Mechanisms of Innate Immune Injury in Arrhythmogenic Cardiomyopathy

Abstract

Inhibition of nuclear factor kappa-B (NF kappa B) signaling prevents disease in Dsg2mut/mut mice, a model of arrhythmogenic cardiomyopathy (ACM). Moreover, NF kappa B is activated in ACM patient-derived iPSC-cardiac myocytes under basal conditions in vitro. Here, we used genetic approaches and sequencing studies to define the relative pathogenic roles of immune signaling in cardiac myocytes vs. inflammatory cells in Dsg2mut/mut mice. We found that NF kappa B signaling in cardiac myocytes drives myocardial injury, contractile dysfunction, and arrhythmias in Dsg2mut/mut mice. It does this by mobilizing cells expressing C-C motif chemokine receptor-2 (CCR2+ cells) to the heart, where they mediate myocardial injury and arrhythmias. Contractile dysfunction in Dsg2mut/mut mice is caused both by loss of heart muscle and negative inotropic effects of inflammation in viable muscle. Single nucleus RNA sequencing and cellular indexing of transcriptomes and epitomes (CITE-seq) studies revealed marked pro-inflammatory changes in gene expression and the cellular landscape in hearts of Dsg2mut/mut mice involving cardiac myocytes, fibroblasts and CCR2+ cells. Changes in gene expression in cardiac myocytes and fibroblasts in Dsg2mut/mut mice were modulated by actions of CCR2+ cells. These results highlight complex mechanisms of immune injury and regulatory crosstalk between cardiac myocytes, inflammatory cells, and fibroblasts in the pathogenesis of ACM.

Inhibition of Soluble Epoxide Hydrolase Reduces Inflammation and Myocardial Injury in Arrhythmogenic Cardiomyopathy

“…3 It also rescues the disease phenotype in Dsg2 mut/mut mice. 3,4 To determine if inhibition of sEH also mitigates the ACM disease phenotype and turns off NFκB signaling in vitro , we incubated cultures of rat myocytes expressing JUP2157del2 with the sEH inhibitor TPPU or the dual COX-2/sEH inhibitor PTUPB. As shown in Figure 4 , TPPU and PTUPB both restored the normal cell surface distribution of plakoglobin and Cx43 in rat myocytes expressing mutant JUP and eliminated nuclear signal for RelA/p65 indicating that NFκB signaling was reduced.…”

“…29 Importantly, both SPMs and EpFAs, synergized by inhibition of sEH, potently down-regulate NFκB pathways, 29,30 which are persistently activated in cardiac myocytes in a cell-autonomous fashion in ACM. [3][4][5][6][7] Increasing evidence suggests that chronic inflammation in ACM is the result of deficient resolution. The consistent presence of inflammatory infiltrates in the hearts of ACM patients obviously implicates immune mechanisms.…”

“…Studies were performed in wild type (WT) mice and mice with homozygous knock-in of a variant in Dsg2 , the gene encoding the desmosomal cadherin desmoglein-2 ( Dsg2 mut/mut mice) as previously described. 3,4,6,7 This variant entails loss of exons 4 and 5 which causes a frameshift and premature termination of translation.…”

“…9-week-old animals underwent echocardiography before being implanted with intraperitoneal osmotic mini-pumps (Alzet, Model 1004) as previously described. 3,4,16 They received either vehicle or drug (TPPU dissolved in DMSO/polyethylene glycol). Drug-treated mice received 5mg/kg/day TPPU via continuous infusion (1µL/hour for 28 days); vehicle-treated mice received an equivalent volume of vehicle for 28 days.…”

“…These data further suggest that cyclooxygenase inhibitors can synergize with sEH inhibitors in reversing disease features in ACM and support previous observations regarding multiple pro-inflammatory pathways in ACM. 3,4 To determine if inhibition of sEH can promote resolution of inflammation and limit myocardial injury in vivo, we treated 9-week old Dsg2 mut/mut mice for 4 weeks with TPPU and monitored effects on left ventricular contractile function and immune signaling. As shown in Figure 5, disease progressed during the 4-week treatment interval in Dsg2 mut/mut mice given vehicle.…”