in relation to infectious, genetic, metabolic, cancer, and age-related diseases. This review aims to describe the current knowledge, challenges and future perspectives to characterize and explore this new intercellular communication system and to design TNT-based therapeutic strategies.
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ELife digest When an embryo develops, it is critical that tissues and organs form properly and at the right time. For this, cells need to be able to communicate over long distances by using signalling molecules called morphogens. Morphogens disperse via extensions that protrude from the surface of a ‘source’ cell. Previous research has shown that these extensions called cytonemes can transport the morphogens to ‘receiver’ cells, and depending on the distance from the source, build a concentration gradient that will either be higher or lower. These gradients then help unspecialized cells to develop into different specialized ones. One of the key morphogens during the development is the Hedgehog protein. Researchers have previously shown that vesicles along cytonemes of cells that produce Hedgehog transport the morphogen to the receiver cells. However, until now it was unclear how the Hedgehog signals are transferred and received. Here, González-Méndez et al. – including researchers involved in the previous studies – investigated the cytonemes located on Hedgehog-receiving cells in the fruit fly. The results showed that these cytonemes are oriented towards the Hedgehog-producing cells and help to create a concentration gradient by varying their length. Moreover, the cytonemes from signal-producing and signal-receiving cells connect at specific sites that are distributed along their lengths. This suggests that the contact sites might help to transfer and receive the morphogens. Thus, the way cells communicate in other tissues of the body could be similar to how nerve cells communicate with each other in the brain. Our next challenges will be to fully understand how cytonemes transfer the Hedgehog signal. This could shed more light on how Hedgehog signaling can be controlled and modulated. DOI:http://dx.doi.org/10.7554/eLife.24045.002
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During development, extracellular cues guiding cell fate determination are provided by morphogens. One mechanism by which morphogens are proposed to traverse extracellular space is by traveling along specialized filopodia called cytonemes. These cellular highways extend between signal producing and receiving cells to enable direct morphogen delivery. Although genetic studies support cytoneme involvement in morphogen transport, mechanistic insight into how they are regulated is limited due to technical challenges associated with performing cell biological analysis of the delicate filopodial structures. Herein, we introduce a fixation method whereby cultured cell cytonemes can be preserved for imaging studies, allowing for interrogation of cytoneme regulation using standard cell biological techniques. Using this method, we examined Hedgehog-containing cytonemes and identified a role for the Hedgehog deployment protein Dispatched in cytoneme stabilization. We demonstrate Hedgehog and Dispatched colocalize in cytonemes, and cholesterol-modified Hedgehog acts through Dispatched to increase cytoneme occurrence. Live imaging suggests this occurs through Dispatched-mediated slowing of cytoneme retraction rates. Dispatched-induced cytoneme modulation was recapitulated in wing imaginal discs of transgenic Drosophila, supporting that cultured cell cytoneme analysis is predictive of in vivo functionality.
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Macrophage interactions with other cells, either locally or at distances, are imperative in both normal and pathological conditions. While soluble means of communication can transmit signals between different cells, it does not account for all long distance macrophage interactions. Recently described tunneling nanotubes (TNTs) are membranous channels that connect cells together and allow for transfer of signals, vesicles, and organelles. However, very little is known about the mechanism by which these structures are formed. Here we investigated the signaling pathways involved in TNT formation by macrophages using multiple imaging techniques including super-resolution microscopy (3D-SIM) and live-cell imaging including the use of FRET-based Rho GTPase biosensors. We found that formation of TNTs required the activity and differential localization of Cdc42 and Rac1. The downstream Rho GTPase effectors mediating actin polymerization through Arp2/3 nucleation, Wiskott-Aldrich syndrome protein (WASP) and WASP family verprolin-homologous 2 (WAVE2) proteins are also important, and both pathways act together during TNT biogenesis. Finally, TNT function as measured by transfer of cellular material between cells was reduced following depletion of a single factor demonstrating the importance of these factors in TNTs. Given that the characterization of TNT formation is still unclear in the field; this study provides new insights and would enhance the understanding of TNT formation towards investigating new markers.
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Tunneling nanotubes (TNTs) are long bridge-like structures that connect eukaryotic cells and mediate intercellular communication. We found earlier that the conserved alphaherpesvirus US3 protein kinase induces long cell projections that contact distant cells and promote intercellular virus spread. In this report, we show that the US3-induced cell projections constitute TNTs. In addition, we report that US3-induced TNTs mediate intercellular transport of information (e.g., green fluorescent protein [GFP]) in the absence of other viral proteins. US3-induced TNTs are remarkably stable compared to most TNTs described in the literature. In line with this, US3-induced TNTs were found to contain stabilized (acetylated and detyrosinated) microtubules. Transmission electron microscopy showed that virus particles are individually transported in membrane-bound vesicles in US3-induced TNTs and are released along the TNT and at the contact area between a TNT and the adjacent cell. Contact between US3-induced TNTs and acceptor cells is very stable, which correlated with a marked enrichment in adherens junction components beta-catenin and E-cadherin at the contact area. These data provide new structural insights into US3-induced TNTs and how they may contribute to intercellular communication and alphaherpesvirus spread. IMPORTANCE Tunneling nanotubes (TNT) represent an important and yet still poorly understood mode of long-distance intercellular communication. We and others reported earlier that the conserved alphaherpesvirus US3 protein kinase induces long cellular protrusions in infected and transfected cells. Here, we show that US3-induced cell projections constitute TNTs, based on structural properties and transport of biomolecules. In addition, we report on different particular characteristics of US3-induced TNTs that help to explain their remarkable stability compared to physiological TNTs. In addition, transmission electron microscopy assays indicate that, in infected cells, virions travel in the US3-induced TNTs in membranous transport vesicles and leave the TNT via exocytosis. These data generate new fundamental insights into the biology of (US3-induced) TNTs and into how they may contribute to intercellular virus spread and communication.
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Morphogen concentration gradients that extend across developmental fields form by dispersion from source cells. In the Drosophila wing disc, Hedgehog (Hh) produced by posterior compartment cells distributes in a concentration gradient to adjacent cells of the anterior compartment. We monitored Hh:GFP after pulsed expression and analyzed movements and co-localization of Hh, Patched (Ptc) and Smoothened (Smo) proteins tagged with GFP or mCherry and expressed at physiological levels from bacterial artificial chromosome transgenes. Hh:GFP moved to basal sub-cellular locations prior to release from posterior compartment cells that express it, and was taken up by basal cytonemes that extend to the source cells. Hh and Ptc were present in puncta that moved along the basal cytonemes and formed characteristic apical-basal distributions in the anterior compartment cells. The basal cytonemes required diaphanous, Scar, neuroglian, and synaptobrevin, and both the Hh gradient and Hh signaling declined under conditions in which the cytonemes were compromised. These findings show that in the wing disc, Hh distributions and signaling are dependent upon basal release and uptake, and on cytoneme-mediated movement. No evidence for apical dispersion was obtained.
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Intercellular communications play a major role in tissue homeostasis and responses to external cues. Novel structures for this communication have recently been described. These tunneling nanotubes (TNTs) consist of thin-extended membrane protrusions that connect cells together. TNTs allow the cell-to-cell transfer of various cellular components, including proteins, RNAs, viruses, and organelles, such as mitochondria. Mesenchymal stem cells (MSCs) are both naturally present and recruited to many different tissues where their interaction with resident cells via secreted factors has been largely documented. Their immunosuppressive and repairing capacities constitute the basis for many current clinical trials. MSCs recruited to the tumor microenvironment also play an important role in tumor progression and resistance to therapy. MSCs are now the focus of intense scrutiny due to their capacity to form TNTs and transfer mitochondria to target cells, either in normal physiological or in pathological conditions, leading to changes in cell energy metabolism and functions, as described in this review.
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Acute lymphoblastic leukemia (ALL) cells create a leukemic niche with mesenchymal stromal cells (MSCs). Cytoskeletal structures called tunneling nanotubes (TNTs) facilitate communication between ALL cells and MSCs by transporting molecules and inducing the secretion of pro-survival cytokines. The identity of the molecules driving these malignant processes are currently unknown. Here we investigate which structures are transported from ALL cells toward MSCs by quantifying the transfer of ectopically expressed fluorescent marker proteins using flow cytometry. Our results indicate that actin, endoplasmatic reticulum, ICAM1, autophagosomes, mitochondria, and endosomes are contact-dependently transferred from leukemic cells toward MSCs. Transfer of mitochondria, adhesion molecule ICAM1, and autophagosomes were significantly reduced (9.6-fold) when TNT signaling was inhibited. Autophagosomes and mitochondria are known inducers of cytokine signaling and hence their transfer might unveil an important mechanism that ALL cells use to affect their microenvironment. Importantly, transfer of autophagosomes was 3.0-fold greater than the other molecules tested in ALL-MSC co-cultures, and has not been previously associated with TNT signaling. These data provide insight into intercellular signaling in the leukemic niche and implicate autophagosomes as novel TNT cargo.
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Tunneling nanotubes (TnTs) are thin channels that temporally connect nearby cells allowing the cell-to-cell trafficking of biomolecules and organelles. The presence or absence of TnTs in human neoplasms and the mechanisms of TnT assembly remains largely unexplored. In this study, we have identified TnTs in tumor cells derived from squamous cell carcinomas (SCC) cultured under bi-dimensional and tri-dimensional conditions and also in human SCC tissues. Our study demonstrates that TnTs are not specific of epithelial or mesenchymal phenotypes and allow the trafficking of endosomal/ lysosomal vesicles, mitochondria, and autophagosomes between both types of cells. We have identified focal adhesion kinase (FAK) as a key molecule required for TnT assembly via a mechanism involving the MMP-2 metalloprotease. We have also found that the FAK inhibitor PF-562271, which is currently in clinical development for cancer treatment, impairs TnT formation. Finally, FAK-deficient cells transfer lysosomes/autophagosomes to FAK-proficient cells via TnTs which may represent a novel mechanism to adapt to the stress elicited by impaired FAK signaling. Collectively, our results strongly suggest a link between FAK, MMP-2, and TnT, and unveil new vulnerabilities that can be exploited to efficiently eradicate cancer cells.
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Parkinson’s disease (PD) is characterized by the presence of inclusions known as Lewy bodies, which mainly consist of α-synuclein (α-syn) aggregates. There is growing evidence that α-syn self-propagates in non-neuronal cells, thereby contributing to the progression and spread of PD pathology in the brain. Tunneling nanotubes (TNTs) are long, thin, F-actin-based membranous channels that connect cells and have been proposed to act as conduits for α-syn transfer between cells. SH-SY5Y cells and primary human brain pericytes, derived from postmortem PD brains, frequently form TNTs that allow α-syn transfer and long-distance electrical coupling between cells. Pericytes in situ contain α-syn precipitates like those seen in neurons. Exchange through TNTs was rapid, but dependent on the size of the protein. Proteins were able to spread throughout a network of cells connected by TNTs. Transfer through TNTs was not restricted to α-syn; fluorescent control proteins and labeled membrane were also exchanged through TNTs. Most importantly the formation of TNTs and transfer continued during mitosis. Together, our results provide a detailed description of TNTs in SH-SY5Y cells and human brain PD pericytes, demonstrating their role in α-syn transfer and further emphasize the importance that non-neuronal cells, such as pericytes play in disease progression.
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Intercellular transfer of organelles via tunneling nanotubes (TNTs) is a novel means of cell-to-cell communication. Here we demonstrate the existence of TNTs between co-cultured RT4 and T24 bladder cancer cells using light microscopy, fluorescence imaging, and scanning electron microscopy (SEM). Spontaneous unidirectional transfer of mitochondria from T24 to RT4 cells was detected using fluorescence imaging and flow cytometry. The distribution of mitochondria migrated from T24 cells was in good agreement with the original mitochondria in RT4 cells, which may imply mitochondrial fusion. We detected cytoskeleton reconstruction in RT4-Mito-T24 cells by observing F-actin redistribution. Akt, mTOR, and their downstream mediators were activated and increased. The resultant increase in the invasiveness of bladder cancer cells was detected in vitro and in vivo. These data indicate that TNTs promote intercellular mitochondrial transfer between heterogeneous cells, followed by an increase in the invasiveness of bladder cancer cells.
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Tunneling nanotubes (TNTs) represent a novel route of intercellular communication. While previous work has shown that TNTs facilitate the exchange of viral or prion proteins from infected to naïve cells, it is not clear whether the viral genome is also transferred via this mechanism and further, whether transfer via this route can result in productive replication of the infectious agents in the recipient cell. Here we present evidence that lung epithelial cells are connected by TNTs, and in spite of the presence of neutralizing antibodies and an antiviral agent, Oseltamivir, influenza virus can exploit these networks to transfer viral proteins and genome from the infected to naïve cell, resulting in productive viral replication in the naïve cells. These observations indicate that influenza viruses can spread using these intercellular networks that connect epithelial cells, evading immune and antiviral defenses and provide an explanation for the incidence of influenza infections even in influenza-immune individuals and vaccine failures.
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Tunneling Nanotubes (TNTs) are actin enriched filopodia-like protrusions that play a pivotal role in long-range intercellular communication. Different pathogens use TNT-like structures as “freeways” to propagate across cells. TNTs are also implicated in cancer and neurodegenerative diseases, making them promising therapeutic targets. Understanding the mechanism of their formation, and their relation with filopodia is of fundamental importance to uncover their physiological function, particularly since filopodia, differently from TNTs, are not able to mediate transfer of cargo between distant cells. Here we studied different regulatory complexes of actin, which play a role in the formation of both these structures. We demonstrate that the filopodia-promoting CDC42/IRSp53/VASP network negatively regulates TNT formation and impairs TNT-mediated intercellular vesicle transfer. Conversely, elevation of Eps8, an actin regulatory protein that inhibits the extension of filopodia in neurons, increases TNT formation. Notably, Eps8-mediated TNT induction requires Eps8 bundling but not its capping activity. Thus, despite their structural similarities, filopodia and TNTs form through distinct molecular mechanisms. Our results further suggest that a switch in the molecular composition in common actin regulatory complexes is critical in driving the formation of either type of membrane protrusion.
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Tunneling nanotubes (TNTs) are ultrafine, filamentous actin-based cytoplasmic extensions which form spontaneously to connect cells at short and long-range distances. We have previously described long-range intercellular communication via TNTs connecting mesothelioma cells in vitro and demonstrated TNTs in intact tumors from patients with mesothelioma. Here, we investigate the ability of TNTs to mediate a viral thymidine kinase based bystander effect after oncolytic viral infection and administration of the nucleoside analog ganciclovir. Using confocal microscopy we assessed the ability of TNTs to propagate enhanced green fluorescent protein (eGFP), which is encoded by the herpes simplex virus NV1066, from infected to uninfected recipient cells. Using time-lapse imaging, we observed eGFP expressed in infected cells being transferred via TNTs to noninfected cells; additionally, increasing fluorescent activity in recipient cells indicated cell-to-cell transmission of the eGFP-expressing NV1066 virus had also occurred. TNTs mediated cell death as a form of direct cell-to-cell transfer following viral thymidine kinase mediated activation of ganciclovir, inducing a unique long-range form of the bystander effect through transmission of activated ganciclovir to nonvirus-infected cells. Thus, we provide proof-of-principle demonstration of a previously unknown and alternative mechanism for inducing apoptosis in noninfected recipient cells. The conceptual advance of this work is that TNTs can be harnessed for delivery of oncolytic viruses and of viral thymidine kinase activated drugs to amplify the bystander effect between cancer cells over long distances in stroma-rich tumor microenvironments.
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A given cell makes exchanges with its neighbors through a variety of means ranging from diffusible factors to vesicles. Cells use also tunneling nanotubes (TNTs), filamentous-actin-containing membranous structures that bridge and connect cells. First described in immune cells, TNTs facilitate HIV-1 transfer and are found in various cell types, including neurons. We show that the microtubule-associated protein Tau, a key player in Alzheimer’s disease, is a bona fide constituent of TNTs. This is important because Tau appears beside filamentous actin and myosin 10 as a specific marker of these fine protrusions of membranes and cytosol that are difficult to visualize. Furthermore, we observed that exogenous Tau species increase the number of TNTs established between primary neurons, thereby facilitating the intercellular transfer of Tau fibrils. In conclusion, Tau may contribute to the formation and function of the highly dynamic TNTs that may be involved in the prion-like propagation of Tau assemblies. Electronic supplementary material The online version of this article (doi:10.1186/s40478-016-0386-4) contains supplementary material, which is available to authorized users.
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The tunneling nanotube (TNT) is a structure used for intercellular communication, and is a thin membrane protrusion mediating transport of various signaling molecules and cellular components. M-Sec has potent membrane deformation ability and induces TNT formation in cooperation with the Ral/exocyst complex. Here, we show that the N-terminal polybasic region of M-Sec directly binds phosphatidylinositol (4,5)-bisphosphate for its localization to the plasma membrane during the initial stage of TNT formation. We further report a crystal structure of M-Sec, which consists of helix bundles arranged in a straight rod-like shape, similar to the membrane tethering complex subunits. A positively charged surface in the C-terminal domains is required for M-Sec interaction with active RalA to extend the plasma membrane protrusions. Our results suggest that the membrane-associated M-Sec recruits active RalA, which directs the exocyst complex to form TNTs.
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The exposure of phosphatidylserine (PS) on the surface membrane of apoptotic cells triggers the recruitment of phagocytic receptors and subsequently results in uptake by phagocytes. Here we describe how apoptotic cells can use intercellular membrane nanotubes to transfer exposed PS to neighboring viable cells, and thus deposit an "eat-me" tag on the viable cells. Tunneling nanotubes (TNTs) connected UV-treated apoptotic rat pheochromocytoma PC12 cells with neighboring untreated cells. These TNTs were composed of PS-exposed plasma membrane and facilitated the transfer of the membrane from apoptotic to viable cells. Other pro-phagocytic signals, such as oxidized phospholipids and calreticulin, were also transferred to viable cells. In addition, anti-phagocytic signal CD47 presenting on the plasma membrane of viable cells was masked by the transferred PS-membrane. Confocal imaging revealed an increase of phagocytosis of viable PC12 cells by murine RAW264.7 macrophages when the viable PC12 cells were cocultured with UV-treated PC12 cells. Treatment with 50 nM cytochalasin D would abolish TNTs and correspondingly inhibit this phagocytosis of the viable cells. Our study indicates that exposed-PS membrane is delivered from apoptotic to viable cells through TNTs. This transferred membrane may act as a pro-phagocytic signal for macrophages to induce phagocytosis of viable cells in a situation where they are in the vicinity of apoptotic cells. This article is protected by copyright. All rights reserved.
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Tunneling membrane nanotubes (TNTs) are thin membrane projections linking cell bodies separated by many micrometers, which are proposed to mediate signaling and even transfer of cytosolic contents between distant cells. Several reports describe propagation of Ca2+ signals between distant cells via TNTs, but the underlying mechanisms remain poorly understood. Utilizing a HeLa M-Sec cell line engineered to upregulate TNTs we replicated previous findings that mechanical stimulation elicits robust cytosolic Ca2+ elevations that propagate to surrounding, physically separate cells. However, whereas this was previously interpreted to involve intercellular communication through TNTs, we found that Ca2+ signal propagation was abolished − even in TNT-connected cells − after blocking ATP-mediated paracrine signaling with a cocktail of extracellular inhibitors. To then establish whether gap junctions may enable cell-cell signaling via TNTs under these conditions, we expressed sfGFP-tagged connexin-43 (Cx43) in HeLa M-Sec cells. We observed robust communication of mechanically-evoked Ca2+ signals between distant but TNT-connected cells, but only when both cells expressed Cx43. Moreover, we also observed communication of Ca2+ signals evoked in one cell by local photorelease of inositol 1,4,5-trisphosphate (IP3). Ca2+ responses in connected cells began after long latencies at intracellular sites several microns from the TNT connection site, implicating intercellular transfer of IP3 and subsequent IP3-mediated Ca2+ liberation, and not Ca2+ itself, as the mediator between TNT-connected, Cx43-expressing cells. Our results emphasize the need to control for paracrine transmission in studies of cell-cell signaling via TNTs and indicate that, in this cell line, TNTs do not establish cytosolic continuity between connected cells but rather point to the crucial importance of connexins to enable communication of cytosolic Ca2+ signals via TNTs.
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Tunnelling nanotubes (TNTs) are increasingly recognized as central players in a multitude of cellular mechanisms and diseases. Although their existence and functions in animal organisms are still elusive, emerging evidence suggests that they are involved in developmental processes, tissue regeneration, viral infections or pathogen transfer, stem cell differentiation, immune responses as well as initiation and progression of neurodegenerative disorders and cancer (see Sisakhtnezhad & Khosravi 2015 Eur. J. Cell Biol. 94, 429- 443. (doi:10.1016/j.ejcb.2015.06.010)). A broader field of vision, including their striking functional and structural resemblance with nanotube-mediated phenomena found throughout the phylogenetic tree, from plants down to bacteria, points to a universal, conserved and tightly regulated mechanism of cellular assemblies. Based on our initial definition of TNTs as open-ended channels mediating membrane continuity between connected cells (Rustom et al. 2004 Science 303, 1007- 1010. (doi:10.1126/science. 1093133)), it is suggested that animal tissues represent supercellular assemblies that-besides opening discrete communication pathways-balance diverse stress factors caused by pathological changes or fluctuating physiological and environmental conditions, such as oxidative stress or nutrient shortage. By combining current knowledge about nanotube formation, intercellular transfer and communication phenomena as well as associated molecular pathways, a model evolves, predicting that the linkage between reactive oxygen species, TNT-based supercellularity and the intercellular shuttling of materials will have significant impact on diverse body functions, such as cell survival, redox/metabolic homeostasis and mitochondrial heteroplasmy. It implies that TNTs are intimately linked to the physiological and pathological state of animal cells and represent a central joint element of diverse diseases, such as neurodegenerative disorders, diabetes or cancer.
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Mesenchymal stromal cells (MSC) have been reported to improve bacterial clearance in pre-clinical models of Acute Respiratory Distress Syndrome (ARDS) and sepsis. The mechanism of this effect is not fully elucidated yet. The primary objective of this study was to investigate the hypothesis that the anti-microbial effect of MSC in vivo depends on their modulation of macrophage phagocytic activity which occurs through mitochondrial transfer. We established that selective depletion of alveolar macrophages (AM) with intranasal (IN) administration of liposomal clodronate resulted in complete abrogation of MSC anti-microbial effect in the in vivo model of E.coli pneumonia. Furthermore, we showed that MSC administration was associated with enhanced AM phagocytosis in vivo. We showed that direct co-culture of MSC with monocyte-derived macrophages (MDMs) enhanced their phagocytic capacity. By fluorescent imaging and flow cytometry we demonstrated extensive mitochondrial transfer from MSC to macrophages which occurred at least partially through TNT-like structures. We also detected that lung macrophages readily acquire MSC mitochondria in vivo, and macrophages which are positive for MSC mitochondria display more pronounced phagocytic activity. Finally, partial inhibition of mitochondrial transfer through blockage of TNT formation by MSC resulted in failure to improve macrophage bioenergetics and complete abrogation of the MSC effect on macrophage phagocytosis in vitro and the anti-microbial effect of MSC in vivo. Collectively, this work for the first time demonstrates that mitochondrial transfer from MSC to innate immune cells leads to enhancement in phagocytic activity and reveals an important novel mechanism for the anti-microbial effect of MSC in ARDS. This article is protected by copyright. All rights reserved.
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Prion diseases are caused by misfolding of the cellular protein PrPC to an infectious conformer, PrPSc. Intercellular PrPSc transfer propagates conversion and allows infectivity to move from the periphery to the brain. However, how prions spread between cells of the central nervous system is unclear. Astrocytes are specialized non-neuronal cells within the brain that have a number of functions indispensable for brain homeostasis. Interestingly, they are one of the earliest sites of prion accumulation in the brain. A fundamental question arising from this observation is whether these cells are involved in intercellular prion transfer and thereby disease propagation. Using co-culture systems between primary infected astrocytes and granule neurons or neuronal cell lines, we provide direct evidence that prion-infected astrocytes can disseminate prion to neurons. Though astrocytes are capable of secreting PrP, this is an inefficient method of transferring prion infectivity. Efficient transfer required co-culturing and direct cell contact. Astrocytes form numerous intercellular connections including tunneling nanotubes, containing PrPSc, often colocalized with endolysosomal vesicles, which may constitute the major mechanism of transfer. Because of their role in intercellular transfer of prions astrocytes may influence progression of the disease.
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