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Pisani, F.

Publications and source records attributed to Pisani, F..

5 recordsLinked to original sources
bioRxiv2026

Intercellular Transfer of PTBP1 Drives Human Neural Stem Cell Fate

During fetal brain development, temporally defined alternative splicing (AS) programs control human neural stem cell (hNSC) self-renewal and differentiation, thereby regulating neurogenesis and gliogenesis. Polypyrimidine tract-binding protein 1 (PTBP1) is a master regulator of AS during neurogenesis; however, its functional role and dynamics in hNSCs remain largely unexplored. Here, we investigate the cellular and molecular functions, nucleocytoplasmic distribution, and intercellular trafficking of PTBP1 in primary hNSCs. We found that PTBP1 knockdown (KD) alters self-renewal capacity, mitochondrial dynamics and membrane potential, lipid droplet abundance, and PTBP2 expression. RNA sequencing analysis revealed that PTBP1 depletion affects the expression profiling of hundreds of coding and non-coding genes, collectively orchestrating a neuronal differentiation program. Super-resolution {tau}-STED microscopy and live-cell imaging demonstrated that PTBP1 localizes not only to the nucleus but also to the cytoplasm, tunneling nanotubes (TNTs), migrasomes, and extracellular vesicles (EVs). Co-culture experiments and EV uptake assays showed that cytosolic PTBP1 can be transferred between hNSCs and delivered to the nuclei of recipient cells via TNTs and EVs. Moreover, EVs were found to contain specific and previously uncharacterized PTBP1 isoforms and were efficiently transferred to PTBP1-KD cells, rescuing their proliferative capacity. Analysis of the mouse brain reveals the presence of PTBP1 in the V-SVZ and within TNT-like structures connecting NSCs, suggesting a role for TNT-mediated PTBP1 trafficking in vivo. Together, these findings uncover previously unrecognized roles for PTBP1 in hNSCs and provide the first evidence that PTBP1 can be transferred between hNSCs via TNTs and EVs, revealing a novel mechanism by which hNSCs may regulate fetal neurogenesis. Graphical AbstractA: PTBP1 regulates hNSC fate by controlling cell proliferation, lipid droplet dynamics, mitochondrial function, and post-transcriptional programs involved in neuronal commitment. B: Cytosolic PTBP1 is transferred between hNSCs via tunneling nanotubes (TNTs) and extracellular vesicles (EVs). Abbreviations: LV, lateral ventricle; aRG, apical radial glial cells; bRG, basal radial glial cells; V-SVZ, ventricular-subventricular zone; EVs, extracellular vesicles; TNTs, tunneling nanotubes.

neuroscience↗
bioRxiv2024

A claudin5-binding peptide enhances the permeability of the blood-brain-barrier

The blood-brain barrier (BBB) is essential to maintain brain homeostasis and healthy conditions but it also prevents drugs from reaching brain cells. In the BBB, tight junctions (TJs) are multi-protein complexes located at the interface between adjacent brain endothelial cells that regulate paracellular diffusion and claudin-5 (CLDN5) is the major component of the TJ portfolio, playing a pivotal role in restricting the paracellular traffic. In view of obtaining fine control over the transport across the BBB, the use of competing peptides able to bind CLDN5 to induce transient and regulated permeabilization of the paracellular passage is emerging as a potentially translatable strategy for clinical applications. In this work, we designed and tested short peptides with improved solubility and biocompatibility using a combined approach that involved structural modeling techniques and in vitro validation, generating a robust workflow for the design, screening, and optimization of peptides for the modulation of the BBB paracellular permeability. We designed a selection of 11- to 16-mer compounds derived from the first CLDN5 extracellular domain and from the CLDN5-binding domain of Clostridium perfringens enterotoxin and determined their efficiency in enhancing BBB permeability. The computational analysis classified all tested peptides based on solubility and affinity to CLDN5, and provided atom-level details of the binding process. From our screening, we identified a novel CLDN5-derived peptide, here called f1-C5C2, which demonstrated good solubility in biological media, efficient binding to CLDN5 subunits, and capability to increase permeability at low concentrations. The peptidomimetic in silico/in vitro strategy described here can achieve a transient and reversible permeabilization of the BBB with potential applications in the pharmacological treatment of brain diseases. HIGHLIGHTSO_LIWater-soluble peptidomimetics are used to competitively bind claudin-5 tight junction proteins and increase the permeability of the blood-brain barrier; C_LIO_LITrans-endothelial electrical resistance and dissociation constant measurements demonstrate the binding affinity of the peptide f1-C5C2 for claudin-5; C_LIO_LIUnbinding free energy calculations correlated with experimental results and provided information on the protein-peptide binding interface. C_LIO_LIIncubation with the peptide f1-C5C2 allows paracellular transport of 4K, but not 70K, dextran. C_LI GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=88 SRC="FIGDIR/small/591687v1_ufig1.gif" ALT="Figure 1"> View larger version (30K): org.highwire.dtl.DTLVardef@70fe54org.highwire.dtl.DTLVardef@496a28org.highwire.dtl.DTLVardef@1d89a81org.highwire.dtl.DTLVardef@187acec_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗
bioRxiv2022

TUNNELING NANOTUBES CONNECT BLOOD-BRAIN BARRIER CELLS: ROLE OF PERICYTES IN BARRIER PRESERVATION DURING ISCHEMIA

Crosstalk mechanisms between pericytes, endothelial cells, and astrocytes preserve integrity and function of the blood-brain-barrier (BBB) under physiological conditions. Long intercellular channels allowing the transfer of small molecules and organelles between distant cells called tunneling nanotubes (TNT) represent a potential substrate for energy and matter exchanges between the tripartite cellular compartments of the BBB. However, the role of TNT across BBB cells under physiological conditions and in the course of BBB dysfunction is unknown. In this work, we analyzed the TNTs role in the functional dialogue between human brain endothelial cells, and brain pericytes co-cultured with human astrocytes under normal conditions or after exposure to ischemia/reperfusion, a condition in which BBB breakdown occurs, and pericytes participate in the BBB repair. Using live time-lapse fluorescence microscopy and laser-scanning confocal microscopy, we found that astrocytes form long TNT with pericytes and endothelial cells and receive functional mitochondria from both cell types through this mechanism. The mitochondrial transfer also occurred in multicellular assembloids of human BBB that reproduce the three-dimensional architecture of the BBB. Under conditions of ischemia/reperfusion, TNT formation is upregulated, and astrocytes exposed to oxygen-glucose deprivation were rescued from apoptosis by healthy pericytes through TNT-mediated transfer of functional mitochondria, an effect that was virtually abolished in the presence of TNT-destroying drugs. The results establish a functional role of TNT in the crosstalk between BBB cells and demonstrate that TNT-mediated mitochondrial transfer from pericytes rescues astrocytes from ischemia/reperfusion-induced apoptosis. Our data confirm that the pericytes might play a pivotal role in preserving the structural and functional integrity of BBB under physiological conditions and participate in BBB repair in brain diseases.

neuroscience↗
bioRxiv2021

AATF/Che-1, a new component of paraspeckles, controls R-loops formation and Interferon activation in Multiple Myeloma.

Multiple myeloma (MM) is a hematological neoplasm of plasma cells characterized by abnormal production of immunoglobulins. Che-1/AATF (Che-1) is an RNA binding protein involved in transcription regulation and is highly expressed in this malignancy. Here we experimentally show that Che-1 interacts with paraspeckle components, including the lncRNA NEAT1_2 (NEAT1), which serves as the seed for the maintenance of these structures. Che-1 and NEAT1 localize on R-loops, three-stranded RNA:DNA hybrids structures involved in DNA transcription and repair. Depletion of Che-1 produces a marked accumulation of RNA:DNA hybrids sustaining activation of a systemic inflammatory response. We provide evidence that high levels of Unfolded Protein Response (UPR) in MM cells induces RNA:DNA hybrids and an interferon (IFN) gene signature. We found that MM patients exhibit elevated R-loops levels and paraspeckle genes mRNAs increase linearly to MM progression. Strikingly, patients showing elevated IFN genes signature are associated with a marked poor prognosis. Overall, these findings delineate that elevated R-loops accumulation and inflammatory signaling may contribute to MM progression and that Che-1/NEAT1 plays an essential role in maintaining R-loops homeostasis by preventing excessive inflammatory signaling.

cell biology↗