Biology subjects

Hoover, P.

Publications and source records attributed to Hoover, P..

8 recordsLinked to original sources

bioRxiv2026

Spatially Distinct Macrophage Subsets Drive Myofibroblast Heterogeneity and Maladaptive Fibrosis in Lupus Nephritis

ObjectivesLupus nephritis (LN) is a severe complication of systemic lupus erythematosus (SLE), leading to progressive renal fibrosis and functional decline. Understanding the interplay between immune cells and stromal cells is needed to develop effective therapeutic strategies. Here, we investigated the landscape of macrophage-fibroblast interactions in human LN and validated these findings in mouse models. MethodsWe characterized distinct fibroblast subsets and their interactions with renal macrophages using single-cell RNA sequencing (scRNAseq) of 156 human LN biopsies and 30 healthy controls from the AMP-SLE cohort, and spatial transcriptomics of biopsies from 6 LN patients. In vitro co-culture studies using mouse models were performed to further define functional consequences of these interactions. ResultsWe identified two myofibroblast subsets: a pro-inflammatory subset (Myofib1) enriched in the tubulointerstitium, and a fibrotic/remodeling subset (Myofib2) in glomeruli, both correlating with the histologic chronicity index. Spatial transcriptomics revealed different colocalization patterns, with Myofib1 interacting with activated resident macrophage (RM) subsets and Myofib2 with glomerular infiltrating disease-associated macrophages. In vitro co-culture studies demonstrated that nephritic RMs promote a pro-inflammatory, remodeling fibroblast phenotype that impairs wound healing and drives a Myofib1-like gene program, whereas disease-associated macrophages generated profibrotic fibroblasts with dysregulated reparative capacity. Cell-cell communication analyses identified key ligand-receptor interactions mediating this crosstalk, including Spp1/integrins, Sema4/PlexinB, and NAMPT/INSR. ConclusionsOur data reveal a spatially and functionally heterogeneous landscape of macrophage-fibroblast crosstalk in LN. These findings advance our understanding of renal fibrogenesis in LN, highlighting specific fibro-inflammatory circuits that may represent therapeutic targets to prevent chronic renal damage.

immunology↗

bioRxiv2026

Early developmental placental defects and pregnancy loss in lupus mice expressing human TLR8

Anti-phospholipid (APL) autoantibodies confer a high risk for adverse pregnancy outcomes, especially in Systemic Lupus Erythematosus (SLE). While human TLR8 (huTLR8) has been implicated in APL antibody-mediated placental injury in vitro, its in vivo role in pregnancy is unexplored. We report a novel mouse model of pregnancy loss in SLE-prone mice expressing huTLR8. Placental analysis revealed early developmental defects starting post-implantation, including a thin junctional zone, impaired vascularization, infarcts, and inflammation. Profound immune dysregulation was evident at E8.5 including increased myeloid cells and CD8 T cells and decreased uterine natural killer (uNK) cells. RNA sequencing revealed downregulated pregnancy-specific glycoproteins, reduced uNK cell-associated genes, and an upregulated myeloid cell signature. Bone marrow chimera studies demonstrated preferential activation of huTLR8-expressing placental Ly6C+ monocytes. Spatial transcriptomics at E9.5 confirmed uNK cell loss, decreased IL15 expression by both stromal and myeloid cells, and discrete inflammatory aggregates in the maternal layers containing myeloid cells and IFN{gamma}-expressing CD8 T cells. We propose that huTLR8, likely through myeloid cell activation and cytolytic T cell recruitment, drives placental injury in the context of SLE and APL autoantibodies. This model provides a valuable platform to dissect early pathogenic events in APL-associated pregnancy loss and identify new therapeutic targets.

immunology↗

bioRxiv2023

Intrarenal myeloid subsets associated with kidney injury are comparable in mice and patients with lupus nephritis

Resident macrophages and infiltrating monocytes in kidneys of patients with lupus nephritis are altered both in frequency and function relative to their counterparts in healthy kidneys. The extent to which mouse models might be useful in developing approaches to target these cells for treating lupus nephritis is poorly understood. Here, we studied four common lupus mouse models that share clinical, serologic, and histopathologic kidney changes with humans. Using single-cell profiling and multiplex spatial imaging to analyze the intrarenal myeloid compartment with the onset of clinical disease in these models, we identified monocyte and macrophage subsets that expand or contract in kidneys with clinical nephritis. A unique subset of classical monocytes expanded with the onset of disease and expressed genes such as CD9, Spp1, Ctsd, Cd63, Apoe, and Trem2 that were previously shown to be induced by tissue injury and play a role in inflammation, lipid metabolism and tissue repair in other organs. Resident macrophages transitioned from a pro-inflammatory to a similar injury-associated state with onset of disease. To test whether these findings in mouse models were also observed in humans, we re-analyzed monocytes and macrophages in a single-cell RNAseq dataset of kidney biopsies from 155 patients with lupus nephritis and 30 healthy donors, collected by the NIH AMP RA/SLE consortium. Human monocytes and macrophages showed conserved changes in gene expression programs associated with lupus nephritis disease indices, and localized to similar kidney microenvironments as in mice. By identifying myeloid subsets and disease-associated alterations in biological processes that are conserved across species, we provide a strong rationale for functional studies of these cells and pathways in mice to uncover mechanisms and find targets relevant to human lupus nephritis. One sentence summaryThis study characterizes intrarenal myeloid cells from four lupus mouse models and 155 patients with lupus nephritis using single-cell RNA-seq and imaging, and identifies novel infiltrating and resident myeloid subsets that are conserved between mouse and human lupus nephritis, thus providing a map and strong rationale for functional studies in mice with relevance to human disease.

immunology↗

bioRxiv2021

An atlas of healthy and injured cell states and niches in the human kidney

Understanding kidney disease relies upon defining the complexity of cell types and states, their associated molecular profiles, and interactions within tissue neighborhoods. We have applied multiple single-cell or -nucleus assays (>400,000 nuclei/cells) and spatial imaging technologies to a broad spectrum of healthy reference (n = 42) and disease (n = 42) kidneys. This has provided a high resolution cellular atlas of 100 cell types that include rare and novel cell populations. The multi-omic approach provides detailed transcriptomic profiles, epigenomic regulatory factors, and spatial localizations for major cell types spanning the entire kidney. We further identify and define cellular states altered in kidney injury, encompassing cycling, adaptive or maladaptive repair, transitioning and degenerative states affecting several segments. Molecular signatures of these states permitted their localization within injury neighborhoods using spatial transcriptomics, and large-scale 3D imaging analysis of [~]1.2 million neighborhoods provided linkages to active immune responses. These analyses further defined biological pathways relevant to injury niches, including signatures underlying the transition from reference to predicted maladaptive states that were associated with a decline in kidney function during chronic kidney disease. This human kidney cell atlas, including injury cell states and neighborhoods, will be a valuable resource for future studies.

genomics↗

bioRxiv2021

Mn2+ coordinates Cap-0-RNA to align substrates for efficient 2'-O-methyl transfer by SARS-CoV-2 nsp16

Capping viral messenger RNAs is essential for efficient translation and prevents their detection by host innate immune responses. For SARS-CoV-2, RNA capping includes 2'-O-methylation of the first ribonucleotide by methyltransferase nsp16 in complex with activator nsp10. The reaction requires substrates, a short RNA and SAM, and is catalyzed by divalent cations, with preference for Mn2+. Crystal structures of nsp16-nsp10 with capped RNAs revealed a critical role of metal ions in stabilizing interactions between ribonucleotides and nsp16, resulting in precise alignment of the substrates for methyl transfer. An aspartate residue that is highly conserved among coronaviruses alters the backbone conformation of the capped RNA in the binding groove. This aspartate is absent in mammalian methyltransferases and is a promising site for designing coronavirus-specific inhibitors.

microbiology↗

bioRxiv2021

Topoisomerase 2b induces DNA breaks to regulate human papillomavirus replication

Topoisomerases regulate higher order chromatin structures through the transient breaking and re-ligating of one or both strands of the phosphodiester backbone of duplex DNA. TOP2{beta} is a type II topoisomerase that induces double strand DNA breaks at topological-associated domains (TADS) to relieve torsional stress arising during transcription or replication. TADS are anchored by CTCF and SMC1 cohesin proteins in complexes with TOP2{beta}. Upon DNA cleavage a covalent intermediate DNA-TOP2{beta} (TOP2{beta}cc) is transiently generated to allow for strand passage. The tyrosyl-DNA phosphodiesterase TDP2 can resolve TOP2{beta}cc but failure to do so quickly can lead to long-lasting DNA breaks. Given the role of CTCF/SMC1 proteins in the HPV life cycle we investigated if TOP2{beta} proteins contribute to HPV pathogenesis. Our studies demonstrated that levels of both TOP2{beta} and TDP2 were substantially increased in cells with high risk HPV genomes and this correlated with high amounts of DNA breaks. Knockdown of TOP2{beta} with shRNAs reduced DNA breaks by over 50% as determined through COMET assays. Furthermore this correlated with substantially reduced formation of repair foci such as {gamma}H2AX, pCHK1 and pSMC1 indicative of impaired activation of DNA damage repair pathways. Importantly, knockdown of TOP2{beta} also blocked HPV genome replication. Our previous studies demonstrated that CTCF /SMC1 factors associate with HPV genomes at sites in the late regions of HPV31 and these correspond to regions that also bind TOP2{beta}. This study identifies TOP2{beta} as responsible for enhanced levels of DNA breaks in HPV positive cells and as a regulator of viral replication.

microbiology↗

bioRxiv2019

A Multimodal and Integrated Approach to Interrogate Human Kidney Biopsies with Rigor and Reproducibility: The Kidney Precision Medicine Project

Comprehensive and spatially mapped molecular atlases of organs at a cellular level are a critical resource to gain insights into pathogenic mechanisms and personalized therapies for diseases. The Kidney Precision Medicine Project (KPMP) is an endeavor to generate 3-dimensional (3D) molecular atlases of healthy and diseased kidney biopsies using multiple state-of-the-art OMICS and imaging technologies across several institutions. Obtaining rigorous and reproducible results from disparate methods and at different sites to interrogate biomolecules at a single cell level or in 3D space is a significant challenge that can be a futile exercise if not well controlled. We describe a "follow the tissue" pipeline for generating a reliable and authentic single cell/region 3D molecular atlas of human adult kidney. Our approach emphasizes quality assurance, quality control, validation and harmonization across different OMICS and imaging technologies from sample procurement, processing, storage, shipping to data generation, analysis and sharing. We established benchmarks for quality control, rigor, reproducibility and feasibility across multiple technologies through a pilot experiment using common source tissue that was processed and analyzed at different institutions and different technologies. A peer review system was established to critically review quality control measures and the reproducibility of data generated by each technology before being approved to interrogate clinical biopsy specimens. The process established economizes the use of valuable biopsy tissue for multi-OMICS and imaging analysis with stringent quality control to ensure rigor and reproducibility of results and serves as a model for precision medicine projects across laboratories, institutions and consortia.

scientific communication and education↗

bioRxiv2018

Multiplexed enrichment and genomic profiling of peripheral immune cell subsets on a microfluidic chip

The human immune system consists of many specialized cell subsets that simultaneously carry out a diverse range of functions using overlapping pathways and signals. Subset-specific immune profiling can resolve immune activity in autoimmune disease, cancer immunity, and infectious disease that may not be discoverable or detectable in analyses of crude blood samples. The activity of specific subsets may help predict the course of disease and response to therapy in certain patient populations. Here, we present a low-input microfluidic system for sorting immune cells into subsets and profiling their cellular states by gene expression analysis using full-length RNA-seq. Our system is robust and has the potential to make multiplexed subset-specific analysis routine in many research laboratories and clinical settings. We validate the devices technical performance by benchmarking its subset enrichment and genomic profiling performance against standard protocols. We make the added value of subset-resolved profiling over crude samples clear through ex vivo experiments that show subset-specific stimulated responses. Finally, we demonstrate the scalability of our device by profiling four immune cell subsets in blood from systemic lupus erythematosus (SLE) patients and matched controls enrolled in a clinical study. The results from our initial cohort confirm the role of type I interferons in lupus pathogenesis and further show that the canonical interferon signature for SLE is prominent in B cells, demonstrating the ability of our integrated analytical platform to identify cell-specific disease signatures.

bioengineering↗