The reduced uterine perfusion pressure (RUPP) rat model of preeclampsia exhibits much of the pathology characterizing this disease, such as hypertension, inflammation, suppressed regulatory T cells (T Regs), reactive oxygen species (ROS), and autoantibodies to the ANG II type I receptor (AT1-AA) during pregnancy. The objective of this study was to determine whether supplementation of normal pregnant (NP) TRegs into RUPP rats would attenuate the pathophysiology associated with preeclampsia during pregnancy. CD4 ϩ /CD25ϩ T cells were isolated from spleens of NP and RUPP rats, cultured, and injected into gestation day (GD) 12 normal pregnant rats that underwent the RUPP procedure on GD 14. On GD 1, mean arterial pressure (MAP) was recorded, and blood and tissues were collected for analysis. One-way ANOVA was used for statistical analysis. MAP increased from 99 Ϯ 2 mmHg in NP (n ϭ 12) to 127 Ϯ 2 mmHg in RUPP (n ϭ 21) but decreased to 118 Ϯ 2 mmHg in RUPPϩNP TRegs (n ϭ 17). Circulating IL-6 and IL-10 were not significantly changed, while circulating TNF-␣ and IL-17 were significantly decreased after supplementation of TRegs. Placental and renal ROS were 339 Ϯ 58.7 and 603 Ϯ 88.1 RLU·min Ϫ1 ·mgϪ1 in RUPP and significantly decreased to 178 Ϯ 27.8 and 171 Ϯ 55.6 RLU·min Ϫ1 ·mg Ϫ1 , respectively, in RUPPϩNP TRegs; AT1-AA was 17.81 Ϯ 1.1 beats per minute (bpm) in RUPP but was attenuated to 0.50 Ϯ 0.3 bpm with NP TRegs. This study demonstrates that NP TRegs can significantly improve inflammatory mediators, such as IL-17, TNF-␣, and AT 1-AA, which have been shown to increase blood pressure during pregnancy.hypertension; pregnancy; inflammation; oxidative stress; regulatory T cells PREECLAMPSIA IS A PREGNANCY-ASSOCIATED disorder that affects 5-8% of pregnancies and is a major cause of maternal, fetal, and neonatal morbidity and mortality (9,29,37). Hallmark characteristics of preeclampsia are new-onset hypertension after 20 wk gestation, proteinuria, chronic immune activation, fetal growth restriction, and maternal endothelial dysfunction. The pathophysiological mechanisms that lead to the development of preeclampsia are poorly understood. It is thought that poor invasion of trophoblasts leads to insufficient spiral artery remodeling, resulting in placental ischemia (4, 10, 31). The hypoxic environment that results from this placental ischemia is suggested to be an important factor in the development of oxidative stress and a shift in the balance of antiangiogenic and proangiogenic factors, which plays a role in endothelial dysfunction of the placenta and maternal vasculature (10). Previous studies from our laboratory in the reduced uterine perfusion pressure (RUPP) rat model of preeclampsia demonstrate that the total population of CD4-positive (CD4 ϩ ) T cells isolated from RUPP spleens and transferred to NP rats causes similar pathology to that seen in RUPP rats (46), demonstrating a role for this population in mediating pathophysiology in response to placental ischemia. Recent data from both clinical and animal model stud...

Women with preeclampsia produce AT1-AA (agonistic autoantibodies to the angiotensin II type 1 receptor), which stimulate reactive oxygen species, inflammatory factors, and hypertensive mechanisms (ET [endothelin] and sFlt-1 [soluble fms-like tyrosine kinase-1]) in rodent models of preeclampsia. The placental ischemic reduced uterine perfusion pressure (RUPP) rat model of preeclampsia exhibits many of these features. In this study, we examined the maternal outcomes of AT1-AA inhibition ('n7AAc') in RUPP rats. Blood pressure was higher in RUPP rats versus normal pregnant (NP) rats (123±2 versus 99±2 mm Hg, <0.05), which was reduced in RUPP+'n7AAc' (105±3 versus 123±2 mm Hg, <0.05 versus RUPP). Uterine artery resistant index was increased in RUPP versus NP rats (0.71±0.02 versus 0.49±0.02, <0.05) and normalized in RUPP+'n7AAc' rats (0.55±0.03). Antiangiogenic factor sFlt-1 was elevated in RUPP versus NP rats (176±37 versus 77±15 pg/mL, <0.05) but normalized in RUPP+'n7AAc' (86±9, =0.05 versus RUPP). Plasma nitrate and nitrite were decreased (14±1 versus 20±1 µMNO, <0.05) and isoprostanes were elevated (20 117±6304 versus 2809±1375 pg/mL, <0.05) in RUPP versus NP rats; and normalized in RUPP+'n7AAc' rats; (18±2 µMNO; 4311±1 pg/mL). PPET-1 (preproendothelin-1) expression increased 4-fold in RUPP versus NP rats which were prevented with 'n7AAc'. Importantly, placental cytolytic natural killer cells were elevated in RUPP versus NP rats (8±2% versus 2±2% gated, <0.05), which was prevented in RUPP+'n7AAc' total (3±1% gated, <0.05) In conclusion, AT1-AA inhibition prevents the rise in maternal blood pressure and several pathophysiological factors associated with preeclampsia in RUPP rats and could be a potential therapy for preeclampsia.

Women with preeclampsia (PE) have increased mean arterial pressure (MAP), natural killer (NK) cells, reactive oxygen species (ROS), and agonistic autoantibodies to the angiotensin II type 1 receptor (AT1-AA). AT1-AA’s administered to pregnant rodents produces a well-accepted model of PE. However, the role of NK cells and mitochondrial reactive oxygen species (mtROS) in AT1-AA mediated hypertension during pregnancy is unknown. We hypothesize that AT1-AA induced model of PE will exhibit elevated MAP, NK cells, and mtROS; while inhibition of the AT1-AA binding to the AT1R would be preventative. Pregnant rats were divided into 4 groups: normal pregnant (NP) (n = 5), NP + AT1-AA inhibitory peptide (NP +‘n7AAc’) (n = 3), NP + AT1-AA infused (NP + AT1-AA) (n = 10), and NP + AT1-AA +‘n7AAc’ (n = 8). Day 13, rats were surgically implanted with mini-pumps infusing either AT1-AA or AT1-AA +‘n7AAc’. Day 19, tissue and blood was collected. MAP was elevated in AT1-AA vs. NP (119 ± 1 vs. 102 ± 2 mmHg, p < 0.05) and this was prevented by ‘n7AAc’ (108 ± 3). There was a 6 fold increase in renal activated NK cells in AT1-AA vs NP (1.2 ± 0.4 vs. 0.2 ± 0.1% Gated, p = 0.05) which returned to NP levels in AT1-AA +‘n7AAc’ (0.1 ± 0.1% Gated). Renal mtROS (317 ± 49 vs. 101 ± 13% Fold, p < 0.05) was elevated with AT1-AA vs NP and was decreased in AT1-AA +‘n7AAc’ (128 ± 16, p < 0.05). In conclusion, AT1-AA’s increased MAP, NK cells, and mtROS which were attenuated by AT1-AA inhibition, thus highlighting new mechanisms of AT1-AA and the importance of drug therapy targeted to AT1-AAs in hypertensive pregnancies.

Preeclampsia is associated with chronic inflammation and an imbalance among T-helper cell subtypes with an increase in T-helper 17 (T17) cells. The objective of this study was to determine a role for T17s, from the reduced uterine perfusion pressure (RUPP) rat model of preeclampsia, in the etiology of hypertension and chronic inflammation during pregnancy. CD4/CD25 T cells were isolated from rat spleens, cultured in T17 media, and were verified as T17s via flow cytometry. On day 12 of gestation, 1×10 T17 cells from RUPP rats were adoptively transferred into NP rats, carotid catheters were inserted on day 18, and on day 19, mean arterial pressure (MAP) was recorded, serum and plasma were collected, and oxidative stress and production of agonistic autoantibodies to the ANG II type I receptor (AT-AA) were analyzed. MAP increased from 100.3 ± 1.7 mmHg in normal pregnant (NP; n = 17) to 124.8 ± 2.1 mmHg in RUPP (n = 22; P < 0.0001) and to 110.8 ± 2.8 mmHg in NP+RUPP T17 (n = 11). Pup weights in NP+RUPP T17s were decreased to 1.92 ± 0.09 g from 2.39 ± 0.14 in NP rats (P < 0.01). AT-AA significantly increased from 0.1 ± 0.2 beats/min in NP to 15.6 ± 0.7 beats/min in NP+RUPP T17s. IL-6 was 22.3 ± 5.7 pg/ml in NP and increased to 60.45 ± 13.8 pg/ml in RUPP (P < 0.05) and 75.9 ± 6.8 pg/ml in NP+RUPP T17 rats (P < 0.01). Placental and renal oxidative stress were 238 ± 27.5 and 411 ± 129.9 relative light units·min·mg in NP and 339 ± 104.6 and 833 ± 331.1 relative light units·min·mg in NP+RUPP T17, respectively. In conclusion, RUPP T17 cells induced intrauterine growth restriction and increased blood pressure, AT-AA, IL-6, and tissue oxidative stress when transferred to NP rats, indicating a role for autoimmune associated T17 cells, to cause much of the pathophysiology associated with preeclampsia.

Preeclampsia is the leading cause of death and morbidity world-wide for the mother and fetus during pregnancy. Preeclampsia does not only effect the mother and the baby during pregnancy, but can also have long-term effects, such as the increase risk of hypertension and cardiovascular disease, on the offspring and the postpartum mother later in life. The exact cause of preeclampsia is unknown, but women with preeclampsia have elevated concentrations of agonistic autoantibodies against the angiotensin II type 1 receptor (AT1-AA). These AT1-AA’s through multiple studies have shown to play a significant role in the pathology and possible genesis of preeclampsia. This review will discuss the discovery of AT1-AAs and the role of AT1-AAs in the pathophysiology of preeclampsia. This review will also discuss future therapeutic approaches towards the AT1-AA to prevent adverse pregnancy outcomes. Furthermore, we will examine the relationship between AT1-AA induced hypertension associated with increase oxidative stress, antiangiogenic factors (such as soluble fms-related tyrosine kinase-1 (sFlt-1), endothelin-1 (ET-1), inflammation, endothelial dysfunction, and reduced renal function. Understanding the pathological role of AT1-AAs in hypertensive pregnancies is important as we search for novel therapies to manage preeclampsia.

Preeclampsia (PE) is associated with altered immune activation during pregnancy. We have previously shown that adoptive transfer of CD4(+) T cells from the reduced uterine perfusion pressure (RUPP) rat model of PE increases blood pressure, oxidative stress (ROS), and inflammation in normal pregnant recipient rats. The objective of this study was to determine if blockade of communication via the CD40-CD40 ligand (CD40L) interaction between placental ischemia-induced CD4(+) T cells with endogenous normal pregnant (NP) cells would improve pathophysiology that was previously observed in NP recipient rats of RUPP CD4(+) T cells. Splenic CD4(+) T lymphocytes were magnetically separated, incubated with 2.5 μg/ml anti-CD40 ligand (αCD40L) overnight, and transferred into NP rats on day 12 of gestation (NP+RUPP CD4(+) T+anti-CD40L). On day 19 of gestation, blood pressure (MAP), blood, and tissues were collected. MAP was 99 ± 2 in NP (n = 13), 116 ± 4 in NP+RUPP CD4(+) T cells (n = 7; P < 0.01); MAP only increased to 104 ± 2 in NP+RUPP CD4(+) T cells+CD40L (n = 24) (P < 0.05 vs. NP+RUPP CD4(+) T cells). Mechanisms of hypertension in response to RUPP CD4(+) T cells include endothelin-1 (ET-1), ROS, and angiotensin II type I receptor (AT1-AA) were analyzed. Inhibition of CD40L binding reduced placental ET-1 to 2.3-fold above NP rats and normalized placental ROS from 318.6 ± 89 in NP+RUPP CD4(+) T cells (P < 0.05) to 118.7 ± 24 in NP+RUPP CD4(+) T+anti-CD40L (P < 0.05). AT1-AA was also normalized with inhibition of CD40L. These data suggest that placental ischemia-induced T-cell communication via the CD40L is one important mechanism leading to much of the pathophysiology of PE.

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The Reduced Uterine Perfusion Pressure (RUPP) rat model and normal pregnant (NP) rat recipients of RUPP CD4+T cells recapitulate many characteristics of preeclampsia (PE) such as hypertension and oxidative stress. We have shown an important hypertensive role for NK cells to cause mitochondrial (mt) dysfunction in RUPP rats, however the role for RUPP CD4+ T cells to stimulate NK cells is unknown. Therefore, we hypothesize that RUPP induced CD4+ T cells activate NK cells to cause mt dysfunction/ROS as mechanisms of hypertension during pregnancy. We tested our hypothesis by adoptive transfer of RUPP CD4+T cells into NP rats or by inhibiting the activation of RUPP CD4+T cells with Orencia (Abatacept) and examining hypertension, NK cells and mt function. RUPP was performed on gestation day 14, splenic CD4+ T cells were isolated on GD19 and injected into NP rats on gestation day (GD) 13. In a separate groups of rats; Orencia was infused and the RUPP procedure performed. MAP and placental and renal mtROS increased in RUPP (n=7, p<0.05) and NP+RUPP CD4+ T cell recipients (n=13, p<0.05) compared to control NP (n=7) and NP+NPCD4+Tcell recipients (n=5), but was reduced with Orencia (n=13, p<0.05). Placental and renal respiration was reduced in RUPP (n=6, p<0.05) and NP+RUPP CD4+ T cells (n=6, state 3-p<0.05) compared to NP (n=5) and NP+NPCD4+Tcell recipients (n=5), but improved with Orencia (n=9, n=8 p<0.05). These data indicate that CD4+ T cells, independent of NK cells, cause mt dysfunction/ROS contributing to hypertension in response to placental ischemia during pregnancy.