Biology subjects

Frey, N.

Publications and source records attributed to Frey, N..

30 records · Page 2Linked to original sources

bioRxiv2023

Tryptophan wasting and disease activity as a systems phenomenon in inflammation - an analysis across 13 chronic inflammatory diseases.

Chronic inflammatory diseases (CID) are systems disorders affecting various organs including the intestine, joint and skin. The essential amino acid tryptophan (Trp) is not only used for protein synthesis but can also be catabolized to various bioactive derivatives that are important for cellular energy metabolism and immune regulation. Increased Trp catabolism via the kynurenine pathway is seen across individual CID entities1-5. Here, we assessed the levels of Trp and tryptophan derivatives across 13 CID to investigate the extent and nature of Trp wasting as a systems phenomenon in CID. We found reduced serum Trp levels across the majority of CID and a prevailing negative relationship between Trp and systemic inflammatory marker C-reactive protein (CRP). Increases in the kynurenine-to-Trp ratio (Kyn:Trp) indicate that the kynurenine pathway is a major route for CID-related Trp wasting. However, the extent of Trp depletion and its relationship with disease activity varies by disease, indicating potential differences in Trp metabolism. In addition, we find that amino acid catabolism in chronic inflammation is specific to tryptophan wasting, whereas other proteinogenic amino acids are not affected. Hence, our results suggest that increased Trp catabolism is a common metabolic occurrence in CID that may directly affect systemic immunity. Grant supportThis work was supported by the DFG Cluster of Excellence 1261 "Precision medicine in chronic inflammation" (KA, SSchr, PR, BH, SWa), the BMBF (e:Med Juniorverbund "Try-IBD" 01ZX1915A and 01ZX2215, the e:Med Network iTREAT 01ZX2202A, and GUIDE-IBD 031L0188A), DFG RU5042 (PR, KA), and Innovative Medicines Initiative 2 Joint Undertakings ("Taxonomy, Treatments, Targets and Remission", No. 831434, "ImmUniverse", grant agreement No. 853995, "BIOMAP", grant agreement No. 821511).

immunology↗

bioRxiv2023

CXCL4-induced PBMCs Modulate Vascular Extracellular Matrix through Wnt5a-dependent Release of Calcific Extracellular Vesicles and Matrix Metalloproteinase-7

BackgroundMacrophage heterogeneity plays an increasing role in the study of vascular inflammatory responses. The CXCL4-induced monocyte/macrophage phenotype has previously been implicated with atherosclerotic plaque destabilization, a key process preceding plaque rupture. Monocyte-derived macrophages were found to exhibit a unique transcriptome in the presence of CXCL4 characterized by upregulation of S100A8 and MMP7. However, the mechanisms involved in CXCL4-induced monocyte-mediated vascular inflammation are unknown. MethodsSingle-cell RNA sequencing data were examined for CXCL4-dependent gene expression signatures in plaque macrophages. Human PBMCs were differentiated with CXCL4 and subsequently characterized in terms of osteogenic gene and protein expression signatures and calcific extracellular vesicle release. Association of the CXCL4-induced phenotype with the Wnt pathway was investigated, and CXCL4-induced PBMC-derived EV were analyzed for their potential to elicit an inflammatory response in vSMC. In-vitro findings were verified histologically in calcified human carotid artery plaques. ResultsIn human plaque macrophages, single-cell sequencing revealed a CXCL4-susceptible subpopulation bearing a distinct proinflammatory gene expression profile. CXCL4-differentiated PBMCs exhibited a marked induction of S100A8, MMP7 and osteogenic marker transcription concomitant with augmented release of calcific EVs enriched with MMP7, S100A8 and alkaline phosphatase. Under osteogenic conditions, increased overt calcification of the extracellular matrix was observed in vitro. Analysis of inflammatory pathway activation identified the paracrine Wnt5a-CaMKII signaling axis to be causally linked to the CXCL4-induced osteogenic PBMC phenotype, S100A8 and MMP7 enrichment as well as calcific potential of secreted EV. Additionally, CXCL4-polarized PBMC-derived EV differentially stimulated osteogenic/inflammatory genotype transition in vSMC. In human carotid artery plaques, occurrence of CXCL4-induced mononuclear cells coincided with Wnt5a-CaMKII pathway activation and progressive plaque calcification. ConclusionsThis study introduces a novel mechanism driving monocyte/macrophage-mediated extracellular matrix remodeling in calcific inflammatory responses through Wnt5a-CaMKII-activated secretion of MMP7+S100A8+ calcifying EV by CXCL4-induced pro-inflammatory monocytes. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=119 SRC="FIGDIR/small/540832v1_ufig1.gif" ALT="Figure 1"> View larger version (31K): org.highwire.dtl.DTLVardef@1f81c66org.highwire.dtl.DTLVardef@107be9org.highwire.dtl.DTLVardef@37042forg.highwire.dtl.DTLVardef@1f3b0f3_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗

bioRxiv2023

Single-cell transcriptomics reveal distinctive patterns of fibroblast activation in murine heart failure with preserved ejection fraction

Inflammation, fibrosis and metabolic stress critically promote heart failure with preserved ejection fraction (HFpEF). Exposure to high-fat diet and nitric oxide synthase inhibitor N[w]-nitro-l-arginine methyl ester (L-NAME) recapitulate features of HFpEF in mice. To identify disease specific traits during adverse remodeling, we profiled interstitial cells in early murine HFpEF using single-cell RNAseq (scRNAseq). Diastolic dysfunction and perivascular fibrosis were accompanied by an activation of cardiac fibroblast and macrophage subsets. Integration of fibroblasts from HFpEF with two murine models for heart failure with reduced ejection fraction (HFrEF) identified a catalog of conserved fibroblast phenotypes across mouse models. Moreover, HFpEF specific characteristics included induced metabolic, hypoxic and inflammatory transcription factors and pathways, including enhanced expression of Angiopoietin-like 4 next to basement membrane compounds. Fibroblast activation was further dissected into transcriptional and compositional shifts and thereby highly responsive cell states for each HF model were identified. In contrast to HFrEF, where myofibroblast and matrifibrocyte activation were crucial features, we found that these cell-states played a subsidiary role in early HFpEF. These disease-specific fibroblast signatures were corroborated in human myocardial bulk transcriptomes. Furthermore, we found an expansion of pro-inflammatory Ly6Chigh macrophages in HFpEF, and we identified a potential cross-talk between macrophages and fibroblasts via SPP1 and TNF[a]. Finally, a marker of murine HFpEF fibroblast activation, Angiopoietin-like 4, was elevated in plasma samples of HFpEF patients and associated with disease severity. Taken together, our study provides a comprehensive characterization of molecular fibroblast and macrophage activation patterns in murine HFpEF, as well as the identification of a novel biomarker for disease progression in patients.

cell biology↗

bioRxiv2023

Amino acid auxotrophies in human gut bacteria are linked to higher microbiome diversity and long-term stability

Amino acid auxotrophies are prevalent among bacteria. They can govern ecological dynamics in microbial communities and indicate metabolic cross-feeding interactions among coexisting genotypes. Despite the ecological importance of auxotrophies, their distribution and impact on the diversity and function of the human gut microbiome remain poorly understood. This study performed the first systematic analysis of the distribution of amino acid auxotrophies in the human gut microbiome using a combined metabolomic, metagenomic, and metabolic modeling approach. Results showed that amino acid auxotrophies are ubiquitous in the colon microbiome, with tryptophan auxotrophy being the most common. Auxotrophy frequencies were higher for those amino acids that are also essential to the human host. Moreover, a higher overall abundance of auxotrophies was associated with greater microbiome diversity and stability, and the distribution of auxotrophs was found to be related to the human hosts metabolome, including trimethylamine oxide, small aromatic acids, and secondary bile acids. Thus, our results suggest that amino acid auxotrophies are important factors contributing to microbiome ecology and host-microbiome metabolic interactions.

microbiology↗

bioRxiv2023

Cardiac-targeted rAAV5-S100A1 gene therapy protects against adverse remodeling and contractile dysfunction in post-ischemic hearts

Toxicity by recombinant adeno-associated viruses (rAAV) in clinical gene therapy trials (e.g., by rAAV9-mediated fatal liver failure) significantly impairs translation of preclinical rAAV-based cardiac gene therapies employing these vectors. For rAAV5 - a capsid that has shown long-term safety in clinical trials - our translational study demonstrates effective transduction of the left ventricle (LV) of healthy pigs via catheter-based retrograde intravenous delivery (CRID) by means of luciferase reporter gene biodistribution analyses. Combination of rAAV5 with the cardioprotective human gene S100A1 (hS100A1) prevents LV myocardial infarct (MI) enlargement and improves LV systolic contractile performance in a porcine model of post-MI chronic cardiac dysfunction. Use of a cardiac-biased promoter ensured the cardiac-directed expression of the therapeutic human transgene without signs of clinical toxicity. The beneficial effects of rAAV5-hS100A1 were linked to an attenuated activity of post-MI inflammatory gene networks and this was further validated in a murine model. These novel data together with proven scalable producibility and low pre-existing immunity against rAAV5 in humans may collectively advance clinical translation of rAAV5-hS100A1 as a gene therapy medicinal product (GTMP) for a common cardiovascular disease, such as chronic heart failure (CHF). HighlightsO_LIRecent fatal adverse events in recombinant adeno-associated virus (AAV)-based clinical gene therapy trials advise the use of rAAV serotypes with proven long-term clinical safety, such as rAAV5, for the pre-clinical development and clinical translation of rAAV-based cardiac gene therapy medicinal products. C_LIO_LIIn a biodistribution and therapeutic proof-of-concept study in farm pigs, rAAV5 was identified as an effective viral vector for cardiac gene transfer and gene therapy for post-ischemic cardiac dysfunction when applied by a standardized cardiac-targeted catheter-based route of administration with the luciferase reporter and cardioprotective human gene S100A1 (hS100A1), respectively. C_LIO_LIA systems biology analysis linked the novel finding of mitigated inflammatory and activated cardioprotective gene network activities in rAAV5-hS100A1 treated postischemic myocardium with improved study left ventricular ejection fraction and prevention of myocardial infarct extension, respectively, which warrants further mechanistic molecular studies. C_LIO_LISince rAAV5 has been recently approved for clinical use in a non-cardiac indication and cardiac-targeted S100A1 gene therapy has been effective in numerous pre-clinical animal models of acute and chronic cardiac dysfunction, our translational data support an expedited developmental path for rAAV5-hS100A1 throughout investigational new drug-enabling studies towards a first-in-human clinical trial for post-myocardial infarction heart failure. C_LI

pathology↗

bioRxiv2023

S100A1ct: a synthetic peptide derived from human S100A1 protein improves cardiac contractile performance and survival in pre-clinical heart failure models

BackgroundThe EF-hand Ca2+ sensor protein S100A1 has been identified as a molecular regulator and enhancer of cardiac performance. S100A1s ability to recognize and modulate the activity of targets such as SERCA2a and RyR2 in cardiomyocytes has mostly been ascribed to its hydrophobic C-terminal -helix (residues 75-94). Objective: We therefore hypothesized that a synthetic peptide consisting of residues 75-94 of S100A1 and an N-terminal solubilization tag (S100A1ct) could mimic the performance enhancing effects of S100A1 and may be suitable as a peptide therapeutic to improve the function of diseased hearts. Methods and Results: Applying an integrative translational research pipeline, ranging from computational molecular modeling to large animal cardiac disease models, we characterize S100A1ct as a cell-penetrating peptide with positive inotropic and antiarrhythmic properties in normal and failing myocardium in vitro and in vivo. This activity translates into improved contractile performance and survival in pre-clinical heart failure models with reduced ejection fraction after S100A1ct systemic administration. Mechanistically, S100A1ct exerts a fast and sustained dose-dependent enhancement of cardiomyocyte Ca2+ cycling and prevents {beta}-adrenergic receptor triggered Ca2+ imbalances by targeting SERCA2a and RyR2 activity. Modeling suggests that S100A1ct may stimulate SERCA2a by interacting with the sarcoplasmic transmembrane segments of the multi-span integral membrane Ca2+ pump. Incorporation of a cardiomyocyte targeting peptide tag into S100A1ct (cor-S100A1ct) further enhanced its biological and therapeutic potency in vitro and in vivo. Conclusion: S100A1ct peptide is a promising lead for the development of a novel peptide-based therapeutic against heart failure with reduced ejection fraction.

pharmacology and toxicology↗

bioRxiv2022

Ythdf2 regulates cardiac remodeling through its m6A-mRNA target transcripts

m6A mRNA methylation controls cardiomyocyte function and increased overall m6A levels are a stereotyping finding in heart failure independent of the underlying etiology. However, it is largely unknown how the information is read by m6A reader proteins in heart failure. Here we show that the m6A reader protein Ythdf2 controls cardiac function and identified a novel mechanism how reader proteins control gene expression and cardiac function. Deletion of Ythdf2 in cardiomyocytes in vivo leads to cardiac hypertrophy, reduced heart function, and increased fibrosis during pressure overload as well as during aging. Similarly, in vitro the knockdown of Ythdf2 results in cardiomyocyte growth and remodeling. Mechanistically, we identified the eucaryotic elongation factor 2 as a major target of Ythdf2 using cell type specific Ribo-seq data. Our study expands our understanding on the regulatory functions of m6A methylation in cardiomyocytes and how cardiac function is controlled by the m6A reader protein Ythdf2.

molecular biology↗

bioRxiv2022

Secreted long non-coding RNAs Gadlor1 and Gadlor2 affect multiple cardiac cell types and aggravate cardiac remodeling during pressure overload

BackgroundPathological overload triggers maladaptive myocardial remodeling that leads to heart failure. Recent studies have shown that long non-coding RNAs (lncRNAs) regulate cardiac remodeling. This study investigates two recently discovered, secreted lncRNAs, Gadlor1 and Gadlor2 (Gadlor 1/2). MethodsWe generated compound Gadlor1/2 knock-out (KO) mice and compared their response to pressure overload by transverse aortic constriction (TAC) to that of wild-type (WT) littermates. Endothelial cells, fibroblasts and cardiomyocytes were isolated from the hearts of both genotypes after TAC and their transcriptome was investigated by RNA sequencing. Gadlor target proteins were identified by RNA antisense purification coupled with mass spectrometry (RAP-MS) in cardiomyocytes. In addition, we investigated the effects of cardiac overexpression of Gadlor1/2. ResultsGadlor1/2 are jointly upregulated in failing mouse hearts as well as in the myocardium of heart failure patients. Cardiac overexpression of Gadlor1 and Gadlor2 aggravated myocardial dysfunction and enhanced hypertrophic and fibrotic remodeling in mice exposed to pressure overload. Compound Gadlor1/2 KO mice, in turn, exerted markedly reduced myocardial hypertrophy, fibrosis and dysfunction, but more angiogenesis during short and long-standing pressure overload. Paradoxically, Gadlor1/2 KO mice suffered from sudden death during prolonged overload, possibly due to cardiac arrhythmia. Gadlor1 and Gadlor2, which are mainly expressed in endothelial cells (ECs) in the heart, where they inhibit pro-angiogenic gene-expression, are strongly secreted within extracellular vesicles (EVs). These EVs transfer Gadlor lncRNAs to cardiomyocytes, where they bind and activate calmodulin-dependent kinase II, induce pro-hypertrophic gene-expression and enhance calcium re-uptake into the sarcoplasmic reticulum. ConclusionGadlor1 and Gadlor2 are lncRNAs that are mainly enriched in EC-derived EVs and are jointly upregulated in mouse and human hearts during pathological overload. We reveal a crucial endothelial cell-cardiomyocyte crosstalk, which aims at restoring calcium homeostasis in cardiomyocytes during overload at the cost of aggravated hypertrophy and fibrosis.

systems biology↗

bioRxiv2022

Transient inhibition of translation improves long-term cardiac function after ischemia/reperfusion by attenuating the inflammatory response

RationaleRapid reperfusion is the most effective treatment for attenuating cardiac injury caused by myocardial ischemia. Yet, reperfusion itself elicits damage to the myocardium through incompletely understood mechanisms, known as ischemia/reperfusion (I/R) injury. The myocardium adapts to I/R by changes in gene expression, which determines the cellular response to reperfusion. Protein translation is a key component of gene expression. However, it is unknown how regulation of translation contributes to cardiac gene expression in response to reperfusion and whether it can be targeted to mitigate I/R injury. MethodsTo examine translation and its impact on gene expression in response to I/R we assessed protein synthesis at different timepoints after ischemia and reperfusion in vitro and in vivo. Pharmacological inhibitors were used to dissect the underlying molecular mechanisms of translational control. Transient inhibition of protein synthesis was undertaken to decipher the effects of the translational response to reperfusion on cardiac function and inflammation. Cell-type-specific ribosome profiling was performed in mice subjected to I/R to determine the impact of translation on the regulation of gene expression in cardiomyocytes. ResultsReperfusion increased translation rates from a previously suppressed state during ischemia in cardiomyocytes, which was associated with the induction of cell death. In vivo, I/R resulted in strong activation of translation in the myocardial border zone. Detailed analysis revealed that the upregulation of translation is mediated by eIF4F complex formation, which was specifically mediated by the mTORC1-4EBP1-eIF4F axis. Short-term pharmacological inhibition of eIF4F complex formation by 4EGI-1 or rapamycin, respectively, attenuated translation, reduced infarct size and improved long-term cardiac function after myocardial infarction. Cardiomyocyte-specific ribosome profiling identified that reperfusion damage increased translation of mRNA networks in cardiomyocytes associated with cardiac inflammation and cell infiltration. Transient inhibition of the mTORC1-4EBP1-eIF4F axis decreased the expression of proinflammatory transcripts such as Ccl2, thereby reducing Ly6Chi monocyte infiltration and myocardial inflammation. ConclusionsMyocardial reperfusion induces protein synthesis in the border zone which contributes to I/R injury by rapidly translating a specific maladaptive mRNA network that mediates immune cell infiltration and inflammation. Transient inhibition of the mTORC1-4EBP1-eIF4F signaling axis during reperfusion attenuates this proinflammatory translational response, protects against I/R injury and improves long-term cardiac function after myocardial infarction. Clinical PerspectiveO_ST_ABSWhat Is New?C_ST_ABSO_LIThis is the first study to investigate the impact of translational regulation on cardiomyocyte gene expression in response to myocardial ischemia/reperfusion. C_LIO_LIWe show that translation regulates approximately two-thirds of differentially expressed genes in cardiomyocytes after ischemia/reperfusion, including many involved in inflammation and immune cell infiltration. C_LIO_LIThe translational response to ischemia/reperfusion is regulated by the mTORC1-4EBP1-eIF4F axis, which determines pro-inflammatory monocyte infiltration via control of the expression of the chemokine Ccl2. C_LI What Are the Clinical Implications?O_LICurrently, there are no specific therapies to prevent ischemia/reperfusion injury, which is mediated, at least in part, by a maladaptive inflammatory response. C_LIO_LIA translationally controlled network regulated by the mTORC1-4EBP1-eIF4F axis can be targeted by a short-term pharmacological intervention to attenuate the inflammatory response and improve cardiac function after ischemia/reperfusion in mice. C_LIO_LIThis study supports the emerging concept of selectively inhibiting maladaptive elements of the inflammatory response to improve outcome in patients after myocardial infarction; in addition, it provides a mechanistic basis for the currently ongoing CLEVER-ACS trial. C_LI

molecular biology↗

bioRxiv2021

Loss of Rnf31 and Vps4b sensitizes pancreatic cancer to T cell-mediated killing

Pancreatic ductal adenocarcinoma (PDA) is an inherently immune cell deprived tumor, characterized by desmoplastic stroma and suppressive immune cells. Here we systematically dissected PDA intrinsic mechanisms of immune evasion by in vitro and in vivo CRISPR screening, and identified Rnf31 and Vps4b as essential factors required for escaping CD8+ T cell-killing. Using murine PDA cells and human PDA organoids, we demonstrate that Rnf31 protects from TNF-mediated caspase 8 cleavage and subsequent apoptosis induction. For Vps4b we found that inactivation impairs autophagy, resulting in increased accumulation of CD8+ T cell-derived granzyme B and subsequent tumor cell lysis. Orthotopic transplantation of Rnf31- or Vps4b deficient pancreatic tumors, moreover, revealed increased CD8+ T cell infiltration and effector function, and markedly reduced tumor growth in mice. Our work uncovers vulnerabilities in PDA that might be exploited to render these tumors more susceptible to the immune system.

cancer biology↗

bioRxiv2020

Predicting base editing outcomes with an attention-based deep learning algorithm trained on high-throughput target library screens

Base editors are chimeric ribonucleoprotein complexes consisting of a DNA-targeting CRISPR-Cas module and a single-stranded DNA deaminase. They enable conversion of C•G into T•A base pairs and vice versa on genomic DNA. While base editors have vast potential as genome editing tools for basic research and gene therapy, their application has been hampered by a broad variation in editing efficiencies on different genomic loci. Here we perform an extensive analysis of adenine- and cytosine base editors on thousands of lentivirally integrated genetic sequences and establish BE-DICT, an attention-based deep learning algorithm capable of predicting base editing outcomes with high accuracy. BE-DICT is a versatile tool that in principle can be trained on any novel base editor variant, facilitating the application of base editing for research and therapy.Competing Interest StatementThe authors have declared no competing interest.View Full Text

genomics↗

bioRxiv2019

Fog signaling has diverse roles in epithelial morphogenesis in insects

The Drosophila Fog pathway represents one of the best-understood signaling cascades controlling epithelial morphogenesis. During gastrulation, Fog induces apical cell constrictions that drive the invagination of mesoderm and posterior gut primordia. The cellular mechanisms underlying primordia internalization vary greatly among insects and recent work has suggested that Fog signaling is specific to the fast mode of gastrulation found in some flies. On the contrary, here we show in the beetle Tribolium, whose development is broadly representative for insects, that Fog has multiple morphogenetic functions. It modulates mesoderm internalization and controls a massive posterior infolding involved in gut and extraembryonic development. In addition, Fog signaling affects blastoderm cellularization, primordial germ cell positioning and cuboidal-to-squamous cell shape transitions in the extraembryonic serosa. Comparative analyses with two other distantly related insect species reveals that Fogs role during cellularisation is widely conserved and therefore might represent the ancestral function of the pathway.

developmental biology↗