Chemokine Receptors CC Chemokine Receptor 5 and C-X-C Motif Chemokine Receptor 4 Are New Therapeutic Targets for Brain Recovery after Traumatic Brain Injury

Recently, chemokine receptor CC chemokine receptor 5 (CCR5) was found to be a negative modulator of learning and memory. Its inhibition improved outcome after stroke and traumatic brain injury (TBI). To better understand its role after TBI and establish therapeutic strategies, we investigated the effect of reduced CCR5 signaling as a neuroprotective strategy and of the temporal changes of CCR5 expression after TBI in different brain cell types. To silence CCR5 expression, ccr5 short hairpin RNA (shRNA) or dsred shRNA (control) was injected into the cornu ammonis (CA) 1 and CA3 regions of the hippocampus 2 weeks before induction of closed-head injury in mice. Animals were then monitored for 32 days and euthanized at different time points to assess lesion area, inflammatory components of the glial response (immunohistochemistry; IHC), cytokine levels (enzyme-linked immunosorbent array), and extracellular signal-regulated kinase (ERK) phosphorylation (western blot). Fluorescence-activated cell sorting (FACS) analysis was performed to study post-injury temporal changes of CCR5 and C-X-C motif chemokine receptor 4 (CXCR4) expression in cortical and hippocampal cell populations (neurons, astrocytes, and microglia). Phosphorylation of the N-methyl-d-aspartate subunit 1 (NR1) subunit of N-methyl-d-aspartate (western blot) and cAMP-response-element-binding protein (CREB; IHC) were also assessed. The ccr5 shRNA mice displayed reduced lesion area, dynamic alterations in levels of inflammation-related CCR5 ligands and cytokines, and higher levels of phosphorylated ERK. The ccr5 shRNA also reduced astrocytosis in the lesioned and sublesioned cortex. FACS analysis revealed increased cortical CCR5 and CXCR4 expression in CD11b-positive cells, astrocytes, and neurons, which was most evident in cells expressing both receptors, at 3 and 11 days post-injury. The lowest levels of phosphorylated NR1 and phosphorylated CREB were found at day 3 post-injury, suggesting that this is the critical time point for therapeutic intervention.