Met-RANTES Preserves the Blood-brain Barrier Through Inhibiting CCR1/SRC/Rac1 Pathway After Intracerebral Hemorrhage in Mice

Background: C-C chemokine receptor type 1 (CCR1) and its endogenous ligand, CCL5, participate in the pathogenesis of neuroinflammatory diseases. However, much remains unknown regarding CCL5/CCR1 signaling in blood-brain barrier (BBB) permeability after intracerebral hemorrhage (ICH).

Methods: A total of 250 CD1 male mice were used and ICH was induced via autologous whole blood injection. Either Met-RANTES, a selective CCR1 antagonist, or Met-RANTES combined with a Rac1 CRISPR activator was administered to the mice 1 h after ICH. Post-ICH assessments included neurobehavioral tests, brain water content, BBB integrity, hematoma volume, Western blot, and immunofluorescence staining. The CCR1 ligand, rCCL5, and SRC CRISPR knockout in naïve mice were used to further elucidate detrimental CCL5/CCR1/SRC signaling.

Results: Brain endogenous CCR1 and CCL5 were upregulated after ICH in mice with a peak at 24 h, and CCR1 was expressed in endothelial cells, astrocytes, and neurons. Met-R treatment reduced brain edema and neurobehavioral impairment, as well as preserved BBB integrity and tight junction protein expression in ICH mice. Met-R treatment decreased expression of p-SRC, Rac1, albumin, and MMP9, but increased claudin-5, occludin, and ZO-1 tight junction proteins after ICH. These effects were regressed using the Rac1 CRISPR activator. Administration of rCCL5 in naïve mice increased expression of p-SRC, Rac1, albumin, and MMP9, but decreased levels of claudin-5, occludin, and ZO-1 tight junction proteins. These effects in naïve mice were reversed with SRC CRISPR (KO).

Conclusions: Our findings demonstrate that CCR5 inhibition by Met-R improves neurological deficits after ICH by preserving BBB integrity through inhibiting CCR1/SRC/Rac1 signaling pathway in mice. Thus, Met-R has therapeutic potential in the management of ICH patients.