Severe preeclampsia: Association of genes polymorphisms and maternal cytokines production in Brazilian population

IntroductionPreeclampsia (PE) is a multi-system disorder of pregnancy characterized by hypertension and proteinuria. Healthy pregnancy is associated with a controlled inflammatory process, which is exacerbated in PE in response to excessive placental stimuli. Gene expression levels can affect inflammation and immune regulation. It is known that differences in cytokine allele frequencies amongst populations may contribute to difference in the incidence of several diseases.ObjectiveThe aim of this study was to investigate the frequency of TNF-α, IL-6, IFN-γ and IL-10 genes polymorphisms and their relationship with the cytokines plasma levels in PE.MethodsA total of 281 women were included in this study; 116 with severe PE, 107 normotensive pregnant and 58 non-pregnant women. Cytokine genotyping was carried out by the polymerase chain reaction. The analyzed polymorphisms were: TNF-α (−308 G → A), IL-10 (−1082 G → A), IL-6 (−174 G → C), and IFN-γ (+874 A → T). Cytokine plasma levels were measured by Cytometric Bead Array method.ResultsA higher frequency of the IFN-γ (+874) T/T genotype in severe PE comparing to normotensive pregnant women was found (P < 0.001). TNF-α, IL-6 and IFN-γ plasma levels were higher in PE women compared to non-pregnant women (P < 0.001; P < 0.001; P = 0.004). IL-6 and IFN-γ levels were also higher in PE women compared to normotensive pregnant (P < 0.001; P = 0.010). IL-10 levels were higher in normotensive pregnant women compared to PE (P < 0.001). IFN-γ and IL-6 genes polymorphisms influenced the genic expression in PE and normotensive pregnant women, respectively.ConclusionsThese results suggest that IFN-γ seems to play a role in PE occurrence.     

硫辛酸对子痫前期患者Endoglin、VEGF及Flt-1的影响及临床意义

目的:探讨硫辛酸联合低分子肝素钠对子痫前期患者胎盘组织Endoglin、VEGF、F1t-1的影响及临床意义。方法:选取我院收治的子痫前期患者108例,每组各54例,对照组予低分子肝素钠100 IU/kg,日一次静点。实验组在对照组的基础上,加以硫辛酸注射液600 mg溶于250 m L生理盐水静脉滴注,日1次,7天为1个疗程,治疗1个疗程。治疗后,观察比较两组患者的血压情况,及胎盘组织中Endoglin、VEGF、F1t-1的水平以及临床疗效。结果:1治疗后,两组患者胎盘组织中Endoglin、Flt-1蛋白表达下降,VEGF蛋白表达升高,且实验组变化更显著,差异有统计学意义(P0.05)。2治疗后,两组患者的临床疗效均有所提高,且实验组明显优于对照组,差异有统计学意义(P0.05)。3治疗后,两组患者的血压情况均有所改善,实验组明显优于对照组,差异有统计学意义(P0.05)。结论:硫辛酸联合低分子肝素钠治疗子痫前期,能够降低胎盘组织中Endoglin、Flt-1蛋白表达,提高VEGF蛋白表达,从而降低血压,改善患者的临床疗效,值得临床推广使用。     

miR-19a在正常妊娠及重度子痫前期患者胎盘中的表达

目的：通过检测正常妊娠及重度子痫前期患者胎盘组织中miR-19a的表达,探讨其与子痫前期发病的关系。方法：收集10例重度子痫前期患者胎盘组织（实验组）和10例正常产妇胎盘组织（对照组）,应用荧光实时定量PCR（Real Time PCR）的方法检测两组miR-19a的表达差异。结果：重度子痫前期患者胎盘组织中miR-19a表达升高（P〈0.05）。结论：子痫前期患者胎盘组织中存在差异表达的miRNA,miR-19a在胎盘组织中的高表达可能与子痫前期的发病有关。     

低分子肝素对子痫前期大鼠炎症反应、肝功能及胎盘组织Bcl-2、Bax蛋白表达的影响

目的:研究低分子肝素对子痫前期大鼠炎症反应、肝功能及胎盘组织Bcl-2、Bax蛋白表达的影响。方法:将90只孕期大鼠以随机数表法分成正常孕组、子痫前期组、治疗组,每组30只。其中子痫前期组和治疗组大鼠于妊娠第13 d开始皮下注射左旋硝基精氨酸甲酯,建立子痫前期大鼠模型,注射剂量为200 mg/(kg·d),正常孕组予以等量生理盐水注射干预。治疗组予以低分子肝素皮下注射干预,注射剂量为40μL/(kg·d),子痫前期组以及正常孕组大鼠予以同等剂量的生理盐水注射处理。比较三组大鼠的血压、24 h蛋白尿,肝功能指标水平,血清炎症因子水平,胎盘组织中Bcl-2及Bax蛋白表达水平。结果:子痫前期组及治疗组大鼠妊娠第15 d、21 d时的血压水平均显著高于正常孕组,且妊娠第21 d时的24 h蛋白尿高于正常孕组,治疗组大鼠妊娠第21 d的血压及24 h蛋白尿均低于子痫前期组(均P<0.05)。妊娠第21 d时子痫前期组、治疗组大鼠的谷丙转氨酶(ALT)、谷草转氨酶(AST)、白细胞介素-2(IL-2)、白细胞介素-6(IL-6)、γ-干扰素(IFN-γ)水平均显著高于正常孕组,且治疗组低于子痫前期组(均P<0.05)。子痫前期组、治疗组大鼠胎盘组织中Bcl-2蛋白表达水平显著低于正常孕组,Bax蛋白表达水平显著高于正常孕组,且治疗组大鼠Bcl-2蛋白表达水平显著高于子痫前期组,Bax蛋白表达水平显著低于子痫前期组(均P<0.05)。结论:低分子肝素对子痫前期大鼠中具有明显的降血压效果,有利于改善大鼠肝功能,其主要作用机制可能与诱导Th1/Th2的平衡朝Th2方向发展,调节Bcl-2/Bax平衡有关。     

Thrombin impairs the angiogenic activity of extravillous trophoblast cells via monocyte chemotactic protein-1 (MCP-1): A possible link with preeclampsia

Cytokines’ secretion from the decidua and trophoblast cells has been known to regulate trophoblast cell functions, such as Extravillous trophoblasts (EVTs) cell migration and invasion and remodeling of spiral arteries. Defective angiogenesis and spiral arteries transformation are mainly caused by proinflammatory cytokines and excessive thrombin generation during preeclampsia. Monocyte chemotactic protein-1 (MCP-1), a crucial cytokine, has a role in maintaining normal pregnancy. In this study, we explored whether thrombin regulates the secretion of MCP-1 in HTR-8/SVneo cells; if yes, what is its function? We used HTR-8/SVneo cells, developed from ?rst trimester villous explants of early pregnancy, as the model of EVTs. MCP-1 gene silencing was performed using gene-specific siRNA. qPCR and ELISA were performed to estimate the expression and secretion of MCP-1. Here, we found that thrombin enhanced the secretion of MCP-1 in HTR-8/SVneo cells. Proteinase-activated receptor-1 (PAR-1) was found as the primary receptor, regulating MCP-1 secretion in these cells. Furthermore, MCP-1 secretion is modulated via protein kinase C (PKC) α, β, and Rho/Rho-kinase-dependent pathways. Thrombin negatively regulates HTR-8/SVneo cells’ ability to mimic tube formation in an MCP-1 dependent manner. In conclusion, we propose that thrombin-controlled MCP-1 secretion may play an essential role in normal placental development and successful pregnancy maintenance. Improper thrombin production and MCP-1 secretion during pregnancy might cause inadequate vascular formation and transformation of spiral arteries, which may contribute to pregnancy disorders, such as preeclampsia.     

A proteomic approach identifies early pregnancy biomarkers for preeclampsia: Novel linkages between a predisposition to preeclampsia and cardiovascular disease

Preeclampsia (PE) is a common, potentially life‐threatening pregnancy syndrome triggered by placental factors released into the maternal circulation, resulting in maternal vascular dysfunction along with activated inflammation and coagulation. Currently there is no screening test for PE. We sought to identify differentially expressed plasma proteins in women who subsequently develop PE that may perform as predictive biomarkers. In seven DIGE experiments, we compared the plasma proteome at 20 wk gestation in women who later developed PE with an appropriate birth weight for gestational age baby (n=27) or a small for gestational age baby (n=12) to healthy controls with uncomplicated pregnancies (n=57). Of the 49 differentially expressed spots associated with PE‐appropriate for gestational age, PE‐small for gestational age or both (p<0.05, false discovery rate corrected), 39 were identified by LC‐MS/MS. Two protein clusters that accurately (>90%) classified women at risk of developing PE were identified. Immunoblots confirmed the overexpression of fibrinogen γ chain and α‐1‐antichymotrypsin in plasma prior to PE. The proteins identified are involved in lipid metabolism, coagulation, complement regulation, extracellular matrix remodeling, protease inhibitor activity and acute‐phase responses, indicating novel synergism between pathways involved in the pathogenesis of PE. Our findings are remarkably similar to recently identified proteins complexed to high‐density lipoprotein and linked to cardiovascular disease.     

Multifrequency Infradian Variation of Blood Pressure During and After Human Pregnancy

We used a chronobiologic approach to explore the possibility that there may be -7-day (circaseptan) and -30-day (circatrigintan) components in blood pressure during a healthy human pregnancy, the amenorrhea of this status notwithstanding. The results were compared with those obtained from data longitudinally monitored on the same subject at a time when she was not pregnant. The woman under study used an ABPM-630 Colin (Komaki, Japan) device to monitor her blood pressures and heart rates at half to 1-h intervals, with few interruptions. During pregnancy, starting during the first gestational week, she monitored herself for 2 of each 6-day span for the entire duration of pregnancy (a total of 76 days of monitoring). Additionally, with a monitoring protocol similar to that during pregnancy, the subject used the same blood pressure monitor for a total of 78 days during 9.6 months and starting 1 year after delivery. The data obtained oscillometrically for both longitudinal profiles were analyzed separately by multiple-component linear least-squares rhythmometry, a procedure used to describe the periodic waveform of nonsinusoidal rhythms. The analysis of blood pressure variability during pregnancy allows the identification not only of the circadian (with a period of 24 h), but also of other statistically significant components with periods of 156 (6.5 days, apparently free-running from the social week) and of 720 h (30 days) for both systolic and diastolic blood pressure. This multiharmonic time structure is somewhat different during menstruation in the same woman and during a similar time span, with statistically significant components of 96 h (4 days), 192 h (8 days), and 960 h (40 days) for both systolic and diastolic blood pressure. Moreover, the ratio between the amplitudes of the infradian components identified during pregnancy in clinical health is reversed from that obtained in women with preeclampsia. The complex time-structure of blood pressure during pregnancy offers new endpoints to be taken into account for an early identification of gestational hypertension or even preeclampsia.     

Concentrations of Pentosidine,an Advanced Glycation End-product,in Umbilical Cord Blood

Advanced glycation end-products (AGEs) are formed over several weeks to months by non-enzymatic glycation and oxidation (“glycoxidation”) reactions between carbohydrate-derived carbonyl groups and protein amino groups, known as the Maillard reaction. Pentosidine is one of the best-characterized AGEs and is accepted as a satisfactory marker for glycoxidation in vivo. The present study was intended to measure pentosidine concentrations in umbilical cord blood from newborns with various gestational ages using our recently established high-performance liquid chromatography method [Tsukahara, H. et al. (2003) Pediatr. Res. 54, 419–424]. Our study demonstrates, for the first time, that pentosidine is detected in most of the umbilical blood samples. This study also shows that the umbilical blood concentrations of pentosidine are considerably lower than normal adult values, but that they increase with gestation progression and fetal growth. Umbilical pentosidine concentrations were significantly elevated in newborns of mothers with preeclampsia compared to those of mothers without preeclampsia. We conclude that accumulation of AGEs and oxidative stress occurs in fetal tissues and organs in utero at the early stage of human life and that their accumulation is augmented in the maternal preeclampsic condition.     

Involvement of placental peptidases associated with renin–angiotensin systems in preeclampsia

Preeclampsia is characterized by pregnancy-induced hypertension accompanied with protein urea and generalized edema. Preeclampsia develops during the second half of pregnancy and resolves postpartum promptly, implicating the placenta as a primary cause in the disorder. Normal pregnancy is associated with reductions in arterial pressure and attenuated pressor response to exogenous infused angiotensin II (ANG II). In contrast, women with preeclampsia show the similar sensitivity to the pressor effect of ANG II as do non-pregnant women. To elucidate the involvement of placental peptidases associated with renin–angiotensin systems, we determined the localization of angiotensin-converting enzyme (ACE) and aminopeptidase A (AP-A), ANG II degrading enzyme, in the placenta and compared the expression of mRNA and protein in uncomplicated and preeclamptic placenta. In addition, AP-A expression in trophoblastic cells treated with ANG II and ACE expression in HUVECs under hypoxic condition were analyzed, respectively. The expression of both peptidases in the preeclamptic placenta was significantly higher than those from uncomplicated. ACE was primarily localized to venous endothelial cells of stem villous whereas AP-A expression was recognized in the trophoblast and pericytes of fetal arterioles and venules within stem villous. Hypoxia induced ACE expression in HUVECs while both hypoxia and ANG II evoked AP-A expression in trophoblast. These results suggested that hypoxic condition in preeclampsia induces ACE activation in feto-placental unit to maintain the fetal hemodynamics and placental AP-A plays a role as a component of the barrier of ANG II between mother and fetus.