The effects of preeclampsia and oxygen environment on endothelial release of matrix metalloproteinase-2.

BACKGROUND: There is evidence of altered vascular endothelial function in women with preeclampsia as well as in the endothelial cells from umbilical vessels of preeclamptic pregnancies. Matrix metalloproteinase (MMP)-2 is elevated in the plasma of preeclamptic women and is a mediator of vascular reactivity; however, whether MMP-2 release is altered in preeclamptic endothelial cells is unknown. We hypothesize that MMP-2 release is enhanced in endothelial cells from preeclamptic compared with uncomplicated pregnancies and that this phenomenon may be mediated by an oxygen-dependent mechanism. Our specific hypothesis is that cells from normal pregnancies will demonstrate enhanced MMP-2 release at low oxygen (< 0.5%, 2%) compared to high oxygen (20%), thus mimicking the behavior of preeclamptic cells. METHODS: Human umbilical vein endothelial cells (HUVECs) from preeclamptic pregnancies (n = 4) and normal pregnancies (n = 4) were incubated for 12 hr in standard culture conditions (20% oxygen). In a separate series of experiments, HUVECs from normal pregnancies (n = 6) were incubated for 12 hr at < 0.5%, 2%, and 20% oxygen. Supernatants were analyzed for MMP-2 and tissue inhibitors of metalloproteinases (TIMP)-1 and -2. RESULTS: The HUVECs from women with preeclampsia demonstrated significantly enhanced release of MMP-2 (p < 0.05), TIMP-1 (p < 0.001), and TIMP-2 (p = 0.01) compared to normal cells. MMP-2 release from HUVECs from uncomplicated pregnancies was significantly elevated at 2% oxygen compared to < 0.5% and 20% oxygen (p < 0.05). TIMP-1 and -2 secretion was not altered with varying oxygen. CONCLUSIONS: Preeclamptic endothelial cells demonstrate significantly enhanced MMP-2, TIMP-1 and TIMP-2 release compared to normal cells. Our data show that there are significant effects of oxygen tension on MMP-2 release from normal cells; however, the magnitude of the enhanced release is small when compared to the differences in MMP-2 release in cells from preeclamptic and normal pregnancies. Furthermore, TIMP-1 and -2 release is not affected by changes in oxygen. It is unlikely that oxygen is a key mediator of the enhanced MMP-2, TIMP-1 and TIMP-2 release observed in preeclamptic cells.     

Vascular Endothelial Growth Factor Induces Endothelin-1 Production via Matrix Metalloproteinase-2 Rather than Endothelin-Converting Enzyme-1

Objective: To investigate the mechanism of vascular endothelial growth factor (VEGF)-induced endothelin-1 production in human umbilical vein endothelial cells (HUVECs). Methods: Endothelin-1 levels were measured in conditioned medium of women with preeclampsia HUVECs were treated with different concentrations of VEGF₁₆₅ and at various time intervals. Next, we measured endothelin-1 levels after HUVECs were also incubated with VEGF and endothelin-converting enzyme-1 (ECE-1) inhibitor or tissue inhibitors of matrix metalloproteinase-2 (TIMP-2). Additionally, the circulating levels of total and free VEGF, matrix metalloproteinase-2 (MMP-2), and endothelin-1 were measured in 20 preeclamptic patients and 20 healthy pregnant controls. Results: HUVECs treated with VEGF increased their endothelin-1 production in a concentration and time-dependent manner. The production of endothelin-1 was inhibited by TIMP-2, but not by the ECE-1 inhibitor. Total VEGF, MMP-2, and endothelin-1 concentrations were higher in preeclampsia and showed significant positive correlations between them. Conclusion: These findings suggest that VEGF-induced endothelin-1 production might be mediated by MMP-2 rather than by ECE-1 upregulation.     

MMP-2 levels are elevated in the plasma of women who subsequently develop preeclampsia.

OBJECTIVE: To determine levels of matrix metalloproteinase (MMP)-2 and MMP-9, and the tissue inhibitors of metalloproteinases (TIMP)-1 and TIMP-2 in the plasma of women destined to develop preeclampsia prior to the onset of clinical disease. STUDY DESIGN: Plasma samples were taken from women whose pregnancies were subsequently complicated by preeclampsia and from normal pregnant women at 22 and 26 weeks and at delivery or diagnosis. Following equal protein loading, MMP-2 and 9 and TIMP-1 and 2 were quantified using zymography and Western blot analysis, respectively. RESULTS: Plasma MMP-2 levels were significantly elevated at 22 weeks (p = 0.02) and at diagnosis (p = 0.003) in the preeclampsia group, but there was no difference at 26 weeks. TIMP-1 levels were significantly reduced in the preeclampsia group at 26 weeks (p = 0.0002), but TIMP-2 levels were not quantifiable. CONCLUSION: At all three gestational time points an imbalance in the MMP-2:TIMP-1 ratio was found in patients who subsequently developed preeclampsia. We speculate that increased net MMP-2 activity may contribute to the endothelial dysfunction that is central to the pathophysiology of preeclampsia.     

The Effects of Preeclampsia and Oxygen Environment on Endothelial Release of Matrix Metalloproteinase‐2

Background: There is evidence of altered vascular endothelial function in women with preeclampsia as well as in the endothelial cells from umbilical vessels of preeclamptic pregnancies. Matrix metalloproteinase (MMP)‐2 is elevated in the plasma of preeclamptic women and is a mediator of vascular reactivity; however, whether MMP‐2 release is altered in preeclamptic endothelial cells is unknown. We hypothesize that MMP‐2 release is enhanced in endothelial cells from preeclamptic compared with uncomplicated pregnancies and that this phenomenon may be mediated by an oxygen‐dependent mechanism. Our specific hypothesis is that cells from normal pregnancies will demonstrate enhanced MMP‐2 release at low oxygen (< 0.5%, 2%) compared to high oxygen (20%), thus mimicking the behavior of preeclamptic cells. Methods: Human umbilical vein endothelial cells (HUVECs) from preeclamptic pregnancies (n = 4) and normal pregnancies (n = 4) were incubated for 12 hr in standard culture conditions (20% oxygen). In a separate series of experiments, HUVECs from normal pregnancies (n = 6) were incubated for 12 hr at < 0.5%, 2%, and 20% oxygen. Supernatants were analyzed for MMP‐2 and tissue inhibitors of metalloproteinases (TIMP)‐1 and ‐2. Results: The HUVECs from women with preeclampsia demonstrated significantly enhanced release of MMP‐2 (p < 0.05), TIMP‐1 (p < 0.001), and TIMP‐2 (p = 0.01) compared to normal cells. MMP‐2 release from HUVECs from uncomplicated pregnancies was significantly elevated at 2% oxygen compared to < 0.5% and 20% oxygen (p < 0.05). TIMP‐1 and ‐2 secretion was not altered with varying oxygen. Conclusions: Preeclamptic endothelial cells demonstrate significantly enhanced MMP‐2, TIMP‐1 and TIMP‐2 release compared to normal cells. Our data show that there are significant effects of oxygen tension on MMP‐2 release from normal cells; however, the magnitude of the enhanced release is small when compared to the differences in MMP‐2 release in cells from preeclamptic and normal pregnancies. Furthermore, TIMP‐1 and ‐2 release is not affected by changes in oxygen. It is unlikely that oxygen is a key mediator of the enhanced MMP‐2, TIMP‐1 and TIMP‐2 release observed in preeclamptic cells.     

Expression of matrix metalloproteinase-2 in serous borderline ovarian tumors is associated with noninvasive implant formation

OBJECTIVE: To investigate matrix metalloproteinase (MMP) proteolytic and vascular endothelial growth factor (VEGF) and receptor (VEGFR-1, VEGFR-2) angiogenetic capacity in serous borderline ovarian tumors (S-BOTs) for women with and without noninvasive implants. METHODS: The population was made up of 99 patients with S-BOTs as the primary diagnosis between 1985 and 1995, 44 of whom had noninvasive implants and 55 without implants. MMP-2, MMP-14, the type-2 tissue inhibitor of MMPs (TIMP-2), and VEGF and receptors (VEGFR-1, VEGFR-2) were examined by immunhistochemistry. RESULTS: Strong positive (+++) MMP-2 staining was found more frequently in women with primary S-BOTs and noninvasive implants (76%) than in those without implants (53%; p < 0.05). In contrast, staining for MMP-14 and TIMP-2 was not significantly different in the two groups. Furthermore, expression of MMP-2, MMP-14, and TIMP-2 was similar in primary tumors and in their noninvasive implants. Most tumors in both groups had no VEGF expression (84% in the noninvasive implant group and 82% in the group without implants), while moderate (++) to strong (+++) expression of VEGFR-1 and VEGFR-2 was detected in 79% and 94% of the two tumor groups, with no significant difference between the groups. CONCLUSIONS: Enhanced MMP-2 was seen in primary S-BOT with noninvasive implants. The presence of noninvasive implants was prognostic for disease-free survival.     

Vascular endothelial growth factor induces endothelin-1 production via matrix metalloproteinase-2 rather than endothelin-converting enzyme-1.

OBJECTIVE: To investigate the mechanism of vascular endothelial growth factor (VEGF)-induced endothelin-1 production in human umbilical vein endothelial cells (HUVECs). METHODS: Endothelin-1 levels were measured in conditioned medium of women with preeclampsia HUVECs were treated with different concentrations of VEGF(165) and at various time intervals. Next, we measured endothelin-1 levels after HUVECs were also incubated with VEGF and endothelin-converting enzyme-1 (ECE-1) inhibitor or tissue inhibitors of matrix metalloproteinase-2 (TIMP-2). Additionally, the circulating levels of total and free VEGF, matrix metalloproteinase-2 (MMP-2), and endothelin-1 were measured in 20 preeclamptic patients and 20 healthy pregnant controls. RESULTS: HUVECs treated with VEGF increased their endothelin-1 production in a concentration and time-dependent manner. The production of endothelin-1 was inhibited by TIMP-2, but not by the ECE-1 inhibitor. Total VEGF, MMP-2, and endothelin-1 concentrations were higher in preeclampsia and showed significant positive correlations between them. CONCLUSION: These findings suggest that VEGF-induced endothelin-1 production might be mediated by MMP-2 rather than by ECE-1 upregulation.     

Raised serum levels of matrix metalloproteinase-9 in women with polycystic ovary syndrome and its association with insulin-like growth factor binding protein-1

Background.?It has been suggested in recent studies that matrix metalloproteinases (MMPs) may be implicated in the pathogenesis of polycystic ovary syndrome (PCOS) through regulating ovarian tissue remodeling. In addition to degrading the extracellular matrix, MMPs exhibit the ability to cleave insulin-like growth factor binding protein-1 (IGFBP-1), the major regulator of insulin-like growth factor-I (IGF-I) in serum. The present study aimed to investigate the possible role of MMPs in the pathophysiology of PCOS.

Methods.?Serum levels of MMP-9, tissue inhibitor of metalloproteinase-1 (TIMP-1), IGF-I and IGFBP-1 were measured in 42 patients with PCOS and 30 healthy women with regular menstruation, matched for age and body mass index. Correlation between IGFBP-1 and other parameters in the PCOS group was analyzed by Pearson's linear correlations.

Results.?Serum MMP-9 concentrations and MMP-9/TIMP-1 ratios were significantly higher in PCOS women than in controls. Serum levels of IGFBP-1 were markedly lower in the PCOS group. There was a negative correlation between serum IGFBP-1 and MMP-9 in women with PCOS.

Conclusion.?Our results raise the possibility that MMPs may be implicated in the pathophysiology of PCOS either by regulating ovarian tissue remodeling or indirectly by facilitating IGF-I bioavailability through proteolysis of IGFBP-1.     

Gelatinase A (MM-2), gelatinase B (MMP-9) and their inhibitors (TIMP 1, TIMP-2) in serum of women with endometriosis: Significant correlation between MMP-2, MMP-9 and their inhibitors without difference in levels of matrix metalloproteinases and tissue inhibitors of metalloproteinases in relation to the severity of endometriosis

Endometriosis is a highly prevalent gynecological condition, where the formation of endometriotic foci is linked with locally increased activity of matrix metalloproteinases (MMPs). In the present study, we tested the hypothesis that raised serum levels of MMPs might reflect the severity of endometriosis. We compared serum levels of MMP-2 and MMP-9, and of their tissue inhibitors TIMP-1 and TIMP-2, in infertile women, matched for age and body mass index, with either mild (stage I, END-I; n = 15) or severe endometriosis (stage IV, END-IV; n = 22). There was no difference in the concentrations of MMP-2, MMP-9, TIMP-1 and TIMP-2 between the analyzed groups. There was, however, a correlation between MMP-9 and TIMP-1 for the combined group (n = 37) (r = 0.48; p = 0.0032) and in women with END-IV (r = 0.51; p = 0.0163), as well as a highly significant correlation between MMP-2 and TIMP-2 for the combined group (r = 0.69; p = 0.0001), END-I (r = 0.51; p = 0.0406) and END-IV groups (r = 0.77; p = 0.0001). There was also a significant correlation between TIMP-1 and TIMP-2 in the combined and END-IV groups (r = 0.39; p = 0.0182 and r = 0.5450; p = 0.0099, respectively). The balance between MMPs and their inhibitors is preserved in the serum of women with endometriosis, but serum concentrations of MMP-2, MMP-9, TIMP-1 and TIMP-2 cannot be considered to represent a valid measure of the severity of endometriosis.     

Serum VEGF and PGF may be significant markers in prediction of severity of preeclampsia

Objective: The aim of this study evaluate the value of vascular endothelial growth factor (VEGF) and placental growth factor (PGF) serum levels in prediction of preeclampsia, severity and onset time of the disease.

Methods: Twenty five placentas of pregnant women diagnosed with preeclampsia (15 severe preeclampsia, 10 mild preeclampsia) and peripheral venous blood samples were collected. The placental and serum levels of VEGF and PGF were measured.

Results: VEGF level was significantly higher in cases and the optimal cut-off point was calculated as 600.5 to differentiate the cases and the controls, with 64% sensitivity and 100% specificity. There was a significant increase in median serum level of VEGF in severe cases compared to the mild cases and the controls. The optimal cut-off point for VEGF was calculated as 673.5 to differentiate mild and severe cases, with 93.3% sensitivity and 90% specificity. Whereas, PGF was significantly lower in severe cases than that in the mild cases and controls. The optimal cut-off point for PGF was calculated as 16.1 to differentiate mild and severe cases, with 66.7% sensitivity and 100% specificity.

Conclusion: VEGF and PGF may be significant markers in prediction of severity of preeclampsia, and VEGF may also be valuable in prediction of preeclampsia.     

子痫前期患者滋养细胞浸润相关基因mRNA及蛋白表达的研究

目的研究子痫前期患者胎盘滋养细胞基质金属蛋白酶(MMP)9、2及其组织抑制物(TIMP)1、2,肿瘤转移抑制基因KiSS-1 mRNA、蛋白的表达变化及其与子痫前期发病的关系。方法采用RT-PCR、免疫印迹(western blot)法对30例正常足月妊娠妇女(正常妊娠组)和10例妊娠期高血压患者(高血压组)及27例子痫前期(子痫前期组)患者胎盘滋养细胞中MMP-9、MMP-2、KiSS-1、TIMP-1和TIMP-2基因mRNA及蛋白表达水平[均以相对吸光度(A)表示]进行检测;应用明胶酶谱分析法,检测3组妇女胎盘孵育液MMP-9、MMP-2活性。结果(1)胎盘滋养细胞浸润相关基因mRNA表达水平:子痫前期组MMP-9、MMP-2 mRNA表达水平分别为0．39±0．05和0．71±0．16,均明显低于正常妊娠组的0．78±0．11和1．63±0．31,两组分别比较,差异有统计学意义(P均<0．05)。高血压组MMP-9 mRNA的表达水平明显高于重度子痫前期患者,差异有统计学意义(P<0．05)。子痫前期组胎盘滋养细胞KiSS-1 mRNA和TIMP-1 mRNA的表达水平分别为1．97±0．21和1．11±0．18,均明显高于正常妊娠组的0．69±0．27和0．65±0．19,差异均有统计学意义(P<0．05);高血压组胎盘滋养细胞KiSS-1 mRNA表达水平低于子痫前期组,但与正常妊娠组比较,差异无统计学意义(P>0．05)。重度子痫前期患者胎盘滋养细胞TIMP-2 mRNA表达水平明显高于正常妊娠组,差异均有统计学意义(P<0．05)。(2)胎盘滋养细胞浸润相关基因的蛋白表达水平:子痫前期组MMP-9、MMP-2基因的蛋白表达水平分别为1．07±0．35和0．74±0．23,均明显低于正常妊娠组的2．43±0．92和1．48±0．78,差异均有统计学意义(P<0．05)。子痫前期组胎盘KISS-1和TIMP-1蛋白表达水平分别为2．46±0．39和1．51±0．40,均明显高于正常妊娠组的0．91±0．35和0．93±0．56,差异均有统计学意义(P<0．05)。子痫前期组胎盘滋养细胞TIMP-2蛋白表达水平与正常妊娠组及高血压组比较,差异均无统计学意义(P>0．05)。(3)胎盘MMP-9和MMP-2酶比活性:子痫前期组分别为(2．67±0．53)和(1．13±0．28)灰度·g-1·L-1,均明显低于正常妊娠组的(8．44±3．70)和(3．87±1．43)灰度·g-1·L-1,差异均有统计学意义(P<0．05)。结论子痫前期患者胎盘滋养细胞促进浸润基因。MMP-9、MMP-2表达降低和抑制浸润基因KiSS-1和TIMP-1表达升高,可能在子痫前期胎盘缺血缺氧中起重要作用。     

Matrix metalloproteinase-13 and membrane type-1 matrix metalloproteinase in peritoneal fluid of women with endometriosis

Objective.?It is becoming more and more evident that different types of matrix metalloproteinases (MMPs) and their tissue inhibitors (TIMPs) are involved in the pathogenesis of endometriosis. The aim of the present study was to measure levels of the active forms of MMP-13 and membrane type-1 matrix metalloproteinase (MT1-MMP)/MMP-14 as well as TIMP-2 in the peritoneal fluid of women with endometriosis.

Study design.?We determined the levels of the active forms MMP-13 and MT1-MMP/MMP-14 as well as TIMP-2 in the peritoneal fluid of 20 women with endometriosis and 18 controls by different types of enzyme-linked immunosorbent assay.

Results.?We found that the concentrations (mean?±?standard deviation) of total active MMP-13 and endogenous active MT1-MMP/MMP-14 in the peritoneal fluid of patients with endometriosis were 1.69?±?0.67 and 3.12?±?1.07?ng/ml, respectively. In control women the corresponding values were 3.02?±?0.43 and 4.45?±?1.03?ng/ml. The differences were statistically significant (p?<?0.0001 and p?<?0.0004 for MMP-13 and MMP-14, respectively). Levels of TIMP-2 did not differ significantly.

Conclusions.?Decreased concentrations of active MMP-13 and MT1-MMP/MMP-14 may imply that the proteolytic activity of the peritoneal milieu of women with endometriosis is disturbed, which may have implications in the pathogenesis of the disease.     

Serum visfatin,vascular endothelial growth factor and matrix metalloproteinase-9 in women with polycystic ovary syndrome

Aim: To evaluate serum concentrations of visfatin, vascular endothelial growth factor (VEGF) and matrix metalloproteinase-9 (MMP-9) in women with polycystic ovary syndrome (PCOS) and to investigate their possible role as early endothelial markers in PCOS.

Methods: Forty-two women with PCOS and 42 controls, matched for age and weight, were included in the study. Serum concentrations of follicle-stimulating hormone (FSH), luteinizing hormone (LH), prolactin, total testosterone (tT), Δ₄-androstenedione (Δ₄A), dehydroepinadrosterone sulphate (DHEA-S), 17-OH-progesterone, sex hormone-binding protein (SHBG), thyroid-stimulating hormone (TSH), free thyroxine (fT₄), visfatin, VEGF and MMP-9 were measured in all women; free androgen index (FAI) was calculated as well. Receiver-operating characteristic (ROC) analysis was performed to examine if visfatin, tT or FAI can predict the clinical status (PCOS or control).

Results: LH, Δ₄A, tT and FAI concentrations were higher in PCOS than in controls (p?=?0.002, 0.029, 0.0005 and 0.014, respectively). Visfatin, VEGF and MMP-9 concentrations were higher in women with PCOS than controls (p?=?0.019, 0.001 and 0.002, respectively). In ROC analysis, area under the curve (AUC) in the prediction of clinical status was 0.641 for visfatin (p?=?0.026), 0.731 for tT (p?=?0.001) and 0.666 for FAI (p?=?0.010), with no difference among them (p?=?0.117).

Conclusions: Visfatin may induce the expression of pro-angiogenic factors, such as VEGF and MMP-9, in women with PCOS, inplying gradually development of endothelial dysfunction. Further studies are required to clarify these findings.     

Matrix metalloproteinase release from placental explants of pregnancies complicated by intrauterine growth restriction

OBJECTIVE: There is evidence of impaired placental development in intrauterine growth restriction (IUGR). Matrix metalloproteinases (MMPs) are extracellular matrix-degrading enzymes that are released by placental cells during tissue remodeling processes. We hypothesized 1) that release of MMP-2 and -9 is decreased and/or release of tissue inhibitors of metalloproteinases (TIMPs) is increased from placental explants in pregnancies complicated by IUGR and 2) that oxygen levels affect such release. METHODS: Placental villous explants from normal (n = 7) and IUGR (n = 7) pregnancies were cultured at high (20%) and low (3%) oxygen levels for 24 hours. Supernatants were analyzed for MMP-2 and MMP-9 by zymography and for TIMP-1 and -2 by western blot analysis. RESULTS:: At 20% oxygen there was significantly reduced MMP-2 (P < .05) and TIMP-1 (P < .01) release and a trend for decreased MMP-9 release (P = .07) in explants from IUGR pregnancies compared with normal pregnancies; however, there were no differences at 3% oxygen. TIMP-2 was below detectable levels in all samples. Although MMP-2 and TIMP-1 release was significantly reduced at 3% compared with 20% oxygen in explants from both normal (P < .001; P < .05) and IUGR (P < .05) pregnancies, MMP-2 release changed less in IUGR compared with normal explant cultures. There were no significant effects of oxygen on MMP-9 release. CONCLUSION: Placental explants from IUGR pregnancies demonstrated reduced MMP-2, MMP-9, and TIMP-1 release compared with explants from normal pregnancies at high (20%) but not low (3%) oxygen.     

Maternal plasma levels of cytokines in normal and preeclamptic pregnancies and their relationship with diastolic blood pressure and fibronectin levels

BACKGROUND: To determine the plasma concentrations of placental growth factor (PLGF), vascular endothelial growth factor (VEGF), transforming growth factor-beta1 (TGF-beta1), soluble tumor necrosis factor alpha receptor (sTNFp55), interleukin-2 receptor (IL-2R), and interleukins 6 and 10 (IL-6, IL-10) in normotensive and preeclamptic women, and to evaluate the correlations between these cytokines and the diastolic blood pressure and fibronectin levels. METHODS: A prospective case-control study. Thirty-five women with preeclampsia were compared with 34 healthy women with uncomplicated pregnancies. Peripheral venous blood samples were obtained and plasma levels of PLGF, VEGF, TGF-beta1, sTNFp55, IL-2R, IL-6 and IL-10 were measured by an enzyme-linked immunoassay and fibronectin by a radial immundiffusion technic. RESULTS: In preeclampsia PLGF and VEGF levels were significantly lower, and TGF-beta1, sTNFp55, IL-2R, IL-6 and IL-10 levels were significantly higher than in normotensive pregnancy (p < 0.001). The plasma levels of PLGF and VEGF significantly decreased, whereas TGF-beta1, sTNFp55, IL-2R, IL-6 and IL-10 levels significantly increased with the increments in diastolic blood pressure and fibronectin levels (p < 0.001). CONCLUSIONS: Altered concentrations of various cytokines might explain the shallow placentation and endothelial cell dysfunction described in preeclampsia. The clinical severity of preeclampsia seems to correlate with the severity of the cytokine abnormalities.     

Regulation of matrix metalloproteinases and their inhibitors in uterine endometrial cells of patients with and without endometriosis

OBJECTIVE: To determine whether alterations in the secretion and regulation of matrix metalloproteinases (MMPs) and their inhibitors are present in uterine endometrial cells from endometriosis patients. STUDY DESIGN: In an in vitro study, uterine endometrial cells from 19 regularly cycling women with and 32 without endometriosis were treated with diethyl stilbestrol, promegestone (R5020), interleukin-1 (IL-1) and tumor necrosis factor a (TNF-alpha). Culture supernatants were assayed for MMPs 1, 2, 3, and 9, and for tissue inhibitors of MMP (TIMP-1 and TIMP-2) by ELISA. RESULTS: MMP-3 was secreted in high concentrations, moderate concentrations were seen for MMP-1 and MMP-2, and very low concentrations for MMP-9. Substantially more TIMP-1 than TIMP-2 was secreted. MMP-1 and MMP-3 were uniformly attenuated by R5020, while MMP-2 was not influenced by hormone treatment. MMP-3 was upregulated by TNF-alpha in all samples while IL-1 only increased secretion in cells from endometriosis patients. CONCLUSION: The upregulation of MMP-3 by IL-1 may contribute to an increased invasiveness of uterine endometrial fragments in endometriosis patients.     

Endocan,a putative endothelial cell marker,is elevated in preeclampsia,decreased in acute pyelonephritis,and unchanged in other obstetrical syndromes

Objective: Endocan, a dermatan sulphate proteoglycan produced by endothelial cells, is considered a biomarker for endothelial cell activation/dysfunction. Preeclampsia is characterized by systemic vascular inflammation, and endothelial cell activation/dysfunction. Therefore, the objectives of this study were to determine whether: (1) plasma endocan concentrations in preeclampsia differ from those in uncomplicated pregnancies; (2) changes in plasma endocan concentration relate to the severity of preeclampsia, and whether these changes are specific or observed in other obstetrical syndromes such as small-for-gestational age (SGA), fetal death (FD), preterm labor (PTL) or preterm prelabor rupture of membranes (PROM); (3) a correlation exists between plasma concentration of endocan and angiogenic (placental growth factor or PlGF)/anti-angiogenic factors (soluble vascular endothelial growth factor receptor or sVEGFR-1, and soluble endoglin or sEng) among pregnancies complicated by preeclampsia; and (4) plasma endocan concentrations in patients with preeclampsia and acute pyelonephritis (both conditions in which there is endothelial cell activation) differ.

Method: This cross-sectional study included the following groups: (1) uncomplicated pregnancy (n?=?130); (2) preeclampsia (n?=?102); (3) pregnant women without preeclampsia who delivered an SGA neonate (n?=?51); (4) FD (n?=?49); (5) acute pyelonephritis (AP; n?=?35); (6) spontaneous PTL (n?=?75); and (7) preterm PROM (n?=?64). Plasma endocan concentrations were determined in all groups, and PIGF, sEng and VEGFR-1 plasma concentrations were measured by ELISA in the preeclampsia group.

Results: (1) Women with preeclampsia had a significantly higher median plasma endocan concentration than those with uncomplicated pregnancies (p?=?0.004); (2) among women with preeclampsia, the median plasma endocan concentration did not differ significantly according to disease severity (p?=?0.1), abnormal uterine artery Doppler velocimetry (p?=?0.7) or whether diagnosis was made before or after 34 weeks gestational age (p?=?0.3); (3) plasma endocan concentration in women with preeclampsia correlated positively with plasma anti-angiogenic factor concentrations [sVEGFR-1: Spearman rho 0.34, p?=?0.001 and sEng: Spearman rho 0.30, p?=?0.003]; (4) pregnancies complicated by acute pyelonephritis with bacteremia had a lower median plasma endocan concentration than pregnancies complicated by acute pyelonephritis without bacteremia (p?=?0.004), as well as uncomplicated pregnancies (p?=?0.001); and (5) there was no significant difference in the median plasma endocan concentration between uncomplicated pregnancies and those complicated by FD, delivery of an SGA neonate, PTL or preterm PROM (other members of the “great obstetrical syndromes”; each p?>?0.05).

Conclusion: Median maternal plasma endocan concentrations were higher preeclampsia and lower in acute pyelonephritis with bacteremia than in uncomplicated pregnancy. No significant difference was observed in the median plasma endocan concentration between other great obstetrical syndromes and uncomplicated pregnancies. The difference in the direction of change of endocan in preeclampsia and acute pyelonephritis with bacteremia may be consistent with the view that both disease entities differ in pathogenic mechanisms, despite their associations with systemic vascular inflammation and endothelial cell activation/dysfunction.     

Matrix metalloproteinases and their inhibitors in the cervical mucus plug at term of pregnancy

OBJECTIVE: This study was undertaken to evaluate the presence of matrix metalloproteinases (MMPs) and their inhibitors in the cervical mucus plug obtained during active labor at term. STUDY DESIGN: Cervical mucus plugs from 17 healthy women in normal active labor were homogenized, extracted, and analyzed by 4 different assays. RESULTS: Gelatin zymography revealed large amounts of MMP-2 and MMP-9, and triple-helical collagen degradation demonstrated collagenase activity (MMP-8). Reverse zymography showed that tissue inhibitor of matrix metalloproteinase 1 (TIMP-1) and TIMP-2 are detectable in the cervical mucus plug. Quantification by enzyme-linked immunosorbent assay confirmed the presence of MMP-2, MMP-8, MMP-9, and TIMP-1. CONCLUSION: Our results suggest that proteolytic or nonproteolytic effects of MMP and TIMP may play a part in the function of the cervical mucus plug of pregnancy.     

Thrombin-dependent regulation of matrix metalloproteinase (MMP)-9 levels in human fetal membranes*

Objective.?Amniochorion matrix metalloproteinase (MMP)-9 levels increase during labor, reaching a maximum in patients with preterm premature rupture of membranes (PPROM). Bleeding is a major risk factor for PPROM. Since such hemorrhage into the tissue factor-enriched decidua induces intense thrombin formation, we determined whether thrombin stimulates MMP levels in amniochorionic membranes.

Study design.?Fetal membrane (amniochorion) cultures were maintained in media with and without thrombin, lipopolysaccharide (LPS), thrombin receptor agonist peptide (TRAP)-14, and the anti-inflammatory steroid, dexamethasone (DEX). Concentrations of MMP-9, MMP-1, and tissue inhibitor of metalloproteinase (TIMP)-1 in culture media were measured by ELISA and normalized to total cell protein.

Results.?The presence of thrombin induced MMP-9 levels. TRAP-14, a thrombin receptor agonist, also significantly increased MMP-9 levels, suggesting that thrombin-induced changes in MMP-9 expression were mediated through the thrombin receptor. Conversely, levels of MMP-1 and TIMP-1 were not affected by thrombin treatment, indicative of specificity of its action. The presence of LPS increased the concentration of MMP-9 and MMP-1. In contrast, DEX treatment significantly reduced MMP-9 levels.

Conclusion.?Our findings clearly demonstrated that thrombin treatment selectively increased the concentration of MMP-9 in culture media of amniochorionic membranes. Our results provide a potential mechanism through which alterations in hemostasis promote PPROM through thrombin-dependent stimulation of MMP-9.     

Placental Growth Factor (PlGF) Is a Surrogate Marker in Preeclamptic Hypertension

Objective: To evaluate plasma levels of angiogenic factors and their association with preeclampsia. Methods: Twenty-three women with preeclampsia and nine normotensive pregnant women from the Maternity of Hospital das Clínicas of Belo Horizonte/MG-Brazil were assessed by National High Blood Pressure Education Program Working Group Creteria (NHBPEPWG). The plasma levels of vascular endothelial growth factor (VEGF) and Placental growth factor (PlGF) were determined by ELISA assay. Results: Plasma concentration of PlGF was 12-fold lower in preeclampsia versus non preeclampsia pregnancies. An inverse correlation was observed between PlGF plasma levels and mean arterial pressure (MAP); a decrease in 1pg/mL of PlGF resulted in 6.18 mm Hg increase in MAP. Conclusion: These results indicate that PlGF is related to MAP in pregnant women.     

Maternal plasma concentrations of sST2 and angiogenic/anti-angiogenic factors in preeclampsia

Abstract

Objectives: Angiogenic/anti-angiogenic factors have emerged as one of the promising biomarkers for the prediction of preeclampsia. Since not all patients with preeclampsia can be identified by these analytes, the search for additional biomarkers continues. The soluble form of ST2 (sST2), a protein capable of binding to interleukin (IL)-33 and thus contributing to a Th1-biased immune response, has been reported to be elevated in maternal plasma of women with preeclampsia. The aims of this study were to examine: (1) differences in maternal plasma concentrations of sST2 and IL-33 between women diagnosed with preeclampsia and those having uncomplicated pregnancies; (2) the relationship between sST2, umbilical and uterine artery Doppler velocimetry, and the severity of preeclampsia; and (3) the performance of sST2 and angiogenic/anti-angiogenic factors in identifying patients with preeclampsia at the time of diagnosis.

Methods: This cross-sectional study included women with preeclampsia (n?=?106) and women with an uncomplicated pregnancy (n?=?131). Plasma concentrations of sST2, IL-33, soluble vascular endothelial growth factor receptor (sVEGFR)-1, soluble endoglin (sEng) and placental growth factor (PlGF) were determined by enzyme linked immune sorbent assay. Area under the receiver operating characteristic curve (AUC) for the identification of preeclampsia was examined for each analyte.

Results: (1) Patients with preeclampsia had a higher mean plasma concentrations of sST2 than those with an uncomplicated pregnancy (p?<?0.0001), while no significant difference in the mean plasma concentration of IL-33 between the two groups was observed; (2) the magnitude of this difference was greater in early-onset, compared to late-onset disease, and in severe compared to mild preeclampsia; (3) sST2 plasma concentrations did not correlate with the results of uterine or umbilical artery Doppler velocimetry (p?=?0.7 and p?=?1, respectively) among women with preeclampsia; (4) sST2 correlated positively with plasma concentrations of sVEGFR1-1 and sEng (Spearman’s Rho?=?0.72 and 0.63; each p?<?0.0001), and negatively with PlGF (Spearman’s Rho?=??0.56, p?<?0.0001); and (5) while the AUC achieved by sST2 and angiogenic/anti-angiogenic factors in identifying women with preeclampsia at the time of diagnosis were non-significantly different prior to term (<37 weeks of gestation), thereafter the AUC achieved by sST2 was significantly less than that achieved by angiogenic/anti-angiogenic factors.

Conclusions: Preeclampsia is associated with increased maternal plasma concentrations of sST2. The findings that sST2 concentrations do not correlate with uterine or umbilical artery Doppler velocimetry in women with preeclampsia suggest that elevated maternal plasma sST2 concentrations in preeclampsia are not related to the increased impedance to flow in the utero-placental circulation. The performance of sST2 in identifying preeclampsia at the time of diagnosis prior to 37 weeks of gestation was comparable to that of angiogenic/anti-angiogenic factors. It remains to be elucidated if an elevation of maternal plasma sST2 concentrations in pregnancy is specific to preeclampsia.