Ana C.T. Palei

Nitric Oxide Formation Is Inversely Related to Serum Levels of Antiangiogenic Factors Soluble Fms-Like Tyrosine Kinase-1 and Soluble Endogline in Preeclampsia

Deficient NO formation has been implicated in hypertensive disorders of pregnancy. However, no previous study has compared the circulating nitrite concentrations in healthy pregnant women with those found in hypertensive disorders of pregnancy. Moreover, 2 antiangiogenic factors produced in the placenta (soluble fms-like tyrosine kinase-1 and soluble endogline) may affect NO formation during pregnancy. Here, we hypothesized that lower concentrations of markers of NO formation exist in hypertensive disorders of pregnancy and that inverse relationships exist between these markers and soluble fms-like tyrosine kinase-1 or soluble endogline. In this cross-sectional study, we compared 58 healthy pregnant women with 56 gestational hypertensive subjects and 45 preeclamptic patients. We measured plasma and whole blood nitrite concentrations using an ozone-based chemiluminescence assay and serum soluble fms-like tyrosine kinase-1 and soluble endogline concentrations using enzyme immunoassays. Whole blood nitrite levels were significantly lower in gestational hypertensive subjects and preeclamptic patients (−36% and −58%, respectively; both P<0.05) compared with healthy pregnant women. The plasma nitrite levels were ≈37% lower in both groups with hypertensive disorders of pregnancy compared with the group with normotensive pregnancies (both P<0.05). As expected, we found higher circulating soluble fms-like tyrosine kinase-1 and soluble endogline concentrations in preeclampsia compared with gestational hypertensive subjects or with healthy pregnancies (both P<0.05). We found negative correlations between antiangiogenic factors and plasma or whole blood nitrite concentrations (Spearman’s r from −0.175 to −0.226; all P<0.05). Our results show clinical evidence for impaired NO formation in preeclampsia or gestational hypertension. The negative correlations between markers of NO formation and antiangiogenic factors in preeclamptic patients suggest an inhibitory effect for these factors on NO formation.

Comparative assessment of matrix metalloproteinase (MMP)-2 and MMP-9, and their inhibitors, tissue inhibitors of metalloproteinase (TIMP)-1 and TIMP-2 in preeclampsia and gestational hypertension

OBJECTIVES To compare the circulating levels of matrix metalloproteinase (MMP)-2, MMP-9, tissue inhibitors of metalloproteinase (TIMP)-1, TIMP-2, and the MMP-9/TIMP-1 and MMP-2/TIMP-2 ratios in preeclampsia and gestational hypertension with those found in normotensive pregnancies. DESIGN AND METHODS We studied 83 pregnant women (30 healthy pregnant women with uncomplicated pregnancies, 26 with gestational hypertension, and 27 with preeclampsia) and 30 healthy nonpregnant women in a cross-sectional study. MMP and TIMP concentrations were measured in plasma samples by gelatin zymography and ELISA, respectively. RESULTS We found higher plasma pro-MMP-9 levels, and higher pro-MMP-9/TIMP-1 ratios in women with gestational hypertension (95%-CI: 1.031 to 2.357, and 0.012 to 0.031, respectively), but not with preeclampsia, compared with those found in normotensive pregnant women (95%-CI: 0.810 to 1.350, and 0.006 to 0.013, respectively; both P<0.05). We found no significant differences in pro-MMP-2 levels (P>0.05). CONCLUSIONS The higher net MMP-9 (but not MMP-2) activity in gestational hypertension compared with normotensive pregnancy suggests that MMP-9 plays a role in the pathophysiology of gestational hypertension. Conversely, the lack of such alterations in preeclampsia is consistent with the notion that different pathophysiological mechanisms are involved in these hypertensive disorders.

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.

Heme Oxygenase-1 Attenuates Hypoxia-Induced sFlt-1 and Oxidative Stress in Placental Villi through Its Metabolic Products CO and Bilirubin

One of the most prevalent complications of pregnancy is preeclampsia, a hypertensive disorder which is a leading cause of maternal and perinatal morbidity and premature birth with no effective pharmacological intervention. While the underlying cause is unclear, it is believed that placental ischemia/hypoxia induces the release of factors into the maternal vasculature and lead to widespread maternal endothelial dysfunction. Recently, HO-1 has been shown to downregulate two of these factors, reactive oxygen species and sFlt-1, and we have reported that HO-1 induction attenuates many of the pathological factors of placental ischemia experimentally. Here, we have examined the direct effect of HO-1 and its bioactive metabolites on hypoxia-induced changes in superoxide and sFlt-1 in placental vascular explants and showed that HO-1 and its metabolites attenuate the production of both factors in this system. These findings suggest that the HO-1 pathway may be a promising therapeutic approach for the treatment of preeclampsia.

Matrix Metalloproteinases as Drug Targets in Preeclampsia

Preeclampsia is an important syndrome complicating pregnancy. While the pathogenesis of preeclampsia is not entirely known, poor placental perfusion leading to widespread maternal endothelial dysfunction is accepted as a major mechanism. It has been suggested that altered placental expression of matrix metalloproteinases (MMPs) may cause shallow cytotrophoblastic invasion and incomplete remodeling of the spiral arteries. MMPs are also thought to link placental ischemia to the cardiovascular alterations of preeclampsia. In fact, MMPs may promote vasoconstriction and surface receptors cleavage affecting the vasculature. Therefore, the overall goal of this review article is to provide an overview of the pathophisiology of preeclampsia, more specifically regarding the role of MMPs in the pathogenesis of preeclampsia and the potential of MMP inhibitors as therapeutic options.

Matrix metalloproteinase (MMP)-9 genotypes and haplotypes in preeclampsia and gestational hypertension

BACKGROUND Abnormal production of matrix metalloproteinases (MMPs), especially MMP-9, may play a role in hypertensive disorders of pregnancy. These alterations may result from functional genetic polymorphisms in the promoter region of MMP-9 gene, which are known to change MMP-9 expression. We examined whether 2 MMP-9 polymorphisms (C(-1562)T and (CA)n) and haplotypes are associated with preeclampsia and/or gestational hypertension. METHODS We studied 476 pregnant women: 176 healthy pregnant (HP), 146 pregnant with gestational hypertension (GH), and 154 pregnant with preeclampsia (PE). Genomic DNA was extracted from whole blood and genotypes for C(-1562)T and (CA)n polymorphisms were determined by PCR-RFLP. Haplotype frequencies were inferred using the PHASE ver. 2.1 program. RESULTS For the g.-90(CA)13-25 polymorphism, no significant differences were found in genotype and allele distributions when PE or GH groups were compared with HP group. However, the CT genotype and T allele for g.-1562C>T polymorphism were more commonly found in GH subjects compared with the HP group (both P<0.05). Conversely, we found no differences in genotypes or allele distributions for the g.-1562C>T polymorphism when the PE and the HP groups were compared. No significant differences were found in overall distributions of haplotype frequencies when the GH or the PE group was compared with the HP group. CONCLUSIONS The C(-1562)T polymorphism in MMP-9 gene is associated with gestational hypertension, but not with preeclampsia. These findings may help to explain the higher plasma MMP-9 levels previously reported in GH compared with HP.

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.

Association between matrix metalloproteinase (MMP)-2 polymorphisms and MMP-2 levels in hypertensive disorders of pregnancy

We examined whether two functional polymorphisms (g.-1306C>T and g.-735C>T) in matrix metalloproteinase (MMP)-2 gene are associated with preeclampsia (PE) or gestational hypertension (GH), and whether they modify MMP-2 or tissue inhibitor of metalloproteinase (TIMP)-2 plasma concentrations in these hypertensive disorders of pregnancy. We studied 130 healthy pregnant (HP), 130 pregnant with GH, and 133 pregnant with PE. Genomic DNA was extracted from whole blood and genotypes for g.-1306C>T and g.-735C>T polymorphisms were determined by Real Time-PCR, using Taqman allele discrimination assays. Haplotypes were inferred using the PHASE program. Plasma MMP-2 and TIMP-2 concentrations were measured by ELISA. The main findings were that pregnant with PE have higher plasma MMP-2 and TIMP-2 concentrations than HP (P<0.05), although the MMP-2/TIMP-2 ratios were similar (P>0.05). Moreover, pregnant with GH have elevated plasma MMP-2 levels and MMP-2/TIMP-2 ratios compared to HP (P<0.05). While MMP-2 genotypes and haplotypes are not linked with hypertensive disorders of pregnancy, MMP-2 genotypes and haplotypes are associated with significant alterations in plasma MMP-2 and TIMP-2 concentrations in preeclampsia (P<0.05). Our findings may help to understand the relevance of MMP-2 and its genetic polymorphisms to the pathophysiology of hypertensive disorders of pregnancy. It is possible that patients with PE and the MMP-2 haplotype combining the C and T alleles for the g.-1306C>T and g.-735C>T polymorphisms may benefit from the use of MMPs inhibitors such as doxycycline. However, this possibility remains to be determined.

Endothelin as a final common pathway in the pathophysiology of preeclampsia: therapeutic implications

Purpose of reviewPreeclampsia remains a major health concern in the United States and worldwide. Recent research has begun to shed light on the underlying mechanisms responsible for the symptoms of preeclampsia, and may provide new avenues for therapy for the preeclamptic patient. Recent findingsThe central role of placental ischemia in the manifestation of preeclampsia has provided new understanding for the origin of pathogenic factors in the preeclamptic patient. The release of soluble factors into the maternal bloodstream from the ischemic placenta is now recognized as a central mechanism in disease manifestation. Specifically, the importance of the vascular endothelial growth factor antagonist soluble fms-like tyrosine kinase and immune factors as factors regulating maternal endothelial dysfunction has become widely acknowledged. Furthermore, mounting evidence implicates the signaling protein endothelin-1 as the final converging factor in the multifaceted cascades that are responsible for the symptomatic manifestation of preeclampsia. Endothelin-1, as a final common pathway in the pathogenic cascade of preeclampsia, presents an intriguing new therapeutic approach for preeclamptic patients. SummaryIdentification of antiangiogenic, autoimmune, and inflammatory factors produced in response to placental ischemia have provided potential new avenues for future research into novel therapies for the preeclamptic patient, and suggest new therapeutic avenues for the treatment of preeclampsia.

Increased circulating cell-free hemoglobin levels reduce nitric oxide bioavailability in preeclampsia

Contrasting with increased nitric oxide (NO) formation during healthy pregnancy, reduced NO bioavailability plays a role in preeclampsia. However, no study has examined whether increased NO consumption by enhanced circulating levels of cell-free hemoglobin plays a role in preeclampsia. We studied 82 pregnant women (38 healthy pregnant and 44 with preeclampsia). To assess NO bioavailability, we measured plasma and whole blood nitrite concentrations using an ozone-based chemiluminescence assay. Plasma ceruloplasmin concentrations and plasma NO consumption (pNOc) were assessed and plasma hemoglobin (pHb) concentrations were measured with a commercial immunoassay. We found lower whole blood and plasma nitrite concentrations in preeclamptic patients (-48 and -39%, respectively; both P<0.05) compared with healthy pregnant women. Plasma samples from preeclamptic women consumed 63% more NO (P=0.003) and had 53% higher pHb and 10% higher ceruloplasmin levels than those found in healthy pregnant women (P<0.01). We found significant positive correlations between pHb and pNOc (r=0.61; P<0.0001), negative correlations between pNOc and whole blood or plasma nitrite concentrations (P=0.02; r=-0.32 and P=0.01; r=-0.34, respectively), and negative correlations between pHb and whole blood or plasma nitrite concentrations (P=0.03; r=-0.36 and P=0.01; r=-0.38, respectively). These findings suggest that increased pHb levels lead to increased NO consumption and lower NO bioavailability in preeclamptic compared with healthy pregnant women.