Abstract MP38: High Salt Activates The NLRP3 Inflammasome In Antigen Presenting Cells Via ENaC To Promote Salt-Sensitive Hypertension

Abstract

Salt-sensitivity is a major risk factor for hypertension and cardiovascular disease (CVD). Reducing dietary Na + decreases blood pressure and CVD risk. However, the precise mechanisms of how Na + leads to hypertension are still not well defined. Recently, we found that dendritic cells (DCs) in response to increases in extracellular [Na + ] exhibit an amiloride-sensitive epithelial Na + channel (ENaC)-dependent activation of NADPH-oxidase, superoxide production, reactive isolevuglandin (IsoLG)-protein adduct formation, and cytokine secretion which promote hypertension. We hypothesized that the NLRP3 inflammasome in antigen-presenting cells (APCs) mediates salt-sensitive hypertension through an ENaC-dependent mechanism. To test this hypothesis, we cultured mouse splenocytes in normal-salt or high-salt (HS) media with or without co-treatment with the ENaC inhibitor, amiloride (20 μM). Using flow cytometry, we found that HS increased monocyte and DC IL-1β production, which was confirmed through an ELISA assay detecting IL-1β release (2.131 ± 0.733 vs 12.75 ± 1.108 pg/mL, p<0.01) into the culture media, and amiloride treatment prevented IL-1β production (12.75 ± 1.108 vs 1.905 ± 0.3495 pg/mL, p<0.01) in these cells. To confirm our in vitro data, we treated salt-sensitive mice on a 129-SvJ background with a HS diet (4% NaCl) for 28 days with or without amiloride (1mg/kg/day in drinking water) or NLRP3 inflammasome inhibitor MCC950 (10mg/kg i.p.). Amiloride or MCC950 treated mice developed blunted hypertension in response to HS (120.4 ± 2.99; 101.0 ± 3.74) compared to vehicle controls (140.5 ± 3.98). Amiloride treated mice also exhibited less expression of NLRP3, pro-IL1β, and IsoLGs in DCs and monocytes compared to controls. Interestingly, MCC950 treated mice exhibited decreased pro-IL1β but not NLRP3 expression or IsoLG production. Using the DOCA-salt model, we found similar increases in NLRP3, pro-IL1β, and IsoLGs expression in DCs and monocytes, which was abolished after treatment with IsoLG scavenger 2-HOBA (1g/L). Our findings suggest a role for ENaC-dependent NLRP3 inflammasome activation in APCs in response to a HS diet, which may represent a promising treatment approach to salt-induced hypertension.