Effect of the IL‐10—IL‐10R pathway on angiogenesis in vivo. (A, C) Matrigel plugs containing ectopic EN‐MSCs mixed with recombinant proteins as indicated (A) or recombinant proteins alone as indicated (C) were implanted into female NUDE mice. Representative photomicrographs of paraformaldehyde‐fixed sections from the Matrigel plugs stained with anti‐mCD31 (brown) and haematoxylin (blue) are shown. The insets (solid box) show two‐fold enlarged images of microvascular structures (dashed box). Arrows indicate microvascular structures. Scale bars = 100 µm. (B, D) Microvessel density was evaluated based on the number of CD31+ haematoxylin+ cells per square millimetre. The experiments were repeated at least three times, and the data are presented as the mean ± SD (n = 3 mice in each group in B; n = 3–6 mice in each group in D). (E) The yolk sac of 72 h post‐fertilisation zebrafish embryos was injected with PBS or with 5 or 25 U/ml rmIL‐10 in PBS. Representative photographs taken 24 h after injection using an epifluorescence microscope show the angiogenesis of the SIV (yellow arrows) in the resulting zebrafish larvae. Scale bars = 50 µm. The number of treated larvae in each group is indicated. (F) The percentage of larvae with angiogenesis of the SIV is shown for each of the three groups. Student's unpaired t‐test was performed in B and D, and Fisher's exact test in F.
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