Fig 1: Iron import proteins were not increased, and no signs of BRB breakdown were found in 20-week-old db/db mouse retinas. (A) Western blot analysis revealed that neither transferrin nor its receptor was increased in db/db mouse retinas. SCARA5, the receptor of L-ferritin, was also not increased in db/db mouse retinas. As a loading control, α-tubulin was used. (B) Analysis of the eye fundus by Scanning Laser Ophthalmoscopy (SLO) showed that injected fluorescein remained inside the retinal blood vessels in db/db mice showing no vascular leakage (n = 16). (C) To assess retinal vessels leakage microscopically, immunohistochemistry against serum albumin and collagen IV, a marker for blood basement membrane, was performed. The results showed that serum albumin was restricted inside the vessel lumen in db/db mice. (D) Western blot analysis of VEGF expression, which induces vascular permeability, showed no statistical differences between db/+ and db/db retinas. As a loading control, α-tubulin was used. GCL, ganglion cell layer; IPL, inner plexiform layer; INL, inner nuclear layer; OPL, outer plexiform layer; ONL, outer nuclear layer. Scale bar: 29.41 µm (inset: 11.49 µm).