RESUMO
Cytotoxic T lymphocytes (CTLs) fight intracellular pathogens and cancer by identifying and destroying infected or transformed target cells1. To kill, CTLs form a specialized cytotoxic immune synapse (IS) with a target of interest and then release toxic perforin and granzymes into the interface to elicit programmed cell death2-5. The IS then dissolves, enabling CTLs to search for additional prey and professional phagocytes to clear the corpse6. While the mechanisms governing IS assembly have been studied extensively, far less is known about target cell release. Here, we applied time-lapse imaging to explore the basis for IS dissolution and found that it occurred concomitantly with the cytoskeletal contraction of apoptotic targets. Genetic and pharmacological perturbation of this contraction response indicated that it was both necessary and sufficient for CTL dissociation. We also found that mechanical amplification of apoptotic contractility promoted faster CTL detachment and serial killing. Collectively, these results establish a biophysical basis for IS dissolution and highlight the importance of mechanosensory feedback in the regulation of cell-cell interactions.
Assuntos
Apoptose , Linfócitos T Citotóxicos , Apoptose/genética , Perforina , GranzimasRESUMO
Fungi are among the most diverse and ecologically important kingdoms in life. However, the distributional ranges of fungi remain largely unknown as do the ecological mechanisms that shape their distributions1,2. To provide an integrated view of the spatial and seasonal dynamics of fungi, we implemented a globally distributed standardized aerial sampling of fungal spores3. The vast majority of operational taxonomic units were detected within only one climatic zone, and the spatiotemporal patterns of species richness and community composition were mostly explained by annual mean air temperature. Tropical regions hosted the highest fungal diversity except for lichenized, ericoid mycorrhizal and ectomycorrhizal fungi, which reached their peak diversity in temperate regions. The sensitivity in climatic responses was associated with phylogenetic relatedness, suggesting that large-scale distributions of some fungal groups are partially constrained by their ancestral niche. There was a strong phylogenetic signal in seasonal sensitivity, suggesting that some groups of fungi have retained their ancestral trait of sporulating for only a short period. Overall, our results show that the hyperdiverse kingdom of fungi follows globally highly predictable spatial and temporal dynamics, with seasonality in both species richness and community composition increasing with latitude. Our study reports patterns resembling those described for other major groups of organisms, thus making a major contribution to the long-standing debate on whether organisms with a microbial lifestyle follow the global biodiversity paradigms known for macroorganisms4,5.
Assuntos
Microbiologia do Ar , Biodiversidade , DNA Fúngico , Fungos , Estações do Ano , Análise Espaço-Temporal , DNA Fúngico/análise , DNA Fúngico/genética , Fungos/genética , Fungos/classificação , Fungos/isolamento & purificação , Micorrizas/genética , Micorrizas/classificação , Micorrizas/isolamento & purificação , Filogenia , Esporos Fúngicos/classificação , Esporos Fúngicos/isolamento & purificação , Temperatura , Clima Tropical , Mapeamento GeográficoRESUMO
Antitumor immunosurveillance is triggered by immune cell recognition of characteristic biochemical signals on the surfaces of cancer cells. Recent data suggest that the mechanical properties of cancer cells influence the strength of these signals, with physically harder target cells (more rigid) eliciting better, faster, and stronger cytotoxic responses against metastasis. Using analogies to a certain electronic music duo, we argue that the biophysical properties of cancer cells and their environment can adjust the volume and tone of the antitumor immune response. We also consider the potential influence of biomechanics-based immunosurveillance in disease progression and posit that targeting the biophysical properties of cancer cells in concert with their biochemical features could increase the efficacy of immunotherapy.
Assuntos
Antineoplásicos , Neoplasias , Biofísica , Humanos , Imunoterapia , Monitorização Imunológica , Neoplasias/imunologiaRESUMO
Immune cells have intensely physical lifestyles characterized by structural plasticity and force exertion. To investigate whether specific immune functions require stereotyped mechanical outputs, we used super-resolution traction force microscopy to compare the immune synapses formed by cytotoxic T cells with contacts formed by other T cell subsets and by macrophages. T cell synapses were globally compressive, which was fundamentally different from the pulling and pinching associated with macrophage phagocytosis. Spectral decomposition of force exertion patterns from each cell type linked cytotoxicity to compressive strength, local protrusiveness, and the induction of complex, asymmetric topography. These features were validated as cytotoxic drivers by genetic disruption of cytoskeletal regulators, live imaging of synaptic secretion, and in silico analysis of interfacial distortion. Synapse architecture and force exertion were sensitive to target stiffness and size, suggesting that the mechanical potentiation of killing is biophysically adaptive. We conclude that cellular cytotoxicity and, by implication, other effector responses are supported by specialized patterns of efferent force.
Assuntos
Sinapses Imunológicas , Análise de Célula Única , Animais , Sinapses Imunológicas/imunologia , Camundongos , Linfócitos T Citotóxicos/imunologia , Fenômenos Biomecânicos/imunologia , Citotoxicidade Imunológica , Macrófagos/imunologia , Camundongos Endogâmicos C57BLRESUMO
Novel methods for sampling and characterizing biodiversity hold great promise for re-evaluating patterns of life across the planet. The sampling of airborne spores with a cyclone sampler, and the sequencing of their DNA, have been suggested as an efficient and well-calibrated tool for surveying fungal diversity across various environments. Here we present data originating from the Global Spore Sampling Project, comprising 2,768 samples collected during two years at 47 outdoor locations across the world. Each sample represents fungal DNA extracted from 24 m3 of air. We applied a conservative bioinformatics pipeline that filtered out sequences that did not show strong evidence of representing a fungal species. The pipeline yielded 27,954 species-level operational taxonomic units (OTUs). Each OTU is accompanied by a probabilistic taxonomic classification, validated through comparison with expert evaluations. To examine the potential of the data for ecological analyses, we partitioned the variation in species distributions into spatial and seasonal components, showing a strong effect of the annual mean temperature on community composition.
Assuntos
Microbiologia do Ar , DNA Fúngico , Esporos Fúngicos , DNA Fúngico/análise , Fungos/genética , Fungos/classificação , BiodiversidadeRESUMO
Immune cells live intensely physical lifestyles characterized by structural plasticity, mechanosensitivity, and force exertion. Whether specific immune functions require stereotyped patterns of mechanical output, however, is largely unknown. To address this question, we used super-resolution traction force microscopy to compare cytotoxic T cell immune synapses with contacts formed by other T cell subsets and macrophages. T cell synapses were globally and locally protrusive, which was fundamentally different from the coupled pinching and pulling of macrophage phagocytosis. By spectrally decomposing the force exertion patterns of each cell type, we associated cytotoxicity with compressive strength, local protrusiveness, and the induction of complex, asymmetric interfacial topographies. These features were further validated as cytotoxic drivers by genetic disruption of cytoskeletal regulators, direct imaging of synaptic secretory events, and in silico analysis of interfacial distortion. We conclude that T cell-mediated killing and, by implication, other effector responses are supported by specialized patterns of efferent force.
RESUMO
Cytotoxic lymphocytes fight pathogens and cancer by forming immune synapses with infected or transformed target cells and then secreting cytotoxic perforin and granzyme into the synaptic space, with potent and specific killing achieved by this focused delivery. The mechanisms that establish the precise location of secretory events, however, remain poorly understood. Here we use single cell biophysical measurements, micropatterning, and functional assays to demonstrate that localized mechanotransduction helps define the position of secretory events within the synapse. Ligand-bound integrins, predominantly the αLß2 isoform LFA-1, function as spatial cues to attract lytic granules containing perforin and granzyme and induce their fusion with the plasma membrane for content release. LFA-1 is subjected to pulling forces within secretory domains, and disruption of these forces via depletion of the adaptor molecule talin abrogates cytotoxicity. We thus conclude that lymphocytes employ an integrin-dependent mechanical checkpoint to enhance their cytotoxic power and fidelity.
Assuntos
Antígeno-1 Associado à Função Linfocitária , Mecanotransdução Celular , Citotoxicidade Imunológica , Granzimas/metabolismo , Antígeno-1 Associado à Função Linfocitária/metabolismo , Perforina/metabolismo , Sinapses/metabolismo , Linfócitos T CitotóxicosRESUMO
Force exertion is an integral part of cellular behavior. Traction force microscopy (TFM) has been instrumental for studying such forces, providing spatial force measurements at subcellular resolution. However, the applications of classical TFM are restricted by the typical planar geometry. Here, we develop a particle-based force sensing strategy for studying cellular interactions. We establish a straightforward batch approach for synthesizing uniform, deformable and tuneable hydrogel particles, which can also be easily derivatized. The 3D shape of such particles can be resolved with superresolution (<50 nm) accuracy using conventional confocal microscopy. We introduce a reference-free computational method allowing inference of traction forces with high sensitivity directly from the particle shape. We illustrate the potential of this approach by revealing subcellular force patterns throughout phagocytic engulfment and force dynamics in the cytotoxic T-cell immunological synapse. This strategy can readily be adapted for studying cellular forces in a wide range of applications.
Assuntos
Comunicação Celular , Linfócitos T Citotóxicos/química , Linfócitos T Citotóxicos/imunologia , Animais , Linhagem Celular , Células Cultivadas , Camundongos , Camundongos Endogâmicos C57BL , Microscopia de Força Atômica , Fagocitose , Linfócitos T Citotóxicos/citologia , TraçãoRESUMO
Psoriasis is a dermatologic disease of immune origins with no definitive cure. We report the Makati Medical Center experience of utilizing autologous mesenchymal stromal cells (MSCs) for one patient with psoriasis vulgaris (PV) and another with psoriatic arthritis (PA). Patients were educated and gave informed consent, according to the principles of the Declaration of Helsinki. The protocol was approved by the Cellular Transplantation Ethics Committee of the Makati Medical Center. Autologous MSCs were cultured from lipoaspirate and expanded in a clean room class 100 facility (Cellular Therapeutics Center, Makati Medical Center). MSCs were infused intravenously at a dose of 0.5-3.1 million cells/kg after complying with quality control parameters. Psoriasis area and severity index (PASI) evaluations were conducted by third-party dermatologists. The PA patient, who was previously unresponsive to standard treatment modalities, demonstrated a decrease in PASI (from 21.6 to 9.0, mild state after two infusions). No improvements were noted in joint pain until further treatment with etanercept and infliximab. The PV patient, who was previously dependent on methotrexate, showed a decrease in PASI from 24.0 to 8.3 after three infusions; this clinical improvement was sustained for 292 days (9.7 months) without methotrexate. The PV patient illustrated a marginal reduction in serum tumor necrosis factor-α (TNF-α), while significant (3.5- to 5-fold) decreases in reactive oxygen species (ROS) activity were noted. The ROS levels correlated with the clinical improvement of the PV patient. No serious adverse events were noted for either patient as a result of MSC infusions. This report demonstrates safe and tolerable transplantation of autologous MSCs for the treatment of psoriasis and warrants large clinical studies to investigate the long-term safety and efficacy of this approach.