Dataset Information

Met-RANTES preserves the blood-brain barrier through inhibiting CCR1/SRC/Rac1 pathway after intracerebral hemorrhage in mice.

ABSTRACT:

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.

SUBMITTER: Yan J

PROVIDER: S-EPMC8781527 | biostudies-literature |

REPOSITORIES: biostudies-literature

ACCESS DATA

Similar Datasets

Project description:Blood-brain barrier (BBB) disruption and subsequent brain edema play important roles in the secondary neuronal death and neurological dysfunction that are observed following intracerebral hemorrhage (ICH). In previous studies, transient receptor potential vanilloid 4 (TRPV4), a calcium-permeable mechanosensitive channel, was shown to induce cytotoxicity in many types of cells and to play a role in orchestrating barrier functions. In the present study, we explored the role of TRPV4 in ICH-induced brain injury, specifically investigating its effect on BBB disruption. Autologous arterial blood was injected into the basal ganglia of rats to mimic ICH. Adult male Sprague Dawley rats were randomly assigned to sham and experimental groups for studies on the time course of TRPV4 expression after ICH. The selective TRPV4 antagonist HC-067047 and TRPV4 siRNA were administered to evaluate the effects of TRPV4 inhibition. GSK1016790A, a TRPV4 agonist, was administered to naive rats to verify the involvement of TRPV4-induced BBB disruption. A PKC inhibitor, dihydrochloride (H7), and a selective RhoA inhibitor, C3 transferase, were administered to clarify the involvement of the PKCα/RhoA/MLC2 pathway following ICH. Post-ICH assessments including functional tests, brain edema measurements, Evans blue extravasation, western blotting and immunohistochemical assays were performed. TRPV4 inhibition remarkably ameliorated neurological symptoms, brain edema, and neuronal death, as well as BBB disruption, 24-72 h following ICH. Meanwhile, TRPV4 blockade preserved the expression of adherens and tight junction proteins, as well as BBB integrity, by inhibiting stress fiber formation, which might be correlated with the regulation of components of the PKCα/RhoA/MLC2 pathway. Furthermore, adherens and tight junction protein degradation induced by GSK1016790A treatment in naive rats was also related to PKCα/RhoA/MLC2-pathway-mediated stress fiber formation. Based on these findings, therapeutic interventions targeting TRPV4 may represent a novel approach to ameliorate secondary brain injury following ICH.

Project description:The activation of C-C chemokine receptor type 1 (CCR1) has been shown to be pro-inflammatory in several animal models of neurological diseases. The objective of this study was to investigate the activation of CCR1 on neuroinflammation in a mouse model of intracerebral hemorrhage (ICH) and the mechanism of CCR1/tetratricopeptide repeat 1 (TPR1)/extracellular signal-regulated kinase 1/2 (ERK1/2) signaling pathway in CCR1-mediated neuroinflammation. Adult male CD1 mice (n = 210) were used in the study. The selective CCR1 antagonist Met-RANTES was administered intranasally at 1 h after autologous blood injection. To elucidate potential mechanism, a specific ERK1/2 activator (ceramide C6) was administered prior to Met-RANTES treatment; CCR1 activator (recombinant CCL5, rCCL5) and TPR1 CRISPR were administered in naïve mouse. Neurobehavioral assessments, brain water content, immunofluorescence staining, and western blot were performed. The endogenous expressions of CCR1, CCL5, TPR1, and p-ERK1/2 were increased in the brain after ICH. CCR1 were expressed on microglia, neurons, and astrocytes. The inhibition of CCR1 with Met-RANTES improved neurologic function, decreased brain edema, and suppressed microglia/macrophage activations and neutrophil infiltration after ICH. Met-RANTES treatment decreased expressions of CCR1, TPR1, p-ERK, TNF-α, and IL-1β, which was reversed by ceramide C6. The brain CCR1 activation by rCCL5 injection in naïve mouse resulted in neurological deficits and increased expressions of CCR1, TPR1, p-ERK, TNF-α, and IL-1β. These detrimental effects of rCCL5 were reversed by TPR1 knockdown using TPR1 CRISPR. Our study demonstrated that CCR1 activation promoted neuroinflammation through CCR1/TPR1/ERK1/2 signaling pathway after ICH in mice. CCR1 inhibition with Met-RANTES attenuated neuroinflammation, thereby reducing brain edema and improving neurobehavioral functions. Targeting CCR1 activation may provide a promising therapeutic approach in the management of ICH patients.