Recovery and Biodistribution of Ex Vivo Expanded Human Erythroblasts Injected into NOD/SCID/IL2Rγnull mice

Ex vivo expanded erythroblasts (EBs) may serve as advanced transfusion products provided that lodgment occurs in the macrophage-niche of the marrow permitting maturation. EBs expanded from adult and cord blood expressed the receptors (CXCR4, VLA-4, and P-selectin ligand 1) necessary for interaction with macrophages. However, 4-days following transfusion to intact NOD/SCID/IL2Rγnull mice, CD235apos EBs were observed inside CD235aneg splenic cells suggesting that they underwent phagocytosis. When splenectomized and intact NOD/SCID/IL2Rγnull mice were transfused using retrovirally labeled human EBs, human cells were visualized by bioluminescence imaging only in splenectomized animals. Four days after injection, human CD235apos cells were detected in marrow and liver of splenectomized mice but only in spleen of controls. Human CD235apos erythrocytes in blood remained low in all cases. These studies establish splenectomized NOD/SCID/IL2Rγnull mice as a suitable model for tracking and quantification of human EBs in vivo

SEMA3A partially reverses VEGF effects through binding to neuropilin-1

Cross-talk between hematopoietic stem cells (HSCs) and bone marrow stromal cells (BMSCs) is essential for HSCs regulation and leukemogenesis. Studying bone marrow of myelodysplasia patients, a pre-leukemic condition, we found mRNA overexpression of vascular endothelial growth factor A (VEGFA) in CD34+ HSCs and semaphorin 3A (SEMA3A) in BMSCs. To better understand the role of VEGFA and SEMA3A in leukemogenesis, we recruited 30 myelodysplastic syndrome (MDS) patients, 29 acute myeloid leukemia (6 secondary to MDS) patients and 12 controls. We found higher VEGFA expression in de novo AML patients (without prior MDS) group (p = 0.0073) and higher SEMA3A expression in all BMSCs patient's samples compared to control group. We then overexpressed VEGFA in an acute myelogenous leukemia cell line, KG1 cells, and in normal CD34+ cells. This overexpression increased KG1 (p = 0.045) and CD34+ cell (p = 0.042) viability and KG1 (p = 0.042) and CD34+ cell (p = 0.047) proliferation. Moreover, KG1 and CD34+ cells overexpressing VEGFA also had increased proliferation when co-cultured with human marrow stromal HS5 cells (p = 0.045 and p = 0.02, respectively). However, co-culture of these transformed cells with HS5 cells overexpressing SEMA3A reduced KG1 (p = 0.004) and CD34+ (p = 0.009) proliferation. Co-culture of KG1 transformed cells with HS27 cells overexpressing SEMA3A reduced KG1 proliferation as well (p = 0.01). To investigate whether the dominant SEMA3A effect over VEGFA could be due to competition for neuropilin1 receptor (NRP1), we performed immunoprecipitation with anti-NRP1 antibody of cell extracts of co-cultured KG1 and HS5 cells, induced or not by VEGFA and SEMA3A recombinant proteins. Results showed a preferential association of NRP1 with SEMA3A, suggesting that SEMA3A can partially reverse the effects caused by the VEGFA preventing its binding with the NRP1 receptor. Since both hematopoietic cells, leukemic and normal, showed similar behavior, we suppose that the attempt to reversion of VEGF effects by SEMA3A is a homeostatic phenomenon in the hematopoietic niche. Finally, we conclude that VEGFA overexpression confers AML cell advantages and SEMA3A may partially reverse this effect; thus, SEMA3A protein combined with VEGFA inhibitors could be beneficial for AML treatment

The Sea Urchin sns5 Insulator Protects Retroviral Vectors From Chromosomal Position Effects by Maintaining Active Chromatin Structure

Silencing and position-effect (PE) variegation (PEV), which is due to integration of viral vectors in heterochromatin regions, are considered significant obstacles to obtaining a consistent level of transgene expression in gene therapy. The inclusion of chromatin insulators into vectors has been proposed to counteract this position-dependent variegation of transgene expression. Here, we show that the sea urchin chromatin insulator, sns5, protects a recombinant γ-retroviral vector from the negative influence of chromatin in erythroid milieu. This element increases the probability of vector expression at different chromosomal integration sites, which reduces both silencing and PEV. By chromatin immunoprecipitation (ChIP) analysis, we demonstrated the specific binding of GATA1 and OCT1 transcription factors and the enrichment of hyperacetylated nucleosomes to sns5 sequences. The results suggest that this new insulator is able to maintain a euchromatin state inside the provirus locus with mechanisms that are common to other characterized insulators. On the basis of its ability to function as barrier element in erythroid milieu and to bind the erythroid specific factor GATA1, the inclusion of sns5 insulator in viral vectors may be of practical benefit in gene transfer applications and, in particular, for gene therapy of erythroid disorders

Lavori atipici ed economia digitale - Prospettiva luso-italo-brasiliana. Contratos atípicos de emprego e economia digital - Perspectiva luso-ítalo-brasileira

Il seminario “Lavori atipici ed economia digitale - Prospettiva luso-italo-brasiliana”, svolto il 28 aprile 2022, presso l'Università La Sapienza di Roma, ha avuto come obiettivo riunire studenti, ricercatori e docenti per confrontarsi su temi legati ai profondi cambiamenti subiti dal mercato del lavoro, presso un momento in cui l'economia globale non riesce a generare sufficienti posti di lavoro. Tra i cambiamenti analizzati durante il seminario si può citare, tra gli altri, la crescita di forme di occupazione diverse dai modelli tradizionalmente utilizzati. Si osserva che le cosiddette “forme atipici di lavoro” possono servire a finalità specifiche, come situazioni di lavoro stagionale, ma possono anche essere utilizzate per regolare nuove attività e forme di occupazione, derivanti dall'evoluzione della società nel nostro tempo. Quest’opera contiene studi relativi al lavoro su piattaforma digitale, contratti a tempo determinato, lavoro temporaneo, somministrazione, telelavoro o smart working, lavoro intermittente, lavoro part-time, tra le altre forme di contratti atipici. In questo modo gli organizzatori, l’università Sapienza di Roma, l’Università Lusófona e l’Instituto Iberoamericano de Estudos Juridícos – IBEROJUR, invitano la comunità accademica, formata da giuristi e non giuristi, giovani ricercatori e professori, a leggere questo libro, che presenta riflessioni critiche degli autori sulle sfide del mondo del lavoro, in una prospettiva globalizzata