Figure 5

The effects of platelet extracts and adenine nucleotides on Matrigel plug vascularization. Matrigel (200 μl per plug) was mixed with platelet extracts (29 μg/ml and 58 μg/ml), ATP (1 mM), mixture of MeSATP, ATPγS, MeSADP, and ADPβS (0.1 mM, 0.5 mM or 1 mM of each), combination of nucleotides and platelet extracts (concentrations as indicated), or vechicle (10 mM HCl in PBS) and injected subcutaneously into ICR mice (n = 4-5 for each group). After 7 days, mice were sacrificed, Matrigel plugs were excised, and 5-7 μm-thick sections were prepared from formalin fixed plugs/paraffin-embedded plugs. H&E staining was performed for identification of plug cellularity and tube formation. Representative images show (a): control plugs, ×40; (b): plugs containing 58 μg/ml platelet extract (×40; insert shows magnified view of an capillary containing red blood cells); (c-d): plugs containing 0.5 mM purine nucleotides (panel c = ×40, panel d = x10 magnification; arrows on the panel c indicate areas of increased plug cellularity); platelet extracts (58 μg/ml), mixture of MeSATP, ATPγS, MeSADP, and ADPβS (0.1 mM, 0.5 mM or 1 mM of each), (e-f): plugs containing platelet extracts (58 μg/ml) and mixture of nucleotides (0.5 mM of each); panel e: plug with increased amount of functional capillaries (enlarged in the insert); panel f: blood cell infiltrates around new formed capillaries (enlarged in the insert); (g-j): Identification of PECAM-positive cells in control plugs (g) and plugs containing 58 μg/ml platelet extract (h), mixture of nucleotides (0.5 mM of each), and combination of 58 μg/ml platelet extract and mixture of nucleotides (0.5 mM of each, j). PECAM-positive cells are shown enlarged in the inserts.