Fig. 1
Experimental workflow and follow-up of ischemic symptoms in CLTI mice in response to cell administration. A Schematic representation of the experimental assay. Mice underwent femoral artery ligation (FAL, biological replicates or BR, n:24) or simulated surgery (Sham group, n:8 BR), and 24 h after, they were administered saline serum (ischemic untreated control, IC group, n:8), AT-ECFCs and MSCs (AT group, n:8) or CB-ECFCs and AT-MSCs (CB group, n:8). Follow-up of ischemic symptoms was done on day 1, 7, 14, and 21 after surgery (n:32). B Representative laser Doppler images from all groups of blood flow (BF) measurements using Pericam PSI HR system: post-surgery, on day 1, day 7, day 14, and day 21 (BR n:32). C Graphical representation of BF changes per group within time. Perfusion (PU) averaged ratios of left (ligated) vs. right (non-ligated) limbs are shown (BR: IC, n:8; AT, n:8; CB, n:8). D Necrosis progression, represented as the percentage of necrotic tissue affected per group, registered on days 1, 7, 14, and 21 BR: IC, n:8; AT, n:8; CB, n:8). E Motility changes (Tarlov score) registered at day 21 (BR: SH, n:8; IC, n:8; AT, n:8; CB, n:8). F Capillary density, the number of blood vessels (vessels/mm2) was measured by staining left limb muscles from all groups (BR: SH, n:8; IC, n:8; AT, n:8; CB, n:8; technical replicates, n:3). G Vessel classification based on abundance percentage of different ranges of internal lumen diameter (μm) (BR: SH, n:8; IC, n:8; AT, n:8; CB, n:8; technical replicates, n:3). Data were presented as mean ± SEM. Statistically significant differences after performing Two-way ANOVA and Tukey’s as post-hoc (C) or Kruskal Wallis and post-hoc Dunn’s test (F) are shown as * p-value < 0.05, ** p-value < 0.01, *** p-value < 0.001. I). H Representative images obtained by immunohistochemistry, used to measure vascular density and diameter size, using anti-mouse smooth muscle α-actin (α-SMA) antibody (red) and DAPI (blue)