Biology subjects

Christensen, S. M.

Publications and source records attributed to Christensen, S. M..

8 recordsLinked to original sources

bioRxiv2025

Myeloid cells contribute to bystander CD8 T cell accumulation in metabolic-associated steatohepatitis and are sufficient for fibrosis

Background & AimsMetabolic dysfunction-associated steatohepatitis (MASH) is a leading cause of liver fibrosis, morbidity, and mortality. It is characterized by the accumulation of fat in hepatocytes causing cell death followed by stromal cell activation and scar deposition at later disease stages. While a marked accumulation of cytotoxic CD8 T cells is observed in MASH in humans and mice, the role of adaptive immune cells in fibrosis progression remains debated. MethodsTranscriptional data were curated from human datasets and preclinical mouse models, including mice deficient in TCR{beta} T cells (Tcrb-/-), which lack conventional CD4 helper and CD8 cytotoxic T cells, in diet-induced fibrosing MASH for single-cell RNA-sequencing. Specific human and mouse liver transcriptomic immune signatures were corroborated by flow cytometry and immunofluorescence. ResultsxMASH was associated with the expansion of lymphocytes and myeloid cells. Auto-aggressive CXCR6+PD1+FASLG+ CD8 T cells were enriched in livers of high-fat-diet-fed mice and correlated with MASH and fibrosis. Notably, Tcrb-deficient mice in a chemical and dietary preclinical MASH model developed fibrosis to the same extent as wild-type mice and exhibited exacerbated pro-fibrotic type 3 inflammation. Loss of conventional CD8 T cells neither impacted myeloid cell number nor phenotype within the fibrotic niche. Neutrophil- and non-conventional lymphocyte-derived GM-CSF and IL-17A were central drivers in the fibrotic niche composed of pathogenic macrophages and activated myofibroblasts. Furthermore, myeloid cells were identified as the main source of CXCL16 in diseased livers, retaining auto-aggressive CD8 T cells to the scar and contributing to their accumulation. Thus, non-adaptive cells, including scar-associated macrophages (SAMs) and myofibroblasts, are sufficient for MASH-driven fibrosis in this model. ConclusionsTargeting myeloid cells and fibroblasts should be prioritized as anti-fibrotic therapies for MASH. Graphical abstractThe immunopathological niche in fibrosis O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=99 SRC="FIGDIR/small/694415v3_ufig1.gif" ALT="Figure 1000"> View larger version (47K): org.highwire.dtl.DTLVardef@122a24org.highwire.dtl.DTLVardef@8ca28borg.highwire.dtl.DTLVardef@103907borg.highwire.dtl.DTLVardef@10280e3_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsAdaptive and innate immune cells localize to the human MASH fibrotic niche CD8 T cells increase with MASH severity while MHC-II+ cells accumulate with fibrosis CXCR6+FASLG+ CD8 auto-aggressive T cells accumulate at the scar CXCL16+ scar-associated macrophages retain CD8 T cells in the fibrotic MASH niche Liver injury, inflammation and fibrosis persist in the absence of conventional T cells Impact and implicationsMetabolic dysfunction-associated steatohepatitis (MASH) is a progressive liver disease in which fibrosis is a key determinant of patient outcomes and survival. This study demonstrates that fibrosis in MASH can develop independent of conventional T cell immunity, driven by myeloid cells including scar-associated macrophages (SAMs) and neutrophils within a localized hepatic niche in mice and humans. This highlights the need to prioritize innate immune mechanisms as key drivers of chronic (late-stage) liver disease for researchers and clinicians developing anti-fibrotic therapies.

immunology↗

bioRxiv2025

A revised model of nuclear actin import: Importin 9 competes with cofilin, profilin, and RanGTP for actin binding

While predominantly studied in cytoplasmic contexts, actin plays critical roles in the nucleus, regulating chromatin accessibility and remodeling, transcription, and DNA damage repair. Cell- based studies have contributed to a widely accepted model in which the import factor Importin 9 (IPO9) acts in concert with the actin filament-severing protein cofilin to transport actin into the nucleus. The classical nuclear localization signal on cofilin is thought to anchor IPO9 to cofilin- bound actin monomers, driving the formation of an import-competent tripartite actin*cofilin*IPO9 complex. In striking contradiction to this established model of actin import, we demonstrate that IPO9 directly binds to monomeric actin with mid-nanomolar affinity and, rather than promoting IPO9*actin complex formation, cofilin competitively inhibits binding of IPO9 to actin. We further report competitive binding for monomeric actin between IPO9 and the canonical actin monomer- sequestering protein, profilin. As cofilin and profilin are both capable of binding actin monomers at the barbed face, our results are consistent with a model in which IPO9 binds an overlapping portion of this interface. In further support, we demonstrate that IPO9 modestly occludes the barbed face of actin monomers, decreasing the rate of filament formation, and exhibits minimal filamentous actin binding. Finally, we identify unexpected affinity between the nuclear import release factor RanGTP and monomeric actin; however, a tripartite IPO9*actin*RanGTP complex does not form. The competitive interactions observed between IPO9 and cytoplasmic actin- binding proteins suggest dynamically coupled equilibria mediate the nuclear transport of actin monomers.

biochemistry↗

bioRxiv2025

BMP9 regulates the endothelial secretome to drive pulmonary hypertension

BMP9, a pleiotropic growth factor cytokine that regulates endothelial function, is implicated in the pathogenesis of pulmonary arterial hypertension (PAH). Loss-of-function mutations in GDF2 are found in heritable PAH, suggesting its function as an endothelial quiescence factor, while agonizing or antagonizing BMP9 signaling are both reported to ameliorate experimental pulmonary hypertension (PH). This study sought to resolve the contribution of BMP9 to pulmonary vascular disease and its status as a potential therapeutic target. The function of BMP9 in experimental PH was interrogated using recombinant BMP9, BMP9/BMP10 ligand trap ALK1-Fc, two anti-BMP9 neutralizing antibodies, and the activin/GDF/BMP ligand trap ACTRIIA-Fc (a.k.a., sotatercept). Disulfide-linked, prodomain complexed BMP9 was not protective in SUGEN-hypoxia or monocrotaline-induced PH models, in contrast to previous studies using incompletely disulfide-linked BMP9. In comparison, selective and non-selective BMP9 antagonism exerted prophylactic and therapeutic effects across PH models. Anti-BMP9 and ACTRIIA-Fc had comparable impact on hemodynamics, RV hypertrophy, and vascular remodeling, while single nucleus RNA-Seq revealed similar inhibition of SMAD1/5 and SMAD2/3 transcriptional activity, and highly overlapping DEGs, particularly in the endothelial compartment (r=0.83, p=2.54e-43, Spearman), suggesting overlap of mechanism in targeting BMP9. A multi-omic approach using lung tissues from human PAH, experimental models of pulmonary hypertension, and transcriptomic analysis of pulmonary microvascular endothelial cells from PAH patients revealed that BMP9 is critical for regulating several endothelial gene products that are overexpressed in human and experimental disease and implicated in disease pathogenesis including CXCL12, PDGF-BB, EDN1, COL18A1, and IGFBP4, and are inhibited by administering anti-BMP9 neutralizing antibodies or ligand traps. Co-culture studies revealed paracrine effects of BMP9-stimulated PMVEC on pulmonary arterial smooth muscle cell (PASMC) phenotypic plasticity, which could be attributed in large part to endothelial-derived CXCL12. In summary, endothelial BMP9 signaling is a key coordinator of vasoactive endothelial gene products that modulate PASMC phenotype and appears to be a shared target of anti-BMP9 and ACTRIIA-Fc. Selective targeting of endothelial BMP9 angiogenic signaling represents a potential therapeutic strategy for human PAH. One sentence summaryInhbiition of BMP9 signaling via different selective and non-selective strategies, including ACTRIIA-Fc/sotatercept, attenuates endothelial expression of vasoactive secreted factors that drive pulmonary vascular remodeling.

physiology↗

bioRxiv2024

Direct observation of assembly and function of trigger responsive lipase biohybrids

Protein-polymer biohybrids mark a cutting edge in the creation of advanced materials designed for high performance under diverse conditions by combining the intrinsic stability and adaptability of synthetic polymers with the remarkable functionality and specificity of proteins. Despite the existence of stimuli responsive polymers, their potential for achieving precise, trigger responsive control over both the function and the assembly of protein-polymer biohybrids has yet to be fully attained. Here, we report the synthesis and thorough characterization of pH and temperature responsive lipase-polymer conjugates at both the supramolecular and the single molecule level. In contrast to conventional ensemble measurements performed in solution, we extensively characterized the stimuli triggered reversibility of the self-assembly of these lipase-based biohybrids and recorded the enzymatic activity at the fundamental level of individual nanoparticles. Our findings unexpectedly demonstrated that the stimuli responsive biohybrids exhibited not only a twofold increased enzymatic activity as compared to native lipases in certain cases but also maintained the trigger responsive control over enzymatic activity by temperature. Direct single particle recordings of biohybrid activity showed a correlation of the biohybrid size to catalytic activity, revealing individual enzymes to display higher activity in smaller nanoparticles. The biohybrids exhibit impressive long-term stability, in some cases almost 95% of the enzymatic activity as compared to [~]40-60% for native lipases after 1 year of storage. The improved stability and activity as compared to native enzymes accompanied with their triggered responsiveness highlight their significant potential for tightly controlled biocatalysis and sustainable industrial applications.

biophysics↗

bioRxiv2023

Microbial Metamorphosis: Symbiotic bacteria and fungi proliferate during diapause and may enhance overwintering survival in a solitary bee

Host-microbe interactions underlie the development and fitness of many macroorganisms including bees. While many social bees benefit from vertically transmitted gut bacteria, solitary bees, which comprise the vast majority of species diversity within bees, lack a specialized gut microbiome. Here we examine the composition and abundance of bacteria and fungi throughout the complete life cycle of a ground-nesting solitary bee Anthophora bomboides standfordiana. In contrast to expectations, immature bee stages maintain a distinct core microbiome consisting of Actinobacteria and fungi in the genus Moniliella. Diapausing larval bees hosted the most abundant and distinctive bacteria and fungi, attaining 33 and 52 times their initial copy number, respectively. We tested two adaptive hypotheses regarding microbial functions for overwintering bees. First, using isolated bacteria and fungi, we found that Streptomyces from brood cells inhibited the growth of multiple pathogenic filamentous fungi, suggesting a role in pathogen protection during the long period of diapause. Second, sugar alcohol composition changed in tandem with major changes in microbial abundance suggesting links with bee metabolism or overwintering biology. Our results suggest that Anthophora bomboides hosts a conserved core microbiome that may provide key fitness advantages through larval development and overwintering, and raises the question of how this microbiome is transmitted between generations. The present work suggests that focus on adult gut microbiomes in solitary bees may overlook microbial symbionts within brood cells that could play diverse roles in bee fitness, and that exploration of microbes associated with immature bees may uncover novel microbial effects on insect hosts.

ecology↗

bioRxiv2023

Stricturing Crohn's disease single-cell RNA sequencing reveals fibroblast heterogeneity and intercellular interactions

BackgroundFibroblasts play a key role in stricture formation in Crohns disease (CD) but understanding its pathogenesis requires a systems-level investigation to uncover new treatment targets. We studied full thickness CD tissues to characterize fibroblast heterogeneity and function by generating the first single cell RNA sequencing (scRNAseq) atlas of strictured bowel and providing proof of principle for therapeutic target validation. MethodsWe performed scRNAseq of 13 fresh full thickness CD resections containing non-involved, inflamed non-strictured, and strictured segments as well as 7 normal non-CD bowel segments. Each segment was separated into mucosa/submucosa or muscularis propria and analyzed separately for a total of 99 tissue samples and 409,001 cells. We validated cadherin-11 (CDH11) as a potential therapeutic target by using whole tissues, isolated intestinal cells, NanoString nCounter, next generation sequencing, proteomics and animal models. ResultsOur integrated dataset revealed fibroblast heterogeneity in strictured CD with the majority of stricture-selective changes detected in the mucosa/submucosa, but not the muscle layer. Cell-cell interaction modeling revealed CXCL14+ as well as MMP/WNT5A+ fibroblasts displaying a central signaling role in CD strictures. CDH11, a fibroblast cell-cell adhesion molecule, was broadly expressed and upregulated, and its pro-fibrotic function was validated by NanoString nCounter, RNA sequencing, tissue target expression, in vitro gain- and loss-of-function experiments, proteomics, and two animal models of experimental colitis. ConclusionA full-thickness bowel scRNAseq atlas revealed previously unrecognized fibroblast heterogeneity and interactions in CD strictures and CDH11 was validated as a potential therapeutic target. These results provide a new resource for a better understanding of CD stricture formation and opens potential therapeutic developments.

immunology↗

bioRxiv2022

Identification of a Broadly Fibrogenic Macrophage Subset Induced by Type 3 Inflammation in Human and Murine Liver and Lung Fibrosis

Macrophages are central orchestrators of the tissue response to injury, with distinct macrophage activation states playing key roles in the progression and resolution of fibrosis. Identifying the unique fibrogenic macrophages that are found in human fibrotic tissues could lead to new and more effective treatments for fibrosis. Here we used human liver and lung single cell RNA sequencing datasets to identify a unique subset of CD9+ TREM2+ macrophages expressing SPP1, GPNMB, FABP5, and CD63 with strong pro-fibrotic activity. This population was validated across orthogonal techniques, species and tissues. These macrophages were enriched at the outside edges of scarring adjacent to activated mesenchymal cells, and in the fibrotic niche across species and organs. Neutrophils producing the type 3 cytokines GM-CSF and IL-17A, and expressing MMP9, which participates in the activation of TGF-{beta}1, clustered with these scar-associated macrophages. Using in vitro primary human cell assays, we determined that GM-CSF, IL-17A and TGF-{beta}1 drive the differentiation of these scar-associated macrophages, and that co-culture of monocyte-derived macrophages with hepatic stellate cells and TGF-{beta}1 augmented type 1 collagen deposition. In vivo blockade of GM-CSF, IL-17A or TGF-{beta}1 with small or large molecules reduced scar-associated macrophage expansion and fibrosis in multiple models of hepatic and pulmonary fibrosis. Our work demonstrates that a specific scar-associated macrophage population is linked with fibrosis across species and tissues. It further provides a strategy for unbiased discovery, triage and preclinical validation of therapeutic targets within this fibrogenic macrophage population.

immunology↗

bioRxiv2021

Nectar bacteria stimulate pollen germination and bursting to enhance their fitness

For many flower visitors, pollen is the primary source of non-carbon nutrition, but pollen has physical defenses that make it difficult for consumers to access nutrients. Nectar-dwelling microbes are nearly ubiquitous among flowers and can reach high densities, despite the fact that floral nectar is nitrogen limited, containing only very low concentrations of non-carbon nutrients. Pollen contains trace micronutrients and high protein content but is protected by a recalcitrant outer shell. Here, we report that a common genus of nectar-dwelling bacteria, Acinetobacter, exploits pollen nutrition by inducing pollen germination and bursting. We use time course germination assays to quantify the effect of Acinetobacter species on pollen germination and pollen bursting. Inoculation with Acinetobacter species resulted in increased germination rates within 15 minutes, and bursting by 45 minutes, as compared to uninoculated pollen. The pollen germination and bursting phenotype is density-dependent, with lower concentrations of A. pollinis SCC477 resulting in a longer lag time before the spike in germination, which is then closely followed by a spike in bursting. Lastly, A. pollinis grows to nearly twice the density with germinable pollen vs ungerminable pollen, indicating that their ability to induce and exploit germination plays an important role in rapid growth. To our knowledge, this is the first direct test of non-plant biological induction of pollen germination, as well as the first evidence of induced germination as a method of nutrient procurement, as the microbes appear to hijack the pollens normally tightly controlled germination mechanisms for their benefit. Our results suggest that further study of microbe-pollen interactions may inform many aspects of pollination ecology, including microbial ecology in flowers, the mechanisms of pollinator nutrient acquisition from pollen, and cues of pollen germination for plant reproduction.

ecology↗