Interleukin‐6 Transactivation of the Mineralocorticoid Receptor

Abstract

Hypertension is characterized by increased sodium (Na+) reabsorption along the aldosterone‐sensitive distal nephron (ASDN) as well as a chronic systemic inflammation. Interleukin‐6 (IL‐6) is thought to be a mediator of this inflammatory process. Interestingly, increased Na+ reabsorption within the ASDN does not always correlate with increases in aldosterone (Aldo), the primary hormone which modulates Na+ reabsorption via the mineralocorticoid receptor (MR). Thus, understanding how increased ASDN Na+ reabsorption may occur independent of Aldo stimulation is critical. We hypothesize that IL‐6 can transactivate the MR, increasing Na+ reabsorption via reactive oxygen species (ROS) generation and activation of the sodium‐chloride cotransporter (NCC). We transfected MR‐EGFP tagged constructs into a model of murine distal convoluted tubule (mDCT15) cells. Cells were then treated with vehicle, Aldo (100–250nM) or IL‐6 (100ng/mL), fixed and visualized with confocal microscopy. MR‐EGFP expression was observed in the cytosol in the vehicle treated cells. However, following IL‐6 treatment (30 min), MR‐EGFP fluorescence was observed translocated into the nucleus, similar to Aldo‐treated positive controls. To confirm activation of downstream transcription factors, mDCT15 cells were transfected with mineralocorticoid response element (MRE)‐luciferase reporter constructs, treated with vehicle, Aldo (100nM, 24hr) or IL‐6 (100ng/mL, 24hr) and luciferase assays performed (48hrs). With IL‐6 treatment, luciferase activity was increased approximately 1.5 fold over MRE‐only transfected cells. Excessive MR stimulation is known to produce ROS generation. Thus, we determined if ROS generation stimulates increased NCC‐mediated Na+ uptake. Using mDCT15 cell monolayers, thiazide‐sensitive 22Na+‐uptake studies were performed. Increased 22Na+ uptake was observed after ROS generation (pyrogallol, 50μM) as compared to vehicle (1834±19 vs. 1506±18nmol/mg, p<0.0001, n=6), which was then reduced with the addition of the superoxide dismutase mimetic tempol (pyrogallol+tempol, 250μM) (1618±27nmol/mg vs. pyrogallol+tempol, p<0.0001, n=6). These data suggest that: IL‐6 can transactivate the MR leading to nuclear translocation and subsequent MRE activation, and ROS generation increases ASDN Na+ reabsorption via NCC providing evidence for alternate mechanisms of ASDN Na+ uptake during conditions where Aldo‐mediated MR stimulation may not occur.Support or Funding InformationNIDDK085097‐RSH2T32DK7656‐21 Emory Renal Division‐BMW