Journal: Journal of the American Heart Association
Article Title: Contribution of Extra‐Cardiac Cells in Murine Heart Valves is Age‐Dependent
doi: 10.1161/jaha.117.007097
Figure Lengend Snippet: Figure 1. In C57/Bl6 wild-type mice, the percentage of CD45+ cells in aortic and mitral valve leaflets increases with age and maintain CD45 expression. A, CD45+ cells represented as a percentage of total cells at each time point in aortic and mitral valves. B, Percentage of CD45-derived (RFP+) cells in aortic valves from CD45-Cre+;Ai9+/ mice at E11.5, PND2, and 6 weeks of age (*P<0.05 compared with E11.5; #P<0.05 compared with PND2). C, Aortic valve from 6-week-old CD45-Cre+;Ai9+/ mice, stained for RFP (red) to detect CD45-derived cells, and for CD45 (green) to detect endogenous CD45 expression. D and E, 6-week-old CD45-Cre+;Ai9+/ aortic valves costained with RFP (red) and immune (CD11b [D] and F4/80) [E]) (green) cell markers. F, F4/80 immunoreactivity in adult liver following irradiation-induced injury. Negative control of no primary antibody (Ab) is shown as inset. G and H, 6-week-old CD45Cre+;Ai9+/ aortic valves costained with RFP (red) and CD31 (endothelial) (G) and Vimentin (interstitial) (H) (green). Arrowheads: double-positive cells. Arrows: CD45-derived cells negative for CD11b, F4/80, CD31, and Vimentin. Notched arrowheads: CD11b-, F4/80-, CD31-, or Vimentin-positive cells that are not CD45 positive. I, Quantification of the number of CD45-derived cells expressing endogenous CD45, CD11b, F4/80, CD31, and Vimentin. n=3. Statistical significance based on *P<0.05 between each group. E indicates embryonic day; PND, postnatal day; RFP, red fluorescent protein.
Article Snippet: Antibodies and Working Concentrations Primary Antibody Vendor Host Dilution GFP Abcam (ab13970) Chicken 1:500 GFP Abcam (ab290) Rabbit 1:1000 CD45 R&D Systems (AF114) Rabbit 1:200 RFP Rockland (600-401-379) Rabbit 1:3000 Vimentin Abcam (ab45939) Rabbit 1:200 Vimentin Novus Bio. (NB300-223) Chicken 1:200 CD31 BD Biosciences (553370) Rat 1:1000 F4/80 Bio Rad (MCA497GA) Rat 1:200 CD11b Bio Rad (MCA711G) Rat 1:200 Cleaved caspase-3 Cell Signaling (9664) Rabbit 1:200 GFP indicates green fluorescent protein; RFP, red fluorescent protein.
Techniques: Expressing, Derivative Assay, Staining, Irradiation, Negative Control
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![Figure 1. In C57/Bl6 wild-type mice, the percentage of <t>CD45+</t> cells in aortic and mitral valve leaflets increases with age and maintain CD45 expression. A, CD45+ cells represented as a percentage of total cells at each time point in aortic and mitral valves. B, Percentage of CD45-derived (RFP+) cells in aortic valves from CD45-Cre+;Ai9+/ mice at E11.5, PND2, and 6 weeks of age (*P<0.05 compared with E11.5; #P<0.05 compared with PND2). C, Aortic valve from 6-week-old CD45-Cre+;Ai9+/ mice, stained for RFP (red) to detect CD45-derived cells, and for CD45 (green) to detect endogenous CD45 expression. D and E, 6-week-old CD45-Cre+;Ai9+/ aortic valves costained with RFP (red) and immune (CD11b [D] and F4/80) [E]) (green) cell markers. F, F4/80 immunoreactivity in adult liver following irradiation-induced injury. Negative control of no primary antibody (Ab) is shown as inset. G and H, 6-week-old CD45Cre+;Ai9+/ aortic valves costained with RFP (red) and CD31 (endothelial) (G) and Vimentin (interstitial) (H) (green). Arrowheads: double-positive cells. Arrows: CD45-derived cells negative for CD11b, F4/80, CD31, and Vimentin. Notched arrowheads: CD11b-, F4/80-, CD31-, or Vimentin-positive cells that are not CD45 positive. I, Quantification of the number of CD45-derived cells expressing endogenous CD45, CD11b, F4/80, CD31, and Vimentin. n=3. Statistical significance based on *P<0.05 between each group. E indicates embryonic day; PND, postnatal day; RFP, red fluorescent protein.](https://doi-unpaywalled-images-cdn.bioz.com/2112/10__1161_slash_jaha__117__007097/10__1161_slash_jaha__117__007097____page5_image1.jpg)