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Glioblastoma (GB) is a lethal brain tumor that rapidly adapts to the dynamic changes of the tumor microenvironment (TME). Mesenchymal stem/stromal cells (MSCs) are one of the stromal components of the TME playing multiple roles in tumor progression. GB progression is prompted by the immunosuppressive microenvironment characterized by high concentrations of the nucleoside adenosine (ADO). ADO acts as a signaling molecule through adenosine receptors (ARs) but also as a genetic and metabolic regulator. Herein, the effects of high extracellular ADO concentrations were investigated in a human glioblastoma cellular model (U343MG) and MSCs. The modulation of the purinome machinery, i.e., the ADO production (CD39, CD73, and adenosine kinase [ADK]), transport (equilibrative nucleoside transporters 1 (ENT1) and 2 (ENT2)), and degradation (adenosine deaminase [ADA]) were investigated in both cell lines to evaluate if ADO could affect its cell management in a positive or negative feed-back loop. Results evidenced a different behavior of GB and MSC cells upon exposure to high extracellular ADO levels: U343MG were less sensitive to the ADO concentration and only a slight increase in ADK and ENT1 was evidenced. Conversely, in MSCs, the high extracellular ADO levels reduced the ADK, ENT1, and ENT2 expression, which further sustained the increase of extracellular ADO. Of note, MSCs primed with the GB-conditioned medium or co-cultured with U343MG cells were not affected by the increase of extracellular ADO. These results evidenced how long exposure to ADO could produce different effects on cancer cells with respect to MSCs, revealing a negative feedback loop that can support the GB immunosuppressive microenvironment. These results improve the knowledge of the ADO role in the maintenance of TME, which should be considered in the development of therapeutic strategies targeting adenosine pathways as well as cell-based strategies using MSCs.
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Multinuclear complexes are metal compounds featured by adjacent bound metal centers that can lead to unconventional reactivity. Some M2L4-type paddlewheel dinuclear complexes with monoanionic bridging ligands feature promising properties, including therapeutic ones. Molybdenum has been studied for the formation of multiple-bonded M2+ compounds due to their unique scaffold, redox, and spectroscopic properties as well as for applications in several fields including catalysis and biology. These latter are much less explored and only sporadic studies have been carried out. Here, a series of four dimolybdenum (II,II) carboxylate paddlewheel complexes were synthesized using different Non-Steroidal Anti-Inflammatory Drugs (NSAIDs) as ligands. The reaction of (NH4)5[Mo2Cl9]·H2O with the selected NSAIDs in methanol produced the complexes Mo2(µ-O2CR)4 where RCO2 is ibuprofen (1), naproxen (2), aspirin (3) and indomethacin (4). The products were obtained in good yields and extensively characterized with integrated techniques. Stability and solution behaviour were studied using a mixed experimental and computational approach. Finally, the biological activity of 1 and 3 (i.e. the most reactive and the most stable compounds of the series, respectively) was preliminarily assessed confirming the disassembling of the molecules in the biological milieu. Overall, some very interesting results emerged for these unconventional compounds from a mechanistic point of view.
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BACKGROUND: Internally displaced people (IDP) in Iraq are 1.2 million (as March 2023). Protracted refugee status endangers the mental health, especially of minorities who survived persecution and conflict, such as the Yazidis. This study aims to identify the mental health needs of Yazidi adolescents and young adults (AYA) in the IDP camp of Bajed Kandala (Iraqi Kurdistan). METHODS: A focus group discussion (FGD) study was conducted between April and August 2022. The FGDs involved AYAs, as well as the staff of the clinic of the Bajed Kandala camp. An inductive approach was adopted referring to the 'theme' as the unit of content analysis of the text. All FGDs were recorded and transcribed. The analysis was carried out independently by two researchers. The inter-rater agreement was assessed through the Cohen's k. RESULTS: A total of 6 FGDs were conducted. The participants were 34 of whom 21 (61.8%) females with a median age of 18.5 years (IQR 17.0-21.0). A total of 156 themes were found as relevant to the objective of this study. Four main areas and twelve subareas of needs in mental health were identified. The interrater agreement over the main area and subareas was good (κ = 0.78 [0.95CI 0.69-0.88], κ = 0.82 [0.95CI 0.73-0.91], respectively). The four areas had a similar frequency: Activities (28.2%), Individual (27.6%), Social relationships (22.4%) and Places/setting (21.8%). The subareas 'community' and 'internal resources' were labelled as negative 85.7% and 61.9% of the time, respectively. These sub-areas referred to stigma and self-stigma towards mental health. The subarea 'female condition' was always considered as negative, as well as the subareas 'camp' and 'tent' referring to housing as an important social determinant of mental health. CONCLUSIONS: Community stigma and self-stigma are two still important factors preventing the achievement of mental well-being. Alongside these, a gender gap in mental health was identified in the FGDs. These factors should be taken into account in order to guide future mental health interventions in refugee camps.
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Grupos Focais , Saúde Mental , Refugiados , Humanos , Feminino , Adolescente , Masculino , Iraque , Adulto Jovem , Refugiados/psicologia , Avaliação das Necessidades , Adulto , Necessidades e Demandas de Serviços de SaúdeRESUMO
Mature oligodendrocytes (OLs) arise from oligodendrocyte precursor cells that, in case of demyelination, are recruited at the lesion site to remyelinate the axons and therefore restore the transmission of nerve impulses. It has been widely documented that exogenously administered steroid molecules are potent inducers of myelination. However, little is known about how neurosteroids produced de novo by OLs can impact this process. Here, we employed a human OL precursor cell line to investigate the role of de novo neurosteroidogenesis in the regulation of OLs differentiation, paying particular attention to the 18 kDa Translocator Protein (TSPO) which controls the rate-limiting step of the neurosteroidogenic process. Our results showed that, over the time of OL maturation, the availability of cholesterol, which is the neurosteroidogenesis initial substrate, and key members of the neurosteroidogenic machinery, including TSPO, were upregulated. In addition, OLs differentiation was impaired following neurosteroidogenesis inhibition and TSPO silencing. On the contrary, TSPO pharmacological stimulation promoted neurosteroidogenic function and positively impacted differentiation. Collectively, our results suggest that de novo neurosteroidogenesis is actively involved in the autocrine and paracrine regulation of human OL differentiation. Moreover, since TSPO was able to promote OL differentiation through a positive modulation of the neurosteroid biosynthetic process, it could be exploited as a promising target to tackle demyelinating diseases.
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Diferenciação Celular , Oligodendroglia , Receptores de GABA , Humanos , Receptores de GABA/metabolismo , Receptores de GABA/genética , Oligodendroglia/metabolismo , Oligodendroglia/efeitos dos fármacos , Oligodendroglia/citologia , Diferenciação Celular/efeitos dos fármacos , Neuroesteroides/metabolismo , Colesterol/metabolismo , Colesterol/biossíntese , Linhagem Celular , Bainha de Mielina/metabolismoRESUMO
BACKGROUND: COVID-19 reduces lung functionality causing a decrease of blood oxygen levels (hypoxemia) often related to a decreased cellular oxygenation (hypoxia). Besides lung injury, other factors are implicated in the regulation of oxygen availability such as pH, partial arterial carbon dioxide tension (PaCO2), temperature, and erythrocytic 2,3-bisphosphoglycerate (2,3-BPG) levels, all factors affecting hemoglobin saturation curve. However, few data are currently available regarding the 2,3-BPG modulation in SARS-CoV-2 affected patients at the hospital admission. MATERIAL AND METHODS: Sixty-eight COVID-19 patients were enrolled at hospital admission. The lung involvement was quantified using chest-Computer Tomography (CT) analysed with automatic software (CALIPER). Haemoglobin concentrations, glycemia, and routine analysis were evaluated in the whole blood, while partial arterial oxygen tension (PaO2), PaCO2, pH, and HCO3- were assessed by arterial blood gas analysis. 2,3-BPG levels were assessed by specific immunoenzymatic assays in RBCs. RESULTS: A higher percentage of interstitial lung disease (ILD) and vascular pulmonary-related structure (VRS) volume on chest-CT quantified with CALIPER had been found in COVID-19 patients with a worse disease outcome (Râ¯=â¯0.4342; and Râ¯=â¯0.3641, respectively). Furthermore, patients with lower PaO2 showed an imbalanced acid-base equilibrium (pH, pâ¯=â¯0.0208; PaCO2, pâ¯=â¯0.0496) and a higher 2,3-BPG levels (pâ¯=â¯0.0221). The 2,3-BPG levels were also lower in patients with metabolic alkalosis (pâ¯=â¯0.0012 vs. no alkalosis; and pâ¯=â¯0.0383 vs. respiratory alkalosis). CONCLUSIONS: Overall, the data reveal a different pattern of activation of blood oxygenation compensatory mechanisms reflecting a different course of the COVID-19 disease specifically focusing on 2,3-BPG modulation.
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The use of platinum-based anticancer drugs, such as cisplatin, oxaliplatin, and carboplatin, is a common frontline option in cancer management, but they have debilitating side effects and can lead to drug resistance. Combination therapy with other chemotherapeutic agents, such as capecitabine and gemcitabine, has been explored. One approach to overcome these limitations is the modification of traditional Pt(II) drugs to obtain new molecules with an improved pharmacological profile, such as Pt(IV) prodrugs. The design, synthesis, and characterization of two novel Pt(IV) prodrugs based on oxaliplatin bearing the anticancer drugs gemcitabine or capecitabine in the axial positions have been reported. These complexes were able to dissociate into their constituents to promote cell death and induce apoptosis and cell cycle blockade in a representative colorectal cancer cell model. Specifically, the complex bearing gemcitabine resulted in being the most active on the HCT116 colorectal cancer cell line with an IC50 value of 0.49 ± 0.04. A pilot study on the encapsulation of these complexes in biocompatible PLGA-PEG nanoparticles is also included to confirm the retention of the pharmacological properties and cellular drug uptake, opening up to the possible delivery of the studied complexes through their nanoformulation.
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Graphene and bidimensional (2D) materials have been widely used in nerve conduits to boost peripheral nerve regeneration. Nevertheless, the experimental and commercial variability in graphene-based materials generates graphene forms with different structures and properties that can trigger entirely diverse biological responses from all the players involved in nerve repair. Herein, we focus on the graphene and tungsten disulfide (WS2) interaction with non-neuronal cell types involved in nerve tissue regeneration. We synthesize highly crystalline graphene and WS2 with scalable techniques such as thermal decomposition and chemical vapor deposition. The materials were able to trigger the activation of a neutrophil human model promoting Neutrophil Extracellular Traps (NETs) production, particularly under basal conditions, although neutrophils were not able to degrade graphene. Of note is that pristine graphene acts as a repellent for the NET adhesion, a beneficial property for nerve conduit long-term applications. Mesenchymal stem cells (MSCs) have been proposed as a promising strategy for nerve regeneration in combination with a conduit. Thus, the interaction of graphene with MSCs was also investigated, and reduced viability was observed only on specific graphene substrates. Overall, the results confirm the possibility of regulating the cell response by varying graphene properties and selecting the most suitable graphene forms.
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Grafite , Células-Tronco Mesenquimais , Humanos , Grafite/química , Neutrófilos , Regeneração NervosaRESUMO
Tendon and ligament injuries are relevant clinical problems in modern society, and the current medical approaches do not guarantee complete recovery of the physiological functionalities. Moreover, they present a non-negligible failure rate after surgery. Failures often occur at the enthesis, which is the area of tendons and ligaments insertion to bones. This area is highly anisotropic and composed of four distinct zones: tendon or ligament, non-mineralized fibrocartilage, mineralized fibrocartilage, and bone. The organization of these regions provides a gradient in mechanical properties, biochemical composition, cellular phenotype, and extracellular matrix organization. Tissue engineering represents an alternative to traditional medical approaches. This work presents a novel biofabrication approach for engineering the enthesis. Gradient-based scaffolds were fabricated by exploiting the combination of electrospinning and three-dimensional (3D) bioprinting technologies. Studies were conducted to evaluate scaffold biocompatibility by seeding bone marrow-derived mesenchymal stem cells (BM-MSCs). Then, the scaffold's ability to promote cellular adhesion, growth, proliferation, and differentiation in both tenogenic and osteogenic phenotypes was evaluated. Fabricated scaffolds were also morphologically and mechanically characterized, showing optimal properties comparable to literature data. The versatility and potentiality of this novel biofabrication approach were demonstrated by fabricating clinical-size 3D enthesis scaffolds. The mechanical characterization highlighted their behavior during a tensile test was comparable to tendons and ligaments in vivo.
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The Apolipoprotein E (ApoE) has been known to regulate cholesterol and ß-amyloid (Aß) production, redistribution, and elimination, in the central nervous system (CNS). The ApoE ε4 polymorphic variant leads to impaired brain cholesterol homeostasis and amyloidogenic pathway, thus representing the major risk factor for Alzheimer's Disease (AD). Currently, less is known about the molecular mechanisms connecting ApoE ε4-related cholesterol metabolism and cholinergic system degeneration, one of the main AD pathological features. Herein, in vitro cholinergic neuron models were developed in order to study ApoE neuronal expression and investigate the possible interplay between cholesterol metabolism and cholinergic pathway impairment prompted by ε4 isoform. Particularly, alterations specifically occurring in ApoE ε4-carrying neurons (i.e. increased intracellular ApoE, amyloid precursor protein (APP) and Aß levels, elevated apoptosis, and reduced cell survival) were recapitulated. ApoE ε4 expression was found to increase intracellular cholesterol accumulation, by regulating the related gene expression, while reducing cholesterol precursor acetyl-CoA, which in turn fuels the acetylcholine (ACh) synthesis route. In parallel, although the ACh intracellular signalling was activated, as demonstrated by the boosted extracellular ACh as well as increased IP3 and Ca2+, the PKCε activation via membrane translocation was surprisingly suppressed, probably explained by the cholesterol overload in ApoE ε4 neuron-like cells. Consequently, the PKC-dependent anti-apoptotic and neuroprotective roles results impaired, reliably adding to other causes of cell death prompted by ApoE ε4. Overall, the obtained data open the way to further critical considerations of ApoE ε4-dependent cholesterol metabolism dysregulation in the alteration of cholinergic pathway, neurotoxicity, and neuronal death.
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Doença de Alzheimer , Apolipoproteína E4 , Humanos , Acetilcolina , Doença de Alzheimer/metabolismo , Apolipoproteína E4/genética , Apolipoproteínas E/genética , Colesterol , Colinérgicos , Neurônios/metabolismo , Proteína Quinase C-épsilon/metabolismoRESUMO
Obstructive sleep apnoea syndrome (OSAS) is a sleep-disordered breathing characterized by nocturnal collapses of the upper airway resulting in cycles of blood oxygen partial pressure oscillations, which lead to tissue and cell damage due to intermittent hypoxia (IH) episodes. Since OSAS-derived IH may lead to cognitive impairment through not fully cleared mechanisms, herein we developed a new in vitro model mimicking IH conditions to shed light on its molecular effects on microglial cells, with particular attention to the inflammatory response. The in vitro model was set-up and validated by measuring the hypoxic state, HIF-1α levels, oxidative stress by ROS production and mitochondrial activity by MTS assay. Then, the mRNA and protein levels of certain inflammatory markers (NF-κB and interleukin 6 (IL-6)) after different IH treatment protocols were investigated. The IH treatments followed by a normoxic period were not able to produce a high inflammatory state in human microglial cells. Nevertheless, microglia appeared to be in a state characterized by increased expression of NF-κB and markers related to a primed phenotype. The microglia exposed to IH cycles and stimulated with exogenous IL-1ß resulted in an exaggerated inflammatory response with increased NF-κB and IL-6 expression, suggesting a role for primed microglia in OSAS-driven neuroinflammation.
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Microglia , Apneia Obstrutiva do Sono , Humanos , Microglia/metabolismo , Interleucina-6/metabolismo , NF-kappa B/metabolismo , Hipóxia/metabolismo , Apneia Obstrutiva do Sono/metabolismoRESUMO
Emerging nanotechnologies offer numerous opportunities in the field of regenerative medicine and have been widely explored to design novel scaffolds for the regeneration and stimulation of nerve tissue. In this review, we focus on peripheral nerve regeneration. First, we introduce the biomedical problem and the present status of nerve conduits that can be used to guide, fasten and enhance regeneration. Then, we thoroughly discuss graphene as an emerging candidate in nerve tissue engineering, in light of its chemical, tribological and electrical properties. We introduce the graphene forms commonly used as neural interfaces, briefly review their applications, and discuss their potential toxicity. We then focus on the adoption of graphene in peripheral nervous system applications, a research field that has gained in the last years ever-increasing attention. We discuss the potential integration of graphene in guidance conduits, and critically review graphene interaction not only with peripheral neurons, but also with non-neural cells involved in nerve regeneration; indeed, the latter have recently emerged as central players in modulating the immune and inflammatory response and accelerating the growth of new tissue.
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Intracellular pathogens, such as Chlamydia trachomatis, have been recently shown to induce degradation of p53 during infection, thus impairing the protective response of the host cells. Therefore, p53 reactivation by disruption of the p53-MDM2 complex could reduce infection and restore pro-apoptotic effect of p53. Here, we report the identification of a novel MDM2 inhibitor with potential antitumoural and antibacterial activity able to reactivate p53. A virtual screening was performed on an in-house chemical library, previously synthesised for other targets, and led to the identification of a hit compound with a benzo[a]dihydrocarbazole structure, RM37. This compound induced p53 up-regulation in U343MG glioblastoma cells by blocking MDM2-p53 interaction and reduced tumour cell growth. NMR studies confirmed its ability to dissociate the MDM2-p53 complex. Notably, RM37 reduced Chlamydia infection in HeLa cells in a concentration-dependent manner and ameliorated the inflammatory status associated with infection.
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Brain-derived neurotrophic factor (BDNF) is a neurotrophin (NT) essential for neuronal development and synaptic plasticity. Dysregulation of BDNF signaling is implicated in different neurological disorders. The direct NT administration as therapeutics has revealed to be challenging. This has prompted the design of peptides mimicking different regions of the BDNF structure. Although loops 2 and 4 have been thoroughly investigated, less is known regarding the BDNF N-terminal region, which is involved in the selective recognition of the TrkB receptor. Herein, a dimeric form of the linear peptide encompassing the 1-12 residues of the BDNF N-terminal (d-bdnf) was synthesized. It demonstrated to act as an agonist promoting specific phosphorylation of TrkB and downstream ERK and AKT effectors. The ability to promote TrkB dimerization was investigated by advanced fluorescence microscopy and molecular dynamics (MD) simulations, finding activation modes shared with BDNF. Furthermore, d-bdnf was able to sustain neurite outgrowth and increase the expression of differentiation (NEFM, LAMC1) and polarization markers (MAP2, MAPT) demonstrating its neurotrophic activity. As TrkB activity is affected by zinc ions in the synaptic cleft, we first verified the ability of d-bdnf to coordinate zinc and then the effect of such complexation on its activity. The d-bdnf neurotrophic activity was reduced by zinc complexation, demonstrating the role of the latter in tuning the activity of the new peptido-mimetic. Taken together our data uncover the neurotrophic properties of a novel BDNF mimetic peptide and pave the way for future studies to understand the pharmacological basis of d-bdnf action and develop novel BDNF-based therapeutic strategies.
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Fator Neurotrófico Derivado do Encéfalo , Zinco , Zinco/farmacologiaRESUMO
A novel gold(I) complex inspired by the known medicinal inorganic compounds auranofin and thimerosal, namely ethylthiosalicylate(triethylphosphine)gold(I) (AFETT hereafter), was synthesized and characterised and its structure was resolved through X-ray diffraction. The solution behavior of AFETT and its interactions with two biologically relevant proteins (i.e. human serum albumin and haemoglobin) and with a synthetic dodecapeptide reproducing the C-terminal portion of thioredoxin reductase were comparatively analyzed through 31P NMR and ESI-MS. Remarkable binding properties toward these biomolecules were disclosed. Moreover, the cytotoxic effects produced by AFETT on two ovarian cancer cell lines (A2780 and A2780 R) and one colorectal cancer cell line (HCT116) were analyzed and found to be strong and nearly superimposable to those of auranofin. Interestingly, for both compounds, the ability to induce downregulation of vimentin expression in A2780 R cells was evidenced. Despite its close similarity to auranofin, AFETT is reported to exhibit some peculiar and distinctive features such as a lower lipophilicity, an increased water solubility and a faster reactivity towards the selected target biomolecules. These differences might confer to AFETT significant pharmaceutical and therapeutic advantages over auranofin itself.
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Antineoplásicos , Neoplasias Ovarianas , Antineoplásicos/química , Auranofina/química , Auranofina/farmacologia , Linhagem Celular Tumoral , Feminino , Ouro/química , HumanosRESUMO
Microglial cells are a component of the innate immune system in the brain that support cell-to-cell communication via secreted molecules and extracellular vesicles (EVs). EVs can be divided into two major populations: large (LEVs) and small (SEVs) EVs, carrying different mediators, such as proteins, lipids, and miRNAs. The microglia EVs cargo crucially reflects the status of parental cells and can lead to both beneficial and detrimental effects in many physiopathological states. Herein, a workflow for the extraction and characterization of SEVs and LEVs from human C20 and HMC3 microglia cell lines derived, respectively, from adult and embryonic microglia is reported. EVs were gathered from the culture media of the two cell lines by sequential ultracentrifugation steps and their biochemical and biophysical properties were analyzed by Western blot, transmission electron microscopy, and dynamic light scattering. Although the C20- and HMC3-derived EVs shared several common features, C20-derived EVs were slightly lower in number and more polydispersed. Interestingly, C20- but not HMC3-SEVs were able to interfere with the proliferation of U87 glioblastoma cells. This correlated with the different relative levels of eight miRNAs involved in neuroinflammation and tumor progression in the C20- and HMC3-derived EVs, which in turn reflected a different basal activation state of the two cell types. Our data fill a gap in the community of microglia EVs, in which the preparations from human cells have been poorly characterized so far. Furthermore, these results shed light on both the differences and similarities of EVs extracted from different human microglia cell models, underlining the need to better characterize the features and biological effects of EVs for therein useful and correct application.
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BACKGROUND: Triple negative breast cancer (TNBC) is the most aggressive subtype of breast cancer (BC). Treatment options for TNBC patients are limited and further insights into disease aetiology are needed to develop better therapeutic approaches. microRNAs' ability to regulate multiple targets could hold a promising discovery approach to pathways relevant for TNBC aggressiveness. Thus, we address the role of miRNAs in controlling three signalling pathways relevant to the biology of TNBC, and their downstream phenotypes. METHODS: To identify miRNAs regulating WNT/ß-catenin, c-Met, and integrin signalling pathways, we performed a high-throughput targeted proteomic approach, investigating the effect of 800 miRNAs on the expression of 62 proteins in the MDA-MB-231 TNBC cell line. We then developed a novel network analysis, Pathway Coregulatory (PC) score, to detect miRNAs regulating these three pathways. Using in vitro assays for cell growth, migration, apoptosis, and stem-cell content, we validated the function of candidate miRNAs. Bioinformatic analyses using BC patients' datasets were employed to assess expression of miRNAs as well as their pathological relevance in TNBC patients. RESULTS: We identified six candidate miRNAs coordinately regulating the three signalling pathways. Quantifying cell growth of three TNBC cell lines upon miRNA gain-of-function experiments, we characterised miR-193b as a strong and consistent repressor of proliferation. Importantly, the effects of miR-193b were stronger than chemical inhibition of the individual pathways. We further demonstrated that miR-193b induced apoptosis, repressed migration, and regulated stem-cell markers in MDA-MB-231 cells. Furthermore, miR-193b expression was the lowest in patients classified as TNBC or Basal compared to other subtypes. Gene Set Enrichment Analysis showed that miR-193b expression was significantly associated with reduced activity of WNT/ß-catenin and c-Met signalling pathways in TNBC patients. CONCLUSIONS: Integrating miRNA-mediated effects and protein functions on networks, we show that miRNAs predominantly act in a coordinated fashion to activate or repress connected signalling pathways responsible for metastatic traits in TNBC. We further demonstrate that our top candidate, miR-193b, regulates these phenotypes to an extent stronger than individual pathway inhibition, thus emphasizing that its effect on TNBC aggressiveness is mediated by the coordinated repression of these functionally interconnected pathways.
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MicroRNAs/metabolismo , Proteínas Proto-Oncogênicas c-met/metabolismo , Neoplasias de Mama Triplo Negativas/genética , Via de Sinalização Wnt/genética , beta Catenina/metabolismo , Linhagem Celular Tumoral , Proliferação de Células , Humanos , Metástase Neoplásica , TransfecçãoRESUMO
Pulmonary fibrosis (PF) is characterised by several grades of chronic inflammation and collagen deposition in the interalveolar space and is a hallmark of interstitial lung diseases (ILDs). Recently, infectious agents have emerged as driving causes for PF development; however, the role of viral/bacterial infections in the initiation and propagation of PF is still debated. In this context, the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the virus responsible for the current coronavirus disease 2019 (COVID-19) pandemic, has been associated with acute respiratory distress syndrome (ARDS) and PF development. Although the infection by SARS-CoV-2 can be eradicated in most cases, the development of fibrotic lesions cannot be precluded; furthermore, whether these lesions are stable or progressive fibrotic events is still unknown. Herein, an overview of the main molecular mechanisms driving the fibrotic process together with the currently approved and newly proposed therapeutic solutions was given. Then, the most recent data that emerged from post-COVID-19 patients was discussed, in order to compare PF and COVID-19-dependent PF, highlighting shared and specific mechanisms. A better understanding of PF aetiology is certainly needed, also to develop effective therapeutic strategies and COVID-19 pathology is offering one more chance to do it. Overall, the work reported here could help to define new approaches for therapeutic intervention in the diversity of the ILD spectrum.
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COVID-19/complicações , Fibrose Pulmonar/imunologia , Fibrose Pulmonar/metabolismo , Animais , COVID-19/etiologia , COVID-19/imunologia , COVID-19/metabolismo , Humanos , Mediadores da Inflamação/imunologia , Mediadores da Inflamação/metabolismo , Neutrófilos/imunologia , Neutrófilos/metabolismo , Fibrose Pulmonar/etiologia , Síndrome de COVID-19 Pós-AgudaRESUMO
BACKGROUND: Regulation of protein output at the level of translation allows for a rapid adaptation to dynamic changes to the cell's requirements. This precise control of gene expression is achieved by complex and interlinked biochemical processes that modulate both the protein synthesis rate and stability of each individual mRNA. A major factor coordinating this regulation is the Ccr4-Not complex. Despite playing a role in most stages of the mRNA life cycle, no attempt has been made to take a global integrated view of how the Ccr4-Not complex affects gene expression. RESULTS: This study has taken a comprehensive approach to investigate post-transcriptional regulation mediated by the Ccr4-Not complex assessing steady-state mRNA levels, ribosome position, mRNA stability, and protein production transcriptome-wide. Depletion of the scaffold protein CNOT1 results in a global upregulation of mRNA stability and the preferential stabilization of mRNAs enriched for G/C-ending codons. We also uncover that mRNAs targeted to the ER for their translation have reduced translational efficiency when CNOT1 is depleted, specifically downstream of the signal sequence cleavage site. In contrast, translationally upregulated mRNAs are normally localized in p-bodies, contain disorder-promoting amino acids, and encode nuclear localized proteins. Finally, we identify ribosome pause sites that are resolved or induced by the depletion of CNOT1. CONCLUSIONS: We define the key mRNA features that determine how the human Ccr4-Not complex differentially regulates mRNA fate and protein synthesis through a mechanism linked to codon composition, amino acid usage, and mRNA localization.
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Regulação da Expressão Gênica , Biossíntese de Proteínas , Estabilidade de RNA , RNA Mensageiro/metabolismo , Fatores de Transcrição/fisiologia , Códon , Técnicas de Silenciamento de Genes , Humanos , Proteínas Nucleares/biossíntese , Proteínas Nucleares/genética , Ribossomos/metabolismo , Fatores de Transcrição/genéticaRESUMO
Microglia are the major component of the innate immune system in the central nervous system. They promote the maintenance of brain homeostasis as well as support inflammatory processes that are often related to pathological conditions such as neurodegenerative diseases. Depending on the stimulus received, microglia cells dynamically change their phenotype releasing specific soluble factors and largely modify the cargo of their secreted extracellular vesicles (EVs). Despite the mechanisms at the basis of microglia actions have not been completely clarified, the recognized functions exerted by their EVs in patho-physiological conditions represent the proof of the crucial role of these organelles in tuning cell-to-cell communication, promoting either protective or harmful effects. Consistently, in vitro cell models to better elucidate microglia EV production and mechanisms of their release have been increased in the last years. In this review, the main microglial cellular models that have been developed and validated will be described and discussed, with particular focus on those used to produce and derive EVs. The advantages and disadvantages of their use will be evidenced too. Finally, given the wide interest in applying EVs in diagnosis and therapy too, the heterogeneity of available models for producing microglia EVs is here underlined, to prompt a cross-check or comparison among them.