Biology subjects

Floyd, D.

Publications and source records attributed to Floyd, D..

5 recordsLinked to original sources

bioRxiv2026

Neutrophil remodeling is associated with human meibomian gland dysfunction and enables IFN-γ- and PAD4-dependent gland obstruction in mice

Meibomian gland dysfunction (MGD), a disorder of the eyelid's modified sebaceous glands, is the leading cause of dry eye disease and ocular surface morbidity, yet the immune mechanisms driving gland obstruction remain poorly defined. In a cross-sectional study of 66 patients with ocular surface inflammation, we used meibography and spectral flow cytometry of tear washes to identify a disease-associated, remodeled neutrophil state whose abundance is associated with gland atrophy. Using single-cell transcriptomics in a murine model of immune-mediated MGD, we revealed a disease-associated neutrophil state that exhibited ocular surface-enrichment, CD14 and ICAM-1 expression, and elevated IFN-{gamma} response and inflammatory signatures. Spatial transcriptomics localized IFN-{gamma} signaling and neutrophil migration signatures to the periglandular compartment. The remodeled neutrophils exhibited PAD4-dependent histone citrullination, with Padi4 deletion reducing NET-associated obstructive plugging, thus identifying PAD4-dependent NETotic activity as their disease-producing output. Inhibition of IFN-{gamma} signaling phenocopied Padi4 deficiency, yet combined disruption of these pathways provided no additive protection, indicating that IFN-{gamma} and PAD4 function as separable required inputs. Remodeled neutrophils accumulated under both conditions, uncoupling disease severity from cell abundance alone. Our findings support immune-mediated obstructive MGD as a mechanistic endotype driven by the IFN-{gamma}- and PAD4-dependent effector output of a remodeled neutrophil state.

immunology↗

bioRxiv2025

Regulation of multiple paralogs of a small subunit ribosomal protein in Francisella tularensis

Francisella tularensis is a highly infectious human pathogen that must replicate inside macrophage to cause disease. The ribosomes of F. tularensis can incorporate one of three different paralogs for the small ribosomal subunit protein bS21. One of these paralogs positively impacts translation of key virulence genes and promotes intramacrophage replication. Although ribosomal bS21 content influences F. tularensis virulence, the factors that control bS21 paralog production are not well understood. Here, we reveal that all three bS21 proteins influence the transcript abundance of the paralog important for virulence, bS21-2. In contrast, the other bS21 paralogs (bS21-1 and bS21-3) do not affect their own production. We further determined that the leader sequence of the bS21-2 mRNA is sufficient for bS21-mediated repression of mRNA abundance, suggesting that bS21-2 is autogenously regulated. Yet we determined that the increase in bS21-2-encoding mRNA is not reflected by increased protein production, suggesting that translation of this transcript is controlled by other factors. Finally, we found that bS21-2 exerts at least some of its effects on the bS21-2 transcript by decreasing its stability. Together, our findings suggest that F. tularensis integrates multiple signals into a regulatory network to control the appropriate production of each bS21 paralog, and particularly the paralog important for virulence, bS21-2. This regulatory network in turn may control ribosomal heterogeneity and virulence gene expression.

microbiology↗

bioRxiv2025

Extracellular vesicles from diverse fungal pathogens induce species-specific and endocytosis-dependent immunomodulation

Microbial pathogens generate extracellular vesicles (EVs) for intercellular communication and quorum sensing. Microbial EVs also induce inflammatory pathways within host innate immune cells. We previously demonstrated that EVs secreted by Candida albicans trigger type I interferon signaling in host cells specifically via the cGAS-STING innate immune signaling pathway. Here, we show that despite sharing similar properties of morphology and internal DNA content, the interactions between EVs and the innate immune system differ according to the parental fungal species. EVs secreted by C. albicans, Saccharomyces cerevisiae, Cryptococcus neoformans, and Aspergillus fumigatus are endocytosed at different rates by murine macrophages triggering varied cytokine responses, innate immune signaling, and subsequent immune cell recruitment. Notably, cell wall constituents that decorate C. neoformans and A. fumigatus EVs inhibit efficient internalization by macrophages and dampen innate immune activation. Our data uncover the transcriptional and functional consequences of the internalization of diverse fungal EVs by immune cells and reveal novel insights into the early innate immune response to distinct clinically significant fungal pathogens.

microbiology↗

bioRxiv2023

AKI and CKD Worsen Outcomes in Experimental Sepsis: Creating a Reverse-translational Model of Acute Peritoneal Infection and Recovery in an Immunocompromised Host.

IntroductionInfection is a leading cause of morbidity and mortality in individuals with kidney disease. Both acute kidney injury (AKI), and chronic kidney disease (CKD) are clinical states that have been associated with higher risk of incident infection, and poor outcomes once infection has been established. A variety of host-and pathogen-specific factors are implicated as potential causes for these disparate outcomes including an altered host microbiome, innate and adaptive immune defects, and poor renal clearance and cytokines. However, there remains significant difficulty in modeling both human kidney disease and infection into an animal host. Likewise, there remains a poor understanding of the mechanisms underlying the unique immunodeficiency imparted by AKI and CKD, and if either condition imparts disparate risk. MethodsC57BL/6J mice were given vehicle or aristolochic acid (AA) to create AKI (control, AKI groups) or CKD (control, CKD groups). Donor mice from all four groups underwent sterile cecal dissection and creation of cecal slurry (CS) preparations, which was later injected into separate mice in a matched host-recipient manner, at either high or lower doses. Animals were clinically monitored for either 24- or 72-hours after inoculation, then euthanized. Animal survival, sepsis severity, temperature, weights, and transcutaneous glomerular filtration rate (tGFR) were tracked longitudinally throughout the study. Histology for kidney injury, peripheral blood flow cytometry for leukocyte counts, plasma cytokines, and typical markers for organ injury were determined. ResultsCompared to controls, animals with AKI experienced much more severe sepsis across virtually all tracked metrics, and no animals with AKI survived high-dose CS injection past 24-hours. AKI mice manifested with a peripheral defect in leukocytes early after sepsis, with severe and persistent cytopenias, and a dramatically heightened early pro-inflammatory cytokine response. Septic CKD mice also had worse outcomes than controls, though less severe, and occurring later than in animals with AKI. Interestingly, animals with AKI had worse clinical outcomes and evidence of organ injury than mice with CKD at any dose or time-point after inoculation, despite a higher mean baseline measured GFR. ConclusionsRodents with established AKI and CKD experience worse clinical outcomes and organ injury versus controls in a CS model intraperitoneal live-bacterial infection. Additionally, mice with AKI experienced earlier and more severe morbidity and mortality than animals with CKD.

immunology↗

bioRxiv2022

Predestined neutrophil heterogeneity in homeostasis varies in transcriptional and phenotypic response to Candida

Once perceived to be homogenous effector cells, neutrophils have since been shown to exhibit population heterogeneity. Here, we established an experimental model of clonal neutrophil heterogeneity using conditionally immortalized clonal granulocyte monocyte progenitors (GMPs) and their mature neutrophil progeny. Transcriptional and epigenetic profiling showed conserved genome-wide signatures of transcription and chromatin accessibility that were specific to individual GMP clones and their paired neutrophil progeny, suggesting that clone specificity is established as early as the GMP stage. Clone-specific genes in vital regulatory pathways were pre-programmed and exhibited delayed expression in the mature neutrophil stage. The clone-specific gene expression in the mature neutrophils paired to enhancer activation in their parental GMPs. To determine whether transcriptional heterogeneity predicted the response to fungal pathogens, neutrophil clones were functionally profiled. Clones demonstrated heterogeneous responses to fungal pathogens in vitro and revealed neutrophil subsets with evidence for tailored functional responses to Candida spp. as well as specific transcriptional and epigenetic patterns that may explain these differences. Together, this work establishes that heterogenous GMP and neutrophil compartments exist under homeostatic conditions and that these represent predefined clusters that are uniquely adapted to control invasive fungal pathogens. Short SummaryClonal neutrophil progenitors demonstrate heterogeneity in transcription and chromatin accessibility which may inform response to later fungal challenges.

immunology↗