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. 2020 Jan 30;11:615. doi: 10.1038/s41467-020-14480-0

Fig. 3. Monocyte/macrophage responses to test groups in a mouse model of hindlimb ischemia.

Fig. 3

a Gene expression (CD3, CD68, and CD31) of infiltrated cells into macro- versus microchannel network hydrogels only after removing surrounding tissues at day 14 post-implantation by qRT-PCR (N = 3). Data presented are mean ± SEM. Statistical significances are determined using a two-tailed Student’s t-test; *p < 0.05 between lined groups. b IVIS images of monocyte infiltration into the implant site (hydrogel + surrounding tissue) of each test group on day 1 post intravenous administration of Vivotrack680-labeled RAW264.7 cells, followed by quantification of the ratio in ischemic to normal hindlimb (N = 3). Data presented are mean ± SEM. Statistical significances are determined using one-way ANOVA with Tukey post-hoc pairwise comparisons; *p < 0.05 and ***p < 0.005 between lined groups. c Representative H&E and immunostaining (CD68) images of ischemic hindlimb tissues at the implant site. Scale bar = 100 µm. d Confocal images of macrophage polarization markers (M1: iNOS versus M2: CD206, both in green), a mouse macrophage marker (F4/80 in red), and nucleus (DAPI in blue) at day 14 post-implantation. The corresponding M1 or M2 cell number % out of the total macrophage number (F4/80+) as well as M1/M2 ratios in the macro- and microchannel groups were determined quantitatively. Scale bar = 100 µm. Data presented are mean ± SEM. Statistical significances are determined using a two-tailed Student’s t-test; *p < 0.05 and ***p < 0.005 between lined groups (N = 4). e Gene expressions of M1 (IL-1β, IL-6, TNF-α, CD80, and NOS2) and M2 (IL-10, arginase-1, CD-163, and CD-206)] markers in mouse macrophages (RAW264.7) post-culture within macro- versus microchannel network hydrogels in vitro by qRT-PCR (N = 5). Data presented are mean ± SEM. Statistical significances are determined using a two-tailed Student’s t-test; *p < 0.05 and ***p < 0.005 between lined groups (N.S.: not significant). Data presented are mean ± SEM. Quantification of endothelial cell (human umbilical vein endothelial cell, HUVEC) f tubulogenesis and g migration induced conditioned media of RAW264.7 cells in vitro (N = 4). Dots represent each replicate of HUVEC-seeded wells in a 24-well plate. Data presented are mean ± SEM. Statistical significances are determined using one-way ANOVA with Tukey post-hoc pairwise comparisons; *p < 0.05 and ***p < 0.005 between lined groups. Source data are provided as a Source Data file.