MAP4K4 induces early blood-brain barrier damage in a murine subarachnoid hemorrhage model

Sterile-20-like mitogen-activated protein kinase kinase kinase kinase 4 (MAP4K4) is expressed in endothelial cells and activates inflammatory vascular damage. Endothelial cells are important components of the blood-brain barrier. To investigate whether MAP4K4 plays a role in the pathophysiology of subarachnoid hemorrhage, we evaluated the time-course expression of MAP4K4 after subarachnoid hemorrhage. A subarachnoid hemorrhage model was established using the intravascular perforation method. The model mice were assigned to four groups: MAP4K4 recombinant protein, scramble small interfering RNA, and MAP4K4 small interfering RNA were delivered by intracerebroventricular injection, while PF-06260933, a small-molecule inhibitor of MAP4K4, was administrated orally. Neurological score assessments, brain water assessments, Evans blue extravasation, immunofluorescence, western blot assay, and gelatin zymography were performed to analyze neurological outcomes and mechanisms of vascular damage. MAP4K4 expression was elevated in the cortex at 24 hours after subarachnoid hemorrhage, and colocalized with endothelial markers. MAP4K4 recombinant protein aggravated neurological impairment, brain edema, and blood-brain barrier damage; upregulated the expression of phosphorylated nuclear factor kappa B (p-p65) and matrix metalloproteinase 9 (MMP9); and degraded tight junction proteins (ZO-1 and claudin 5). Injection with MAP4K4 small interfering RNA reversed these effects. Furthermore, administration of the MAP4K4 inhibitor PF-06260933 reduced blood-brain barrier damage in mice, promoted the recovery of neurological function, and reduced p-p65 and MMP9 protein expression. Taken together, the results further illustrate that MAP4K4 causes early blood-brain barrier damage after subarachnoid hemorrhage. The mechanism can be confirmed by inhibiting the MAP4K4/NF-κB/MMP9 pathway. All experimental procedures and protocols were approved by the Experimental Animal Ethics Committee of General Hospital of Northern Theater Command (No. 2018002) on January 15, 2018.