Purpose: Microglia and astrocytes are crucial neuroimmune cells that modulate the neuroinflammatory response following TBI. Using C34, a novel pharmacologic TLR4 inhibitor, we investigated the role of these cells in a murine TBI model.
Methods: A murine controlled cortical impact model was utilized, and the results were analyzed on post-injury days (PID) 1, 7, 28 and 35. The experimental groups are: (1) Sham, (2) Sham+C34, (3) TBI and (4) TBI+C34. Real-time PCR (qRT-PCR) was used to quantify gene expression associated with the different polarization states of microglia (M1 and M2) and astrocytes (A1 and A2). Fixed frozen cortical samples were sectioned, stained for MBP, NG2 and DAPI then imaged. Neurocognitive outcomes were evaluated using the Morris Water Maze (MWM) test. Student’s t-test and one-way ANOVA were used for statistical analysis with significance achieved when p< 0.05.
Results: On qRT-PCR, C34 treated groups showed a significant decrease in the expression of proinflammatory A1 and M1 markers and a significant increase in the expression of anti-inflammatory A2 markers during acute inflammation. Our results also showed a substantial downregulation in the expression of A1, A2 and M2 markers with C34 treatment during chronic inflammation. On immunohistochemistry (IHC), C34 treated groups demonstrated increased MBP staining into the lesion by PID28. C34 treated groups also showed more platform entries on MWM on PID7 and 28 when compared to untreated groups.
Conclusion: Following TBI, TLR4 inhibition attenuates the secondary dysregulated neuroinflammatory response and improves neurocognitive outcomes by promoting myelin regeneration. Moreover, although A2 astrocytes and M2 microglia have been consistently shown as inducers of neuroprotection, our results clearly demonstrate their detrimental role during the chronic phases of healing post-TBI.
