Abstract Body: Salt-Sensitive hypertension is associated with an increase in pro-inflammatory cytokines and immune cells. We previously showed an acute increase in systolic blood pressure following IL-6 + high salt (HS) after 3 days, coupled with an immediate (1 day) reduction in sodium excretion. Renal resident tissue macrophages (rRTMs) form an intricate network within the kidney and function to both maintain homeostasis and secrete pro-inflammatory cytokines. We hypothesize that rRTMs are primary contributors to the immediate increases in IL-6 following high salt.

To test this hypothesis, male wild-type mice lacking a functional CX₃CR₁ chemokine receptor (CX₃CR₁-^EGFP+/+) or wild-type (Wt, C57Bl6) were used. CX₃CR₁ is required for RTM localization to the kidney; thus, these mice are rRTM-depleted. Mice were fed HS (1% saline or 4% HS chow) or normal chow (NC) for 6 days and kidneys harvested. Kidneys were digested and FACS analysis performed on total immune cell populations or used for RT-PCR (n=4-5 Wt & rRTM-depleted). Cells were gated on: Live + CD45⁺ + MHCII + Lyc6C^- + B220^- + CD64⁺ + F4/F80^hi + CD11b^lo. Or Lyc6C^hi. Lyc6C^hi and rRTMs were pre-stimulated (LPS, 1mg/mL, 3hrs) before staining with intracellular IL-6 antibody. FACS data were analyzed as a percent increase (mean fluorescence intensity, MFI) of IL-6 over control. RT-PCR data were analyzed as DCt/housekeeping.

Our data show that rRTMs are the most abundant immune cell in the kidney (23.4% of total CD45⁺) and predominantly reside in the cortex and outer medulla (>99%). HS feeding (4%) significantly increased whole kidney IL-6 mRNA (12.0 ± 0.5 vs. 15.3 ± 0.6 DCt Wt NC, **p<0.01). This increase was completely prevented in rRTM-depleted mice. While intracellular IL-6 levels were generally increased in LPS-stimulated Lyc6C^hi inflammatory monocytes, the rRTMs showed a preferential increase in intracellular IL-6 with both hyperosmolar saline (1%, >13% increase) and HS chow (4%, >50%).

Here, we show that rRTM-depletion prevents the HS-induced increase in kidney IL-6. We additionally show that rRTMs are preferentially increasing intracellular levels of IL-6 following an acute HS load of hyperosmolar saline (1%) or HS chow (4%). We previously demonstrated that IL-6 contributes to increased blood pressure and sodium retention, and these data suggest that rRTMs preferentially contribute to the increased IL-6.