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Identification of an Angiotensin-(1–7)-Producing fusion protein in the brain of transgenic rats Reveals a hypotensive effect mediated through modulation of the GABA–nNOS–NO pathway and highlighting Astrocyte–Neuron crosstalk
No full textThis study explores the anatomical distribution of Angiotensin-(Ang)-(1–7) fusion protein within the central nervous system of the novel transgenic rat model (TG7371). The Ang-(1–7)/Mas pathway of the renin-angiotensin system (RAS) plays a key role in cardiovascular regulation and influences higher brain functions, including cognition and emotion. TG7371 expresses a transgenic Ang-(1–7)-producing fusion protein which resulted in a hypotensive phenotype. Here, we assessed the expression of Ang-(1–7) fusion mRNA and protein in primary cortical cells from neonates and identified their distribution in the brain of adult rats using qPCR, WB, ISH, and immunolabeling. In neonates, Ang-(1–7) mRNA was mainly found in proliferating cells, whereas in adults, it was primarily identified in GFAP-positive astrocytes. The Ang-(1–7) fusion protein, however, was predominantly found in neurons, including GABAergic interneurons and specific pyramidal cells. High protein levels were particularly noted in cardiovascular control regions like the medulla, as well as in other non-cardiovascular areas. TG7371 displayed twofold increase in brain levels of Ang-(1–7) compared to Ang II vs. Control, which remained unchanged, alongside significant changes in the expression of RAS components and nNOS. These findings indicate that the Ang-(1–7) fusion protein modulates the GABA-nNOS-NO-pathway, contributing to the low blood pressure phenotype of these rats, and promotes a mode of astrocytes-neurons-communication. The widespread expression of the fusion protein in the brain also suggests a potential role in modulating mood, cognition, and neurological disorders. Overall, TG7371 presents a valuable model to explore the long-term cardiovascular and neurobehavioral effects of Ang-(1–7), highlighting promising therapeutic implications and neural crosstalk
Changes in sputum viscoelastic properties and airway inflammation in primary ciliary dyskinesia are comparable to cystic fibrosis on elexacaftor/tezacaftor/ivacaftor therapy
No full textivacaftor (ETI) therapy. Methods: We measured and compared sputum rheology, inflammation markers and the proteome in 42 clinically stable adolescent and adult patients with PCD, 40 patients with CF with at least one F508del allele before and 3 months after initiation of ETI and 15 age-matched healthy controls. Results: Patients with PCD showed increased viscoelastic properties (G’ and G’’ P<0.001) of airway mucus and elevated levels of airway inflammation markers, including free neutrophil serine protease activity (free NE, CatG, PR3 P<0.001), IL-1β and IL-8 (P<0.001), compared to healthy controls, but lower than observed in CF patients at baseline (free proteases, IL-1β, G’ and G’’ P<0.001, IL-8 P<0.05). Significant differences in sputum viscoelastic properties, airway inflammation and the sputum proteome between PCD patients and CF patients on ETI therapy were no longer observed. Conclusions: Clinically stable patients with PCD exhibit abnormalities in sputum viscoelastic properties, inflammation markers and the proteome that are less severe than in patients with CF without ETI, but similar to observed under ETI therapy
Small extracellular vesicle-mediated adipocyte-cardiomyocyte crosstalk exacerbates heart failure with preserved ejection fraction
No full textAims: Although growing evidence suggests that obesity/central adiposity predispose to development and exacerbation of heart failure with preserved ejection fraction (HFpEF), it remains to be clarified whether there is a causal relationship between adiposity and HFpEF pathogenesis. Methods and results: HFpEF was induced in male, C57BL/6N mice using high-fat diet + Nω-nitro-l-arginine methyl ester. Resection or transplantation of visceral adipose tissue (VAT) blunted or exacerbated HFpEF phenotypes, respectively, the in mice. VAT from HFpEF mice displayed greater weight and secreted more small extracellular vesicles (sEVs) than those from chow-fed mice. Either systemic inhibition of sEV secretion or VAT-specific knockdown of Rab27b (an indispensable GTPase for sEV secretion) protected against HFpEF. Discovery-driven experiments identified miR-2953p within sEVs as a possible mediator of the VAT-heart axis, which impaired cardiac autophagy by binding to Ulk1 mRNA. MiR-295-3p antagomir treatment mitigated HFpEF phenotypes. Additionally, neonatal mouse cardiomyocytes (NMCMs) manifested blunted autophagic flux after treatment with plasma sEVs from HFpEF mice. Notably, HFpEF patients displayed downregulated cardiac Ulk1 and autophagy compared with healthy individuals. Restoration of cardiac autophagy with rapamycin or ULK1 overexpression via AAV-9 attenuated the HFpEF phenotype in mice. Conclusions: The present work unveils a mechanism whereby obesity promotes HFpEF progression, emphasizing the role of VAT-heart crosstalk. Specifically, VATderived sEVs, miR-295-3p, and the resultant disruption of cardiac autophagy contribute significantly to the pathogenesis of HFpEF
Indole-3 acetate limits dysbiosis-driven diastolic failure via Hcrt neurons
No full textBACKGROUND: The nervous, gastrointestinal, renal, and cardiovascular systems orchestrate ion-fluid homeostasis and impose reciprocal adaptations to hypertensive challenges. Mechanistic insight into the interorgan crosstalk is fundamental for tackling pathogenesis of hypertensive heart disease. METHODS: We integrated gut microbiome profiling and targeted metabolomics in a zebrafish model of ion dyshomeostasis-induced diastolic dysfunction to identify microbial metabolites linked to hypertensive cardiac remodeling. To dissect the gut-brain-heart axis, we depleted microbiota, supplemented specific microbial metabolites, and chemogenetically ablated hypothalamic neurons. Neuronal activity was monitored using in vivo calcium imaging and immunohistochemistry, and cardiovascular function was assessed by live imaging. Patient serum metabolic profiles were analyzed to evaluate relevance to human hypertension. RESULTS: Zebrafish larvae exposed to ion dyshomeostasis exhibited gut dysbiosis, marked by reduced microbial richness and diversity, particularly among indole- and indole-3-producing taxa. Functionally, commensal microbiota protected against cardiovascular structural and functional remodeling during hypertensive challenge, whereas antibiotic-induced perturbation worsened hemodynamic parameters of arterial hypertension and impaired ventricular relaxation. Gut metabolomics identified a lower abundance of indole-3 acetic acid as a key signature of the hypertensive response, a pattern conserved in serum metabolome from patients with hypertension. Indole-3 acetic acid supplementation, acting via the aryl hydrocarbon receptor, mitigated cardiac concentric hypertrophy and diastolic dysfunction. These effects involved hypothalamic hypocretin neurons, with indole-3 acetic acid suppressing their overactivation and the associated sympathetic overdrive in cardiac-projecting paravertebral ganglia during the hypertensive challenge. Indole-3 acetic acid also prevented renin-angiotensin-aldosterone system upregulation, indicating that it operates upstream of both autonomic and hormonal pathways. CONCLUSIONS: Our findings uncover a gut-brain-heart crosstalk where hypertensive gut dysbiosis signals to the central nervous system to drive diastolic remodeling. Modulation of indole-3 acetic acid signaling and hypocretin neuron activity represents a promising strategy to counter the multisystemic pathogenesis of hypertensive heart disease
Complex human hear bearing skin organoids reveal cell type specific susceptibility and innate immune responses to Herpes Simplex Virus 1
Full text linkEpithelial surfaces are the initial site of Herpes Simplex Virus 1 (HSV-1) infection. After latency establishment in the peripheral nervous system, reactivation can lead to skin pathologies. To reflect the complexity of skin we used complex human hair bearing skin organoids as a new model system for HSV-1 infection. We analyzed spread of the infection through the three-dimensional tissue and the cell type specific host response in a spatial temporal manner. Single-cell-resolution spatial transcriptomics and imaging shows a restricted viral infection in keratinocytes of the epidermis and hair follicles. The host factor IER3 is upregulated in papillary fibroblasts of the dermis as the infection progresses through the tissue. In keratinocytes of the stratum basale, we find IRF3 upregulated in infected but also virus negative cells, suggesting a signaling mechanism to initiate antiviral responses. Furthermore, we find a cell type specific inflammatory response with an upregulation of TNFSF9 in dermal fibroblasts and TNF in keratinocytes. TNFα target genes are induced in neighboring uninfected cells suggesting the induction of a localized inflammatory reaction to suppress viral infection. In summary, our findings position human hair-bearing skin organoids as a highly physiological model system for studying HSV-1 pathogenesis and reveal spatially organized host responses
Spatial proteomics of ovarian cancer precursors delineates early disease changes and drug targets
Full text linkHigh-grade serous ovarian cancer (HGSOC) is often detected at an advanced stage, where curative treatment options are limited. Recent advances in ultrasensitive mass spectrometry-based spatial proteomics have provided a unique opportunity to uncover molecular drivers of early tumorigenesis and novel therapeutic targets. Here, we present a comprehensive proteomic analysis of serous tubal intraepithelial carcinoma (STIC), the HGSOC precursor lesion, and concurrent invasive carcinoma, covering more than 10,000 proteins from ultra-low input archival tissue. STIC and HGSOC showed highly similar proteomes, clustering into two subtypes with distinct tumor-immune microenvironments and common remodeling of the extracellular matrix. We discovered cell-of-origin signatures from secretory fallopian tube epithelial cells in STICs and identified early dysregulated pathways of therapeutic relevance. Targeting cholesterol biosynthesis by inhibiting the terminal steps via DHCR7 showed therapeutic effects in ovarian cancer cell lines and synergized with standard-of-care carboplatin treatment. This study demonstrates the power of spatially resolved quantitative proteomics in understanding early carcinogenesis and provides a rich resource for biomarker and drug target research
Metabolic inflammation, brain age and cognitive functioning in short- and long-term clinical weight loss trials
Full text linkBACKGROUND: Observational studies suggest that metabolic inflammation in obesity can impair brain health, but studies on beneficial effects of weight loss-induced improvements in such markers on brain health and their consequences for clinical outcomes are scarce. METHODS: Consequently, we investigated 53 obese participants in a short-term dietary weight loss trial (up to 4 months, 137 samples; "Muscle Metabolism Study" or "MMS") and 30 in an independent long-term trial (up to 39 months, 100 samples; "Maintain"). For each participant and visit, brain health was characterised in terms of the "brain-predicted age difference" ("brain-PAD"; the difference of the age of a person predicted with machine learning from structural brain MRI minus their chronological age). Increasingly positive brain-PAD scores indicate increasingly poorer brain health. Further, we determined the HOMA index, leptin, fetuin B and CRP levels as markers collectively reflecting low-grade inflammation and impaired metabolic signalling. Finally, we evaluated the relevance of these parameters for brain-PAD and the association of brain-PAD alterations for cognition, which was measured in the MMS with neuropsychological tests. FINDINGS: Weight loss led to improved brain-PAD scores (MMS: t = -2.02, p = 0.023, effect size partial η(2) (η(2)(p)) = 0.03; Maintain: t = -7.37, p = 4.2·10(-11), η(2)(p) = 0.38). According to a False Discovery Rate (FDR) method-corrected threshold (α(FDR) = 0.05), HOMA index (MMS: t = 2.28, p(FDR) = 0.024, η(2)(p) = 0.04; Maintain: t = 2.33, p(FDR) = 0.023, η(2)(p) = 0.08), and leptin (MMS: t = 4.43, p(FDR) = 4.3·10(-5), η(2)(p) = 0.14; Maintain: t = 1.91, p(FDR) = 0.041, η(2)(p) = 0.06), showed significant positive links to brain-PAD in both trials, fetuin B did so in Maintain (t = 2.57, p(FDR) = 0.023, η(2)(p) = 0.11). Brain-PAD variations were associated with a neuropsychological test of psychomotor speed and visual attention (t = 2.32, p(FDR) = 0.022, η(2)(p) = 0.05). Application of explainable artificial intelligence methods showed that this link was parallelled by widespread brain age-related tissue alterations in white and grey matter involved in these functions. INTERPRETATION: Analyses of two independent weight loss trials suggest that weight loss-induced improvements in metabolic-inflammatory markers have beneficial effects on brain-PAD and the latter were associated with enhancements in cognitive functioning, underscoring the potential clinical relevance of metabolic brain age regulation
Lactococcus A phages predict ACLF while Enterococcus B phages predict bacterial infection in decompensated cirrhosis
Full text linkBACKGROUND & AIMS: As portal hypertension progresses in cirrhosis, bacterial translocation across a compromised gut barrier leads to endotoxemia, systemic inflammation and immune dysfunction. Gut phages play a key role in these processes by influencing bacteria-host interactions. This study explores the role of the human gut virome in acute decompensation of cirrhosis and acute-on-chronic liver failure (ACLF). METHODS: The fecal virome was longitudinally assessed by metagenomic sequencing in two independent cohorts: 93 patients (292 samples) with acute decompensation or ACLF from the PREDICT study, and 94 patients (94 samples) with decompensated cirrhosis undergoing TIPS (transjugular intrahepatic portosystemic shunt) surgery collected in a tertiary care setting. Besides descriptive analysis, phages were grouped according to their predicted bacterial host and lifestyle, and associated with clinical parameters. RESULTS: Phage alpha-diversity was higher in patients with ACLF and correlated with ACLF severity. In the absence of ACLF, the phageome was dominated by virulent phages, but in ACLF, temperate phages became more prevalent. Genus-level analysis showed that phageomes were highly patient-specific. Lactococcus A phages were the only phage-host group predicting ACLF development (odds ratio [OR] = 14; Fisher test p = 0.0129). Enterococcus B phages (OR = 14.7; p = 0.0015; adj. p = 0.037) and their bacterial hosts (OR = 2.8; p = 0.020) were significantly more prevalent in cases of proven systemic bacterial infection. The presence of both phage families was linked to increased 90-day mortality rates. CONCLUSION: ACLF is characterized by increased fecal virome diversity and a shift from virulent toward temperate phages at disease onset. Our study links Lactococcus A phages to ACLF development, and Enterococcus B phages to bacterial infection, while both are associated with increased 90-day mortality
Mitigation and detection of putative microbial contaminant reads from long-read metagenomic datasets
Full text linkMetagenomic sequencing of clinical samples has significantly enhanced our understanding of microbial communities. However, microbial contamination and host-derived DNA remain a major obstacle to accurate data interpretation. Here, we present a methodology called 'Stop-Check-Go' for detecting and mitigating contaminants in metagenomic datasets obtained from neonatal patient samples (nasal and rectal swabs). This method incorporates laboratory and bioinformatics work combining a prevalence method, coverage estimation and microbiological reports. We compared the 'Stop-Check-Go' decontamination system with other published decontamination tools and commonly found poor performance in decontaminating microbiologically negative patients (false positives). We emphasize that host DNA decreased by an average of 76% per sample using a lysis method and was further reduced during post-sequencing analysis. Microbial species were classified as putative contaminants and assigned to 'Stop' in nearly 60% of the dataset. The 'Stop-Check-Go' system was developed to address the specific need of decontaminating low-biomass samples, where existing tools primarily designed for short-read metagenomic data showed limited performance