Treatment of hyperglycaemia improves VEGFR2 expression in ischaemic hind limb muscles of Type 1 DM mice, and neutralizing VEGFR2 abolishes the treatment-related improved perfusion recovery. (A) A fold increase in VEGFR2 protein is comparable in treated Type 1 DM and non-DM controls in Day 3 post-ischaemic hind limb muscles, but impaired in untreated Type 1 DM mice (treated Type 1 DM, n = 7; non-DM controls n = 5; untreated Type 1 DM n = 6, *P < 0.05, NS = not significant, P > 0.05). (B) VEGFR2 mRNA expression by quantitative PCR showed no difference in the level of expression between non-DM, untreated Type 1 DM, and treated Type 1 DM in Day 3 post-ischaemic hind limbs (n = 6, 7, and 5, respectively, NS, P > 0.05). (C) Treatment with a VEGFR2-neutralizing antibody results in impaired perfusion recovery in treated Type 1 DM. The Y-axis shows the perfusion ratio (ischaemic-to-non-ischaemic limb) normalized to values immediately post-surgery, while the X-axis shows time post-surgery [non-DM controls n = 6–7, treated (Tx) type 1 DM n = 8–10, *P = 0.008 at W3 and 0.01 at W, NS, P = 0.99]. (D) Treatment with isotype matched control antibody had no effect on perfusion recovery in treated Type 1 DM mice [non-DM controls n = 7, treated (Tx) Type 1 DM n = 5]. (E, F, and G) Immunostaining of sections from Week 5 post-ischaemic hind limb muscles of non-diabetic control (E), untreated Type 1 DM (F), and Treated Type 1 DM mice (G). It shows co-localization of VEGFR2 expression with CD31-expressing cells (CD31 = green, VEGFR2 = pink, actin = red, blue = nuclear staining, green + pink = white) consistent with endothelial cell expression.