Biology subjects

Duarte, I. F.

Publications and source records attributed to Duarte, I. F..

5 recordsLinked to original sources

bioRxiv2024

Distinct metabolic profiles of circulating plasmacytoid dendritic cells in systemic sclerosis patients stratified by clinical phenotypes

BackgroundPlasmacytoid dendritic cells (pDCs) play a key role in systemic sclerosis (SSc) pathophysiology. However, despite the recognised importance of metabolic reprogramming for pDC function, their metabolic profile in SSc remains to be elucidated. Thus, our study aimed to investigate the role of pDC metabolism in SSc. MethodsPeripheral blood mononuclear cells (PBMCs) were isolated from the blood of healthy donors and SSc patients. SCENITH, a single-cell flow cytometry-based method, was applied to infer the metabolic profile of circulating pDCs from patients with SSc. pDCs (CD304+ Lin-) at steady-state or stimulated with CpG A were analysed. Toll-like receptor (TLR)9 activation was confirmed by ribosomal protein S6 phosphorylation. ResultsCirculating pDCs from 10 healthy donors and 14 SSc patients were analysed. pDCs from anti-centromere antibody-positive (ACA+) patients displayed higher mitochondrial dependence and lower glycolytic capacity than those from anti-topoisomerase I antibody- positive (ATA+) patients. Furthermore, cells from both ACA+ patients and limited cutaneous SSc (lcSSc) patients showed a stronger response towards TLR9 activation than cells from ATA+, anti-RNA polymerase III antibody-positive (ARA+) or diffuse cutaneous SSc (dcSSc) patients. ConclusionsOur results show that pDCs from ACA+ patients rely more on oxidative phosphorylation (OXPHOS) and are more responsive to external stimuli, suggesting that pDCs from ATA+ patients may be more activated or exhausted. These findings point for the possible contribution of pDC metabolism to SSc clinical course, unveiling new potential targets for therapeutic approaches in SSc.

immunology↗

bioRxiv2024

VPS34-IN1 inhibits cap-mediated translation and synergizes with STING to drive type-I IFN expression in human plasmacytoid DCs

Inhibition of the phosphatidylinositol kinase vacuolar protein sorting 34 (VPS34) with the pharmacological compound VPS34-IN1 has a range of effects on the dynamics of endosomes. While VPS34 inhibition has been suggested as a potential therapeutic approach for treating certain cancers, our findings indicate that it has minimal cytotoxic effects on leukemic blastic plasmacytoid dendritic cell neoplasms (BPDCN). VPS34-IN1, however, interferes with plasmacytoid dendritic cells (pDCs) function by blocking the recruitment of serum and glucocorticoid-regulated kinase 3 (SGK3) to endosomes, which is shown to be necessary for Toll-like receptor 7 (TLR7) signaling. In a contrasting parallel, VPS34-IN1 triggers the activation of the stimulator of interferon genes (STING) and significantly enhances pDCs response to the STING agonist 23-cyclic guanosine monophosphate-adenosine monophosphate (23-cGAMP). This cooperative action with VPS34-IN1 leads to strongly increased expression of type-I interferons (IFNs), associated with an alteration of STING degradation and importantly, inhibition of cap-mediated mRNA translation. Inhibition of protein synthesis by VPS34-IN1 appears to be central to this synergy with STING activation, notably by compromising the expression of IFIT1/ISG56, a negative regulator of innate signaling. Thus, despite their limited toxicity towards different cancer lines, inhibitors targeting VPS34 and SGK3 may present promising compounds for controlling the expression of type-I IFNs in response to various microbial stimuli and pathological contexts. One-sentence summaryPharmacological inhibition of VPS34 affects multiple signaling pathways downstream of innate immunity receptors and consequently can inhibit or potentiate type-I Interferon induction according to the danger or microbial stimuli received by plasmacytoid DCs.

immunology↗

bioRxiv2024

Genetically diverse Mycobacterium tuberculosis isolates manipulate inflammasome activation and IL-1β secretion independently of macrophage metabolic rewiring

The natural diversity of Mycobacterium tuberculosis is gaining relevance in dictating the outcome of tuberculosis (TB). We previously revealed a link between TB severity and M. tuberculosis-driven evasion of the macrophage cytosolic surveillance systems, with isolates from severe TB cases reducing inflammasome activation and interleukin (IL)-1{beta} production by infected cells. IL-1{beta} production and inflammasome activation are commonly associated with the metabolic reprogramming of stimulated macrophages. Thus, we questioned whether the differential modulation of the inflammasome and IL-1{beta} by M. tuberculosis isolates depended on distinct macrophage metabolic reprogramming. Using metabolic inhibitors, mice deficient for key metabolic regulators, and a metabolomics approach, we found that the macrophage metabolic landscape was similar regardless of the infecting M. tuberculosis isolate. Paralleling single-TLR activated macrophages, inhibition of glycolysis during infection impaired IL-1{beta} secretion. However, departing from TLR based models, in M. tuberculosis-infected macrophages IL-1{beta} secretion was independent of macrophage mitochondrial metabolic changes and the transcription factor hypoxia-inducible factor (HIF)-1. Additionally, we found a previously unappreciated impact of host metabolic inhibitors on the pathogen, and show that inhibition of the mycobacteria metabolism dampened both inflammasome activation and IL-1{beta} production. Collectively, our study raises awareness of the potential confounding effect of host metabolic inhibitors acting on the pathogen itself and demonstrates that the modulation of the inflammasome by M. tuberculosis may be uncoupled from the host metabolic reprogramming. Author SummaryMycobacterium tuberculosis is the causative agent of tuberculosis and one of the top infectious killers in the world, with around 1.3 million deaths reported annually. The genetic variability of this pathogen can shape its interaction with the host and modulate disease outcomes. We previously found that M. tuberculosis clinical isolates from patients with severe forms of tuberculosis evade cytosolic surveillance systems in macrophages. Here, we explored whether this evasion tactic was linked to metabolic alterations in the infected macrophages. We found that different M. tuberculosis isolates induced similar metabolic changes in infected macrophages. Additionally, we demonstrate that both host glycolysis and pathogens metabolism were pivotal for maximum IL-1{beta} production. These findings highlight the complexity of macrophage-pathogen interactions and emphasize that bacterial metabolism should be considered in metabolic studies and may be amenable to therapeutic intervention against tuberculosis.

immunology↗

bioRxiv2022

Camel urine limits proliferation and modifies cell morphology in human renal tumoral and non-tumoral cells

The widespread ethnomedical practice of dromedary urinotherapy as a remedy against various illnesses is well recognized in traditional dromedary countries, and multiple researchers tried to unravel its bioactive potential and provide scientific evidence through in vivo and in vitro experiments. None of these studies (i) measured urine osmolarity prior to bioactivity testing, which could deeply influence the results of in vitro tests, nor (ii) addressed issues related to cells morphological changes after exposure to camel urines. Taken together, the above aspects point to the need for a "good practice" to be shared by researchers in this field, in order to reduce the variability of in vitro testing of camel urine bioactivity. In this work, using a set of biological samples from animals differing in sex, age, and physiological status, we investigated, the antiproliferative activity of camel urine towards human non-tumoral (HK2) and tumoral renal cells (Caki-1), through cell viability and microscopy analysis, and taking the possible influence of osmolarity into account. We employed cell lines commonly used in toxicological research which, to the best of our knowledge, have not been previously exposed to camel urine. HK2 and Caki-1 cells tolerated well mannitol-induced hyperosmolarity up to 500 mOsm/L. Significant antiproliferative effects were observed only in Caki-1 cells, when exposed to urine solutions (diluted to <500 mOsm/L) from two males out of the ten tested samples, while effects on cell morphology (elongation) were observed only in HK2 cells, when exposed to urine solutions from six samples. The significant antiproliferative effect observed only in tumoral cells looks promising for forthcoming developments in the cancer treatment field. Finally, the presented approach may serve as a guide for future research in this specific, multidisciplinary field.

cell biology↗

bioRxiv2021

Aspergillus fumigatus acetate utilisation impacts virulence traits and pathogenicity

Aspergillus fumigatus is a major opportunistic fungal pathogen of immunocompromised and immunocompetent hosts. To successfully establish an infection, A. fumigatus needs to use host carbon sources, such as acetate, present in the body fluids and peripheral tissues. However, utilisation of acetate as a carbon source by fungi in the context of infection has not been investigated. This work shows that acetate is metabolised via different pathways in A. fumigatus and that acetate utilisation is under the regulatory control of a transcription factor (TF), FacB. A. fumigatus acetate utilisation is subject to carbon catabolite repression (CCR), although this is only partially dependent on the TF and main regulator of CCR CreA. The available extracellular carbon source, in this case glucose and acetate, significantly affected A. fumigatus virulence traits such as secondary metabolite secretion and cell wall composition, with the latter having consequences for resistance to oxidative stress, to anti-fungal drugs and to human neutrophil-mediated killing. Furthermore, deletion of facB significantly impaired the in vivo virulence of A. fumigatus in both insect and mammalian models of invasive aspergillosis. This is the first report on acetate utilisation in A. fumigatus and this work further highlights the importance of available host-specific carbon sources in shaping fungal virulence traits and subsequent disease outcome, and a potential target for the development of anti-fungal strategies. ImportanceAspergillus fumigatus is an opportunistic fungal pathogen in humans. During infection, A. fumigatus is predicted to use host carbon sources, such as acetate, present in body fluids and peripheral tissues, to sustain growth and promote colonisation and invasion. This work shows that A. fumigatus metabolises acetate via different pathways, a process that is dependent on the transcription factor FacB. Furthermore, the type and concentration of the extracellular available carbon source were determined to shape A. fumigatus virulence determinants such as secondary metabolite secretion and cell wall composition. Subsequently, interactions with immune cells are altered in a carbon source-specific manner. FacB is required for A. fumigatus in vivo virulence in both insect and mammalian models of invasive aspergillosis. This is the first report that characterises acetate utilisation in A. fumigatus and highlights the importance of available host-specific carbon sources in shaping virulence traits and potentially subsequent disease outcome.

microbiology↗