Abstract
Aims
Acute myocardial infarction rapidly increases blood neutrophils (<2 h). Release from bone marrow, in response to chemokine elevation, has been considered their source, but chemokine ...levels peak up to 24 h after injury, and after neutrophil elevation. This suggests that additional non-chemokine-dependent processes may be involved. Endothelial cell (EC) activation promotes the rapid (<30 min) release of extracellular vesicles (EVs), which have emerged as an important means of cell–cell signalling and are thus a potential mechanism for communicating with remote tissues.
Methods and results
Here, we show that injury to the myocardium rapidly mobilizes neutrophils from the spleen to peripheral blood and induces their transcriptional activation prior to arrival at the injured tissue. Time course analysis of plasma-EV composition revealed a rapid and selective increase in EVs bearing VCAM-1. These EVs, which were also enriched for miRNA-126, accumulated preferentially in the spleen where they induced local inflammatory gene and chemokine protein expression, and mobilized splenic-neutrophils to peripheral blood. Using CRISPR/Cas9 genome editing, we generated VCAM-1-deficient EC-EVs and showed that its deletion removed the ability of EC-EVs to provoke the mobilization of neutrophils. Furthermore, inhibition of miRNA-126 in vivo reduced myocardial infarction size in a mouse model.
Conclusions
Our findings show a novel EV-dependent mechanism for the rapid mobilization of neutrophils to peripheral blood from a splenic reserve and establish a proof of concept for functional manipulation of EV-communications through genetic alteration of parent cells.
Graphical Abstract
Graphical Abstract
Dendritic cells (DCs) are key directors of tolerogenic and immunogenic immune responses. During the steady state, DCs maintain T cell tolerance to self-antigens by multiple mechanisms including ...inducing anergy, deletion, and Treg activity. All of these mechanisms help to prevent autoimmune diseases or other hyperreactivities. Different DC subsets contribute to pathogen recognition by expression of different subsets of pattern recognition receptors, including Toll-like receptors or C-type lectins. In addition to the triggering of immune responses in infected hosts, most pathogens have evolved mechanisms for evasion of targeted responses. One such strategy is characterized by adopting the host's T cell tolerance mechanisms. Understanding these tolerogenic mechanisms is of utmost importance for therapeutic approaches to treat immune pathologies, tumors and infections. Transcriptional profiling has developed into a potent tool for DC subset identification. Here, we review and compile pathogen-induced tolerogenic transcriptional signatures from mRNA profiling data of currently available bacterial- or helminth-induced transcriptional signatures. We compare them with signatures of tolerogenic steady-state DC subtypes to identify common and divergent strategies of pathogen induced immune evasion. Candidate molecules are discussed in detail. Our analysis provides further insights into tolerogenic DC signatures and their exploitation by different pathogens.
Intracellular bacterial pathogens can exhibit large heterogeneity in growth rate inside host cells, with major consequences for the infection outcome. If and how the host responds to this ...heterogeneity remains poorly understood. Here, we combined a fluorescent reporter of bacterial cell division with single-cell RNA-sequencing analysis to study the macrophage response to different intracellular states of the model pathogen Salmonella enterica serovar Typhimurium. The transcriptomes of individual infected macrophages revealed a spectrum of functional host response states to growing and non-growing bacteria. Intriguingly, macrophages harbouring non-growing Salmonella display hallmarks of the proinflammatory M1 polarization state and differ little from bystander cells, suggesting that non-growing bacteria evade recognition by intracellular immune receptors. By contrast, macrophages containing growing bacteria have turned into an anti-inflammatory, M2-like state, as if fast-growing intracellular Salmonella overcome host defence by reprogramming macrophage polarization. Additionally, our clustering approach reveals intermediate host functional states between these extremes. Altogether, our data suggest that gene expression variability in infected host cells shapes different cellular environments, some of which may favour a growth arrest of Salmonella facilitating immune evasion and the establishment of a long-term niche, while others allow Salmonella to escape intracellular antimicrobial activity and proliferate.
Since the approval of ibrutinib for relapsed/refractory mantle cell lymphoma (MCL), the treatment of this rare mature B-cell neoplasm has taken a great leap forward. Despite promising efficacy of the ...Bruton tyrosine kinase inhibitor, resistance arises inevitably and the underlying mechanisms remain to be elucidated. Here, we aimed to decipher the response of a sensitive MCL cell line treated with ibrutinib using time-resolved single-cell RNA sequencing. The analysis uncovered five subpopulations and their individual responses to the treatment. The effects on the B cell receptor pathway, cell cycle, surface antigen expression, and metabolism were revealed by the computational analysis and were validated by molecular biological methods. The observed upregulation of B cell receptor signaling, crosstalk with the microenvironment, upregulation of
, and metabolic reprogramming towards dependence on oxidative phosphorylation favor resistance to ibrutinib treatment. Targeting these cellular responses provide new therapy options in MCL.
Nasal xenobiotic metabolizing enzymes (XMEs) are important for the sense of smell because they influence odorant availability and quality. Since the major part of the human nasal cavity is lined by a ...respiratory mucosa, we hypothesized that this tissue contributed to nasal odorant metabolism through XME activity. Thus, we built human respiratory tissue models and characterized the XME profiles using single-cell RNA sequencing. We focused on the XMEs dicarbonyl and l-xylulose reductase, aldehyde dehydrogenase (ALDH) 1A1, and ALDH3A1, which play a role in food odorant metabolism. We demonstrated protein abundance and localization in the tissue models and showed the metabolic activity of the corresponding enzyme families by exposing the models to the odorants 3,4-hexandione and benzaldehyde. Using gas chromatography coupled with mass spectrometry, we observed, for example, a significantly higher formation of the corresponding metabolites 4-hydroxy-3-hexanone (39.03 ± 1.5%, p = 0.0022), benzyl alcohol (10.05 ± 0.88%, p = 0.0008), and benzoic acid (8.49 ± 0.57%, p = 0.0004) in odorant-treated tissue models compared to untreated controls (0 ± 0, 0.12 ± 0.12, and 0.18 ± 0.18%, respectively). This is the first study that reveals the XME profile of tissue-engineered human respiratory mucosa models and demonstrates their suitability to study nasal odorant metabolism.
Aging is a major risk factor for the development of nervous system functional decline, even in the absence of diseases or trauma. The axon-myelin units and synaptic terminals are some of the neural ...structures most vulnerable to aging-related deterioration
, but the underlying mechanisms are poorly understood. In the peripheral nervous system, macrophages-important representatives of the innate immune system-are prominent drivers of structural and functional decline of myelinated fibers and motor endplates during aging
. Similarly, in the aging central nervous system (CNS), microglial cells promote damage of myelinated axons and synapses
. Here we examine the role of cytotoxic CD8
T lymphocytes, a type of adaptive immune cells previously identified as amplifiers of axonal perturbation in various models of genetically mediated CNS diseases
but understudied in the aging CNS
. We show that accumulation of CD8
T cells drives axon degeneration in the normal aging mouse CNS and contributes to age-related cognitive and motor decline. We characterize CD8
T-cell population heterogeneity in the adult and aged mouse brain by single-cell transcriptomics and identify aging-related changes. Mechanistically, we provide evidence that CD8
T cells drive axon degeneration in a T-cell receptor- and granzyme B-dependent manner. Cytotoxic neural damage is further aggravated by systemic inflammation in aged but not adult mice. We also find increased densities of T cells in white matter autopsy material from older humans. Our results suggest that targeting CD8
CNS-associated T cells in older adults might mitigate aging-related decline of brain structure and function.
Innate lymphoid cells (ILCs) participate in tissue homeostasis, inflammation, and early immunity against infection. It is unclear how ILCs acquire effector function and whether these mechanisms ...differ between organs. Through multiplexed single-cell mRNA sequencing, we identified cKit
CD127
TCF-1
early differentiation stages of T-bet
ILC1s. These cells were present across different organs and had the potential to mature toward CD127
TCF-1
and CD127
TCF-1
ILC1s. Paralleling a gradual loss of TCF-1, differentiating ILC1s forfeited their expansion potential while increasing expression of effector molecules, reminiscent of T cell differentiation in secondary lymphoid organs. The transcription factor Hobit was induced in TCF-1
ILC1s and was required for their effector differentiation. These findings reveal sequential mechanisms of ILC1 lineage commitment and effector differentiation that are conserved across tissues. Our analyses suggest that ILC1s emerge as TCF-1
cells in the periphery and acquire a spectrum of organ-specific effector phenotypes through a uniform Hobit-dependent differentiation pathway driven by local cues.
Abstract
The human gastric epithelium forms highly organized gland structures with different subtypes of cells. The carcinogenic bacterium
Helicobacter pylori
can attach to gastric cells and ...subsequently translocate its virulence factor CagA, but the possible host cell tropism of
H. pylori
is currently unknown. Here, we report that
H. pylori
preferentially attaches to differentiated cells in the pit region of gastric units. Single-cell RNA-seq shows that organoid-derived monolayers recapitulate the pit region, while organoids capture the gland region of the gastric units. Using these models, we show that
H. pylori
preferentially attaches to highly differentiated pit cells, marked by high levels of GKN1, GKN2 and PSCA. Directed differentiation of host cells enable enrichment of the target cell population and confirm
H. pylori
preferential attachment and CagA translocation into these cells. Attachment is independent of MUC5AC or PSCA expression, and instead relies on bacterial TlpB-dependent chemotaxis towards host cell-released urea, which scales with host cell size.
Many cellular processes involve the recruitment of proteins to specific membranes, which are decorated with distinctive lipids that act as docking sites. The phosphoinositides form signaling hubs, ...and we examine mechanisms underlying recruitment. We applied a physiological, quantitative, liposome microarray-based assay to measure the membrane-binding properties of 91 pleckstrin homology (PH) domains, the most common phosphoinositide-binding target. 10,514 experiments quantified the role of phosphoinositides in membrane recruitment. For most domains examined, the observed binding specificity implied cooperativity with additional signaling lipids. Analyses of PH domains with similar lipid-binding profiles identified a conserved motif, mutations in which—including some found in human cancers—induced discrete changes in binding affinities in vitro and protein mislocalization in vivo. The data set reveals cooperativity as a key mechanism for membrane recruitment and, by enabling the interpretation of disease-associated mutations, suggests avenues for the design of small molecules targeting PH domains.
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•The lipid-binding properties of 91 pleckstrin homology domains have been quantified•Over 10,000 individual protein-lipid interaction experiments have been performed•The binding specificity and affinity imply cooperativity between signaling lipids•Mutations found in cancer biopsies fine-tune lipid-binding specificities
Vonkova et al. systematically quantify the lipid-binding properties of 91 pleckstrin homology (PH) domains using a physiological, quantitative, liposome microarray-based assay. The data set reveals that cooperativity between lipids is a key mechanism for membrane recruitment of PH domains.
Infection research largely relies on classical cell culture or mouse models. Despite having delivered invaluable insights into host-pathogen interactions, both have limitations in translating ...mechanistic principles to human pathologies. Alternatives can be derived from modern Tissue Engineering approaches, allowing the reconstruction of functional tissue models
. Here, we combined a biological extracellular matrix with primary tissue-derived enteroids to establish an
model of the human small intestinal epithelium exhibiting
-like characteristics. Using the foodborne pathogen
serovar Typhimurium, we demonstrated the applicability of our model to enteric infection research in the human context. Infection assays coupled to spatio-temporal readouts recapitulated the established key steps of epithelial infection by this pathogen in our model. Besides, we detected the upregulation of olfactomedin 4 in infected cells, a hitherto unrecognized aspect of the host response to
infection. Together, this primary human small intestinal tissue model fills the gap between simplistic cell culture and animal models of infection, and shall prove valuable in uncovering human-specific features of host-pathogen interplay.
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