Type I Interferon Response Is Mediated by NLRX1-cGAS-STING Signaling in Brain Injury

Abstract

BackgroundInflammation is a significant contributor to neuronal death and dysfunction following traumatic brain injury (TBI). Recent evidence suggests that interferons may be a key regulator of this response. Our studies evaluated the role of the Cyclic GMP-AMP Synthase-Stimulator of Interferon Genes (cGAS-STING) signaling pathway in a murine model of TBI.MethodsMale, 8-week old wildtype, STING knockout (−/−), cGAS−/−, and NLRX1−/− mice were subjected to controlled cortical impact (CCI) or sham injury. Histopathological evaluation of tissue damage was assessed using non-biased stereology, which was complemented by analysis at the mRNA and protein level using qPCR and western blot analysis, respectively.ResultsWe found that STING and Type I interferon-stimulated genes were upregulated after CCI injury in a bi-phasic manner and that loss of cGAS or STING conferred neuroprotection concomitant with a blunted inflammatory response at 24 h post-injury. cGAS−/− animals showed reduced motor deficits 4 days after injury (dpi), and amelioration of tissue damage was seen in both groups of mice up to 14 dpi. Given that cGAS requires a cytosolic damage- or pathogen-associated molecular pattern (DAMP/PAMP) to prompt downstream STING signaling, we further demonstrate that mitochondrial DNA is present in the cytosol after TBI as one possible trigger for this pathway. Recent reports suggest that the immune modulator NLR containing X1 (NLRX1) may sequester STING during viral infection. Our findings show that NLRX1 may be an additional regulator that functions upstream to regulate the cGAS-STING pathway in the brain.ConclusionsThese findings suggest that the canonical cGAS-STING-mediated Type I interferon signaling axis is a critical component of neural tissue damage following TBI and that mtDNA may be a possible trigger in this response.

Commentary: Type I Interferon Response Is Mediated by NLRX1-cGAS-STING Signaling in Brain Injury

“…Finally, Fritsch et al found greater gene expression of cGAS and STING in microglia compared to other brain cells, suggesting that microglia may be central to cGAS-STING activation (Supplementary Figure 5, Fritsch et al, 2022), which is an important observation consistent with the central role of microglia in neuroinflammation (Shao et al, 2022). However, others have also shown that astrocytes are involved in TBI-related responses (Burda et al, 2016;Michinaga and Koyama, 2021) and cGAS-STING activation (Jeffries and Marriott, 2017), and the current data do not rule out the contribution of other glial cells-especially since cell isolation protocols themselves can contribute to inflammatory microglial activation (Cadiz et al, 2022) and the authors did not report on cGAS-STING levels in adherent cells other than microglia.…”

“…STING phosphorylation can be inhibited by nucleotide-binding oligomerization domain leucine-rich repeat containing X1 (NLRX1; Guo et al, 2016), but until recently, the role of cGAS-STING activation and NLRX1 in TBI in vivo was unknown. Fritsch et al (2022) addressed this gap in knowledge using an in vivo model of TBI. Mice were subjected to a controlled cortical impact (CCI) injury, and the authors found that CCI injury increased IFN and pro-inflammatory transcripts, including STING transcripts, which remained elevated 24 h after injury.…”

“…For example, studies in humans have shown that nuclear dsDNA is implicated in TBI (Schwab et al, 2019 ) and cGAS-STING signaling (Li and Chen, 2018 ). Fritsch et al did measure cytosolic HMGB1 (Figure 2F, Fritsch et al, 2022 ), a nuclear dsDNA protein reported to be involved in TBI (Paudel et al, 2018 ), but they saw no difference between ipsilateral and contralateral cortices after CCI injury. However, this is only one marker of nuclear dsDNA, and its absence does not definitively confirm that nuclear dsDNA is not involved in cGAS-STING activation.…”

“…This ensures that all mice receive the same TBI, and it allows the contralateral side of the brain to be used as an “in-mouse” control. In fact, in Supplementary Figure 3, Fritsch et al showed no difference between sham (no injury) ipsilateral and CCI injury contralateral cortices (Fritsch et al, 2022 ). Thus, any observed changes were the result of TBI-related effects and not “inter-mouse” differences.…”

“…Fritsch et al ( 2022 ) addressed this gap in knowledge using an in vivo model of TBI. Mice were subjected to a controlled cortical impact (CCI) injury, and the authors found that CCI injury increased IFN and pro-inflammatory transcripts, including STING transcripts, which remained elevated 24 h after injury.…”