Gene-gene interactions in the NAMPT pathway, plasma visfatin/NAMPT levels, and antihypertensive therapy responsiveness in hypertensive disorders of pregnancy.

Nicotinamide phosphorybosil transferase (NAMPT) polymorphisms affect visfatin/NAMPT levels and may affect the responsiveness to therapy in hypertensive disorders of pregnancy. We examined whether NAMPT polymorphisms (rs1319501 T>C and rs3801266 A>G), or haplotypes, and gene-gene interactions in the NAMPT pathway affect plasma visfatin/NAMPT levels and the response to antihypertensive therapy in 205 patients with preeclampsia (PE) and 174 patients with gestational hypertension. We also studied 207 healthy pregnant controls. Plasma visfatin/NAMPT levels were measured by ELISA. Non-responsive PE patients with the TC+CC genotypes for the rs1319501 T>C, and with the AG+GG genotypes for the rs3801266 A>G polymorphism had lower and higher visfatin/NAMPT levels, respectively. The 'C, A' haplotype was associated with response to antihypertensive therapy, and with lower visfatin/NAMPT levels in PE. Interactions among NAMPT, TIMP-1 and MMP-2 genotypes were associated with PE and with lack of response to antihypertensive therapy in PE. Our results suggest that NAMPT polymorphisms affect plasma visfatin/NAMPT levels in nonresponsive PE patients, and that gene-gene interactions in the NAMPT pathway not only promote PE but also decrease the response to antihypertensive therapy in PE.

Tissue inhibitor of matrix metalloproteinase-1 polymorphism, plasma TIMP-1 levels, and antihypertensive therapy responsiveness in hypertensive disorders of pregnancy.

Tissue inhibitor of metalloproteinase (TIMP)-1 is a major endogenous inhibitor of matrix metalloproteinase (MMP)-9, which may affect the responsiveness to therapy in hypertensive disorders of pregnancy. We examined whether TIMP-1 polymorphism (g.-9830T>G, rs2070584) modifies plasma MMP-9 and TIMP-1 levels and the response to antihypertensive therapy in 596 pregnant: 206 patients with preeclampsia (PE), 183 patients with gestational hypertension (GH) and 207 healthy pregnant controls. We also studied the TIMP-3 polymorphism (g.-1296T>C, rs9619311). Plasma MMP-9 and TIMP-1 levels were measured by ELISA. GH patients with the GG genotype for the TIMP-1 polymorphism had lower MMP-9 levels and MMP-9/TIMP-1 ratios than those with the TT genotype. PE patients with the TG genotype had higher TIMP-1 levels. The G allele and the GG genotype were associated with PE and responsiveness to antihypertensive therapy in PE, but not in GH. Our results suggest that the TIMP-1 g.-9830T>G polymorphism not only promotes PE but also decreases the responses to antihypertensive therapy.

Polymorphisms of the adiponectin gene in gestational hypertension and pre-eclampsia.

Adiponectin is a hormone involved in energy homeostasis by regulating glucose and lipid metabolism. In addition, the adiponectin gene (ADIPOQ) has polymorphisms that can modulate the circulating concentration of adiponectin. Abnormal adiponectin levels have been associated with pre-eclampsia (PE); however, the influence of genetic polymorphisms on the development of hypertensive disorders of pregnancy is unclear. The aim of this study was to examine whether ADIPOQ polymorphisms are associated with gestational hypertension (GH) and/or PE. We studied 401 pregnant women: 161 healthy pregnant (HP), 113 pregnant with GH and 127 pregnant with PE. ADIPOQ polymorphisms -11391G>A (rs17300539), -11377C>G (rs266729), 45T>G (rs2241766) and 276G>T (rs1501299) were genotyped by allelic discrimination assays using real-time PCR. Haplotypes were inferred using the PHASE 2.1 program. We observed that the genotypic frequencies of the -11377C>G polymorphism were different in PE compared with HP (P<0.0125), with the CT genotype being more commonly found in PE patients than in HP women (P<0.0125). However, allelic frequencies of this single-nucleotide polymorphism were similar between PE and HP (P>0.0125). No difference was observed when GH and HP groups were compared (both P>0.0125). In addition, we found no difference in genotype or allele distributions for the -11391G>A, 45T>G and 276G>T polymorphisms when we compared GH or PE with HP (all P>0.0125). In conclusion, we found a modest association between the CG genotype of the -11377C>G polymorphism and PE.

Assessment of oxidative status markers and NO bioavailability in hypertensive disorders of pregnancy.

The systemic oxidative status in hypertensives disorders of pregnancy (HDP) and its association with endothelial dysfunction is controversial. In the present study, we evaluated systemic plasma levels of oxidative stress markers (TBARS (thiobarbituric acid-reactive substances) and carbonyl) and total antioxidant status (FRAP (ferric reducing ability of plasma (ferric reducing/antioxidant power) and reduction of MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide))), as well as assessed the impact these markers have on nitric oxide (NO) status in healthy pregnant (HP, n=38), gestational hypertensive (GH, n=33) and preeclamptic pregnant women (PE, n=28). We found similar values of TBARS among all groups, and reduced carbonyl levels in HDP between the PE and GH. Conversely, significant increases in plasma activity of antioxidant status were observed in the GH and PE groups compared to the HP group (using both MTT or FRAP method). Importantly, HDP present significantly lower nitrite levels compared to HP women. In Conclusion, our findings show a compensatory antioxidant mechanism against reactive oxygen species (ROS) generation in HDP, which is not associated with nitrite levels restoration.

Haptoglobin polymorphism affects nitric oxide bioavailability in preeclampsia.

Studies showed elevated cell-free hemoglobin (Hb) in preeclampsia (PE), and Hb reacts with nitric oxide (NO), decreasing its bioavailability. Haptoglobin (Hp) is a polymorphic protein (Hp1-1, Hp2-1 and Hp2-2) that binds Hb to form a complex that is removed from circulation, thus preventing Hb-driven oxidative stress and NO scavenging. Hp protein products differ in biochemical and biophysical properties, which reflects on the Hb-Hp complex clearance rate. We hypothesized that Hp phenotypes modulate NO bioavailability by influencing NO consumption in PE. We studied 92 PE subjects and 105 normal pregnant women (NP). Hp genotypes were determined using real-time PCR. To assess NO bioavailability, we measured plasma nitrite using an ozone-based chemiluminescence assay. Plasma Hb and Hp were assessed with commercial immunoassays. A NO consumption assay was used to measure NO consumption. We found no differences in Hp genotype frequencies between PE and NP groups. Hp genotypes had no effects on plasma heme levels, NO consumption and plasma nitrite in NP. However, in PE, Hp2-1 and Hp2-2 were associated with higher plasma heme levels (48 and 55% higher, respectively; P<0.05), increased NO consumption (42 and 44% more, respectively; P<0.05) and lower plasma nitrite (39% less for Hp2-2; P<0.05) compared with Hp1-1. These findings indicate that although Hp genotype does not affect the risk of PE, Hp1-1 genotype may exert a protective role in PE by reducing NO scavenging, whereas Hp2-1 and Hp2-2 further may aggravate PE by reducing NO bioavailability.

Maternal iNOS genetic polymorphisms and hypertensive disorders of pregnancy.

Increased expression and activity of inducible nitric oxide synthase (iNOS) may contribute to the pathogenesis of pre-eclampsia (PE) and gestational hypertension (GH). However, no previous study has examined whether genetic polymorphisms in the iNOS gene are associated with PE or GH. We examined whether two functional, clinically relevant iNOS genetic polymorphisms (the C(-1026)A polymorphism, rs2779249, in the promoter region, and the G2087A polymorphism, rs2297518, in exon 16) are associated with GH or with PE. We studied 565 pregnant women: 212 healthy pregnant (HP), 166 pregnant with GH and 187 pregnant with PE. Genotypes were determined by real-time PCR, using the Taqman allele discrimination assay. The PHASE 2.1 program was used to estimate haplotype distributions in the three study groups. We found no significant association between the C(-1026)A polymorphism and PE or GH (P>0.05). However, we found the GA genotype and the A allele for the G2087A polymorphism at higher frequency in PE, but not in GH, compared with HP (P<0.05). The haplotype analysis showed no significant intergroup differences (P>0.05). These findings suggest that iNOS genetic variants may affect the susceptibility to PE, but not to GH.

Matrix metalloproteinase-9 polymorphisms affect plasma MMP-9 levels and antihypertensive therapy responsiveness in hypertensive disorders of pregnancy.

Abnormal matrix metalloproteinase (MMP)-9 levels may have a role in hypertensive disorders of pregnancy. We examined whether MMP-9 genetic polymorphisms (g.-1562C >T and g.-90(CA)13-25) modify plasma MMP-9 and tissue inhibitor of metalloproteinase (TIMP)-1 levels and the responses to antihypertensive therapy in 214 patients with preeclampsia (PE), 185 patients with gestational hypertension (GH) and a control group of 214 healthy pregnant (HP). Alleles for the g.-90(CA)13-25 polymorphism were grouped L (low) (< 21 CA repeats) or H (high) (≥ 21 CA repeats). Plasma MMP-9 and TIMP-1 concentrations were measured by enzyme-linked immunosorbent assay. Plasma MMP-9 concentrations were not affected by genotypes or haplotypes in HP and PE groups, except for the g.-90(CA)13-25 polymorphism: GH patients with the LH genotype for this polymorphism have higher MMP-9 levels than those with other genotypes. The T allele for the g.-1562C > T polymorphism and the H4 haplotype (combining T and H alleles) are associated with GH and lack of responsiveness to antihypertensive therapy in GH. The H2 haplotype (combining C and H alleles) was associated with lack of responsiveness to antihypertensive therapy in PE, but not in GH. In conclusion, our results show that MMP-9 genetic variants are associated with GH and suggest that MMP-9 haplotypes affect the responsiveness to antihypertensive therapy in hypertensive disorders of pregnancy.

PP002. Study of polymorphisms of the adiponectin gene in women with gestational hypertension and preeclampsia.

INTRODUCTION: Adiponectin is involved in energy homeostasis by regulating glucose and lipid metabolism. Additionally, it presents anti-inflammatory and anti-atherosclerotic functions. Polymorphisms in adiponectin gene (ADIPOQ) can modulate the concentrations of adiponectin. The influence of these polymorphisms on the development of gestational hypertension (GH) and preeclampsia (PE) is unknown. OBJECTIVES: The aim of this work was to examine the influence of polymorphisms in the gene ADIPOQ on the development of gestational hypertension and preeclampsia. PATIENTS AND METHODS: We studied 401 pregnant women: 161 healthy pregnant (HP), 113 pregnant with gestational hypertension (GH) and 127 pregnant with preeclampsia (PE). Polymorphisms ADIPOQ -11391G>A (rs17300539), -11377C>G (rs266729), 45T>G (rs2241766) and 276G>T (rs1501299) were genotyped by allelic discrimination by PCR in real time. Haplotypes were inferred using the PHASE 2.1 program. RESULTS: There were no statistically significant differences in allele and genotype frequencies of the polymorphisms studied. In the analysis of haplotypes, we observed small differences in haplotype frequencies between groups, however, none of these differences was statistically significant (P>0.05). CONCLUSION: We found no association between the genotypic and allelic variants of the ADIPOQ gene polymorphisms with the development of gestational hypertension and preeclampsia.

PP070. Maternal flow-mediated dilation and nitrite concentration during third trimester of pregnancy and postpartum period.

INTRODUCTION: The vascular endothelium is thought to be responsible for cardiovascular adaptations in gestation, such as the decrease in peripheral vascular resistance and the decrease in arterial pressure. There is an increase of nitric oxide (NO) serum levels in normal gestation due to an increment in the activity of the enzyme endothelial nitric oxide synthase (eNOS). OBJECTIVES: To compare maternal flow-mediated dilation of the brachial artery (FMD) and the nitrite concentration between the third trimester of pregnancy and postpartum period. Additionally we want to evaluate whether FMD correlates with nitrite concentration. METHODS: Eligibility criteria were healthy pregnant women with single fetus, gestational age greater than 28 weeks, nonsmokers, and without personal or family history of vascular disease. Each pregnant woman was examined in the third trimester of pregnancy (3(rd)T) and between 8 and 12 weeks postpartum (PP) to evaluate FMD and nitrite concentrations in the whole blood. We excluded women who were not examined in both periods. We compared the values between the two periods using paired t tests. The correlation between FMD and nitrite concentration was examined by Pearson correlation coefficient. Significance level was set at p<0.05. RESULTS: 42 pregnant women were invited for the study. 35 healthy women were elected and 7 of them were excluded for not attending the postpartum evaluation. We found a trend of decreased FMD in the PP period (10.39±5.57 % vs. 8.42±4.21 %, p=0.11; 3(rd)T vs. PP respectively). No significant change was observed in the nitrite concentration (257.41±122.95nmol/L vs.237.16±90.01nmol/L, p=0.28). We did not observe significant correlation between FMD and nitrite during 3(rd)T (r=-0.13, p=0.50) or PP (r=0.14, p=0.48). CONCLUSION: Although our sample size did not permit sufficient precision, FMD seems to decrease between the third trimester and postpartum period. Nitrite concentration did not change between the third trimester of pregnancy and the postpartum period, and it was not correlated to FMD. Studies evaluating larger samples are necessary to confirm these findings.

PP071. Evaluation of maternal-fetal doppler parameters and of whole blood nitrite throughout gestation.

INTRODUCTION: The Doppler method is extensively applied today for the evaluation of pregnancies with involvement of the uteroplacental blood flow. Although increased nitric oxide (NO) formation plays an important role in regulation of systemic vascular resistance during pregnancy, growing evidence indicates that reduced NO formation is associated with hypertensive disorders of pregnancy, especially preeclampsia. OBJECTIVES: The studies were to assess the maternal and fetal Doppler parameters and to determine the whole blood nitrite levels during pregnancy. METHODS: Thirty-three healthy pregnant women were evaluated during the first (11-14 weeks), second (20-24 weeks) and third trimesters (34-36 weeks) of pregnancy. The maternal (uterine arteries) and fetal (cerebral and umbilical arteries) vessels were evaluated by Doppler velocimetry. venous blood was collected(15mL) for the determination of plasma nitrite by chemiluminescence. RESULTS: Regarding the Doppler parameters of the uterine arteries the mean pulsatility index was 1.73, 1.06 and 0.73 in the first, second and third trimesters of pregnancy, respectively. Fetal Doppler showed a mean resistance index of 0.82 and 0.81 for the middle cerebral artery, 0.73 and 0.60 for the umbilical artery in the second and third trimesters, respectively. The mean plasma nitrite concentration was 189.10, 178.28 and 199.57 nmol/ml in the first, second and third trimesters of pregnancy, respectively. CONCLUSION: The study demonstrated that a fall in flow resistance occurs in the uteroplacental vessels without changes in plasma nitrite concentrations during pregnancy.

eNOS haplotypes affect the responsiveness to antihypertensive therapy in preeclampsia but not in gestational hypertension.

Variations of the endothelial nitric oxide synthase (eNOS) gene have been associated with hypertensive disorders of pregnancy. We examined whether eNOS polymorphisms affect the therapeutic responses of women with gestational hypertension (GH) or preeclampsia (PE). We studied 304 hypertensive pregnant women (152 GH and 152 PE), who were stratified according to clinical and laboratorial parameters of therapeutic responsiveness. We compared the frequencies of three eNOS genetic polymorphisms (T-786C, Glu298Asp and b/a intron 4) in responsive and nonresponsive PE and GH patients. We found no significant differences in genotype or allele distributions when responsive and nonresponsive groups were compared (both PE or GH; all P>0.05). However, the eNOS haplotype distribution differed in PE (but not in GH)-responsive and -nonresponsive groups (P=0.0003). The 'C-Glu-a' and 'T-Asp-a' hapotypes were associated with responsiveness and nonresponsiveness to therapy, respectively (both P<0.001), thus suggesting that eNOS haplotypes affect the responsiveness to antihypertensive therapy in PE.