Figure 6.

Metformin promotes vascular regeneration in aged mice post-SCI.

(A) Representative image of spinal cord blood vessels immunostained for CD31 (green-Alexa Fluor® 488) in aged mice treated with metformin (100 mg/kg) or PBS. Metformin markedly enhanced vascular density in aged mice after metformin administration compared with the PBS group. Red lines indicate the epicenter of the injury site. (B) Quantification of the CD31-labeled blood vessels. (C) Representative images of blood vessels immunostained with CD31 (green-Alexa Fluor® 488) and Ki67 (red-Alexa Fluor® 594) in the sham mice and in aged metformin-treated and PBS-treated mice 7 days post-SCI. Metformin promoted more vascular regeneration in aged mice after metformin administration compared with the PBS group. Scale bars: 150 μm in A, 50 μm in C. (D) Quantification of CD31⁺Ki67⁺ cells in the different groups. (E) Representative western blot images of p-AMPK, AMPK, p-eNOS, and eNOS expression in aged mice 7 days post-SCI. (F, G) Quantification of p-AMPK/AMPK and p-eNOS/eNOS expression in aged mice or the sham group treated with metformin or PBS. The data are presented as the mean ± SEM (n = 3 per group). #P < 0.05, ##P < 0.01 (two-tailed Student’s t-test [B]), or one-way analysis of variance followed by Tukey’s multiple comparisons test [D, E–G]). PBS: Phosphate buffer saline; SCI: spinal cord injury.