Brain immune cells undergo cGAS/STING-dependent apoptosis during herpes simplex virus type 1 infection to limit type I IFN production
Line S. Reinert, … , Georges MGM Verjans, Søren R. Paludan
Line S. Reinert, … , Georges MGM Verjans, Søren R. Paludan
Published September 29, 2020
Citation Information: J Clin Invest. 2021;131(1):e136824. https://doi.org/10.1172/JCI136824.
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Research Article Immunology Infectious disease

Brain immune cells undergo cGAS/STING-dependent apoptosis during herpes simplex virus type 1 infection to limit type I IFN production

Abstract

Protection of the brain from viral infections involves the type I IFN (IFN-I) system, defects in which render humans susceptible to herpes simplex encephalitis (HSE). However, excessive cerebral IFN-I levels lead to pathologies, suggesting the need for tight regulation of responses. Based on data from mouse models, human HSE cases, and primary cell culture systems, we showed that microglia and other immune cells undergo apoptosis in the HSV-1–infected brain through a mechanism dependent on the cyclic GMP-AMP synthase/stimulator of interferon genes (cGAS/STING) pathway, but independent of IFN-I. HSV-1 infection of microglia induced cGAS-dependent apoptosis at high viral doses, whereas lower viral doses led to IFN-I responses. Importantly, inhibition of caspase activity prevented microglial cell death and augmented IFN-I responses. Accordingly, HSV-1–infected organotypic brain slices or mice treated with a caspase inhibitor exhibited lower viral load and an improved infection outcome. Collectively, we identify an activation-induced apoptosis program in brain immune cells that downmodulates local immune responses.

Authors

Line S. Reinert, Ahmad S. Rashidi, Diana N. Tran, Georgios Katzilieris-Petras, Astrid K. Hvidt, Mette Gohr, Stefanie Fruhwürth, Chiranjeevi Bodda, Martin K. Thomsen, Mikkel H. Vendelbo, Ahmad R. Khan, Brian Hansen, Petra Bergström, Lotta Agholme, Trine H. Mogensen, Maria H. Christensen, Jens R. Nyengaard, Ganes C. Sen, Henrik Zetterberg, Georges MGM Verjans, Søren R. Paludan

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Figure 2

The cGAS-dependent cell death in the HSV-1–infected brain is primarily apoptosis.

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The cGAS-dependent cell death in the HSV-1–infected brain is primarily a...
(A–C, E, F, and H) Immunoblots for the indicated PCD markers in the brainstem from HSV-1–infected mice at day 5 after infection; vinculin was used as a loading control. (A and B) Both full-length gasdermin D (GSDMD) (fl) and the cleaved (CGSDMD) (cl*) product in the brainstem were detected with a polyclonal antibody (A), while the monoclonal antibody used (B) detects predominately the CGSDMD (cl*) product. The positive control used was bone marrow–derived macrophages (BMDM) treated in vitro with LPS (1 μg/mL) for 4 hours followed by an additional treatment with nigericin for 1 hour (10 μM). (C) Immunoblotting for caspase-1 showing the full-length caspase-1 (C1-fl) and the cleaved caspase-1 (P20)*. BMDM cells were treated as in B. (D, G, and J) Organotypic brain slices from WT and cGas–/– mice were cultured and infected with 1 × 104 PFU of HSV-1 for 20 hours, fixed and stained for HSV-1 (DP5) (red), GSDMD (monoclonal) antibody, P-MLKL, or cleaved caspase-3 (CC3), as indicated (all shown in green). Scale bar: 10 μm. (E and F) Immunoblots for the necroptosis markers P-MLKL and P-RIPK3 together with vinculin, total MLKL, and total RIPK3. (H and I) Immunoblotting for and quantification of CC3 bands normalized to vinculin bands presented as mean ± SEM per group (n = 8–10), P values were calculated by Wilcoxon rank-sum test. ***0.0001 < P < 0.001. (K) Levels of CC3 in all cells present in the brainstem were analyzed by flow cytometry and presented as mean ± SEM. n = 1–6 per group, P values were calculated by 1-way ANOVA with Tukey’s multiple-comparison test. ***0.0001 < P < 0.001; ****P < 0.0001. All results presented in this figure are representative for at least 3 independent experiments.