Priming adult stem cells by hypoxic pretreatments for applications in regenerative medicine

The efficiency of regenerative medicine can be ameliorated by improving the biological performances of stem cells before their transplantation. Several ex-vivo protocols of non-damaging cell hypoxia have been demonstrated to significantly increase survival, proliferation and post-engraftment differentiation potential of stem cells. The best results for priming cultured stem cells against a following, otherwise lethal, ischemic stress have been obtained with brief intermittent episodes of hypoxia, or anoxia, and reoxygenation in accordance with the extraordinary protection afforded by the conventional maneuver of ischemic preconditioning in severely ischemic organs. These protocols of hypoxic preconditioning can be rather easily reproduced in a laboratory; however, more suitable pharmacological interventions inducing stem cell responses similar to those activated in hypoxia are considered among the most promising solutions for future applications in cell therapy. Here we want to offer an up-to-date review of the molecular mechanisms translating hypoxia into beneficial events for regenerative medicine. To this aim the involvement of epigenetic modifications, microRNAs, and oxidative stress, mainly activated by hypoxia inducible factors, will be discussed. Stem cell adaptation to their natural hypoxic microenvironments (niche) in healthy and neoplastic tissues will be also considered

Strategies Affording Prevascularized Cell-Based Constructs for Myocardial Tissue Engineering

The production of a functional cardiac tissue to be transplanted in the injured area of the infarcted myocardium represents a challenge for regenerative medicine. Most cell-based grafts are unviable because of inadequate perfusion; therefore, prevascularization might be a suitable approach for myocardial tissue engineering. To this aim, cells with a differentiation potential towards vascular and cardiac muscle phenotypes have been cocultured in 2D or 3D appropriate scaffolds. In addition to these basic approaches, more sophisticated strategies have been followed employing mixed-cell sheets, microvascular modules, and inosculation from vascular explants. Technologies exerting spatial control of vascular cells, such as topographical surface roughening and ordered patterning, represent other ways to drive scaffold vascularization. Finally, microfluidic devices and bioreactors exerting mechanical stress have also been employed for high-throughput scaling-up production in order to accelerate muscle differentiation and speeding the endothelialization process. Future research should address issues such as how to optimize cells, biomaterials, and biochemical components to improve the vascular integration of the construct within the cardiac wall, satisfying the metabolic and functional needs of the myocardial tissue

Circulating miR-320b and miR-483-5p levels are associated with COVID-19 in-hospital mortality

none28noThe stratification of mortality risk in COVID-19 patients remains extremely challenging for physicians, especially in older patients. Innovative minimally invasive molecular biomarkers are needed to improve the prediction of mortality risk and better customize patient management. In this study, aimed at identifying circulating miRNAs associated with the risk of COVID-19 in-hospital mortality, we analyzed serum samples of 12 COVID-19 patients by small RNA-seq and validated the findings in an independent cohort of 116 COVID-19 patients by qRT-PCR. Thirty-four significantly deregulated miRNAs, 25 downregulated and 9 upregulated in deceased COVID-19 patients compared to survivors, were identified in the discovery cohort. Based on the highest fold-changes and on the highest expression levels, 5 of these 34 miRNAs were selected for the analysis in the validation cohort. MiR-320b and miR-483-5p were confirmed to be significantly hyper-expressed in deceased patients compared to survived ones. Kaplan-Meier and Cox regression models, adjusted for relevant confounders, confirmed that patients with the 20% highest miR-320b and miR-483-5p serum levels had three-fold increased risk to die during in-hospital stay for COVID-19. In conclusion, high levels of circulating miR-320b and miR-483-5p can be useful as minimally invasive biomarkers to stratify older COVID-19 patients with an increased risk of in-hospital mortality.restrictedGiuliani, Angelica; Matacchione, Giulia; Ramini, Deborah; Di Rosa, Mirko; Bonfigli, Anna Rita; Sabbatinelli, Jacopo; Monsurrò, Vladia; Recchioni, Rina; Marcheselli, Fiorella; Marchegiani, Francesca; Piacenza, Francesco; Cardelli, Maurizio; Galeazzi, Roberta; Pomponio, Giovanni; Ferrarini, Alessia; Gabrielli, Armando; Baroni, Silvia Svegliati; Moretti, Marco; Sarzani, Riccardo; Giordano, Piero; Cherubini, Antonio; Corsonello, Andrea; Antonicelli, Roberto; Procopio, Antonio Domenico; Ferracin, Manuela; Bonafè, Massimiliano; Lattanzio, Fabrizia; Olivieri, FabiolaGiuliani, Angelica; Matacchione, Giulia; Ramini, Deborah; Di Rosa, Mirko; Bonfigli, Anna Rita; Sabbatinelli, Jacopo; Monsurrò, Vladia; Recchioni, Rina; Marcheselli, Fiorella; Marchegiani, Francesca; Piacenza, Francesco; Cardelli, Maurizio; Galeazzi, Roberta; Pomponio, Giovanni; Ferrarini, Alessia; Gabrielli, Armando; Baroni, Silvia Svegliati; Moretti, Marco; Sarzani, Riccardo; Giordano, Piero; Cherubini, Antonio; Corsonello, Andrea; Antonicelli, Roberto; Procopio, Antonio Domenico; Ferracin, Manuela; Bonafè, Massimiliano; Lattanzio, Fabrizia; Olivieri, Fabiol

Peripheral blood cellular profile at pre-lymphodepletion is associated with CD19-targeted CAR-T cell-associated neurotoxicity

BackgroundInfusion of second generation autologous CD19-targeted chimeric antigen receptor (CAR) T cells in patients with R/R relapsed/refractory B-cell lymphoma (BCL) is affected by inflammatory complications, such as Immune Effector Cell-Associated Neurotoxicity Syndrome (ICANS). Current literature suggests that the immune profile prior to CAR-T infusion modifies the chance to develop ICANS.MethodsThis is a monocenter prospective study on 53 patients receiving approved CAR T-cell products (29 axi-cel, 24 tisa-cel) for R/R-BCL. Clinical, biochemical, and hematological variables were analyzed at the time of pre-lymphodepletion (pre-LD). In a subset of 21 patients whose fresh peripheral blood sample was available, we performed cytofluorimetric analysis of leukocytes and extracellular vesicles (EVs). Moreover, we assessed a panel of soluble plasma biomarkers (IL-6/IL-10/GDF-15/IL-15/CXCL9/NfL) and microRNAs (miR-146a-5p, miR-21-5p, miR-126-3p, miR-150-5p) which are associated with senescence and inflammation.ResultsMultivariate analysis at the pre-LD time-point in the entire cohort (n=53) showed that a lower percentage of CD3+CD8+ lymphocytes (38.6% vs 46.8%, OR=0.937 [95% CI: 0.882-0.996], p=0.035) and higher levels of serum C-reactive protein (CRP, 4.52 mg/dl vs 1.00 mg/dl, OR=7.133 [95% CI: 1.796-28], p=0.005) are associated with ICANS. In the pre-LD samples of 21 patients, a significant increase in the percentage of CD8+CD45RA+CD57+ senescent cells (median % value: 16.50% vs 9.10%, p=0.009) and monocytic-myeloid derived suppressor cells (M-MDSC, median % value: 4.4 vs 1.8, p=0.020) was found in ICANS patients. These latter also showed increased levels of EVs carrying CD14+ and CD45+ myeloid markers, of the myeloid chemokine CXCL-9, as well of the MDSC-secreted cytokine IL-10. Notably, the serum levels of circulating neurofilament light chain, a marker of neuroaxonal injury, were positively correlated with the levels of senescent CD8+ T cells, M-MDSC, IL-10 and CXCL-9. No variation in the levels of the selected miRNAs was observed between ICANS and no-ICANS patients.DiscussionOur data support the notion that pre-CAR-T systemic inflammation is associated with ICANS. Higher proportion of senescence CD8+ T cells and M-MDSC correlate with early signs of neuroaxonal injury at pre-LD time-point, suggesting that ICANS may be the final event of a process that begins before CAR-T infusion, consequence to patient clinical history

Nitric oxide regulates multiple functions and fate of adult progenitor and stem cells

Nitric oxide is an endogenous gas which exerts autocrine/paracrine actions by different signaling pathways and/or direct interactions with intracellular compounds and structures. Several processes are regulated by nitric oxide in stem cells including self-renewal, viability, migration, proliferation, and differentiation. The modulation of cell functions depends on its concentrations because opposite effects can be observed when low and high levels of nitric oxide are compared. Here, the responses to nitric oxide of adult stem/progenitor cells which are often used in regenerative medicine, including mesenchymal stem cells, hematopoietic stem cells, neural stem cells, endothelial progenitor cells, satellite cells, and fibro-adipogenic precursor cells, are reviewed. Therapeutic strategies which employ drugs releasing nitric oxide or modulating nitric oxide intracellular pathways are suggested to perform new ex vivo preconditioning or in vivo treatments suitable for stem/progenitor cell therapy and tissue engineering applications

Cancer stem cells and mesenchymal stem cells in the hypoxic tumor niche: Two different targets for one only drug

Putative cancer stem cells (CSCs) reside in a hypoxic microenvironment where mesenchymal stem cells (MSCs) are also present. In this niche MSCs seem to promote the generation of CSCs and sustain tumor progression. Therefore, it may assume clinical relevance to produce a drug which kills not only CSCs but also MSCs. We hypothesized that bifunctional nanoparticles, loaded with a HIF-1 alpha inhibitor and conjugated with an aptamer targeting a common receptor of CSCs and MSCs, may fulfill this strategy. The nanoparticle should ensure that: (1) the conveyed drug is less susceptible to degradation, (2) the common receptor of CSCs and MSCs is recognized by a superselective aptamer, and (3) receptor-mediated internalization is the main process to enter target cells. Small RNA or DNA aptamers represent an advantage over antibodies because do not cause immune reactions, are better internalized into the target cell, are more resistant to degradation, their cost of production are lower, and the purity of the oligonucleotide ligand is extremely elevated. Concerning the drugs to be delivered, we suggest to employ those exerting an anti-HIF-1 alpha activity because they should be harmful for hypoxic CSCs and MCSs in their tumor niche but provide very limited toxicity, or even none, to well-oxygenated normal cells. Corresponding experimental approaches to perform pre-clinical studies and verify this hypothesis are also addressed

The role of extracellular DNA in COVID-19: clues from inflamm-aging

Epidemiological data convey severe prognosis and high mortality rate for COVID-19 in elderly men affected by age-related diseases. These subjects develop local and systemic hyper-inflammation, which are associated with thrombotic complications and multi-organ failure. Therefore, understanding SARS-CoV-2 induced hyper-inflammation in elderly men is a pressing need. Here we focus on the role of extracellular DNA, mainly mitochondrial DNA (mtDNA) and telomeric DNA (telDNA) in the modulation of systemic inflammation in these subjects. In particular, extracellular mtDNA is regarded as a powerful trigger of the inflammatory response. On the contrary, extracellular telDNA pool is estimated to be capable of inhibiting a variety of inflammatory pathways. In turn, we underpin that telDNA reservoir is progressively depleted during aging, and that it is scarcer in men than in women. We propose that an increase in extracellular mtDNA, concomitant with the reduction of the anti-inflammatory telDNA reservoir may explain hyper-inflammation in elderly male affected by COVID-19. This scenario is reminiscent of inflamm-aging, the portmanteau word that depicts how aging and aging related diseases are intimately linked to inflammation

Sex/gender-related differences in inflammaging

: Geroscience puts mechanisms of aging as a driver of the most common age-related diseases and dysfunctions. Under this perspective, addressing the basic mechanisms of aging will produce a better understanding than addressing each disease pathophysiology individually. Worldwide, despite greater functional impairment, life expectancy is higher in women than in men. Gender differences in the prevalence of multimorbidity lead mandatory to the understanding of the mechanisms underlying gender-related differences in multimorbidity patterns and disability-free life expectancy. Extensive literature suggested that inflammaging is at the crossroad of aging and age-related diseases. In this review, we highlight the main evidence on sex/gender differences in the mechanisms that foster inflammaging, i.e. the age-dependent triggering of innate immunity, modifications of adaptive immunity, and accrual of senescent cells, underpinning some biomarkers of inflammaging that show sex-related differences. In the framework of the "gender medicine perspective", we will also discuss how sex/gender differences in inflammaging can affect sex differences in COVID-19 severe outcomes