Role of Natural Killer Cells as Mediators of Mitochondrial Dysfunction Induced by Placental Ischemia in the Reduced Uterine Perfusion Pressure (RUPP) Rat Model of Preeclampsia
Authors: Vaka Venkata Ramana , McMaster Kristen M , Cunningham Mark W , Amaral Lorena M , Ibrahim Tarek , Cornelius Denise C , LaMarca Babbette 
Published: November 2023

Introduction

Preeclampsia (PE), is characterized by new onset hypertension and is associated with immune activation and placental oxidative stress. Placental ischemia is believed to be the initial event in the development of PE. Cytolytic Natural Killer (NK) cells are elevated in the placentas of PE women. One mechanism of cytotoxicity of NK cells is release of proteins modulating mitochondrial dysfunction and oxidative stress which may play a role in the pathophysiology of PE. We have shown that placental ischemia induces NK cell activation in the reduced uterine perfusion pressure (RUPP) rat model of PE. Thus, we hypothesize that NK cells depletion could improve oxidative stress, mitochondrial function and blood pressure in RUPP rats.

Methods

Pregnant Sprague Dawley rats were divided into three groups; normal pregnant (NP) and RUPP and RUPP+NK cell depletion rats (RUPP+NKD). On gestational day (GD) 14, RUPP surgery was performed, and NK cells were depleted with Anti‐asialo GM1 antibodies (7μg/100μL, i.p) on Gday 15 and Gday 17. On GD19 conscious blood pressure (MAP) was measured, placentas were collected and mitochondria were isolated. Mitochondrial function was assessed by studying respiration and Complex I activities. The reduced rates of respiration and complex I activity indicates mitochondrial dysfunction. Respiration measurements included: basal state (isolated mitochondria with no substrates), state 2 (glutamate and malate as complex I substrates), state 3 (ADP stimulated), leak state (oligomycin induced ATP synthase inhibition), and maximal state (FCCP stimulated uncoupled) were obtained using Oxygraph‐2k. Complex I activity was measured using the spectrophotometer, data are expressed as mean±SEM, statistical analysis included one way ANOVA and Bonferroni post hoc test.

Results

MAP was elevated in RUPP (n=9) compared to NP rats (n=10) (125±3 mmHg vs. 109±2 mmHg, p<0.05) which was normalized in RUPP+NKD (n=3; 106±6 mmHg). Placental state 3 (313±16 vs. 423±15 pmol/sec/mg, p<0.05) and uncoupled (244±13 vs. 300±11 pmol/sec/mg, p<0.05) respiration rates were significantly reduced in RUPP (n=7) vs. NP (n=8) but improved in RUPP+NKD (n=3; 398±28 pmol/sec/mg, p<0.05 vs RUPP). However, there was no change in uncoupled respiration. Placental mitochondrial respiratory control ratio (RCR) (state 3/ state 4) was significantly reduced in RUPP (n=7) vs. NP (n=8) (7±1 vs 11±1, p<0.05) but was significantly improved in RUPP+NKD (n=3; 12±2, p<0.05 vs. RUPP). Complex I (12±3 vs. 23 ± 2 nmol e‐/min/mg, P<0.05) was drastically reduced in RUPP compared to NPs but was increased in RUPP+NKD (n=2; 27±10, p=0.1614 vs RUPPs) but did not reach statistical significance.

Conclusion

The reduction in mitochondrial respiration respiratory and activity were improved with NK cell depletion in RUPP rats indicating the importance NK cells play in causing placental mitochondrial dysfunction and pathophysiology in response to placental ischemia of pregnancy.

Support or Funding Information

NIH grant RO1HD067541 (BL)