Eosinophils, but not neutrophils, exibit an efficient DNA repair machinary and high nucleolar activity

BACKGROUND AND OBJECTIVES: Traditionally eosinophils have been considered terminally differentiated cells that play a role in host protection against parasites. However, there is some evidence showing that eosinophils are, in fact, multifunctional leukocytes involved in inflammatory responses, as well as in tissue homeostasis. We characterized the transcriptome profile of human eosinophils, and, for the purpose of comparison, the transcriptome profile of neutrophils, monocytes and hematopoietic progenitor cells. Moreover, we studied the activation of selected cellular processes for which a significant differential expression was demonstrated. DESIGN AND METHODS: We profiled gene expression using Affymetrix GeneChips. DNA repair capacity was tested using the comet assay. Nucleoli and their activity were characterized by transmission electron microscopy analysis, silver staining of nucleolus regions (AgNOR) and RNA staining. RESULTS: Gene expression profiling showed that eosinophils appear hierarchically closer to monocytes than to neutrophils. Gene ontology mapping of differentially expressed genes revealed that eosinophils express categories very similar to those expressed by monocytes, related to DNA repair and nucleolar functions. Moreover, our data show that eosinophils and monocytes maintain the ability to repair both double and single strand DNA breaks, whereas neutrophils lack this capacity. Furthermore, eosinophils exhibit nucleolar activity, which is lacking in neutrophils, but resembles that in monocytes. INTERPRETATION AND CONCLUSIONS: The presence of large, active nucleoli in eosinophils, coupled to marked activity of DNA repair systems, suggests that eosinophils are not terminally differentiated cells. Indeed, their transcriptome profile and functional properties are more similar to those of non-terminally differentiated cells such as monocytes, rather than to neutrophils

A candidate gene for fire blight resistance in Malus × robusta 5 is coding for a CC-NBS-LRR

Fire blight is the most important bacterial disease in apple (Malus ×  domestica) and pear (Pyrus communis) production. Today, the causal bacterium Erwinia amylovora is present in many apple- and pear-growing areas. We investigated the natural resistance of the wild apple Malus ×  robusta 5 against E. amylovora, previously mapped to linkage group 3. With a fine-mapping approach on a population of 2,133 individuals followed by phenotyping of the recombinants from the region of interest, we developed flanking markers useful for marker-assisted selection. Open reading frames were predicted on the sequence of a BAC spanning the resistance locus. One open reading frame coded for a protein belonging to the NBS-LRR family. The in silico investigation of the structure of the candidate resistance gene against fire blight of M. ×  robusta 5, FB_MR5, led us hypothesize the presence of a coiled-coil region followed by an NBS and an LRR-like structure with the consensus ‘LxxLx[IL]xxCxxLxxL'. The function of FB_MR5 was predicted in agreement with the decoy/guard model, that FB_MR5 monitors the transcribed RIN4_MR5, a homolog of RIN4 of Arabidopsis thaliana that could interact with the previously described effector AvrRpt2EA of E. amylovor

A candidate gene for fire blight resistance in Malus × robusta 5 is coding for a CC-NBS-LRR

Erworben im Rahmen der Schweizer Nationallizenzen (http://www.nationallizenzen.ch)Fire blight is the most important bacterial disease in apple (Malus ×  domestica) and pear (Pyrus communis) production. Today, the causal bacterium Erwinia amylovora is present in many apple- and pear-growing areas. We investigated the natural resistance of the wild apple Malus ×  robusta 5 against E. amylovora, previously mapped to linkage group 3. With a fine-mapping approach on a population of 2,133 individuals followed by phenotyping of the recombinants from the region of interest, we developed flanking markers useful for marker-assisted selection. Open reading frames were predicted on the sequence of a BAC spanning the resistance locus. One open reading frame coded for a protein belonging to the NBS–LRR family. The in silico investigation of the structure of the candidate resistance gene against fire blight of M. ×  robusta 5, FB_MR5, led us hypothesize the presence of a coiled-coil region followed by an NBS and an LRR-like structure with the consensus ‘LxxLx[IL]xxCxxLxxL’. The function of FB_MR5 was predicted in agreement with the decoy/guard model, that FB_MR5 monitors the transcribed RIN4_MR5, a homolog of RIN4 of Arabidopsis thaliana that could interact with the previously described effector AvrRpt2EA of E. amylovora

A technology evaluation method for assessing the potential contribution of energy technologies to decarbonisation of the Italian production system

A methodology focused on technology evaluation is proposed to give a footprint of the development potential of energy technologies in Italy. The approach focuses on the impact on climate, the potential in terms of R&D, the competitiveness of Italian companies and their diffusion on the territory. A reference Catalogue was realised in the framework  of the ‘Technical Board on Decarbonisation of the Economy’, established by the Italian Presidency of the Council of Ministers. 36 datasheets, containing quantitative and qualitative information on Technology Readiness Level (TRL), efficiency, environmental and economic impacts and policy aspects were filled by 70 experts for each technology. Some data were extracted from the Catalogue - TRL, CO2 emissions, developers, and centres of excellence - and further analysed with other information relating to the Italian production and innovation system collected from the National Enterprise Registry (ASIA). Companies and research centres  are involved in development of technologies based on Renewable Energy Sources (RES) and Energy Storage (ES) with different levels of TRL and high potential for mitigating effects on climate. However, their distribution shows a rather inhomogeneous presence at territorial level. This evaluation provided useful elements to elaborate policy measures to support the diffusion of energy technologies

Polysaccharides on gelatin-based hydrogels differently affect chondrogenic differentiation of human mesenchymal stromal cells

Selection of feasible hybrid-hydrogels for best chondrogenic differentiation of human mesenchymal stromal cells (hMSCs) represents an important challenge in cartilage regeneration. In this study, three-dimensional hybrid hydrogels obtained by chemical crosslinking of poly (ethylene glycol) diglycidyl ether (PEGDGE), gelatin (G) without or with chitosan (Ch) or dextran (Dx) polysaccharides were developed. The hydrogels, namely G-PEG, G-PEG-Ch and G-PEG-Dx, were prepared with an innovative, versatile and cell-friendly technique that involves two preparation steps specifically chosen to increase the degree of crosslinking and the physical-mechanical stability of the product: a first homogeneous phase reaction followed by directional freezing, freeze-drying and post-curing. Chondrogenic differentiation of human bone marrow mesenchymal stromal cells (hBM-MSC) was tested on these hydrogels to ascertain whether the presence of different polysaccharides could favor the formation of the native cartilage structure. We demonstrated that the hydrogels exhibited an open pore porous morphology with high interconnectivity and the incorporation of Ch and Dx into the G-PEG common backbone determined a slightly reduced stiffness compared to that of G-PEG hydrogels. We demonstrated that G-PEG-Dx showed a significant increase of its anisotropic characteristic and G-PEG-Ch exhibited higher and faster stress relaxation behavior than the other hydrogels. These characteristics were associated to absence of chondrogenic differentiation on G-PEG-Dx scaffold and good chondrogenic differentiation on G-PEG and G-PEG-Ch. Furthermore, G-PEG-Ch induced the minor collagen proteins and the formation of collagen fibrils with a diameter like native cartilage. This study demonstrated that both anisotropic and stress relaxation characteristics of the hybrid hydrogels were important features directly influencing the chondrogenic differentiation potentiality of hBM-MSC

Frailty trajectories in community-dwelling older adults during COVID-19 pandemic: The PRESTIGE study

Background Frailty has been recognized as potential surrogate of biological age and relevant risk factor for COVID-19 severity. Thus, it is important to explore the frailty trajectories during COVID-19 pandemic and understand how COVID-19 directly and indirectly impacts on frailty condition. Methods We enrolled 217 community-dwelling older adults with available information on frailty condition as assessed by multidimensional frailty model both at baseline and at one-year follow-up using Multidimensional Prognostic Index (MPI) tools. Pre-frail/frail subjects were identified at baseline as those with MPI score >0.33 (MPI grades 2-3). Frailty worsening was defined by MPI difference between 12 months follow-up and baseline >= 0.1. Multivariable logistic regression was modelled to identify predictors of worsening of frailty condition. Results Frailer subjects at baseline (MPI grades 2-3 = 48.4%) were older, more frequently female and had higher rates of hospitalization and Sars-CoV-2 infection compared to robust ones (MPI grade 1). Having MPI grades 2-3 at baseline was associated with higher risk of further worsening of frailty condition (adjusted odd ratio (aOR): 13.60, 95% confidence interval (CI): 4.01-46.09), independently by age, gender and Sars-CoV-2 infection. Specifically, frail subjects without COVID-19 (aOR: 14.84, 95% CI: 4.26-51.74) as well as those with COVID-19 (aOR: 12.77, 95% CI: 2.66-61.40, p = 0.001) had significantly higher risk of worsening of frailty condition. Conclusions Effects of COVID-19 pandemic among community-dwelling frailer individuals are far beyond the mere infection and disease, determining a significant deterioration of frailty status both in infected and non-infected subjects

Calreticulin Ins5 and Del52 mutations impair unfolded protein and oxidative stress responses in K562 cells expressing CALR mutants

Somatic mutations of calreticulin (CALR) have been described in approximately 60-80% of JAK2 and MPL unmutated Essential Thrombocythemia and Primary Myelofibrosis patients. CALR is an endoplasmic reticulum (ER) chaperone responsible for proper protein folding and calcium retention. Recent data demonstrated that the TPO receptor (MPL) is essential for the development of CALR mutant-driven Myeloproliferative Neoplasms (MPNs). However, the precise mechanism of action of CALR mutants haven't been fully unraveled. In this study, we showed that CALR mutants impair the ability to respond to the ER stress and reduce the activation of the pro-apoptotic pathway of the unfolded protein response (UPR). Moreover, our data demonstrated that CALR mutations induce increased sensitivity to oxidative stress, leading to increase oxidative DNA damage. We finally demonstrated that the downmodulation of OXR1 in CALR-mutated cells could be one of the molecular mechanisms responsible for the increased sensitivity to oxidative stress mediated by mutant CALR. Altogether, our data identify novel mechanisms collaborating with MPL activation in CALR-mediated cellular transformation. CALR mutants negatively impact on the capability of cells to respond to oxidative stress leading to genomic instability and on the ability to react to ER stress, causing resistance to UPR-induced apoptosis

Hematopoiesis and immune reconstitution after CD19 directed chimeric antigen receptor T‐cells (CAR‐T): A comprehensive review on incidence, risk factors and current management

Impaired function of hematopoiesis after treatment with chimeric antigen T-cells (CAR-T) is a frequent finding and can interest a wide range of patients, regardless of age and underlying disease. Trilinear cytopenias, as well as hypogammaglobulinemia, B-cell aplasia, and T-cell impairment, can severely affect the infectious risk of CAR-T recipients, as well as their quality of life. In this review, we provide an overview of defects in hematopoiesis after CAR-T, starting with a summary of different definitions and thresholds. We then move to summarize the main pathogenetic mechanisms of cytopenias, and we offer insight into cytomorphological aspects, the role of clonal hematopoiesis, and the risk of secondary myeloid malignancies. Subsequently, we expose the major findings and reports on T-cell and B-cell quantitative and functional impairment after CAR-T. Finally, we provide an overview of current recommendations and leading experiences regarding the management of cytopenias and defective B- and T-cell function

Calreticulin affects hematopoietic stem/progenitor cell fate by impacting erythroid and megakaryocytic differentiation

Calreticulin (CALR) is a chaperone protein that localizes primarily to the endoplasmic reticulum (ER) lumen where it is responsible for the control of proper folding of neo-synthesized glycoproteins and for the retention of calcium. Recently, mutations affecting exon 9 of the CALR gene have been described in approximately 40% of patients with myeloproliferative neoplasms (MPNs). Although the role of mutated CALR in the development of MPNs has begun to be clarified, there are still no data available on the function of wild-type (WT) CALR during physiological hematopoiesis. In order to shed light on the role of WT CALR during normal hematopoiesis, we performed gene silencing and overexpression experiments in Hematopoietic Stem Progenitor Cells (HSPCs). Our results showed that CALR overexpression is able to affect physiological hematopoiesis by enhancing both erythroid and megakaryocytic (MK) differentiation. In agreement with overexpression data, CALR silencing caused a significant decrease in both erythroid and MK differentiation of human HSPCs. Gene expression profiling (GEP) analysis showed that CALR is able to affect the expression of several genes involved in HSPCs differentiation towards both the erythroid and MK lineages. Moreover, GEP data also highlighted the modulation of several genes involved in ER stress response, unfolded protein response (UPR), DNA repair and of several genes already described to play a role in MPN development, such as pro-inflammatory cytokines and hematological neoplasms-related markers. Altogether, our data unraveled a new and unexpected role for CALR in the regulation of normal hematopoietic differentiation. Moreover, by showing the impact of CALR on the expression of genes involved in several biological processes already described in cellular transformation, our data strongly suggest a more complex role for CALR in MPN development that goes beyond the activation of the THPO receptor and involves ER stress response, UPR and DNA repair