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Silvestrelli, G.

Publications and source records attributed to Silvestrelli, G..

2 recordsLinked to original sources

Lineage origin of spinal cord cell type diversity

The complexity and specificity of movement in vertebrates is driven by a rich diversity of spinal motor and interneuron cell types. During development, eleven spinal cord progenitor domains generate an equivalent number of cardinal neuron types. How progenitor domains, individual progenitors, and post-mitotic diversity relate is still unknown. We performed high-resolution, single-progenitor cell lineage tracing in the embryonic mouse spinal cord using mosaic analysis with double markers (MADM). Our quantitative study of lineage progression revealed that spinal cord progenitors undergo highly variable numbers of proliferative, neurogenic, and gliogenic cell divisions. The nascent clonally-related neurons migrate radially over large distances, span the dorsoventral axis, and even cross the midline, demonstrating striking bilaterality. Molecular and morphometric analysis indicate high levels of progenitor multipotency, with an individual progenitor capable of producing several molecularly and morphologically distinct neuron types, as well as astrocytes. These findings redefine spinal cord development as a process in which lineage variability--rather than rigid progenitor identity--drives the generation of cellular diversity.

neuroscience↗

Assessing extracellular vesicles from bovine mammary gland epithelial cells cultured in FBS-free medium

AimMammary gland extracellular vesicles (EVs) are found in both human and livestock milk. Our knowledge of the role of EVs in the mammary gland development, breast cancer and mastitis derives mainly from in vitro cell culture models. However, a commonly shared limitation is the use of foetal bovine serum (FBS) as a supplement, which naturally contains EVs. For this reason, the purpose of the study is to establish a novel tool to investigate mammary gland EVs in vitro and in an FBS-free system. MethodsPrimary bovine mammary epithelial cells (pbMECs) and a mammary gland alveolar epithelial cell line (MAC-T) were cultured in a chemically defined EV-free medium. To find a reliable EVs isolation protocol from a starting cell conditioned medium (10 mL), we compared eight different methodologies by combining ultracentrifugation (UC), chemical precipitation (CP), size exclusion chromatography (SEC), and ultrafiltration (UF). ResultsThe medium formula sustained both pbMECs and MAC-T cell growth and did not alter MAC-T cell identity. Transmission electron microscopy revealed that we obtained EV-like particles in five out of eight protocols. The cleanest samples with the highest particles amount and detectable amounts of RNA were obtained by using UF-SEC-UC and UC-SEC-UC. ConclusionOur chemically defined, EV-free medium sustains the growth of both pbMECs and MAC-T and allows the isolation of EVs that are free from any contamination by UF-SEC-UC and UC-SEC-UC. In conclusion, we propose a new culture system and EVs isolation protocols for further research on mammary epithelial EVs.

physiology↗