Extracellular adenosine oppositely regulates the purinome machinery in glioblastoma and mesenchymal stem cells.

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.

Blood oxygenation state in COVID-19 patients: Unexplored role of 2,3-bisphosphoglycerate.

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 (R = 0.4342; and R = 0.3641, respectively). Furthermore, patients with lower PaO2 showed an imbalanced acid-base equilibrium (pH, p = 0.0208; PaCO2, p = 0.0496) and a higher 2,3-BPG levels (p = 0.0221). The 2,3-BPG levels were also lower in patients with metabolic alkalosis (p = 0.0012 vs. no alkalosis; and p = 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.

Interaction of graphene and WS2 with neutrophils and mesenchymal stem cells: implications for peripheral nerve regeneration.

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.

Apolipoprotein E ε4 triggers neurotoxicity via cholesterol accumulation, acetylcholine dyshomeostasis, and PKCε mislocalization in cholinergic neuronal cells.

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.