Abstract Body: Introduction: Infarct-induced neurodegeneration occurs chronically after stroke and can be prevented by inhibiting B cell influx to the brain. Vascular cell adhesion molecule 1 (VCAM1) facilitates immune cell diapedesis by binding very late antigen 4 (VLA4), and VCAM1 plasma levels are elevated after stroke in people and mice. We hypothesized that chronic treatment with anti-VCAM1 or anti-VLA4 would reduce lymphocyte trafficking and prevent cognitive decline.

Methods: Adult (3 month old) and middle-aged (10 month old) C57BL/6J male & female mice (N=9-10/group) underwent permanent middle cerebral artery occlusion and were dosed with anti-VCAM1, anti-VLA4 or control IgG every 3 days beginning 4 days after stroke. Sham mice received IgG only. Barnes maze and novel object were performed at -1, 1 and 6 weeks, and single-cell RNA sequencing was performed on immune and endothelial cells at 10 weeks. We quantified pericyte (CD13) vascular (CD31) coverage and extravascular fibrinogen leakage with immunostaining.

Results: Blocking either VCAM1 or VLA4 reduced B cells 67% or 55%, respectively, compared to IgG treated stroked mice three weeks after stroke. Stroked mice treated with IgG developed a cognitive deficit in both Barnes maze and novel object by 6 weeks, while both anti-VCAM1 and anti-VLA4 treated mice performed comparably to sham animals in the Barnes maze (p=0.398; p=0.972, respectively). Similarly, sham IgG (p<0.0001), stroke anti-VCAM1 (p<0.0001) and VLA4 (p=0.0059) could all perform the novel object task. Single-cell sequencing of immune and brain endothelial cells demonstrated that most transcriptional changes occur in endothelial cells and not immune cells. Anti-VCAM1 and anti-VLA4 both increase expression of BBB maintenance genes (cdh5, cldn5, ocln, pecam1, lama2) and decrease inflammatory genes CD74 and IL1r1. Blocking either VCAM1 or VLA4 increased pericyte vascular coverage by 67% (p=0.0113) and 65% (p=0.0215), respectively. Finally, extravascular fibrinogen leakage was also reduced 18% by anti-VCAM1 (p=0.031) and 15% by anti-VLA4 (p=0.0493).

Conclusion: Blocking either VCAM1 or VLA4 prevents infarct-induced neurodegeneration in middle-aged mice. BBB restoration via increased pericyte coverage and decreased immune cell trafficking into the brain likely both contribute to these therapeutic effects. Together, these findings establish the VCAM1-VLA4 axis as a promising target to restore BBB integrity and prevent infarct-induced neurodegeneration.