Biology subjects

Drnevich, J.

Publications and source records attributed to Drnevich, J..

11 recordsLinked to original sources

bioRxiv2026

Paradoxical Th1 activation and CTLA-4 regulation is beneficial during latent cryptococcosis

Cryptococcus neoformans is the predominant causative agent of cryptococcal meningitis in immunocompromised individuals. Conversely in immunocompetent individuals, C. neoformans establishes a latent pulmonary infection characterized by a paucity of clinical symptoms. Using a mouse inhalation model of latent C. neoformans infection, we previously showed that CD4 T-cells are necessary for preventing fungal proliferation in the lungs. In the current study, we performed single cell RNA sequencing (scRNAseq) and found that the CD4 T-cell response was both highly heterogenous and dichotomous during pulmonary C. neoformans infection, with concomitant expression of genes related to Th1 polarization (Tbx21, Ifng) and immune regulation (Ctla4). First, we demonstrated that cells with Th1-like phenotypes are necessary and sufficient to control latent infection via adoptive transfer of T-bet positive cells into infection-matched CD4-depleted recipient mice. Second, scRNAseq analysis revealed the subpopulation of effector CD4 T-cells that co-expressed Ctla4 and Gata3 was significantly higher than a subpopulation that co-expressed Ctla4 and Tbx21. Furthermore, our data suggested that CTLA-4 upregulation is beneficial against C. neoformans infection, as CTLA-4 blockade promoted fungal proliferation. Thus, we propose a model wherein Th1 control of latent C. neoformans infection is supported by CTLA-4 suppression of detrimental Th2 activation. IMPORTANCECD4 T-cells are a critical part of the adaptive immune response in many infections. Upon inhalation of the fungal pathogen Cryptococcus neoformans, CD4 T-cells are necessary to control the initial latent infection and prevent disease. In immunocompromised individuals that lack CD4 T-cells, the initial infection in the lungs can spread to the brain and cause lethal meningitis. In this study, we show that the CD4 T-cells present during latent C. neoformans infection are heterogeneous with expression of diverse polarization and immune regulation markers that function in concert to prevent disease. Our studies highlight the previously unappreciated diversity and complex regulation of the CD4 T-cell response that is required to prevent disease in this important fungal pathogen.

genomics↗

bioRxiv2026

Loss of Mast cells and histaminergic signaling link diet to platelet-mediated NETosis and mammary cancer recurrence

Breast cancer recurrence remains a clinical challenge. The period after the treatment of the primary tumor while cancer cells that evaded initial treatment lay dormant, provides a unique window of opportunity for interventions to prevent recurrence. Specific modifiable factors such as consumption of high fat diets or elevated circulating cholesterol are associated with decreased time to recurrence. Mechanistically, oxidized cholesterol and lipid species have been implicated in the regulation of the tumor microenvironment. This suggests that consumption of food prepared under oxidizing conditions such as pan-frying, may be an underappreciated risk. Using murine models of mammary cancer dormancy, we found that a diet enriched with fat from fried, cured bacon (cfBF) decreased dormancy latency times. Resulting lesions had fewer mast cells (MCs). Loss of MCs alone resulted in reemergence from dormancy. Elevated expression of a MC gene signature in breast tumors was associated with improved progression free and overall survival, highlighting the human relevance of these findings. MCs are a major source of tissue histamine, and lesions from mice fed cfBF had decreased concentrations. Importantly, antagonists of the histamine receptor 2 (H2R) sparked reemergence from dormancy. H2R antagonists are over-the-counter drugs are taken to alleviate gastroesophageal reflux disease. Chronic treatment of mice with H2R-antagonists sensitized platelets towards activation and crosstalk with neutrophils, and subsequent formation of neutrophil extracellular traps (NETs). The loss of platelet or NETosis activity mitigated the H2R-antagonist stimulated reemergence from dormancy. Therefore, we establish a novel metastatic axis which links diet to recurrence via MCs, histaminergic signaling and NETosis: Diet -- MC -- H2R -- (decreased) Platelet Activity -- (decreased) Neutrophil-NETosis -- (decreased) Reemergence from Dormancy. Our data reveal several potential intervention strategies: lifestyle, MC stabilization, histaminergic signaling, and neutrophil and platelet activity.

cancer biology↗

bioRxiv2025

LRH-1 is a novel regulator of neutrophil-driven immune responses within the tumor microenvironment.

Elevated plasma cholesterol levels have been linked to worse outcomes in breast and ovarian cancer. Prior work including our own has demonstrated that myeloid immune cells are highly responsive to cholesterol fluctuations and to proteins involved in cholesterol regulation. However, the specific roles of Liver Receptor Homolog-1 (LRH-1, or NR5A2), a key transcriptional regulator of cholesterol homeostasis, within myeloid cells remains largely undefined. Interestingly, LRH-1 mRNA levels are reduced in both breast and ovarian tumors compared to normal tissue. Its elevated expression within tumors is associated with increased survival time. These clinical correlations prompted us to explore the role of LRH-1 in myeloid cells particularly in the context of breast and ovarian cancer progression. Initial analyses confirmed LRH-1 expression in various myeloid cell types, with particularly high levels in neutrophils. We therefore focused on how LRH-1 influences neutrophil behaviors relevant to cancer, including migration, NETosis, phagocytosis, and interactions with T cells. Small molecule ligands for LRH-1 regulated neutrophil migration towards cancer cells. Phagocytosis was also regulated by LRH-1 small molecule ligands, the extent being dependent on type of bait (e. coli vs. cancer cells). In T cell co-cultures, neutrophils pretreated with an LRH-1 agonist promoted greater T cell expansion - particularly in CD4 cells - while LRH-1 inhibition suppressed this response. Moreover, LRH-1 activation reduced NETosis, while treatment with an antagonist or inverse agonist enhanced NETosis. Treatment of mice with an inverse agonist of LRH-1 increased the growth of 4T1 and E0771 mammary tumors. Given prior evidence that neutrophils contribute to the recurrence of dormant lesions, we further examined the role of LRH-1 in this context. Mice harboring dormant D2.0R mammary cancer lesions treated with an LRH-1 inverse agonist exhibited earlier tumor recurrence and enhanced metastatic progression compared to vehicle-treated controls. In contrast, treatment with BL001, a small chemical LRH-1 agonist, delayed recurrence in D2.0R-grafted mice. Collectively, these findings reveal that LRH-1 is a novel regulator of neutrophil-driven immune responses within the tumor microenvironment. LRH-1 thus remerges as a promising therapeutic target to suppress metastasis or prevent recurrence in breast cancer.

cancer biology↗

bioRxiv2024

Lipid Nanoparticle Barcoding for Multiplexed Single-Cell RNA Sequencing

Sample multiplexing is an emerging method in single-cell RNA sequencing (scRNA-seq) that addresses high costs and batch effects. Current multiplexing schemes use DNA labels to barcode cell samples but are limited in their stability and extent of labeling across heterogeneous cell populations. Here, we introduce Nanocoding using lipid nanoparticles (LNPs) for high barcode labeling density in multiplexed scRNA-seq. LNPs reduce dependencies on cell surface labeling mechanisms due to multiple controllable means of cell uptake, amplifying barcode loading 10-100-fold and allowing both protection and efficient release by dissolution. In cultured cell lines and heterogeneous cells from tissue digests, Nanocoding occurs in 40 minutes with stability after sample mixing and requires only commercially available reagents without novel chemical modifications. In spleen digests, 6-plex barcoded samples show minimal unlabeled cells, with all barcodes giving bimodal count distributions. Challenging samples containing lipid-rich debris and heterogeneous cells from adipose tissue of obese rodents show more than 95% labeling with all known subtypes identified. Using Nanocoding, we investigate gene expression changes related to aging in adipose tissue, profiling cells that could not be readily identified with current direct conjugate methods using lipid or antibody conjugates. This ease of generating and tuning these constructs may afford efficient and robust whole-sample multiplexing with minimal sample crosstalk.

bioengineering↗

bioRxiv2024

Neutrophils exposed to a cholesterol metabolite secrete extracellular vesicles that promote epithelial-mesenchymal transition and stemness in breast cancer cells.

Small extracellular vesicles (sEVs) are emerging as critical mediators of intercellular communication in the tumor microenvironment (TME). Here, we investigate the mechanisms by which sEVs derived from neutrophils treated with the cholesterol metabolite, 27-hydroxycholesterol (27HC), influence breast cancer progression. sEVs released from 27HC treated neutrophils enhance epithelial-mesenchymal transition (EMT) and stem-like properties in breast cancer cells, resulting in loss of adherence, increased migratory capacity and resistance to cytotoxic chemotherapy. Decreased microRNAs (miRs) within the sEVs resulted in activation of the WNT/{beta}-catenin signaling pathway in recipient cells and suggest that this may be a predominant pathway for stem-like phenotype and EMT. Our findings underscore a novel mechanism by which 27HC-modulated neutrophils contribute to breast cancer pathophysiology through EV-mediated intercellular communication, suggesting potential therapeutic targets in cancer treatment.

cancer biology↗

bioRxiv2024

Murine aortic valve cell heterogeneity at birth

Heart valve function requires a highly organized extracellular matrix (ECM) network that provides the necessary biomechanical properties needed to withstand pressure changes during each cardiac cycle. Lay down of the valve ECM begins during embryogenesis and continues throughout postnatal stages when it is remodeled into stratified layers and arranged according to blood flow. Alterations in this process can lead to dysfunction and if left untreated, heart failure. Despite this, the mechanisms that establish structure-function relationships of the valve, particularly during postnatal maturation, are poorly understood. To address this, single cell transcriptomics was performed on murine aortic valve structures at postnatal day one (PND1). Overall, 18 clusters of 7 diverse cell populations were identified, including a novel VEC subpopulation unique to PND1, and three previously unappreciated VIC subpopulations defined as "primitive", "remodeling" and "bioactive". Additional lineage tracing of the "primitive" VIC subpopulation in mice uncovered a temporal and spatial trajectory throughout postnatal maturation. In summary, this work highlights the heterogeneity of cell types within the aortic valve structure at birth that contribute to establishing and maintaining structure and function throughout life.

cell biology↗

bioRxiv2022

Myofibroblast Ccn3 is regulated by Yap and Wwtr1 and contributes to adverse cardiac outcomes

While Yap and Wwtr1 regulate resident cardiac fibroblast to myofibroblast differentiation following cardiac injury, their role specifically in activated myofibroblasts remains unexplored. Here we assess the pathophysiological and cellular consequence of genetic depletion of Yap alone (Yapfl/fl;PostnMCM) or Yap and Wwtr1 (Yapfl/fl;Wwtr1fl/+;PostnMCM) in adult mouse myofibroblasts following myocardial infarction and identify and validate novel downstream factors specifically in cardiac myofibroblasts that mediate pathological remodeling. Following myocardial infarction, depletion of Yap in myofibroblasts had minimal effect on heart function while depletion of Yap/Wwtr1 resulted in smaller scars, reduced interstitial fibrosis, and improved ejection fraction and fractional shortening. Single cell RNA sequencing of interstitial cardiac cells 7 days post infarction showed suppression of pro-fibrotic genes in fibroblasts derived from Yapfl/fl,Wwtr1fl/+;PostnMCM hearts. In vivo myofibroblast depletion of Yap/Wwtr1 as well in vitro knockdown of Yap/Wwtr1 dramatically decreased RNA and protein expression of the matricellular factor Ccn3. Administration of recombinant CCN3 to adult mice following myocardial infarction remarkably aggravated cardiac function and scarring. CCN3 administration drove myocardial gene expression of pro-fibrotic genes in infarcted left ventricles implicating CCN3 as a novel driver of cardiac fibrotic processes following myocardial infarction.

physiology↗

bioRxiv2022

Mevalonate metabolites contribute to granulocyte chemotaxis and mortality in murine endotoxemia

Statins appear to dramatically increase sepsis survival but only when treatment is initiated prior to hospitalization. This implies that statins exhibit a delayed-onset pro-survival benefit in septic patients that results in clinical futility of statin-therapy for patients already diagnosed with sepsis. Identification of mechanisms that contribute to increased sepsis-survival following statin-pretreatment may reveal novel therapeutic targets that do not suffer similar delayed onset benefits. Statins are used to treat hypercholesterolemia and function by inhibiting the production of the rate-limiting metabolite mevalonate. This indirectly inhibits the de novo synthesis of not only cholesterol but also isoprenoids that are involved in prenylation, the post-translational lipid modification of proteins. Mirroring clinical observations, chronic but not acute treatment with simvastatin significantly increased survival in a murine endotoxemia model. This was associated with reduced systemic granulocyte chemotaxis that occurred in a cell-intrinsic manner. In vitro modeling showed that simvastatin abolished chemoattractant responses and that this could be reversed by restoring geranylgeranyl pyrophosphate (GGPP) but not farnesyl pyrophosphate (FPP) nor cholesterol. Treatment with prenyltransferase inhibitors showed that chemoattractant responses were dependent on geranylgeranylation. Proteomic analysis of C15AlkOPP-prenylated proteins identified geranylgeranylated proteins involved in chemoattractant responses, including RHOA, RAC1, CDC42, and GNG2. Given the kinetic problems with initiating statin treatment after sepsis onset, prenyltransferases and geranylgeranylated proteins, such as RAC1 and GNG2, are promising interventional candidates for sepsis and critical inflammatory illness.

immunology↗

bioRxiv2021

Targeting metabolic adaptations in the breast cancer liver metastatic niche using dietary approaches to improve endocrine therapy efficacy

Estrogen receptor-positive (ER+) metastatic tumors contribute to nearly 70% of breast cancer-related deaths. Most patients with ER+ metastatic breast cancer (MBC) undergo treatment with the estrogen receptor antagonist fulvestrant (Fulv) as standard-of-care. Yet, among such patients, metastasis in liver is associated with reduced overall survival compared to other metastasis sites. The factors underlying the reduced responsiveness of liver metastases to ER-targeting agents remain unknown, impeding the development of more effective treatment approaches to improve outcomes for patients with ER+ liver metastases. We therefore evaluated site-specific changes in MBC cells and determined the mechanisms through which the liver metastatic niche specifically influences ER+ tumor metabolism and drug resistance. We characterized ER activity of MBC cells both in vitro, using a novel system of tissue-specific extracellular matrix hydrogels representing the stroma of ER+ tumor metastatic sites (liver, lung and bone), and in vivo, in liver and lung metastasis mouse models. ER+ metastatic liver tumors and MBC cells grown in liver hydrogels displayed upregulated expression of glucose metabolism enzymes in response to Fulv. Furthermore, differential ER activity, but not expression, was detected in liver hydrogels. In vivo, increased glucose metabolism led to increased glycogen deposition in liver metastatic tumors, while a fasting-mimicking diet increased efficacy of Fulv treatment to reduce the metastatic burden. ImplicationsOur findings identify a novel mechanism of endocrine resistance driven by the liver tumor microenvironment. These results may guide the development of dietary strategies to circumvent drug resistance in liver metastasis, with potential applicability in other metastatic diseases.

cancer biology↗

bioRxiv2020

The evolution of extreme fertility defied ancestral gonadotropin mediated brain-reproduction tradeoff

Gonadotropic hormones coordinate processes in diverse tissues regulating animal reproductive physiology and behavior. Juvenile hormone (JH) is the ancient and most common gonadotropin in insects, but not in advanced eusocial honey bees and ants. To probe the evolutionary basis of this change, we combined endocrine manipulations, transcriptomics, and behavioral analyses to study JH regulated processes in a bumble bee showing an intermediate level of sociliality. We found that in the fat body, more JH-regulated genes were upregulated and enriched for metabolic and biosynthetic pathways. This transcriptomic pattern is consistent with earlier evidence that JH is the major gonadotropin in the bumble bee. In the brain, most JH-regulated genes were downregulated and enriched for protein turnover pathways. Brain ribosomal protein gene expression was similarly downregulated in dominant workers, which naturally have high JH titers. In other species, similar downregulation of protein turnover is found in aging brains or under stress, and is associated with compromised long-term memory and health. These findings suggest a previously unknown gonadotropin-mediated tradeoff. We did not find a similar downregulation of protein turnover pathways in the brain of honey bees in which JH is not a gonadotropin but rather regulates division of labor. These differences between JH effets in the bumble bee and in the advanced eusocial honey bee suggest that the evolution of advanced eusociality was associated with modifications in hormonal signaling supporting extended and extremely high fertility while reducing the ancient costs of high gonadotropin titers to the brain.

evolutionary biology↗

bioRxiv2019

A common pattern of influenza A virus single cell gene expression heterogeneity governs the innate antiviral response to infection

Viral infection outcomes are governed by the complex and dynamic interplay between the infecting virus population and the host response. It is increasingly clear that both viral and host cell populations are highly heterogeneous, but little is known about how this heterogeneity influences infection dynamics or viral pathogenicity. To dissect the interactions between influenza A virus (IAV) and host cell heterogeneity, we examined the combined host and viral transcriptomes of thousands of individual, single virion-infected cells. We observed complex patterns of viral gene expression and the existence of multiple distinct host transcriptional responses to infection at the single cell level. Our analyses reveal that viral NS segment gene expression diverges from that of the rest of the viral genome within a subset of infected cells, and that this unique pattern of NS segment expression can play a dominant role in shaping the host cell response to infection. Finally, we show that seasonal human H1N1 and H3N2 strains differ significantly in patterns of host anti-viral gene transcriptional heterogeneity at the single cell level. Altogether, these data reveal a common pattern of viral gene expression heterogeneity across human IAV subtypes that can serve as a major determinant of antiviral gene activation.

microbiology↗