Bihui Hu

Decreased Seizure Threshold in an Eclampsia-Like Model Induced in Pregnant Rats with Lipopolysaccharide and Pentylenetetrazol Treatments

Objective Eclampsia is a poorly understood but potentially fatal complication of pregnancy. Research to date on this disorder has been hampered by the lack of a suitable animal model. To correct this deficiency, this report describes the generation of a rat eclampsia-like model using pentylenetetrazol (PTZ) in a previously established rat preeclampsia model. Method Rats were administered lipopolysaccharide (1.0 µg/kg) by tail vein injection on gestational day 14 to establish preeclampsia (PE). PE and control rats (non-pregnant, NP; normal-pregnant, P) were injected intraperitoneally (i.p.) with PTZ (40 mg/kg) to induce seizures. In separate experiments, MgSO4 (270 mg/kg IP) was injected in advance of PTZ into PE rats to observe its effect on PTZ-induced seizures. Results PE conditions were verified in rats after LPS administration by significantly higher blood pressure (P<0.01) and urinary albumin excretion (P<0.05), elevated sFlt-1 (P<0.05) and decreased PlGF serum levels (P<0.05), and evidence of hepatic dysfunction compared to control groups. PTZ successfully induced seizure activity in all groups studied. Latency to seizure was significantly (P<0.01) less in the PE-PTZ group (73.2±6.6 sec.) than in PTZ-treated controls (107.0±7.4 sec.). Pretreatment with MgSO4 prolonged (P<0.05) latency to seizure, shortened seizure duration and decreased seizure rates. Significant increased (P<0.05) in the serum levels of the inflammatory cytokines TNF-α and IL-1β in PE and PE-PTZ groups, and decreased (P<0.05) in their levels following MgSO4 administration. Conclusion This PTZ-induced eclampsia-like rat model is comparable to the human condition of eclampsia and may serve as a useful research tool for future studies of this disease. The increased inflammatory cytokines in preeclampsia are coincident with a decreased threshold for PTZ-induced seizures, suggesting that an inflammatory mechanism may contribute to the susceptibility to seizure activity and inflammation might have an important role in eclampsia.

Cyclosporin A significantly improves preeclampsia signs and suppresses inflammation in a rat model.

Preeclampsia is associated with an increased inflammatory response. Immune suppression might be an effective treatment. The aim of this study was to examine whether Cyclosporin A (CsA), an immunosuppressant, improves clinical characteristics of preeclampsia and suppresses inflammation in a lipopolysaccharide (LPS) induced preeclampsia rat model. Pregnant rats were randomly divided into 4 groups: group 1 (PE) rats each received LPS via tail vein on gestational day (GD) 14; group 2 (PE+CsA5) rats were pretreated with LPS (1.0 μg/kg) on GD 14 and were then treated with CsA (5mg/kg, ip) on GDs 16, 17 and 18; group 3 (PE+CsA10) rats were pretreated with LPS (1.0 μg/kg) on GD 14 and were then treated with CsA (10mg/kg, ip) on GDs 16, 17 and 18; group 4 (pregnant control, PC) rats were treated with the vehicle (saline) used for groups 1, 2 and 3. Systolic blood pressure, urinary albumin, biometric parameters and the levels of serum cytokines were measured on day 20. CsA treatment significantly reduced LPS-induced systolic blood pressure and the mean 24-h urinary albumin excretion. Pro-inflammatory cytokines IL-6, IL-17, IFN-γ and TNF-α were increased in the LPS treatment group but were reduced in (LPS+CsA) group (P<0.05). Anti-inflammatory cytokine IL-4 was decreased in the LPS group but was increased in (LPS+CsA) group (P<0.05). Cyclosporine A improved preeclampsia signs and attenuated inflammatory responses in the LPS induced preeclampsia rat model which suggests that immunosuppressant might be an alternative management option for preeclampsia.

PP024. Effects of intravenous magnesium sulfate on the characteristics of eclamptic seizures induced by electrical stimuli in a rat preeclampsia/eclampsia model

BACKGROUND/AIMS Eclampsia is a serious complication of pregnancy and remains a leading cause of maternal mortality worldwide. Magnesium sulfate is commonly used in the prophylaxis and treatment of eclampsia. However, uncertainty remain regarding its anticonvulsant mechanism(s) of action. This study examined the effects of intravenous magnesium sulfate on the characteristics of eclamptic seizures in a rat preeclampsia/eclampsia model. METHODS All rats were implanted with stainless nickel-cadmium alloy bipolar electrodes one week before fertilization. Next, an experimental rat preeclampsia (PE) model was induced on gestational day 14 by anaesthetising rats and infusing over 1 hour into their tail veins lipopolysaccharide (LPS) (1.0μg/kg body weight) (with control rats receiving normal saline). The rats were then divided into three groups: a normal pregnancy (NP) group (n=6) which received a continuous infusion of saline; a control PE model group (n=7) (which had previously received the LPS treatment) which also received a continuous infusion of saline; and a treated PE model group (n=8) (which had previously received the LPS treatment) which received a continuous infusion of magnesium sulfate (60mg/kg/day). The continuous infusions in all three groups were delivered by implanted osmotic minipumps . Measurements were made of blood pressure, albuminuria, serum ALT, AST, and creatinine, BUN and serum magnesium concentrations. On gestational day 18, all experimental rats received a standardized electrical stimulus. Seizure activity was assessed using electroencephalogram (EEG) recordings. Terminations of pregnancy were performed on gestational day 21. Resorptions and pup birth weights were recorded. RESULTS The pregnant LPS treated rats developed many features of human PE (e.g. hypertension, proteinuria, liver and kidney dysfunctions). The mean concentration of Mg(2+) in the magnesium sulfate therapy group (0.86±0.24mmol/L) was significantly higher (p<0.05) than in both the control PE model group (0.61±0.12mmol/L) and the NP group (0.62±0.09mmol/L). The magnesium sulfate therapy group had a significantly (p<0.05) increased latency period (21.7±8.9min) to evoke a full motor seizure compared to both the NP group (4.8±2.2min)and the control PE model group (3.3±1.4min), there being no significant difference (p>0.05) between the latency periods of the NP group and the control PE model group. Overall, the magnesium sulfate therapy regimen completely prevented seizure activity in 3/8 (37.5%) of the treated PE model rats compared to 6/6 (100%) of the NP rats and 7/7 (100%) of the control PE rats. The treated PE model group also had significantly (p<0.05) reduced seizure duration (26±4s) compared to both the NP (40±7s) and the control PE model (45±9s) groups. As well, there was a significantly (p<0.05) shorter EEG seizure amplitude change in the treated PE model group (58±6μv). CONCLUSION In this rat preeclamsia/eclampsia model, the anticonvulsant characteristics of magnesium sulfate have been shown to include significantly increasing seizure latency period, reducing seizure duration and decreasing seizure EEG amplitude.

Changes in the Expression of AQP4 and AQP9 in the Hippocampus Following Eclampsia-Like Seizure

Eclampsia is a hypertensive disorder of pregnancy that is defined by the new onset of grand mal seizures on the basis of pre-eclampsia. Until now, the mechanisms underlying eclampsia were poorly understood. Brain edema is considered a leading cause of eclamptic seizures; aquaporins (AQP4 and AQP9), the glial water channel proteins mainly expressed in the nervous system, play an important role in brain edema. We studied AQP4 and AQP9 expression in the hippocampus of pre-eclamptic and eclamptic rats in order to explore the molecular mechanisms involved in brain edema. Using our previous animal models, we found several neuronal deaths in the hippocampal CA1 and CA3 regions after pre-eclampsia and that eclampsia induced more neuronal deaths in both areas by Nissl staining. In the current study, RT-PCR and Western blotting data showed significant upregulation of AQP4 and AQP9 mRNA and protein levels after eclamptic seizures in comparison to pre-eclampsia and at the same time AQP4 and AQP9 immunoreactivity also increased after eclampsia. These findings showed that eclamptic seizures induced cell death and that upregulation of AQP4 and AQP9 may play an important role in this pathophysiological process.

PP023. Soluble Fms-like tyrosine kinase-1 and placental growth factor expression in a rat model of pre-eclampsia.

BACKGROUND/AIMS Soluble Fms-like tyrosine kinase-1(sFlt-1) and placental growth factor (PIGF) have been used clinically to predict preeclampsia (PE). This study investigated these factors in a rat model of preeclampsia induced by ultra-low-dose endotoxin. METHODS The experimental PE rat model was generated on gestational day 14. Rats were anesthetized and divided into a normal pregnancy group (NP, n=7) (which received a normal saline infusion) and a PE model group (n=9) (which received an infusion of lipopolysaccharide (LPS) endotoxin (1.0μg/kg body weight). Infusions were given through the tail vein for 1 hour. Blood pressure was monitored and albuminuria, serum ALT, AST, creatinine and BUN were measured. As well, concentrations of sFlt-1 and PIGF in serum and amniotic fluid were measured by enzyme-linked immuno sorbent assay (ELISA). RESULTS Arterial pressure was increased (135±7 versus 116±3mmHg; P<0.03) in the PE model rats compared with the NP rats. The concentration of sFlt-1 in the PE model group (162.7±73.9pg/ml) was significantly higher than NP group (123±64pg/ml) (P<0.05). The serum level of PIGF in PE model group (9.7±6.2pg/ml) was significantly lower compared to the NP group (23.4±12.4) (P<0.05). The plasma sFlt-1/PIGF ratio in the PE model group (18.3±5.6) was greater than that in the NP group (6.7±2.1) (P<0.05). Similar changes were also present in the amniotic fluid. CONCLUSION sFlt-1 and PIGF levels in this rat PE model induced by ultra-low-dose endontoxin showed changes consistent with findings in preeclamptic patients, indicating that this animal model mimics human preeclampsia well in these aspects of the disorder.

PP022. An animal model for eclampsia

BACKGROUND/AIMS Eclampsia, a leading cause of maternal mortality. Little is known about the causes of eclampsia and there is no effective treatment. Development of an animal model may help to expand our understanding and may hold great potential for the design of effective treatment. METHODS Experimental preeclampsia model was build first, rats were received a infusion of endotoxin (LPS) (1.0μg/kg body weight) or saline solution through the tail vein during 1h on gestational day 14, blood pressure and albuminuria were measured. On gestational day 18 all experimental rats were received electric stimulation, the seizure behaviors were observed and EEG were recorded in these rats. Terminations of pregnancy were performed on day 21 of gestation, resorptions and pups birth weights were recorded. RESULTS PE model rats develop many features of human PE that correlates with bad pregnancy outcomes. Both clinical behavior and EEG records documented seizure activity developed in 100% of pregnancy rats and 58.3% of non-pregnancy rats (P=0.01). The PE model rats had a 31.25% decrease of the latency (3.3±1.4min) to evoke a full motor seizure compared with normal pregnancy rats (4.8±2.2min), and had a significant decrease contrast to non-pregnancy rats (10.6±7.1min), £¨P<0.05). EEG recordings showed seizure activities when rats had clinically generalized tonic-clonic convulsions. CONCLUSION We described a rat model of eclampsia, were the relevant predominant features of human eclampsia.