Elevated Levels of Homocysteine Compromise Blood-Brain Barrier Integrity in Mice.

Elevated levels of plasma homocysteine (Hcy) correlate with increased risks of cardiovascular and Alzheimer’s diseases. We studied the effect of elevated Hcy on the blood-brain barrier (BBB) to explore the possibility of a vascular link between the two diseases. On a methionine-enriched diet, cystathionine beta-synthase (CBS) heterozygous mice develop hyperhomocysteinemia (HHcy). To evaluate the extent of HHcy induced by an 8-week methionine-enriched diet (Diet), we determined plasma Hcy concentration. Concentrations were 23.5 ± 5 μM for wild type (WT) and 98.4 ± 22 μM for CBS+/− on Diet and 4.1 ± 0.20 μM for WT on normal chow. Inflammation can lead to BBB breakdown, therefore we first determined whether we could detect systemic endothelial activation/increased leukocyte adherence in the CBS+/− mice as an indicator of inflammation. We observed mesenteric venules (200–300 μm) in WT and CBS +/− mice fed Diet and compared them with WT on chow. Leukocyte rolling velocity was slower in CBS+/− than in WT mice indicating an increase in the density of adhesion molecules on the endothelium. In CBS+/− mice, 60% of leukocytes rolled at a velocity of < 10 μm/s as compared with only 8% in WT mice on Diet (P<0.002) and 3% WT on chow. The slower leukocyte velocity was due at least in part to higher expression of P-selectin on the endothelium. Infusion of fluorescent beads coated with antibody to P-selectin showed a several fold increase in their binding to venules of CBS +/− mice with HHcy as compared with WT on Diet (P<0.05). We also observed increased leukocyte adherence in CBS +/− mice (P<0.05). We then compared BBB function in mice after the 8-week Diet. We injected Evans blue (EB) dye intraperitoneally. EB binds to albumin and does not readily cross the BBB. The CBS+/− mice exhibited 25% greater EB in cortex compared with WT. EB leakage in WT mice was 3.19 ± 0.09, while CBS+/− leakage was 3.99 ± 0.29 (P < 0.03). WT mice on Diet showed slightly higher plasma Hcy levels than on normal chow; but this increase was not sufficient to break the BBB, as the BBB leakage in the WT mice on Diet was similar to that of WT mice on normal chow. Our study suggests an important toxic effect of elevated Hcy on brain microvessels and implicates Hcy in the disruption of the BBB.