CD151 Maintains Endolysosomal Protein Quality to Inhibit Vascular Inflammation.

BACKGROUND: Tetraspanin CD151 is highly expressed in endothelia and reinforces cell adhesion, but its role in vascular inflammation remains largely unknown. METHODS: In vitro molecular and cellular biological analyses on genetically modified endothelial cells, in vivo vascular biological analyses on genetically engineered mouse models, and in silico systems biology and bioinformatics analyses on CD151-related events. RESULTS: Endothelial ablation of Cd151 leads to pulmonary and cardiac inflammation, severe sepsis, and perilous COVID-19, and endothelial CD151 becomes downregulated in inflammation. Mechanistically, CD151 restrains endothelial release of proinflammatory molecules for less leukocyte infiltration. At the subcellular level, CD151 determines the integrity of multivesicular bodies/lysosomes and confines the production of exosomes that carry cytokines such as ANGPT2 (angiopoietin-2) and proteases such as cathepsin-D. At the molecular level, CD151 docks VCP (valosin-containing protein)/p97, which controls protein quality via mediating deubiquitination for proteolytic degradation, onto endolysosomes to facilitate VCP/p97 function. At the endolysosome membrane, CD151 links VCP/p97 to (1) IFITM3 (interferon-induced transmembrane protein 3), which regulates multivesicular body functions, to restrain IFITM3-mediated exosomal sorting, and (2) V-ATPase, which dictates endolysosome pH, to support functional assembly of V-ATPase. CONCLUSIONS: Distinct from its canonical function in strengthening cell adhesion at cell surface, CD151 maintains endolysosome function by sustaining VCP/p97-mediated protein unfolding and turnover. By supporting protein quality control and protein degradation, CD151 prevents proteins from (1) buildup in endolysosomes and (2) discharge through exosomes, to limit vascular inflammation. Also, our study conceptualizes that balance between degradation and discharge of proteins in endothelial cells determines vascular information. Thus, the IFITM3/V-ATPase-tetraspanin-VCP/p97 complexes on endolysosome, as a protein quality control and inflammation-inhibitory machinery, could be beneficial for therapeutic intervention against vascular inflammation.

Inhibition of LTßR signalling activates WNT-induced regeneration in lung.

Lymphotoxin ß-receptor (LTßR) signalling promotes lymphoid neogenesis and the development of tertiary lymphoid structures1,2, which are associated with severe chronic inflammatory diseases that span several organ systems3-6. How LTßR signalling drives chronic tissue damage particularly in the lung, the mechanism(s) that regulate this process, and whether LTßR blockade might be of therapeutic value have remained unclear. Here we demonstrate increased expression of LTßR ligands in adaptive and innate immune cells, enhanced non-canonical NF-κB signalling, and enriched LTßR target gene expression in lung epithelial cells from patients with smoking-associated chronic obstructive pulmonary disease (COPD) and from mice chronically exposed to cigarette smoke. Therapeutic inhibition of LTßR signalling in young and aged mice disrupted smoking-related inducible bronchus-associated lymphoid tissue, induced regeneration of lung tissue, and reverted airway fibrosis and systemic muscle wasting. Mechanistically, blockade of LTßR signalling dampened epithelial non-canonical activation of NF-κB, reduced TGFß signalling in airways, and induced regeneration by preventing epithelial cell death and activating WNT/ß-catenin signalling in alveolar epithelial progenitor cells. These findings suggest that inhibition of LTßR signalling represents a viable therapeutic option that combines prevention of tertiary lymphoid structures1 and inhibition of apoptosis with tissue-regenerative strategies.

Sfrp1 inhibits lung fibroblast invasion during transition to injury-induced myofibroblasts.

BACKGROUND: Fibroblast-to-myofibroblast conversion is a major driver of tissue remodelling in organ fibrosis. Distinct lineages of fibroblasts support homeostatic tissue niche functions, yet their specific activation states and phenotypic trajectories during injury and repair have remained unclear. METHODS: We combined spatial transcriptomics, multiplexed immunostainings, longitudinal single-cell RNA-sequencing and genetic lineage tracing to study fibroblast fates during mouse lung regeneration. Our findings were validated in idiopathic pulmonary fibrosis patient tissues in situ as well as in cell differentiation and invasion assays using patient lung fibroblasts. Cell differentiation and invasion assays established a function of SFRP1 in regulating human lung fibroblast invasion in response to transforming growth factor (TGF)ß1. MEASUREMENTS AND MAIN RESULTS: We discovered a transitional fibroblast state characterised by high Sfrp1 expression, derived from both Tcf21-Cre lineage positive and negative cells. Sfrp1 + cells appeared early after injury in peribronchiolar, adventitial and alveolar locations and preceded the emergence of myofibroblasts. We identified lineage-specific paracrine signals and inferred converging transcriptional trajectories towards Sfrp1 + transitional fibroblasts and Cthrc1 + myofibroblasts. TGFß1 downregulated SFRP1 in noninvasive transitional cells and induced their switch to an invasive CTHRC1+ myofibroblast identity. Finally, using loss-of-function studies we showed that SFRP1 modulates TGFß1-induced fibroblast invasion and RHOA pathway activity. CONCLUSIONS: Our study reveals the convergence of spatially and transcriptionally distinct fibroblast lineages into transcriptionally uniform myofibroblasts and identifies SFRP1 as a modulator of TGFß1-driven fibroblast phenotypes in fibrogenesis. These findings are relevant in the context of therapeutic interventions that aim at limiting or reversing fibroblast foci formation.

Transcriptomic and Proteomic Changes Driving Pulmonary Fibrosis Resolution in Young and Old Mice.

Bleomycin-induced pulmonary fibrosis in mice mimics major hallmarks of idiopathic pulmonary fibrosis. Yet in this model, it spontaneously resolves over time. We studied molecular mechanisms of fibrosis resolution and lung repair, focusing on transcriptional and proteomic signatures and the effect of aging. Old mice showed incomplete and delayed lung function recovery 8 weeks after bleomycin instillation. This shift in structural and functional repair in old bleomycin-treated mice was reflected in a temporal shift in gene and protein expression. We reveal gene signatures and signaling pathways that underpin the lung repair process. Importantly, the downregulation of WNT, BMP, and TGFß antagonists Frzb, Sfrp1, Dkk2, Grem1, Fst, Fstl1, and Inhba correlated with lung function improvement. Those genes constitute a network with functions in stem cell pathways, wound, and pulmonary healing. We suggest that insufficient and delayed downregulation of those antagonists during fibrosis resolution in old mice explains the impaired regenerative outcome. Together, we identified signaling pathway molecules with relevance to lung regeneration that should be tested in-depth experimentally as potential therapeutic targets for pulmonary fibrosis.

Climate change-related growth improvements in a wide niche-breadth tree species across contrasting environments.

BACKGROUND AND AIMS: The vulnerability and responsiveness of forests to drought are immensely variable across biomes. Intraspecific tree responses to drought in species with wide niche breadths that grow across contrasting climatically environments might provide key information regarding forest resistance and changes in species distribution under climate change. Using a species with an exceptionally wide niche breath, we tested the hypothesis that tree populations thriving in dry environments are more resistant to drought than those growing in moist locations. METHODS: We determined temporal trends in tree radial growth of 12 tree populations of Nothofagus antarctica (Nothofagaceae) located across a sharp precipitation gradient (annual precipitation of 500-2000 mm) in Chile and Argentina. Using dendrochronological methods, we fitted generalized additive mixed-effect models to predict the annual basal area increment as a function of year and dryness (De Martonne aridity index). We also measured carbon and oxygen isotope signals (and estimated intrinsic water-use efficiency) to provide potential physiological causes for tree growth responses to drought. KEY RESULTS: We found unexpected improvements in growth during 1980-1998 in moist sites, while growth responses in dry sites were mixed. All populations, independent of site moisture, showed an increase in their intrinsic water-use efficiency in recent decades, a tendency that seemed to be explained by an increase in the photosynthetic rate instead of drought-induced stomatal closure, given that δ18O did not change with time. CONCLUSIONS: The absence of drought-induced negative effects on tree growth in a tree species with a wide niche breadth is promising because it might relate to the causal mechanisms tree species possess to face ongoing drought events. We suggest that the drought resistance of N. antarctica might be attributable to its low stature and relatively low growth rate.

The discovAIR project: a roadmap towards the Human Lung Cell Atlas.

The Human Cell Atlas (HCA) consortium aims to establish an atlas of all organs in the healthy human body at single-cell resolution to increase our understanding of basic biological processes that govern development, physiology and anatomy, and to accelerate diagnosis and treatment of disease. The Lung Biological Network of the HCA aims to generate the Human Lung Cell Atlas as a reference for the cellular repertoire, molecular cell states and phenotypes, and cell-cell interactions that characterise normal lung homeostasis in healthy lung tissue. Such a reference atlas of the healthy human lung will facilitate mapping the changes in the cellular landscape in disease. The discovAIR project is one of six pilot actions for the HCA funded by the European Commission in the context of the H2020 framework programme. discovAIR aims to establish the first draft of an integrated Human Lung Cell Atlas, combining single-cell transcriptional and epigenetic profiling with spatially resolving techniques on matched tissue samples, as well as including a number of chronic and infectious diseases of the lung. The integrated Human Lung Cell Atlas will be available as a resource for the wider respiratory community, including basic and translational scientists, clinical medicine, and the private sector, as well as for patients with lung disease and the interested lay public. We anticipate that the Human Lung Cell Atlas will be the founding stone for a more detailed understanding of the pathogenesis of lung diseases, guiding the design of novel diagnostics and preventive or curative interventions.

Rivaroxaban Reduces Arterial Thrombosis by Inhibition of FXa-Driven Platelet Activation via Protease Activated Receptor-1.

RATIONALE: A reduced rate of myocardial infarction has been reported in patients with atrial fibrillation treated with FXa (factor Xa) inhibitors including rivaroxaban compared with vitamin K antagonists. At the same time, low-dose rivaroxaban has been shown to reduce mortality and atherothrombotic events in patients with coronary artery disease. Yet, the mechanisms underlying this reduction remain unknown. OBJECTIVE: In this study, we hypothesized that rivaroxaban's antithrombotic potential is linked to a hitherto unknown rivaroxaban effect that impacts on platelet reactivity and arterial thrombosis. METHODS AND RESULTS: In this study, we identified FXa as potent, direct agonist of the PAR-1 (protease-activated receptor 1), leading to platelet activation and thrombus formation, which can be inhibited by rivaroxaban. We found that rivaroxaban reduced arterial thrombus stability in a mouse model of arterial thrombosis using intravital microscopy. For in vitro studies, atrial fibrillation patients on permanent rivaroxaban treatment for stroke prevention, respective controls, and patients with new-onset atrial fibrillation before and after first intake of rivaroxaban (time series analysis) were recruited. Platelet aggregation responses, as well as thrombus formation under arterial flow conditions on collagen and atherosclerotic plaque material, were attenuated by rivaroxaban. We show that rivaroxaban's antiplatelet effect is plasma dependent but independent of thrombin and rivaroxaban's anticoagulatory capacity. CONCLUSIONS: Here, we identified FXa as potent platelet agonist that acts through PAR-1. Therefore, rivaroxaban exerts an antiplatelet effect that together with its well-known potent anticoagulatory capacity might lead to reduced frequency of atherothrombotic events and improved outcome in patients.

Stable isotope relationships between apatite phosphate (δ18 O), structural carbonate (δ18 O, δ13 C), and collagen (δ2 H, δ13 C, δ15 N, δ34 S) in modern human dentine.

RATIONALE: The use of multi-isotopic analysis (δ2 H, δ13 C, δ15 N, δ18 O, and δ34 S values) of modern human body tissues for provenancing of unknown individuals in forensics is increasing. Tooth dentine develops during childhood and adolescence, therefore providing geographical information from that period of life. Tooth apatite δ18 O values are commonly used for the reconstruction of drinking water values, and H-C-N-S isotope ratios in collagen supply additional information about the composition of diet. We tested if dentine collagen δ2 H values provide similar information to apatite δ18 O values with a proof-of-concept study. METHODS: Tooth samples were taken from modern-day individuals born in different regions of the world. Apatite and collagen were prepared from dentine. Stable isotope analyses were performed on apatite phosphate oxygen (δ18 Ophos ); oxygen and carbon of the structural carbonate (δ18 Ocarb , δ13 Ccarb ); and hydrogen, carbon, nitrogen, and sulfur of the collagen (δ2 Hcoll, δ13 Ccoll , δ15 N, δ34 S). RESULTS: δ18 Ophos , δ18 Ocarb , and δ2 Hcoll values are highly correlated in modern human dentine. There are significant relationships of δ18 O values in the apatite fraction and δ2 H values in the collagen fraction with local δ18 O and δ2 H precipitation values, respectively. Pearson correlation coefficients indicate no direct relationship between δ15 N values and the isotope ratios of any other element. Weak relationships exist between collagen δ34 S values and δ18 Ocarb or δ18 Ophos values. CONCLUSIONS: The highly significant correlation of δ18 Ophos , δ18 Ocarb , and δ2 Hcoll values in the modern human dentine implies that measurement of δ2 H values in collagen or δ18 O values in bioapatite will provide reliable information about the climate at the person's whereabouts.

Multi-isotope provenancing of archaeological skeletons including cremations in a reference area of the European Alps.

RATIONALE: Due to the spatial heterogeneity of stable isotope ratios of single elements measured in attempts to georeference bioarchaeological finds, multi-isotope fingerprints are frequently employed under the assumption that similar isotopic signatures are indicative of similar shared environments by the individuals studied. The extraction of the spatial information from multi-isotope datasets, however, is challenging. METHODS: Gaussian mixture clustering of six- to seven-dimensional isotopic fingerprints measured in archaeological animal and human bones was performed. Uncremated animal bones served for an isotopic mapping of a specific reference area of eminent archaeological importance, namely the Inn-Eisack-Adige passage across the European Alps. The fingerprints consist of 87 Sr/86 Sr, 208 Pb/204 Pb, 207 Pb/204 Pb, 206 Pb/204 Pb, 208 Pb/207 Pb, and 206 Pb/207 Pb ratios, and δ18 Ophosphate values in uncremated bone apatite, while the thermally unstable δ18 O values of human cremations from this region were discarded. RESULTS: The bone finds were successfully decontaminated. Animal and human isotope clusters not only reflect individual similarities in the multi-isotopic fingerprints, but also permit a spatial allocation of the finds. This holds also for cremated finds where the δ18 Ophosphate value is no longer informative. To our knowledge, for the first time Pb stable isotopes have been systematically studied in cremated skeletal remains and proved significant in a region that was sought after for its ore deposits in prehistory. CONCLUSIONS: Gaussian mixture clustering is a promising method for the interpretation of multi-isotopic fingerprints aiming at detecting and quantifying migration and trade.

Deep Proteome Profiling Reveals Common Prevalence of MZB1-Positive Plasma B Cells in Human Lung and Skin Fibrosis.

RATIONALE: Analyzing the molecular heterogeneity of different forms of organ fibrosis may reveal common and specific factors and thus identify potential future therapeutic targets. OBJECTIVES: We sought to use proteome-wide profiling of human tissue fibrosis to (1) identify common and specific signatures across end-stage interstitial lung disease (ILD) cases, (2) characterize ILD subgroups in an unbiased fashion, and (3) identify common and specific features of lung and skin fibrosis. METHODS: We collected samples of ILD tissue (n = 45) and healthy donor control samples (n = 10), as well as fibrotic skin lesions from localized scleroderma and uninvolved skin (n = 6). Samples were profiled by quantitative label-free mass spectrometry, Western blotting, or confocal imaging. MEASUREMENTS AND MAIN RESULTS: We determined the abundance of more than 7,900 proteins and stratified these proteins according to their detergent solubility profiles. Common protein regulations across all ILD cases, as well as distinct ILD subsets, were observed. Proteomic comparison of lung and skin fibrosis identified a common upregulation of marginal zone B- and B1-cell-specific protein (MZB1), the expression of which identified MZB1+/CD38+/CD138+/CD27+/CD45-/CD20- plasma B cells in fibrotic lung and skin tissue. MZB1 levels correlated positively with tissue IgG and negatively with diffusing capacity of the lung for carbon monoxide. CONCLUSIONS: Despite the presumably high molecular and cellular heterogeneity of ILD, common protein regulations are observed, even across organ boundaries. The surprisingly high prevalence of MZB1-positive plasma B cells in tissue fibrosis warrants future investigations regarding the causative role of antibody-mediated autoimmunity in idiopathic cases of organ fibrosis, such as idiopathic pulmonary fibrosis.

Anxiety, depression, and health-related quality of life in aging people living with HIV compared to diabetes patients and patients with minor health conditions: a longitudinal study.

HIV infection has evolved from a fatal to a treatable condition, leading to an increase in the rate of elderly People Living with HIV (PLWH). However, little is known about the psychosocial burden of elderly PLWH. Thus, the aim of this longitudinal multi-center cohort study was to investigate whether elderly PLWH experience more anxiety and depression and reduced health related quality of life (HRQOL) compared to elderly patients with other chronic conditions. PLWH were compared to diabetes patients (DM) and patients with minor health conditions (MHC), e.g. patients with hypertension or allergic conditions. All patients were over 50 years old. Anxiety and depression (HADS) as well as HRQOL (SF-36) were assessed at baseline and after 12 months. 218 PLWH, 249 DM and 254 MHC were included. At baseline, the study groups did not differ in anxiety, depression, and physical HRQOL. However, PLWH indicated lower mental HRQOL than DM and MHC patients (p = 0.001). We did not obtain any moderating effects showing a differential effect of patient characteristics on anxiety, depression, and HRQOL in the three patient groups. At follow-up, the level of anxiety, depression, and HRQOL did not change significantly. The prevalence of anxiety ranged between 27 and 35%, and that of depression between 17 and 28%. Thus, the results of our investigation tentatively suggest that the psychosocial adaptation to HIV among elderly PLWH resembles those of other chronic diseases. There may be some subtle impairments, though, as PLWH experienced lower mental HRQOL.

Effects of organic matter on carbonate stable isotope ratios (δ13C, δ18O values)--implications for analyses of bulk sediments.

RATIONALE: Stable isotope ratio (δ(13)C, δ(18)O values) analyses of carbonates can be biased by CO(2) release from organic impurities. This is most critical for carbonate isotope analyses from bulk sediments containing comparably high amounts of organic matter (OM). Several methods have been developed to remove OM prior to analyses, but none of them can be universally applied. Moreover, pretreatment methods cause isotopic bias in themselves and should probably best be avoided. Thus, it is essential to have indicators for reliable isotope values of untreated carbonate-OM mixtures. METHODS: Artificial mixtures of organic compounds with a standard carbonate were analyzed to investigate the bias on carbonate isotope ratios caused by OM in the sample. The total-inorganic-carbon to total-organic-carbon ratio (TIC/TOC) was used as a measure for the " organic impurity" of the sample. The target was to evaluate TIC/TOC as a measure for sample quality and to define TIC/TOC thresholds for reliable isotope measurements of mixtures between calcium carbonate and organic compounds. RESULTS: The effect of organic impurities on carbonate stable isotope values depended on the specific OM compound and the respective TIC/TOC ratio. Different CO(2) release rates were determined for the pure OM compounds. A sample TIC/TOC ratio ≥0.3 was found to be a threshold for reliable measurements of the isotope composition of calcium carbonate. CONCLUSIONS: Bulk carbonate analyses from carbonate-OM mixtures are reliable only if the TIC/TOC values do not fall below certain thresholds. This has implications for carbonate isotope studies from bulk sediments for which the TIC/TOC ratios should be considered as an easy-to-determine measure for sample-quality assessment.

Multi-isotope fingerprints of recent environmental samples from the Baltic coast and their implications for bioarchaeological studies.

Stable isotopes in coastal regions are influenced by the so-called sea spray effect which masks the actual terrestrial isotope fingerprint with a marine isotope signal. The sea spray impact on plants was investigated by analyzing different stable isotope systems (δ13Ccellulose, δ18Ocellulose, δ18Osulfate, δ34Ssulfate, δ34Stotal S, δ34Sorganic S, 87Sr/86Sr) in recent environmental samples (plants, soil, water) collected close to the Baltic Sea. All these isotopic systems are influenced by the sea spray, either by the uptake of ions (HCO3-, SO42-, Sr2+) of marine origin, thus exhibiting a marine isotopic signature, or by biochemical reactions associated with, e.g., salinity stress. A shift towards seawater values is observed for δ18Osulfate, δ34S, and 87Sr/86Sr. Cellulose becomes enriched in 13C and 18O due to sea spray, further enhanced (δ13Ccellulose) or mitigated (δ18Ocellulose) by salinity stress. The effect differs both regionally and seasonally, probably as a result of, e.g., differences in wind strength or prevailing wind direction, as well as between plants collected only few meters apart, in either the open field or at more wind-protected sites, reflecting samples more or less influenced by sea spray. The stable isotope data of recent environmental samples are compared to previously analyzed archaeological bone samples of animals from the Viking Haithabu and Early Medieval Schleswig sites located close to the Baltic Sea. Potential regions of origin can be predicted based on the magnitude of the (recent) local sea spray effect. This enables the identification of probably non-local individuals. The insights into sea spray mechanisms, biochemical reactions in plants, as well as seasonal, regional, and small-scale differences in stable isotope data will help to interpret multi-isotope fingerprints at coastal sites. Our study demonstrates the usefulness of environmental samples for bioarchaeological studies. Moreover, the detected seasonal and small-scale differences require adjusted sampling strategies for, e.g., isotopic baselines in coastal areas.

Identification and quantification of the sea spray effect on isotopic systems in α-cellulose (δ13C, δ18O), total sulfur (δ34S), and 87Sr/86Sr of European beach grass (Ammophila arenaria, L.) in a greenhouse experiment.

The sea spray effect can severely influence the isotopic signature of terrestrial individuals in coastal regions. To further specify this effect, beach grass was grown in a greenhouse under controlled environmental conditions and sprayed with mineral salt solution containing different mineral salts but only traces of NaCl (group 1). Another group of plants was sprayed with salty water from the Schlei inlet and the Baltic Sea, respectively (group 2). Control plants were only sprayed with tap water. Isotope analyses were conducted on the unwashed and washed plants (δ13Ccellulose, δ18Ocellulose, δ34Stotal S, 87Sr/86Sr), soil (δ18Osulfate, δ34Ssulfate, 87Sr/86Sr), and spray as well as irrigation water (δ18Osulfate, δ34Ssulfate, 87Sr/86Sr). Moreover, elemental analyses were performed on the water samples. The sea spray effect was visible in all isotopic systems under study. The uptake of SO42-, HCO3-, and Sr2+ directly affected plants of group 1, while plants of group 2, sprayed with salty water, additionally showed salinity stress in the case of α-cellulose and total sulfur due to biochemical reactions of the plants. Very high concentrations in HCO3- or SO42- also affected the plants' isotopic signatures. The impact of the sea spray and additional stress reactions were quantified. Our study is the first experiment creating an artificial sea spray effect in a greenhouse. This experiment for the first time enables the identification and quantification of the sea spray effect in environmental samples. The marine signature taken up by the plants and recorded by the investigated isotopic systems is apparently high and should have an impact on the isotopic fingerprints of animal consumers at the coast, as evidenced for archaeological animals from the Viking Haithabu and the early medieval Schleswig sites located close to the Baltic Sea. This result demonstrates the potential of greenhouse experiments as an isotopic predictor of the past local sea spray effect.

Mass spectrometry-based autoimmune profiling reveals predictive autoantigens in idiopathic pulmonary fibrosis.

Autoimmunity plays a role in certain types of lung fibrosis, notably connective tissue disease-associated interstitial lung disease (CTD-ILD). In idiopathic pulmonary fibrosis (IPF), an incurable and fatal lung disease, diagnosis typically requires clinical exclusion of autoimmunity. However, autoantibodies of unknown significance have been detected in IPF patients. We conducted computational analysis of B cell transcriptomes in published transcriptomics datasets and developed a proteomic Differential Antigen Capture (DAC) assay that captures plasma antibodies followed by affinity purification of lung proteins coupled to mass spectrometry. We analyzed antibody capture in two independent cohorts of IPF and CTL-ILD patients over two disease progression time points. Our findings revealed significant upregulation of specific immunoglobulins with V-segment bias in IPF across multiple cohorts. We identified a predictive autoimmune signature linked to reduced transplant-free survival in IPF, persisting over time. Notably, autoantibodies against thrombospondin-1 were associated with decreased survival, suggesting their potential as predictive biomarkers.

Ex vivo tissue perturbations coupled to single-cell RNA-seq reveal multilineage cell circuit dynamics in human lung fibrogenesis.

Pulmonary fibrosis develops as a consequence of failed regeneration after injury. Analyzing mechanisms of regeneration and fibrogenesis directly in human tissue has been hampered by the lack of organotypic models and analytical techniques. In this work, we coupled ex vivo cytokine and drug perturbations of human precision-cut lung slices (hPCLS) with single-cell RNA sequencing and induced a multilineage circuit of fibrogenic cell states in hPCLS. We showed that these cell states were highly similar to the in vivo cell circuit in a multicohort lung cell atlas from patients with pulmonary fibrosis. Using micro-CT-staged patient tissues, we characterized the appearance and interaction of myofibroblasts, an ectopic endothelial cell state, and basaloid epithelial cells in the thickened alveolar septum of early-stage lung fibrosis. Induction of these states in the hPCLS model provided evidence that the basaloid cell state was derived from alveolar type 2 cells, whereas the ectopic endothelial cell state emerged from capillary cell plasticity. Cell-cell communication routes in patients were largely conserved in hPCLS, and antifibrotic drug treatments showed highly cell type-specific effects. Our work provides an experimental framework for perturbational single-cell genomics directly in human lung tissue that enables analysis of tissue homeostasis, regeneration, and pathology. We further demonstrate that hPCLS offer an avenue for scalable, high-resolution drug testing to accelerate antifibrotic drug development and translation.

An integrated cell atlas of the lung in health and disease.

Single-cell technologies have transformed our understanding of human tissues. Yet, studies typically capture only a limited number of donors and disagree on cell type definitions. Integrating many single-cell datasets can address these limitations of individual studies and capture the variability present in the population. Here we present the integrated Human Lung Cell Atlas (HLCA), combining 49 datasets of the human respiratory system into a single atlas spanning over 2.4 million cells from 486 individuals. The HLCA presents a consensus cell type re-annotation with matching marker genes, including annotations of rare and previously undescribed cell types. Leveraging the number and diversity of individuals in the HLCA, we identify gene modules that are associated with demographic covariates such as age, sex and body mass index, as well as gene modules changing expression along the proximal-to-distal axis of the bronchial tree. Mapping new data to the HLCA enables rapid data annotation and interpretation. Using the HLCA as a reference for the study of disease, we identify shared cell states across multiple lung diseases, including SPP1+ profibrotic monocyte-derived macrophages in COVID-19, pulmonary fibrosis and lung carcinoma. Overall, the HLCA serves as an example for the development and use of large-scale, cross-dataset organ atlases within the Human Cell Atlas.

Genetic variation in IL28B and treatment-induced clearance of hepatitis C virus in HIV-positive patients with acute and chronic hepatitis C.

Recently, a IL28B (rs 12979860) gene polymorphism was identified as a predictor for response to hepatitis C virus-specific treatment in human immunodeficiency virus (HIV)-uninfected and -infected patients with chronic hepatitis C. In an analysis of HIV-infected patients with acute hepatitis C, we found that the IL28B genotype was associated with serum levels of hepatitis C virus RNA, g-GT, and CD4 cell count. In contrast to HIV-infected patients with chronic hepatitis C, the IL28B genotype was not significantly associated with treatment response rates in patients with acute hepatitis C. Thus, effects of the IL28B single-nucleotide polymorphism may differ in HIV-infected patients with chronic and acute hepatitis C.