IL‐17 Signaling Mediates Cytolytic Natural Killer Cell Activation in Placental‐Ischemic Rats

Abstract

The Reduced Uterine Perfusion Pressure (RUPP) rat model of placental ischemia mimics many characteristics of preeclampsia (PE) including hypertension (HTN), fetal growth restriction, and increased TH17s, IL‐17, and cytolytic natural killer cells (cNKs). IL‐17 stimulates NK cytotoxicity in vitro. This study tested the hypothesis that blockade of IL‐17 signaling would inhibit activation of cNKs to improve blood pressure and fetal growth in RUPP rats. On gestation day (GD) 14, osmotic pumps infusing IL‐17 RC (100 pg/d), a soluble receptor for IL‐17, were implanted into pregnant rats undergoing RUPP (RUPP+IL17 RC). On GD 19, circulating and placental cNKs were quantified via flow cytometry in normal pregnant (NP, n=9), RUPP (n=9), and RUPP+IL17 RC (n=9) rats. cNK activation, placental ROS, fetal and placental weights, circulating and placental cytokines, and MAP were also assessed. Placental cNKs (% gated), increased from 7.2±2.8% in NP to 16.6±3.3% in RUPP (p<0.05 vs NP), and were normalized to 6.8±1.7% after IL‐17 blockade (p<0.05 vs RUPP). Similarly, circulating cNKs increased from 6.1±1.4% in NP to 16.7±3.2% in RUPP (p<0.05 vs NP) and decreased to 8.1±1.9% in RUPP+IL‐17 RC (p<0.05 vs RUPP). Placental levels of the cNK cytokine, TNFα, increased from 25.9±3.0 pg/mg in NP to 44.8±5.0 pg/mg in RUPP (p<0.05 vs NP) and was normalized to 28.5±4.6 pg/mg in RUPP+IL‐17 RC (p<0.05 vs RUPP). Similarly, placental levels of MIP3a, a cNK chemokine, increased from 0.84±0.1 pg/mg in NP to 1.4±0.2 pg/mg in RUPP (p<0.05 vs NP) and decreased to 0.76±0.1 pg/mg in RUPP+IL‐17 RC (p<0.05 vs RUPP). Assessment of in vitro cytotoxic activity of placental NKs from rats in each group demonstrated a 5‐fold increase in cytotoxicity by RUPP NKs (n=6) compared to NP NKs (n=6) (p<0.05 vs NP). Cytotoxicity of NKs from RUPP+IL‐17 RC rats (n=6) was not significantly higher than NP NKs (3‐fold higher, p>0.05 vs NP or RUPP). Circulating levels of VEGF decreased from 163±11 pg/mL in NP to 88±16 pg/mL RUPP (p<0.05 vs NP) and increased to 195±12 pg/mL in RUPP+IL‐17 RC (p<0.05 vs RUPP). Soluble fms‐like tyrosine kinase 1 (sFlt‐1), a VEGF antagonist, increased from 196±100 pg/mL in NP to 963±114 pg/mL in RUPP (p<0.05 vs NP) and decreased to 249±78 pg/mL in RUPP+IL‐17 RC (p<0.05 vs RUPP). Placental ROS (RLU/min/mg) was increased from 890±218 in NP (n=6) to 2240±319 in RUPP (n=6, p<0.05 vs NP) and decreased to 1145±137 with IL‐17 blockade (n=5, p<0.05 vs RUPP). Fetal weight decreased from 2.4±0.04 g in NP to 2.0±0.06 g in RUPP (p<0.05 vs NP) and increased to 2.2±0.04 g in RUPP+IL17 RC (p<0.05 vs RUPP). Placental weights followed a similar trend. MAP increased from 90±2 mmHg in NP to 122±3 mmHg in RUPP (p<0.05 vs NP), and was decreased to 103±3 mmHg in RUPP+IL17 RC (p<0.05 vs RUPP, p<0.05 vs NP). These data demonstrate a direct role for IL‐17 signaling to mediate cNK activation in response to placental ischemia during pregnancy. By better understanding this stimulus, novel therapeutics can be developed to inhibit cytolytic NK activation while also retaining the critical roles of normal NKs in pregnancy maintenance and immune defense. Targeting IL‐17 signaling may be effective in ameliorating several pathophysiologies of PE and could improve both maternal and fetal outcomes.Support or Funding InformationR00HL130456, T32HL105324