IL-1 mediates microbiome-induced inflammaging of hematopoietic stem cells in mice.

Aging is associated with impaired hematopoietic and immune function caused in part by decreased fitness in the hematopoietic stem cell (HSC) population and an increased myeloid differentiation bias. The reasons for this aging-associated HSC impairment are incompletely understood. Here we demonstrate that older specific pathogen free (SPF) wild-type (WT) mice in contrast to young SPF mice produce more interleukin-1a and interleukin-1b (IL-1a/b) in steady-state bone marrow (BM), with most of the IL-1a/b being derived from myeloid BM cells. Furthermore, blood from steady-state older SPF WT mice contains higher levels of microbe-associated molecular patterns, specifically TLR4 and TLR8 ligands. In addition, BM myeloid cells from older mice produce more IL-1b in vitro, and older mice show higher and more durable IL-1a/b responses upon stimulation with lipopolysaccharide in vivo. To test whether HSC aging is driven by IL-1a/b, we evaluated HSCs from IL-1 receptor 1 (IL-1R1) knockout (KO) mice. Indeed, older HSCs from IL-1R1KO mice show significantly mitigated aging-associated inflammatory signatures. Moreover, HSCs from older IL-1R1KO and from germ-free mice maintain unbiased lymphomyeloid hematopoietic differentiation upon transplantation, thus resembling this functionality of young HSCs. Importantly, in vivo antibiotic suppression of microbiota or pharmacologic blockade of IL-1 signaling in older WT mice was similarly sufficient to reverse myeloid-biased output of their HSC populations. Collectively, our data define the microbiome/IL-1/IL-1R1 axis as a key, self-sustaining and also therapeutically partially reversible driver of HSC inflammaging.

In vivo single-cell labeling by confined primed conversion.

Spatially confined green-to-red photoconversion of fluorescent proteins with high-power, pulsed laser illumination is negligible, thus precluding optical selection of single cells in vivo. We report primed conversion, in which low-power, dual-wavelength, continuous-wave illumination results in pronounced photoconversion. With a straightforward addition to a conventional confocal microscope, we show confined primed conversion in living zebrafish and reveal the complex anatomy of individual neurons packed between neighboring cells.

Nitrile-functionalized Poly(siloxane) as Electrolytes for High-Energy-Density Solid-State Li Batteries.

In the quest to replace liquid Li-ion electrolytes with safer and non-toxic solid counterparts for Li-ion batteries, polysiloxane polymers have attracted considerable attention as they offer low glass transition temperatures, stability with metallic lithium, and versatility in chemical functionalization of the backbone. Herein, we present the synthesis of Li-ion conductive polysiloxane-based polymers functionalized with 60 % nitrile groups per chain unit. The synthesis procedure is based on the reaction of poly-(dimethylsiloxane-co-methylvinylsiloxane) polymer with 2-cyanoethanethiol, followed by the addition of lithium bis (trifluoromethanesulfonyl) imide. The presented polysiloxane-based polymers exhibit exceptionally high ionic conductivity up to 0.375 mS cm-1 at 60 °C and Li+ ion transfer number of 0.73, one of the highest reported for polymer Li-ion conducting electrolytes. Their electrochemical performance was evaluated in both symmetrical and full-cell configurations to test the utility of synthesized polymers as electrolytes in Li-ion batteries.

Modality Attention and Sampling Enables Deep Learning with Heterogeneous Marker Combinations in Fluorescence Microscopy.

Fluorescence microscopy allows for a detailed inspection of cells, cellular networks, and anatomical landmarks by staining with a variety of carefully-selected markers visualized as color channels. Quantitative characterization of structures in acquired images often relies on automatic image analysis methods. Despite the success of deep learning methods in other vision applications, their potential for fluorescence image analysis remains underexploited. One reason lies in the considerable workload required to train accurate models, which are normally specific for a given combination of markers, and therefore applicable to a very restricted number of experimental settings. We herein propose Marker Sampling and Excite - a neural network approach with a modality sampling strategy and a novel attention module that together enable (i) flexible training with heterogeneous datasets with combinations of markers and (ii) successful utility of learned models on arbitrary subsets of markers prospectively. We show that our single neural network solution performs comparably to an upper bound scenario where an ensemble of many networks is naïvely trained for each possible marker combination separately. In addition, we demonstrate the feasibility of this framework in high-throughput biological analysis by revising a recent quantitative characterization of bone marrow vasculature in 3D confocal microscopy datasets and further confirm the validity of our approach on an additional, significantly different dataset of microvessels in fetal liver tissues. Not only can our work substantially ameliorate the use of deep learning in fluorescence microscopy analysis, but it can also be utilized in other fields with incomplete data acquisitions and missing modalities.

Chronic viral infections persistently alter marrow stroma and impair hematopoietic stem cell fitness.

Chronic viral infections are associated with hematopoietic suppression, bone marrow (BM) failure, and hematopoietic stem cell (HSC) exhaustion. However, how persistent viral challenge and inflammatory responses target BM tissues and perturb hematopoietic competence remains poorly understood. Here, we combine functional analyses with advanced 3D microscopy to demonstrate that chronic infection with lymphocytic choriomeningitis virus leads to (1) long-lasting decimation of the BM stromal network of mesenchymal CXCL12-abundant reticular cells, (2) proinflammatory transcriptional remodeling of remaining components of this key niche subset, and (3) durable functional defects and decreased competitive fitness in HSCs. Mechanistically, BM immunopathology is elicited by virus-specific, activated CD8 T cells, which accumulate in the BM via interferon-dependent mechanisms. Combined antibody-mediated inhibition of type I and II IFN pathways completely preempts degeneration of CARc and protects HSCs from chronic dysfunction. Hence, viral infections and ensuing immune reactions durably impact BM homeostasis by persistently decreasing the competitive fitness of HSCs and disrupting essential stromal-derived, hematopoietic-supporting cues.

Imaging and spatial analysis of hematopoietic stem cell niches.

Hematopoietic stem cells (HSCs) have been long proposed to reside in defined anatomical locations within bone marrow (BM) tissues in direct contact or close proximity to nurturing cell types. Imaging techniques that allow the simultaneous mapping of HSCs and interacting cell types have been central to the discovery of basic principles of these so-called HSC niches. Despite major progress in the field, a quantitative and comprehensive model of the cellular and molecular components that define these specialized microenvironments is lacking to date, and uncertainties remain on the preferential localization of HSCs in the context of complex BM tissue landscapes. Recent technological breakthroughs currently allow for the quantitative spatial analysis of BM cellular components with extraordinary precision. Here, we critically discuss essential technical aspects related to imaging approaches, image processing tools, and spatial statistics, which constitute the three basic elements of rigorous quantitative spatial analyses of HSC niches in the BM microenvironment.

Combined single-cell and spatial transcriptomics reveal the molecular, cellular and spatial bone marrow niche organization.

The bone marrow constitutes the primary site for life-long blood production and skeletal regeneration. However, its cellular and spatial organization remains controversial. Here, we combine single-cell and spatially resolved transcriptomics to systematically map the molecular, cellular and spatial composition of distinct bone marrow niches. This allowed us to transcriptionally profile all major bone-marrow-resident cell types, determine their localization and clarify sources of pro-haematopoietic factors. Our data demonstrate that Cxcl12-abundant-reticular (CAR) cell subsets (Adipo-CAR and Osteo-CAR) differentially localize to sinusoidal and arteriolar surfaces, act locally as 'professional cytokine-secreting cells' and thereby establish peri-vascular micro-niches. Importantly, the three-dimensional bone-marrow organization can be accurately inferred from single-cell transcriptome data using the RNA-Magnet algorithm described here. Together, our study reveals the cellular and spatial organization of bone marrow niches and offers a systematic approach to dissect the complex organization of whole organs.

Global Transcriptomic Profiling of the Bone Marrow Stromal Microenvironment during Postnatal Development, Aging, and Inflammation.

Bone marrow (BM) stromal cells provide the regulatory framework for hematopoiesis and contribute to developmental stage-specific niches, such as those preserving hematopoietic stem cells. Despite advances in our understanding of stromal function, little is known about the transcriptional changes that this compartment undergoes throughout lifespan and during adaptation to stress. Using RNA sequencing, we perform transcriptional analyses of four principal stromal subsets, namely CXCL12-abundant reticular, platelet-derived growth factor receptor (PDGFR)-α+Sca1+, sinusoidal, and arterial endothelial cells, from early postnatal, adult, and aged mice. Our data reveal (1) molecular fingerprints defining cell-specific anatomical and functional features, (2) a radical reprogramming of pro-hematopoietic, immune, and matrisomic transcriptional programs during the transition from juvenile stages to adulthood, and (3) the aging-driven progressive upregulation of pro-inflammatory gene expression in stroma. We further demonstrate that transcriptomic pathways elicited in vivo by prototypic microbial molecules are largely recapitulated during aging, thereby supporting the inflammatory basis of age-related adaptations of BM hematopoietic function.

Platelet GPIbα is a mediator and potential interventional target for NASH and subsequent liver cancer.

Non-alcoholic fatty liver disease ranges from steatosis to non-alcoholic steatohepatitis (NASH), potentially progressing to cirrhosis and hepatocellular carcinoma (HCC). Here, we show that platelet number, platelet activation and platelet aggregation are increased in NASH but not in steatosis or insulin resistance. Antiplatelet therapy (APT; aspirin/clopidogrel, ticagrelor) but not nonsteroidal anti-inflammatory drug (NSAID) treatment with sulindac prevented NASH and subsequent HCC development. Intravital microscopy showed that liver colonization by platelets depended primarily on Kupffer cells at early and late stages of NASH, involving hyaluronan-CD44 binding. APT reduced intrahepatic platelet accumulation and the frequency of platelet-immune cell interaction, thereby limiting hepatic immune cell trafficking. Consequently, intrahepatic cytokine and chemokine release, macrovesicular steatosis and liver damage were attenuated. Platelet cargo, platelet adhesion and platelet activation but not platelet aggregation were identified as pivotal for NASH and subsequent hepatocarcinogenesis. In particular, platelet-derived GPIbα proved critical for development of NASH and subsequent HCC, independent of its reported cognate ligands vWF, P-selectin or Mac-1, offering a potential target against NASH.

Quantitative spatial analysis of haematopoiesis-regulating stromal cells in the bone marrow microenvironment by 3D microscopy.

Sinusoidal endothelial cells and mesenchymal CXCL12-abundant reticular cells are principal bone marrow stromal components, which critically modulate haematopoiesis at various levels, including haematopoietic stem cell maintenance. These stromal subsets are thought to be scarce and function via highly specific interactions in anatomically confined niches. Yet, knowledge on their abundance, global distribution and spatial associations remains limited. Using three-dimensional quantitative microscopy we show that sinusoidal endothelial and mesenchymal reticular subsets are remarkably more abundant than estimated by conventional flow cytometry. Moreover, both cell types assemble in topologically complex networks, associate to extracellular matrix and pervade marrow tissues. Through spatial statistical methods we challenge previous models and demonstrate that even in the absence of major specific interaction forces, virtually all tissue-resident cells are invariably in physical contact with, or close proximity to, mesenchymal reticular and sinusoidal endothelial cells. We further show that basic structural features of these stromal components are preserved during ageing.