Journal: Science Advances

Article Title: Regulation of blood-brain barrier integrity by Dmp1 -expressing astrocytes through mitochondrial transfer

doi: 10.1126/sciadv.adk2913

Figure Lengend Snippet: ( A ) Work flow of 2/2 μM A/R treatment on bEnd.3 endothelial cells followed by mitochondrial transplantation and H 2 DCFDA staining for flow cytometry analysis. ( B ) Representative histogram overlays and quantitative results of ROS (H 2 DCFDA) intensity in healthy (vehicle), 2/2 μM A/R-damaged bEnd.3 endothelial cells, and 2/2 μM A/R-damaged bEnd.3 endothelial cells after transplanted with 21.25-, 42.5-, and 85-μg astrocytes-derived mitochondria (Mito-T) ( n = 4). One-way ANOVA followed by Tukey’s post hoc test. Data are presented as means ± SEM. ( C ) Top 10 up-regulated GO enrichment terms of cellular components (CC) in bEnd.3 cells transplanted with primary WT astrocytes-derived mitochondria compared with normal control bEnd.3 cells. ( D ) Workflow for 4/4 μM A/R treatment on bEnd.3 endothelial cells followed by mitochondrial transplantation and cell scratch assay. ( E ) Representative images of scratched bEnd.3 endothelial cells without A/R treatment (vehicle), with 4/4 μM A/R treatment, with 4/4 μM A/R treatment and subsequent transplantation of approximately 170 μg of mitochondria isolated from primary astrocytes (up panel), followed by 24 hours of normal culture (below panel). Scale bar, 200 μm. ( F ) Quantification of wound healing rate after 24 hours of normal culture in bEnd.3 endothelial cells without 4/4 μM A/R treatment (vehicle), with 4/4 μM A/R treatment, with 4/4 μM A/R treatment and subsequent transplantation of approximately 170 μg of mitochondria ( n = 4). One-way ANOVA followed by Tukey’s post hoc test. Data are presented as means ± SEM.

Article Snippet: For the assessment of the alleviation of ROS accumulation after mitochondrial transplantation, 2/2 μM A/R-treated bEnd.3 endothelial cells were coincubated with mitochondria isolated from primary WT astrocytes in 12-well plates and stained with 10 μM H 2 DCFDA (Solarbio) for 20 min at 37°C.

Techniques: Transplantation Assay, Staining, Flow Cytometry, Derivative Assay, Control, Wound Healing Assay, Isolation