Fig 1: OVA-releasing scaffolds implanted in vaccinated mice enhance TH2 responses after ischemic injury.(A) Experimental setup with (left) BALB/c mice undergoing (right) timeline of procedures involving vaccination and boost with OVA/ALUM, followed by hindlimb ischemia induction and scaffold implantation. (B to J) In these figures, naïve + OVA denotes mice receiving no vaccination and implanted with an OVA-containing scaffold, vacc + blank denotes mice receiving vaccination with OVA/ALUM and a blank scaffold, and vacc + OVA denotes mice receiving vaccination with OVA/ALUM and an OVA-containing scaffold. (B) Number of CD4+ T cells (TCR-β+/CD4+ cells) per milligram of tissue in the ischemic upper leg at days 4 and 7 after ischemic ligation in the various treated groups. (C) Representative images of IL-5 enzyme-linked immunospot (ELISPOT) assay, where cells derived from the ischemic upper leg were cultured in the presence of splenocytes presenting either OVA or CT26-gp70 peptide as an irrelevant peptide control. (D) Quantification of OVA-specific IL-5 spot-forming cells per milligram of tissue determined by subtracting the number of IL-5 spot-forming cells in the CT26-gp70 condition from that in the OVA condition. (E to G) Concentrations of (E) IL-5, (F) IL-10, and (G) IFN-γ produced from OVA-stimulated cells isolated from the ischemic upper leg 7 days after ischemic ligation (n = 6 mice for naïve + OVA; n = 4 mice for vacc + blank; and n = 4 mice for vacc + OVA). (H to J) Number of (H) macrophages (F4/80+ cells), (I) M2a macrophages (F4/80+/CD206+ cells), and (J) eosinophils (Siglec-F+/CD11b+ cells) per milligram of tissue in the ischemic upper leg at days 4 and 7 after ischemic ligation in the various treated groups. For day 4 data in (B) to (D) and (H) to (J), n = 5 mice for naïve + OVA, n = 6 mice for vacc + blank, and n = 6 mice for vacc + OVA. For day 7 data in (B) to (D) and (H) to (J), n = 3 mice for naïve + OVA, n = 6 mice for vacc + blank, and n = 5 mice for vacc + OVA. Data are presented as means ± SD. Significance is denoted by *P ≤ 0.05, **P ≤ 0.01, or ***P ≤ 0.001 by one-way analysis of variance (ANOVA) with Bonferroni’s post hoc test.