Barrier tissue dysfunction is a fundamental feature of chronic human inflammatory diseases. Specialized subsets of epithelial cells-including secretory and ciliated cells-differentiate from basal stem cells to collectively protect the upper airway. Allergic inflammation can develop from persistent activation of type 2 immunity in the upper airway, resulting in chronic rhinosinusitis, which ranges in severity from rhinitis to severe nasal polyps. Basal cell hyperplasia is a hallmark of severe disease, but it is not known how these progenitor cells contribute to clinical presentation and barrier tissue dysfunction in humans. Here we profile primary human surgical chronic rhinosinusitis samples (18,036 cells, n = 12) that span the disease spectrum using Seq-Well for massively parallel single-cell RNA sequencing, report transcriptomes for human respiratory epithelial, immune and stromal cell types and subsets from a type 2 inflammatory disease, and map key mediators. By comparison with nasal scrapings (18,704 cells, n = 9), we define signatures of core, healthy, inflamed and polyp secretory cells. We reveal marked differences between the epithelial compartments of the non-polyp and polyp cellular ecosystems, identifying and validating a global reduction in cellular diversity of polyps characterized by basal cell hyperplasia, concomitant decreases in glandular cells, and phenotypic shifts in secretory cell antimicrobial expression. We detect an aberrant basal progenitor differentiation trajectory in polyps, and propose cell-intrinsic, epigenetic and extrinsic factors that lock polyp basal cells into this uncommitted state. Finally, we functionally demonstrate that ex vivo cultured basal cells retain intrinsic memory of IL-4/IL-13 exposure, and test the potential for clinical blockade of the IL-4 receptor α-subunit to modify basal and secretory cell states in vivo. Overall, we find that reduced epithelial diversity stemming from functional shifts in basal cells is a key characteristic of type 2 immune-mediated barrier tissue dysfunction. Our results demonstrate that epithelial stem cells may contribute to the persistence of human disease by serving as repositories for allergic memories.
Mast cells are evolutionarily ancient sentinel cells. Like basophils,
mast cells express the high-affinity IgE receptor and are implicated in host
defense and diverse immune-mediated diseases. To better characterize the
function of these cells, we assessed the transcriptional profiles of mast cells
isolated from peripheral connective tissues and basophils isolated from spleen
and blood. We found that mast cells were transcriptionally distinct, clustering
independently from all other profiled cells, and that mast cells demonstrated
considerably greater heterogeneity across tissues than previously appreciated.
We observed minimal homology between mast cells and basophils, which share more
overlap with other circulating granulocytes than with mast cells. Derivation of
mast cell and basophil transcriptional signatures underscores their differential
capacity to detect environmental signals and influence the inflammatory
milieu.
Respiratory epithelial cells (EpCs) orchestrate airway mucosal inflammation in response to diverse environmental stimuli,
but how distinct EpC programs are regulated remains poorly understood. Here, we report that inhalation of aeroallergens leads to
expansion of airway brush cells (BrCs), specialized chemosensory EpCs and the dominant epithelial source of interleukin-25
(IL-25). BrC expansion was attenuated in mice lacking either LTC4 synthase, the biosynthetic enzyme required for
cysteinyl leukotriene (CysLT) generation, or the EpC receptor for leukotriene E4 (LTE4), CysLT3R.
LTE4 inhalation was sufficient to elicit CysLT3R-dependent BrC expansion in the murine airway through an
IL-25–dependent but STAT6-independent signaling pathway. Last, blockade of IL-25 attenuated both aeroallergen and
LTE4-elicited CysLT3R-dependent type 2 lung inflammation. These results demonstrate that
CysLT3R senses the endogenously generated lipid ligand LTE4 and regulates airway BrC number and
function.
Contained within the adult lung are differentiated mesenchymal cell types (cartilage, smooth muscle, and myofibrobasts) that provide structural support for airways and vessels. Alterations in the number and phenotype of these cells figure prominently in the pathogenesis of a variety of lung diseases. While these cells are thought to arise locally, progenitors have yet to be purified. In previous work, we developed a method for isolating progenitors from lung tissue: this technique takes advantage of the unique ability of cell populations enriched for somatic stem and progenitor activity to efflux the vital dye Hoechst 33342, a feature that permits isolation by flow cytometry-based procedures. Using this method, we determined that a rare population of mesenchymal progenitors resides within the CD45- CD31- Hoechst low fraction of the adult murine lung. Similar to other mesenchymal progenitors, these cells express Sca-1, CD106, and CD44; can be serially passaged; and can differentiate to smooth muscle, cartilage, bone, and fat. Overall, these findings demonstrate that a phenotypically distinct mesenchymal progenitor resides within the adult murine lung, and provide a scheme for their isolation and study.
Adiponectin (APN) is an adipocyte-derived factor that exists at high concentrations in serum and has anti-inflammatory and systemic vascular-protective properties. In this study, we investigated the role of APN in pulmonary vascular homeostasis. We found that APN localizes to the luminal side of blood vessels in lung and acts in vitro to block TNF-alpha-induced E-selectin upregulation in pulmonary artery endothelial cells. Targeted deletion of the APN gene in mice leads to a vascular phenotype in lung characterized by E-selectin upregulation and age-dependent increases in perivascular inflammatory cell infiltration and pulmonary arterial pressures. Taken together, these findings demonstrate an important role for APN in lung vascular homeostasis and suggest that APN-deficient states may contribute to the pathogenesis of inflammatory pulmonary vascular disease and to the development of pulmonary hypertension.
CD4
+
effector lymphocytes (Teff) are traditionally classified by the cytokines they produce. To determine the states that Teff actually adopt in frontline tissues
in vivo
, we applied single-cell transcriptome and chromatin analysis on colonic Teff cells, in germ-free or conventional mice, or after challenge with a range of phenotypically biasing microbes. Subsets were marked by expression of interferon-signature or myeloid-specific transcripts, but transcriptome or chromatin structure could not resolve discrete clusters fitting classic T
H
subsets. At baseline or at different times of infection, transcripts encoding cytokines or proteins commonly used as T
H
markers distributed in a polarized continuum, which was also functionally validated. Clones derived from single progenitors gave rise to both IFN-γ and IL17-producing cells. Most transcriptional variance was tied to the infecting agent, independent of the cytokines produced, and chromatin variance primarily reflected activity of AP1 and IRF transcription factor families, not the canonical subset master regulators T-bet, GATA3, RORγ.
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