The pathogenesis of radiation (XRT) induced cardiac toxicity remains poorly understood. We sought to identify the molecular mechanisms of late cardiac toxicity using a mouse model with goal of elucidating potential therapeutic targets. Protocols were approved by our institution’s animal care and use committee. Cardiac XRT was delivered to adult wild type C57BL/6 mice aged 8-12 weeks via a precision small animal irradiator (X-Rad 225Cx). Principal cardiac tissues were acutely harvested and enzymatically isolated into cell type specific populations and analyzed by RNAseq and quantitative RT-PCR. Subsequent studies were carried out using Cgas-/-, Tmem173-/-, Ifnar-/-, and Mavs-/- strains (Jackson Laboratories). Acutely harvested heart tissues following survival studies were examined by histopathologic evaluation. Statistical analyses were carried out using t-test with multiple comparison adjustment where appropriate, and log-rank method for survival analysis. RNAseq analysis of cardiac fibroblasts, myocytes and endothelium after cardiac XRT with 12 Gy revealed strong activation of Irf7, an interferon activating transcription factor, in fibroblasts (p < 10-78) and to a lesser degree in myocytes one month after XRT, but not in endothelial cells. GSEA analysis of all upregulated genes 28 days after XRT revealed enrichment for multiple Ifn transcription factors in fibroblasts (p < 10-21), but not in endothelium. Because radiation might elicit specific damage associated molecular pattern (DAMP) signaling, we hypothesized that this Ifn signaling resulted from cGAS/STING activation following DNA damage. Indeed, RT-PCR for Irf7 in cardiac fibroblasts from wild-type and Mavs-/- (which are deficient in RNA mediated DAMP recognition) mice showed strong activation (p < 0.001 compared to sham XRT), but Cgas-/- animals lacked this response (p = NS compared to sham XRT, p < 0.001 compared to wild-type and Mavs-/- after XRT). Cgas-/- (HR = 0.40, p = 0.034) and Tmem173-/- (HR = 0.15, p = 0.002) animals had significantly improved survival after cardiac XRT as compared to wild-type mice, while Ifnar-/- animals showed a trend for improvement as well (HR = 0.47, p = 0.094). Histopathologic analysis of cardiac tissues at time of death revealed consistent widespread atrial and ventricular mural thrombus formation and cardiomyocyte atrophy in irradiated wild-type animals, but markedly reduced in Tmem173-/- counterparts. Our system establishes a contemporary, genetically defined mouse model of XRT-induced cardiac toxicity. In doing so, we find cardiac irradiation elicits cGAS/STING- (and not MAVS-) dependent, delayed, chronic interferon activation in cardiac fibroblasts. Eliminating this signaling rescues radiation induced cardiac toxicity and improves survival. Inhibitors the cGAS/STING pathway are soon to begin early stage clinical trials and may be candidate drugs for mitigation of late cardiac toxicity following radiotherapy.
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