Z. B. R. and T. R. contributed equally to this paper.
Differential effects of paracrine factors on the survival of cells of the neurovascular unit during oxygen glucose deprivation
Version of Record online: 10 NOV 2013
© 2013 The Authors. International Journal of Stroke © 2013 World Stroke Organization
International Journal of Stroke
Volume 10, Issue 3, pages 407–414, April 2015
How to Cite
Redzic, Z. B., Rabie, T., Sutherland, B. A. and Buchan, A. M. (2015), Differential effects of paracrine factors on the survival of cells of the neurovascular unit during oxygen glucose deprivation. International Journal of Stroke, 10: 407–414. doi: 10.1111/ijs.12197
Conflicts of interest: None declared.
- Issue online: 16 MAR 2015
- Version of Record online: 10 NOV 2013
- Manuscript Accepted: 5 AUG 2013
- Manuscript Received: 15 MAY 2013
- Henry Smith Charity
- Fondation Leducq
Fig. S1. Immunostaining of primary brain endothelial cells (BECs) with rabbit polyclonal anti-von Willebrand factor antibody (a), rat astrocytes with rabbit polyclonal anti-glial fibrillary acidic protein (GFAP) antibody (b), pericytes with rabbit monoclonal anti-platelet-derived growth factor receptor beta (PDGFRb) and mouse monoclonal anti-neuronal-glial 2 (NG2) proteoglycan antibodies (c) and microglia with rabbit polyclonal anti-CD11b antibody (d).
Fig. S2. Cell necrosis and late apoptosis of astrocytes measured by Annexin PE/7-AAD staining.
Primary astrocytes were exposed to six-hour control conditions (a) or six-hour OGD (b). Cells were detached and an aliquot of ≈500 000 cells were stained with Annexin V – Phycoerythrin (PE) and 7-Amino-actinomycin (7-AAD) according to manufacturer's instruction (BD Pharmigen), giving a total volume of 0·5 ml. Quantification of stained cells was done in Beckman Cytmoics FC 500 flow cytometer. 7-AAD fluorescence was plotted versus Annexin PE fluorescence. Cells located in the B4 quadrant were early, apoptotic, 7-AAD-negative/Annexin PE-positive. Cells in the B3 quadrant were viable, 7-AAD-negative/Annexin PE-negative. Cells in the upper quadrants B1 (7-AAD-positive/annexin PE-negative) and B2 (7AAD-positive/annexin PE-positive) were necrotic and late apoptotic, respectively. A representative panel from each group is shown.
Table S1. Effect of time on concentration of angiogenic factors in the cell culture medium during OGD and in controls – one-way ANOVA.
Table S2. Cell necrosis and late apoptosis of astrocytes measured by Annexin PE/7-AAD staining and a comparison of these data with percentage of dead cells estimated by LDH assay.
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