FTO Regulates Microglia-Induced Inflammation by Stabilizing ADAM17 Expression After Experimental Traumatic Brain Injury

Background The neuroinflammatory response mediated by microglial polarization plays an important role in the secondary nerve injury of traumatic brain injury (TBI). The post-transcriptional modification of n6-methyladenosine (m6A) is ubiquitous in the immune response of the central nervous system. The fat mass and obesity (FTO)-related protein can regulate the splicing process of pre-mRNA. However, after experimental traumatic brain injury (TBI), the role of FTO in microglial polarization and the subsequent neuroinflammatory response is still unclear. Methods TBI mice model was established by the Feeney weight-drop method. Neurological severity score, brain water content measurement and Nissl staining were used to detect the role of FTO in microglial polarization and the molecular mechanism of targeted RNA epigenetic modification. In vitro and in vivo experiments were conducted to evaluate microglial polarization and the neuroinflammatory response by down-regulation of FTO expression. The level of m6A modification in M1 activated microglia was detected by qRT-PCR, m6A-MeRIP and m6A high-throughput sequencing. Fluorescent in situ hybridization combined with immunofluorescence imaging were used to detect the epigenetic regulation of ADAM17 mediated by an FTO-m6A-dependent mechanism. Results The expression of FTO was significantly down-regulated in BV2 cells treated with lipopolysaccharide and mice with TBI. Down-regulation of FTO expression increased the level of m6A in M1 microglia at the level of the entire transcriptome. Meanwhile, after FTO interference, M1/M0 phenotype detection experiments revealed the BV2 cells shifted from an M0 to M1 phenotype as the population rate of CD11b+/CD86+ increased and secretion of pro-inflammatory cytokines was enhanced. Methylated RNA immunoprecipitation assay showed that the m6A peaks located in the ADAM17 and TNF-α genes increased. Taken together, the results indicated that FTO can affect the transcription modification of ADAM17 and the expression of the downstream TNF-α/NF-kB pathway. In turn, ADAM17 can block the M1-phenotypic transition of microglia driven by FTO-m6A modification. Conclusions The down-regulation of FTO expression leads to the abnormally high expression of ADAM17 in microglia. The activation of microglia and neuroinflammatory response regulated by FTO-related m6A modification play an important role in the early pro-inflammatory process of TBI secondary injury.

Angiotensin II Receptor Type 2 Activation Does Not Influence Brain Damage, Neurological Impairment and Inflammation After Experimental Traumatic Brain Injury

Antagonism of the angiotensin II type 1 receptor (AT1) improves neurological function and reduces brain damage after experimental traumatic brain injury (TBI), which may be partly a result of enhanced indirect angiotensin II type 2 receptor (AT2) stimulation. AT2 stimulation was demonstrated to act neuroprotective via anti-inflammatory, vasodilatory, and neuroregenerative mechanisms in experimental cerebral pathology models. We recently demonstrated an upregulation of AT2 after TBI suggesting a protective mechanism. The present study investigated the effect of post-traumatic (5 days after TBI) AT2 activation via high and low doses of a selective AT2 agonist, compound 21 (C21), compared to vehicle-treated controls. No differences in the extent of the TBI-induced lesions were found between both doses of C21 and the controls. We then tested AT2-knockdown animals for secondary brain damage after experimental TBI. Lesion volume and neurological outcomes in AT2-deficient mice were similar to those in wild-type control mice at both 24 hours and 5 days post-trauma. Thus, in contrast to AT1 antagonism, AT2 modulation does not influence the initial pathophysiological mechanisms of TBI in the first 5 days after the insult, indicating that AT2 plays only a minor role in the early phase following trauma-induced brain damage.

Mutated neuronal voltage-gated CaV2.1 channels causing familial hemiplegic migraine 1 increase the susceptibility for cortical spreading depolarization and seizures and worsen outcome after experimental traumatic brain injury

Patients suffering from familial hemiplegic migraine type 1 (FHM1) may have a disproportionally severe outcome after head trauma, but the underlying mechanisms are unclear. Hence, we subjected knock-in mice carrying the severer S218L or milder R192Q FHM1 gain-of-function missense mutation in the CACNA1A gene that encodes the α1A subunit of neuronal voltage-gated CaV2.1 (P/Q-type) calcium channels and their wild-type (WT) littermates to experimental traumatic brain injury (TBI) by controlled cortical impact (CCI) and investigated cortical spreading depolarizations (CSDs), lesion volume, brain edema formation, and functional outcome. After TBI, all mutant mice displayed considerably more CSDs and seizures than WT mice, while S218L mutant mice had a substantially higher mortality. Brain edema formation and the resulting increase in intracranial pressure was more pronounced in mutant mice, while only S218L mutant mice had larger lesion volumes and worse functional outcome. Here we show that gain of CaV2.1 channel function worsens histopathological and functional outcome after TBI in mice. This phenotype was associated with a higher number of CSDs, increased seizure activity, and more pronounced brain edema formation. Hence, our results suggest increased susceptibility for CSDs and seizures as potential mechanisms for bad outcome after TBI in FHM1 mutation carriers.