Biology subjects

Ziegler, C. G. K.

Publications and source records attributed to Ziegler, C. G. K..

7 recordsLinked to original sources

bioRxiv2024

Impact of variants and vaccination on nasal immunity across three waves of SARS-CoV-2

SARS-CoV-2 infection and COVID-19 disease vary with respect to viral variant and host vaccination status. However, how vaccines, emergent variants, and their intersection shift host responses in the human nasal mucosa remains uncharacterized. We and others have shown during the first SARS-CoV-2 wave that a muted nasal epithelial interferon response at the site of infection underlies severe COVID-19. We sought to further understand how upper airway cell subsets and states associate with COVID-19 phenotypes across viral variants and vaccination. Here, we integrated new single-cell RNA-sequencing (scRNA-seq) data from nasopharyngeal swabs collected from 67 adult participants during the Delta and Omicron waves with data from 45 participants collected during the original (Ancestral) wave in our prior study. By characterizing detailed cellular states during infection, we identified changes in epithelial and immune cells that are both unique and shared across variants and vaccination status. By defining SARS-CoV-2 RNA+ cells for each variant, we found that Delta samples had a marked increase in the abundance of viral RNA+ cells. Despite this dramatic increase in viral RNA+ cells in Delta cases, the nasal cellular compositions of Delta and Omicron exhibit greater similarity, driven partly by myeloid subsets, than the Ancestral landscapes associated with specialized epithelial subsets. We found that vaccination prior to infection was surprisingly associated with nasal macrophage recruitment and activation rather than adaptive immune cell signatures. While patients with severe disease caused by Ancestral or Delta variants had muted interferon responses, Omicron-infected patients had equivalent interferon responses regardless of disease severity. Our study defines the evolution of cellular targets and signatures of disease severity in the upper respiratory tract across SARS-CoV-2 variants, and suggests that intramuscular vaccines shape myeloid responses in the nasal mucosa upon SARS-CoV-2 infection.

immunology↗

bioRxiv2022

A single-nucleus and spatial transcriptomic atlas of the COVID-19 liver reveals topological, functional, and regenerative organ disruption in patients

The molecular underpinnings of organ dysfunction in acute COVID-19 and its potential long-term sequelae are under intense investigation. To shed light on these in the context of liver function, we performed single-nucleus RNA-seq and spatial transcriptomic profiling of livers from 17 COVID-19 decedents. We identified hepatocytes positive for SARS-CoV-2 RNA with an expression phenotype resembling infected lung epithelial cells. Integrated analysis and comparisons with healthy controls revealed extensive changes in the cellular composition and expression states in COVID-19 liver, reflecting hepatocellular injury, ductular reaction, pathologic vascular expansion, and fibrogenesis. We also observed Kupffer cell proliferation and erythrocyte progenitors for the first time in a human liver single-cell atlas, resembling similar responses in liver injury in mice and in sepsis, respectively. Despite the absence of a clinical acute liver injury phenotype, endothelial cell composition was dramatically impacted in COVID-19, concomitantly with extensive alterations and profibrogenic activation of reactive cholangiocytes and mesenchymal cells. Our atlas provides novel insights into liver physiology and pathology in COVID-19 and forms a foundational resource for its investigation and understanding.

genomics↗

bioRxiv2021

A single-cell and spatial atlas of autopsy tissues reveals pathology and cellular targets of SARS-CoV-2

The SARS-CoV-2 pandemic has caused over 1 million deaths globally, mostly due to acute lung injury and acute respiratory distress syndrome, or direct complications resulting in multiple-organ failures. Little is known about the host tissue immune and cellular responses associated with COVID-19 infection, symptoms, and lethality. To address this, we collected tissues from 11 organs during the clinical autopsy of 17 individuals who succumbed to COVID-19, resulting in a tissue bank of approximately 420 specimens. We generated comprehensive cellular maps capturing COVID-19 biology related to patients demise through single-cell and single-nucleus RNA-Seq of lung, kidney, liver and heart tissues, and further contextualized our findings through spatial RNA profiling of distinct lung regions. We developed a computational framework that incorporates removal of ambient RNA and automated cell type annotation to facilitate comparison with other healthy and diseased tissue atlases. In the lung, we uncovered significantly altered transcriptional programs within the epithelial, immune, and stromal compartments and cell intrinsic changes in multiple cell types relative to lung tissue from healthy controls. We observed evidence of: alveolar type 2 (AT2) differentiation replacing depleted alveolar type 1 (AT1) lung epithelial cells, as previously seen in fibrosis; a concomitant increase in myofibroblasts reflective of defective tissue repair; and, putative TP63+ intrapulmonary basal-like progenitor (IPBLP) cells, similar to cells identified in H1N1 influenza, that may serve as an emergency cellular reserve for severely damaged alveoli. Together, these findings suggest the activation and failure of multiple avenues for regeneration of the epithelium in these terminal lungs. SARS-CoV-2 RNA reads were enriched in lung mononuclear phagocytic cells and endothelial cells, and these cells expressed distinct host response transcriptional programs. We corroborated the compositional and transcriptional changes in lung tissue through spatial analysis of RNA profiles in situ and distinguished unique tissue host responses between regions with and without viral RNA, and in COVID-19 donor tissues relative to healthy lung. Finally, we analyzed genetic regions implicated in COVID-19 GWAS with transcriptomic data to implicate specific cell types and genes associated with disease severity. Overall, our COVID-19 cell atlas is a foundational dataset to better understand the biological impact of SARS-CoV-2 infection across the human body and empowers the identification of new therapeutic interventions and prevention strategies.

genomics↗

bioRxiv2021

Impaired local intrinsic immunity to SARS-CoV-2 infection in severe COVID-19

Infection with SARS-CoV-2, the virus that causes COVID-19, can lead to severe lower respiratory illness including pneumonia and acute respiratory distress syndrome, which can result in profound morbidity and mortality. However, many infected individuals are either asymptomatic or have isolated upper respiratory symptoms, which suggests that the upper airways represent the initial site of viral infection, and that some individuals are able to largely constrain viral pathology to the nasal and oropharyngeal tissues. Which cell types in the human nasopharynx are the primary targets of SARS-CoV-2 infection, and how infection influences the cellular organization of the respiratory epithelium remains incompletely understood. Here, we present nasopharyngeal samples from a cohort of 35 individuals with COVID-19, representing a wide spectrum of disease states from ambulatory to critically ill, as well as 23 healthy and intubated patients without COVID-19. Using standard nasopharyngeal swabs, we collected viable cells and performed single-cell RNA-sequencing (scRNA-seq), simultaneously profiling both host and viral RNA. We find that following infection with SARS-CoV-2, the upper respiratory epithelium undergoes massive reorganization: secretory cells diversify and expand, and mature epithelial cells are preferentially lost. Further, we observe evidence for deuterosomal cell and immature ciliated cell expansion, potentially representing active repopulation of lost ciliated cells through coupled secretory cell differentiation. Epithelial cells from participants with mild/moderate COVID-19 show extensive induction of genes associated with anti-viral and type I interferon responses. In contrast, cells from participants with severe lower respiratory symptoms appear globally muted in their anti-viral capacity, despite substantially higher local inflammatory myeloid populations and equivalent nasal viral loads. This suggests an essential role for intrinsic, local epithelial immunity in curbing and constraining viral-induced pathology. Using a custom computational pipeline, we characterized cell-associated SARS-CoV-2 RNA and identified rare cells with RNA intermediates strongly suggestive of active replication. Both within and across individuals, we find remarkable diversity and heterogeneity among SARS-CoV-2 RNA+ host cells, including developing/immature and interferon-responsive ciliated cells, KRT13+ "hillock"-like cells, and unique subsets of secretory, goblet, and squamous cells. Finally, SARS-CoV-2 RNA+ cells, as compared to uninfected bystanders, are enriched for genes involved in susceptibility (e.g., CTSL, TMPRSS2) or response (e.g., MX1, IFITM3, EIF2AK2) to infection. Together, this work defines both protective and detrimental host responses to SARS-CoV-2, determines the direct viral targets of infection, and suggests that failed anti-viral epithelial immunity in the nasal mucosa may underlie the progression to severe COVID-19.

immunology↗

bioRxiv2020

Spatially-Resolved Live Cell Tagging and Isolation Using Protected Photoactivatable Cell Dyes

Whether cultured in vitro or part of a complex tissue in vivo, a cells phenotype and function are significantly influenced by dynamic interactions with its microenvironment. To explicitly examine how a cells spatiotemporal activity impacts its behavior, we developed and validated a strategy termed SPACECAT--Spatially PhotoActivatable Color Encoded Cell Address Tags--to annotate, track, and isolate specific cells in a non-destructive, viability-preserving manner. In SPACECAT, a biological sample is immersed in a photocaged fluorescent molecule, and cells within a location of interest are labeled for further study by uncaging that molecule with user-patterned near-UV light. SPACECAT offers high spatial precision and temporal stability across diverse cell and tissue types, and is compatible with common downstream assays, including flow cytometry and single-cell RNA-Seq. Illustratively, we leveraged this approach in patient-derived intestinal organoids, a spatially complex system less amenable to genetic manipulations, to select for crypt-like regions enriched in stem-like and actively mitotic cells. Moreover, we demonstrate its applicability and utility on ex vivo tissue sections from four healthy organs and an autochthonous lung tumor model, uncovering spatially-biased gene expression patterns among immune cell subsets and identifying rare myeloid phenotypes enriched around tumor/healthy border regions. In sum, our method provides a minimally invasive and broadly applicable approach to link cellular spatiotemporal features and/or behavioral phenotypes with diverse downstream assays, enabling fundamental insights into the connections between tissue microenvironments and biological (dys)function.

systems biology↗

bioRxiv2019

Sensory Neurons Innervate Peripheral Lymph Nodes and Locally Regulate Gene Expression in Postsynaptic Endothelium, Stromal Cells, and Innate Leukocytes

Immune responses within barrier tissues are regulated, in part, by nociceptors, specialized peripheral sensory neurons that detect noxious stimuli. Previous work has shown that nociceptor ablation not only alters local responses to immune challenge at peripheral sites, but also within draining lymph nodes (LNs). The mechanisms and significance of nociceptor-dependent modulation of LN function are unknown. Indeed, although sympathetic innervation of LNs is well documented, it has been unclear whether the LN parenchyma itself is innervated by sensory neurons. Here, using a combination of high-resolution imaging, retrograde viral tracing, single-cell transcriptomics (scRNA-seq), and optogenetics, we identified and functionally tested a sensory neuro-immune circuit that is preferentially located in the outermost cortex of skin-draining LNs. Transcriptomic profiling revealed that there are at least four discrete subsets of sensory neurons that innervate LNs with a predominance of peptidergic nociceptors, and an innervation pattern that is distinct from that in the surrounding skin. To uncover potential LN-resident communication partners for LN-innervating sensory neurons, we employed scRNA-seq to generate a draft atlas of all murine LN cells and, based on receptor-ligand expression patterns, nominated candidate target populations among stromal and immune cells. Using selective optogenetic stimulation of LN-innervating sensory axons, we directly experimentally tested our inferred connections. Acute neuronal activation triggered rapid transcriptional changes preferentially within our top-ranked putative interacting partners, principally endothelium and other nodal stroma cells, as well as several innate leukocyte populations. Thus, LNs are monitored by a unique population of sensory neurons that possesses immunomodulatory potential.

immunology↗

bioRxiv2019

Integrated Single-Cell Analysis of Multicellular Immune Dynamics during Hyper-Acute HIV-1 Infection

Cellular immunity is critical for controlling intracellular pathogens, but the dynamics and cooperativity of the evolving host response to infection are not well defined. Here, we apply single-cell RNA-sequencing to longitudinally profile pre- and immediately post-HIV infection peripheral immune responses of multiple cell types in four untreated individuals. Onset of viremia induces a strong transcriptional interferon response integrated across most cell types, with subsequent pro-inflammatory T cell differentiation, monocyte MHC-II upregulation, and cytolytic killing. With longitudinal sampling, we nominate key intra- and extracellular drivers that induce these programs, and assign their multi-cellular targets, temporal ordering, and duration in acute infection. Two individuals studied developed spontaneous viral control, associated with initial elevated frequencies of proliferating cytotoxic cells, inclusive of a previously unappreciated proliferating natural killer (NK) cell subset. Our study presents a unified framework for characterizing immune evolution during a persistent human viral infection at single-cell resolution, and highlights programs that may drive response coordination and influence clinical trajectory.

immunology↗