Deletion of GIT1 Impacts eNOS Activity To Aggravate sFlt-1-Induced Preeclampsia Phenotype in Mice

Abstract

Preeclampsia, a serious multisystem disorder specific to human pregnancy, remains a considerable burden of disease worldwide. Reduced nitric oxide bioavailability is proved to be crucial in the maternal and fetal pathophysiology of preeclampsia. G-protein-coupled Receptor Kinase Interactor-1 (GIT1) is a novel endothelial nitric oxide synthases (eNOS) interactor mediator. The aim of this paper is to investigate the effect of GIT1 on preeclampsia. Blood pressure (BP) was measured using a carotid catheter-calibrated eight-chamber tail-cuff system (CODA) at the same time daily. Urinary albumin excretion (UAE) was determined using Albuwell-M kits (Exocell Inc) and creatinine clearance (CCr) was determined by measuring urinary creatinine concentration with tandem liquid chromatography-mass spectrometry. The release of nitrite was analyzed to detect nitric oxide (NO) production using a Sievers Chemiluminescence NO Analyzer. NOS activity was examined by measuring the conversion of ³H-labeled l-arginine to ³H-labeled l-citrulline. BP was significantly increased in GIT1^-/- mice with or without sFIT-1 treatment. In addition, GIT1-/- mice possessed higher UAE and lower CCr. Depletion of GIT1 impedes the NO production and placenta eNOS activity. Additional GIT1 attenuates sFlt-1-induced preeclampsia phenotypes. Our findings suggest that GIT1 significantly extenuates the sFlt-1-induced preeclampsia phenotypes by inhibiting eNOS activity, indicating a crucial role of GIT1 in the progression of preeclampsia.

Keywords: G-protein-coupled Receptor Kinase Interactor-1 (GIT1); NO production; blood pressure (BP); endothelial nitric oxide synthases (eNOS); preeclampsia; urinary albumin excretion (UAE).

Figure 1

Blood pressures were increased in GIT1^−/− mice with or without treated with sFIT-1. (A) Systolic blood pressure (B) Diastolic blood pressure. The tests were started at day 2 (-2) before administration of sFlt-1 and finished at day 6 (6) after administration of sFlt-1, day 0(0) was the day administrated with sFlt-1. N = 6 mice in each experimental group. **P < 0.01 vs. WT mice.

Figure 2

Renal function was impacted in GIT1^−/− mice. (A) Urinary Albumin Excretion (B) Creatinine Clearance. The tests were done at day 6 after administration of sFlt-1/Fc(control). N = 6 mice in each experimental group. **P < 0.01 vs. WT/control mice, #P < 0.01 vs. WT/ sFlt-1 mice.

Figure 3

Serum NO level and placenta eNOS activity were impacted in GIT1^−/− mice. (A) Serum NO level (B) Placenta eNOS expression (C) Placenta eNOS activity. The tests were done at day 6 after administration of sFlt-1/Fc (control). N = 6 mice in each experimental group. *P < 0.05, **P < 0.01 vs. WT/control mice, #P < 0.01 vs. WT/ sFlt-1 mice.

Figure 4

Additional GIT1 attenuated sFlt-1-induced preeclampsia phenotype in WT mice. (A) Systolic blood pressure (B) Diastolic blood pressure. The tests were started at day 2(-2) before administration of sFlt-1 and finished at day 6(6) after administration of sFlt-1, day 0(0) was the day administrated with sFlt-1, day 1 was the day administrated with GIT1. (C) Urinary Albumin Excretion (D) Creatinine Clearance. The tests were done at day 6 after administration of sFlt-1. Vehicle group was WT mice treated with sFlt-1, GIT1 group was WT mice treated with sFlt-1 and GIT1. N = 6 mice in each experimental group. P < 0.05,**P < 0.01 vs. Vehicle mice.

Figure 5

Additional GIT1 upregulated serum NO level and placenta eNOS activity. (A) Serum NO level (B) Placenta eNOS activity. The tests were done at day 6 after administration of sFlt-1/Fc(control). N = 6 mice in each experimental group. *P < 0.05, **P < 0.01 vs. Vehicle mice.