B Cell Depletion During Pregnancy Improves Hypertension, Natural Killer Cell Activation, and May Not Worsen Fetal Outcomes in Response to Placental Ischemia

Abstract

Preeclampsia (PE), new onset hypertension during pregnancy, is the leading cause of death and morbidity world-wide for the mother and fetus. Women with PE have elevated inflammatory cytokines, cytolytic natural killer (cNK) cells, and B cells producing agonistic antibodies to the angiotensin II type I receptor (AT1-AA). The Reduced Uterine Perfusion Pressure Rat Model of PE (RUPP) exhibits many characteristics of PE. We have shown Rituximab, a chimeric monoclonal antibody used clinically for B cell depletion, lowers blood pressure, AT1-AA, and cytokines in RUPP rats. However, we don't know the effect of maternal B cell depletion on placental cNK cells, or offspring survival and growth. Moreover, recent studies have touted that Rituximab doesn't work in rodents nor are B cells important for hypertension in RUPP rats. Therefore, we hypothesize that Rituximab will deplete circulating B cells and lower AT1-AA stimulated cNK cells and blood pressure in response to placental ischemia in RUPP rats. Rituximab (250 mcg/kg) was given on gestation day (GD) 14 via mini-osmotic pump. Blood and tissues were collected, blood pressure (MAP), pup weight, and NK cells were measured by flow cytometry in the blood and placenta on GD 19. A separate group of dams were allowed to deliver and birth weight of pups were recorded within 12 hours. Weights were measured weekly until 12 weeks. A one-way ANOVA was used for statistical analysis. On GD19, circulating B cells were 15.67±2.28 % gate (n=14) in RUPP rats which was elevated compared to NP rats, 7.04±2.3 % gate (n=6, p<0.05), and was normalized with Rituximab (4.21±0.61 % gate (n=3, p<0.05)). MAP increased in RUPP 123±2 mmHg (n=19, p<0.05) compared to NP controls 101±1 mmHg (n=18) and was normalized with Rituximab 106±3 mmHg (n=8, p<0.05). Circulating and placental cNK cells were 0.22±0.14, 0.64±0.48 % gate in NP rats (n=9, n=6), 1.36±0.32, 1.83±0.49 % gate in RUPP rats (n=16 p<0.05, n=10), which was decreased to 0.01±0.01, 0.81±0.50 % gate with Rituximab (n=5, p<0.05, n=5). RUPP pup weight, 1.78±0.06 g (n=19), was reduced compared to NP pup weight, 2.29±0.14 g (n=18, p<0.05) and was unchanged by Rituximab, 1.82±0.16 g (n=7). RUPP and Rituximab male and female offspring were smaller at birth than NP offspring. Although there no differences among the female offspring, RUPP and Rituximab male offspring remained smaller at 12 weeks of age compared to NP offspring. Our findings indicate that Rituximab indeed lowers maternal circulating B cells, cNK cells and blood pressure in response to placental ischemia in pregnant dams and may not exacerbate adverse fetal outcomes in response to placental ischemia.