Journal: Biomedicines

Article Title: Modulation of Small RNA Signatures in Schwann-Cell-Derived Extracellular Vesicles by the p75 Neurotrophin Receptor and Sortilin

doi: 10.3390/biomedicines8110450

Figure Lengend Snippet: Primary Schwann cell (SC) characterization. ( a ) Schematic representation of the cell isolation strategy used. ( b ) Depiction of SC primary culture with immunocytochemistry against S100 and p75 NTR (both in green) in primary SCs from wild-type (WT) or Ngfr −/− neonatal rats. Images show lack of p75 NTR staining in Ngfr −/− derived cells. Nuclei are labeled in blue with Hoechst. Scale bar 50 μm. ( c ) Double immunocytochemistry between sortilin (red) and S100 (green) for both WT and Sort1 −/− derived primary SCs. Hoechst was used to label the cell nuclei (blue). Scale bar 50 μm. ( d ) Representative immunoblot for sortilin and p75 NTR from primary SC lysates, validating sortilin or p75 NTR protein absence in Sort1 −/− or Ngfr −/− primary SCs, respectively. ( e ) Live and death assay where viable cells were identified in SC cultures by Calcein green fluorescence, while dead cells were labeled red by ethidium homodimer-1. Scale bar 100 μm ( f ) Quantification shows that viability of Ngfr −/− or Sort1 −/− SCs is similar to that of WT SCs.

Article Snippet: After 1 h blocking with tris buffer saline containing 2% skimmed milk and 2% Tween-20, membranes were incubated overnight at 4 °C with primary antibodies against Sortilin (1:250; R&D Systems) and p75 NTR (1:500; Abcam, Cambridge, UK).

Techniques: Cell Isolation, Immunocytochemistry, Staining, Derivative Assay, Labeling, Western Blot, Fluorescence

Journal: Biomedicines

Article Title: Modulation of Small RNA Signatures in Schwann-Cell-Derived Extracellular Vesicles by the p75 Neurotrophin Receptor and Sortilin

doi: 10.3390/biomedicines8110450

Figure Lengend Snippet: The most differentially expressed miRNAs in EVs from rat primary Schwann cells lacking p75 NTR or sortilin.

Article Snippet: After 1 h blocking with tris buffer saline containing 2% skimmed milk and 2% Tween-20, membranes were incubated overnight at 4 °C with primary antibodies against Sortilin (1:250; R&D Systems) and p75 NTR (1:500; Abcam, Cambridge, UK).

Techniques:

Journal: Biomedicines

Article Title: Modulation of Small RNA Signatures in Schwann-Cell-Derived Extracellular Vesicles by the p75 Neurotrophin Receptor and Sortilin

doi: 10.3390/biomedicines8110450

Figure Lengend Snippet: Transgenic Schwann cell EV cargo characterization (lacking p75 NTR or sortilin expression). ( a ) UpSet plot showing the number of predicted target genes found to be targeted by individual miRNAs or by multiple miRNAs. The plot is made using the R package “UpSetR”. The blue bars on the left side of the plot show the number of targets predicted for each miRNA. The right-side bar plot shows the number of genes targeted by only a single miRNA (single black dot in the lower right panel) or multiple miRNAs (multiple linked dots in the lower right panel). ( b ) Pathway analysis for all genes predicted to be targets of 3 or more miRNAs. All hits from KEGG pathways with p -value < 0.05. ( c ) Pathway analysis for all genes predicted to be targets of 3 or more miRNAs. The top 20 hits from Gene Ontology Biological Process.

Article Snippet: After 1 h blocking with tris buffer saline containing 2% skimmed milk and 2% Tween-20, membranes were incubated overnight at 4 °C with primary antibodies against Sortilin (1:250; R&D Systems) and p75 NTR (1:500; Abcam, Cambridge, UK).

Techniques: Transgenic Assay, Expressing