- Email: [email protected]
Serum from preeclamptic women induces caspase-3 activation in first-trimester trophoblast cells
SMFM Abstracts S95
Volume 189, Number 6 Am J Obstet Gynecol 109
EXTREMELY ELEVATED SECOND-TRIMESTER HUMAN CHORIONIC GONADOTROPIN LEVELS ARE ASSOCIATED WITH ADVERSE PREGNANCY OUTCOME NISHA VYAS1, CAROL NOREM2, DAVID WALTON1, EDGAR SCHOEN2, JENNIFER O’KEEFE2, ROBYN KRIEGER2, TRINH TO2, 1The Permanente Medical Group, Obstetrics and Gynecology, Oakland, CA 2The Permanente Medical Group, Genetics, Oakland, CA OBJECTIVE: It is exceedingly rare to find an extremely elevated human chorionic gonadotropin (hCG) level (>4.0 multiples of the median) at the time of second-trimester maternal serum screening. The purpose of this report is to assess the clinical value of extremely elevated second-trimester hCG levels as an indicator of pregnancy complications and adverse outcomes. STUDY DESIGN: We studied serum hCG levels in 48,028 gravidas undergoing California Expanded Alpha-Fetoprotein serum screening at 15-20 weeks’ gestation during a 2-year period at a large health maintenance organization. Women who were screened too early, who had an indeterminate screening result, or who had multiple gestations were excluded. RESULTS: Of the 48,028 women, 166 (0.35%) had hCG levels greater than 4.0 multiples of the median (MoM) for gestational age. The Table depicts the rate of specific pregnancy outcomes occurring in women with an extremely elevated hCG. CONCLUSION: Pregnancies complicated by second-trimester hCG serum screening levels greater than 4.0 MoM have a markedly higher rate of stillbirth, pregnancy-related hypertension, abruption, and second-trimester fetal loss. These findings are in sharp contrast to prior studies of elevated hCG less than 4.0 MoM, which demonstrated little clinical value in predicting adverse pregnancy outcome.
111
SERUM FROM PREECLAMPTIC WOMEN INDUCES CASPASE-3 ACTIVATION IN FIRST-TRIMESTER TROPHOBLAST CELLS SHAWN STRASZEWSKI1, GIL MOR2, ROBERTO ROMERO3, DONNA NEALE4, 1 Yale University, Molecular, Cellular and Developmental Biology, New Haven, CT 2Yale University, Obstetrics and Gynecology, New Haven, CT 3National Institute of Child Health and Human Development, Detroit, MI 4Yale University, Obstetrics and Gynecology; Division of Maternal-Fetal Medicine, New Haven, CT OBJECTIVE: Preeclampsia is associated with insufficient trophoblast invasion, which may be a result of increased trophoblast apoptosis during the first trimester of pregnancy. Recently, we demonstrated that serum obtained from women with preeclampsia decreased trophoblast cell viability and rendered first-trimester trophoblast cells sensitive to Fas-mediated apoptosis. In order to further investigate these results, we evaluated whether preeclamptic serum activates effector caspase-3, an indicator of apoptosis, in first-trimester trophoblast cells. STUDY DESIGN: The H8 first-trimester trophoblast cell line was cultured in the presence of 10% serum obtained from either preeclamptic women or normotensive pregnant controls for 48 hours. Trophoblast cell viability was evaluated using the Cell Titer 96 assay. Activation of capase-3 was determined by Western blot analysis. RESULTS: Similar to our previous study, a significant decrease in trophoblast cell viability was observed following treatment with serum from preeclamptic patients in comparison to the normotensive controls. Only the pro-form of caspase-3 (p33) was detected in trophoblast cells treated with serum from normotensive patients. In contrast, the active forms of caspase-3 (17/19 kDa) were observed in trophoblast cells treated with serum obtained from preeclamptic patients. CONCLUSION: The findings described in this study suggest that a link exists between factor(s) in maternal serum and the induction of trophoblast apoptosis. The identification of the active forms of caspase-3 in trophoblast cells treated with serum obtained from preeclamptic patients, but not from the normotensive controls, supports the hypothesis that factors circulating in preeclamptic women reduce trophoblast cell viability through activation of the apoptotic cascade.
112
A PROTEOMIC APPROACH FOR THE IDENTIFICATION OF BIOLOGICAL MARKERS OF PREECLAMPSIA TINNAKORN CHAIWORAPONGSA1, RICARDO GOMEZ1, SAMUEL EDWIN1, GIL MOR2, TIMOTHY BARDER3, ROBERTO ROMERO1, 1Perinatology Research Branch, NICHD/ NIH/DHHS, Detroit, MI 2Yale University, Obstetrics and Gynecology, New Haven, CT 3Epogen, Inc, Darien, IL OBJECTIVE: Obstetrics is moving beyond genomics and into proteomics. The Plasma Proteome Project aims to comprehensively and quantitatively analyze plasma proteins to elucidate biomarkers for disease. The characterization of plasma proteins presents multiple challenges because of protein diversity and the large dynamic range of expression. The purpose of this study was to use a novel approach for proteomic analysis to identify differentially expressed proteins in the plasma of normal women and those with preeclampsia. STUDY DESIGN: Two-dimensional chromatography was employed to separate and analyze samples from normal women (n = 41) and those with preeclampsia (n = 41). The first dimension employed high-performance chromatofocusing to identify the isoelectric point, and the second dimension, high-resolution reverse-phase high-pressure liquid chromatography to provide protein hydrophobicity of each fraction. A 2D protein map was generated using software that displays the isoelectric point and the degree of hydrophobicity. The maps were used as pattern recognition tools to discern unique combinations of proteins expressed in plasma. RESULTS: The following two partial 2D protein maps represent the differential plasma proteome of normal pregnant women and those with preeclampsia. Dramatic differences between normal and preeclamptic plasma are observed. Interface with mass spectrometry using the liquid phase output allows protein analysis and identification. CONCLUSION: Plasma proteomic analysis using 2D chromatography revealed unique differential expression in preeclampsia. This novel approach can be used for the identification of biomarkers and patterns of biomarkers predictive of disease.
Complications and outcomes associated with extremely elevated hCG hCG >4.0 MoM, N = 166 Stillbirth Second-trimester loss Hypertensive disease Abruption IUGR Postpartum hemorrhage Preterm delivery
110
n n n n n n n
= = = = = = =
5; (3%) 11; (7%) 3; (19%) 10; (6%) 7; (4%) 8; (5%) 12; (7%)
SOLUBLE FMS-LIKE TYROSINE KINASE 1 LEVELS ARE ELEVATED IN PREECLAMPISA AND CORRELATE WITH DISEASE SEVERITY EUGENE CHANG1, CHRISTOPHER ROBINSON1, DONNA JOHNSON1, MICHAEL ARMSTRONG1, 1Medical University of South Carolina, Obstetrics and Gynecology, Charleston, SC OBJECTIVE: Soluble Flt-1 (sFlt1) is a receptor tyrosine kinase that binds and antagonizes the angiogenic/permeability factors, vascular endothelial (VEGF) and placental growth factor (PlGF). Excessive sFlt-1 causes hypertension and proteinuria in rats. The purpose of this study was to determine whether sFlt-1 levels correlate with the severity of preeclampsia. STUDY DESIGN: After IRB approval, blood samples from normotensive controls and from patients with preeclampsia were collected. Patients were classified with mild or severe disease based on ACOG criteria. Patients with chronic hypertension, collagen vascular disease, diabetes, and multiple gestations were excluded. Serum was stored at ÿ708C. sFlt-1 concentration (pg/mL) was measured by ELISA. Statistical analysis was performed with ANOVA and Student-t test. RESULTS: 32 controls and 47 patients with preeclampsia were enrolled. 23 patients had mild preeclampsia and 24 had severe. Mean gestational age (GA) (weeks) was similar for both controls and mildly affected patients (37.2 ± 2.0 vs 35.8 ± 2.7, P = NS) but GA was more premature for severely affected patients (31.6 ± 3.0 P < 0.03) When compared to controls, sFlt-1 levels were increased in preeclamptic patients (100.3 ± 142.0 vs 507.7 ± 441.5, P < 0.05). sFlt-1 was significantly higher in patients with severe preeclampsia compared to patients with mild disease (644.9 ± 457.9 vs 344.3 ± 367.9, P < 0.05). sFlt-1 was also significantly higher in patients with more than 1 gm of proteinuria than less than 1 gm (320.4 ± 408 vs 631.7 ± 406.7, P < 0.04) CONCLUSION: sFLt-1 levels are increased in patients with preeclampsia. sFlt-1 levels correlate with the severity of disease. s-Flt-1 may antagonize the normal effects of PlGF and VEGF.
normal (left) and PE (right)
Volume 189, Number 6 Am J Obstet Gynecol 109
EXTREMELY ELEVATED SECOND-TRIMESTER HUMAN CHORIONIC GONADOTROPIN LEVELS ARE ASSOCIATED WITH ADVERSE PREGNANCY OUTCOME NISHA VYAS1, CAROL NOREM2, DAVID WALTON1, EDGAR SCHOEN2, JENNIFER O’KEEFE2, ROBYN KRIEGER2, TRINH TO2, 1The Permanente Medical Group, Obstetrics and Gynecology, Oakland, CA 2The Permanente Medical Group, Genetics, Oakland, CA OBJECTIVE: It is exceedingly rare to find an extremely elevated human chorionic gonadotropin (hCG) level (>4.0 multiples of the median) at the time of second-trimester maternal serum screening. The purpose of this report is to assess the clinical value of extremely elevated second-trimester hCG levels as an indicator of pregnancy complications and adverse outcomes. STUDY DESIGN: We studied serum hCG levels in 48,028 gravidas undergoing California Expanded Alpha-Fetoprotein serum screening at 15-20 weeks’ gestation during a 2-year period at a large health maintenance organization. Women who were screened too early, who had an indeterminate screening result, or who had multiple gestations were excluded. RESULTS: Of the 48,028 women, 166 (0.35%) had hCG levels greater than 4.0 multiples of the median (MoM) for gestational age. The Table depicts the rate of specific pregnancy outcomes occurring in women with an extremely elevated hCG. CONCLUSION: Pregnancies complicated by second-trimester hCG serum screening levels greater than 4.0 MoM have a markedly higher rate of stillbirth, pregnancy-related hypertension, abruption, and second-trimester fetal loss. These findings are in sharp contrast to prior studies of elevated hCG less than 4.0 MoM, which demonstrated little clinical value in predicting adverse pregnancy outcome.
111
SERUM FROM PREECLAMPTIC WOMEN INDUCES CASPASE-3 ACTIVATION IN FIRST-TRIMESTER TROPHOBLAST CELLS SHAWN STRASZEWSKI1, GIL MOR2, ROBERTO ROMERO3, DONNA NEALE4, 1 Yale University, Molecular, Cellular and Developmental Biology, New Haven, CT 2Yale University, Obstetrics and Gynecology, New Haven, CT 3National Institute of Child Health and Human Development, Detroit, MI 4Yale University, Obstetrics and Gynecology; Division of Maternal-Fetal Medicine, New Haven, CT OBJECTIVE: Preeclampsia is associated with insufficient trophoblast invasion, which may be a result of increased trophoblast apoptosis during the first trimester of pregnancy. Recently, we demonstrated that serum obtained from women with preeclampsia decreased trophoblast cell viability and rendered first-trimester trophoblast cells sensitive to Fas-mediated apoptosis. In order to further investigate these results, we evaluated whether preeclamptic serum activates effector caspase-3, an indicator of apoptosis, in first-trimester trophoblast cells. STUDY DESIGN: The H8 first-trimester trophoblast cell line was cultured in the presence of 10% serum obtained from either preeclamptic women or normotensive pregnant controls for 48 hours. Trophoblast cell viability was evaluated using the Cell Titer 96 assay. Activation of capase-3 was determined by Western blot analysis. RESULTS: Similar to our previous study, a significant decrease in trophoblast cell viability was observed following treatment with serum from preeclamptic patients in comparison to the normotensive controls. Only the pro-form of caspase-3 (p33) was detected in trophoblast cells treated with serum from normotensive patients. In contrast, the active forms of caspase-3 (17/19 kDa) were observed in trophoblast cells treated with serum obtained from preeclamptic patients. CONCLUSION: The findings described in this study suggest that a link exists between factor(s) in maternal serum and the induction of trophoblast apoptosis. The identification of the active forms of caspase-3 in trophoblast cells treated with serum obtained from preeclamptic patients, but not from the normotensive controls, supports the hypothesis that factors circulating in preeclamptic women reduce trophoblast cell viability through activation of the apoptotic cascade.
112
A PROTEOMIC APPROACH FOR THE IDENTIFICATION OF BIOLOGICAL MARKERS OF PREECLAMPSIA TINNAKORN CHAIWORAPONGSA1, RICARDO GOMEZ1, SAMUEL EDWIN1, GIL MOR2, TIMOTHY BARDER3, ROBERTO ROMERO1, 1Perinatology Research Branch, NICHD/ NIH/DHHS, Detroit, MI 2Yale University, Obstetrics and Gynecology, New Haven, CT 3Epogen, Inc, Darien, IL OBJECTIVE: Obstetrics is moving beyond genomics and into proteomics. The Plasma Proteome Project aims to comprehensively and quantitatively analyze plasma proteins to elucidate biomarkers for disease. The characterization of plasma proteins presents multiple challenges because of protein diversity and the large dynamic range of expression. The purpose of this study was to use a novel approach for proteomic analysis to identify differentially expressed proteins in the plasma of normal women and those with preeclampsia. STUDY DESIGN: Two-dimensional chromatography was employed to separate and analyze samples from normal women (n = 41) and those with preeclampsia (n = 41). The first dimension employed high-performance chromatofocusing to identify the isoelectric point, and the second dimension, high-resolution reverse-phase high-pressure liquid chromatography to provide protein hydrophobicity of each fraction. A 2D protein map was generated using software that displays the isoelectric point and the degree of hydrophobicity. The maps were used as pattern recognition tools to discern unique combinations of proteins expressed in plasma. RESULTS: The following two partial 2D protein maps represent the differential plasma proteome of normal pregnant women and those with preeclampsia. Dramatic differences between normal and preeclamptic plasma are observed. Interface with mass spectrometry using the liquid phase output allows protein analysis and identification. CONCLUSION: Plasma proteomic analysis using 2D chromatography revealed unique differential expression in preeclampsia. This novel approach can be used for the identification of biomarkers and patterns of biomarkers predictive of disease.
Complications and outcomes associated with extremely elevated hCG hCG >4.0 MoM, N = 166 Stillbirth Second-trimester loss Hypertensive disease Abruption IUGR Postpartum hemorrhage Preterm delivery
110
n n n n n n n
= = = = = = =
5; (3%) 11; (7%) 3; (19%) 10; (6%) 7; (4%) 8; (5%) 12; (7%)
SOLUBLE FMS-LIKE TYROSINE KINASE 1 LEVELS ARE ELEVATED IN PREECLAMPISA AND CORRELATE WITH DISEASE SEVERITY EUGENE CHANG1, CHRISTOPHER ROBINSON1, DONNA JOHNSON1, MICHAEL ARMSTRONG1, 1Medical University of South Carolina, Obstetrics and Gynecology, Charleston, SC OBJECTIVE: Soluble Flt-1 (sFlt1) is a receptor tyrosine kinase that binds and antagonizes the angiogenic/permeability factors, vascular endothelial (VEGF) and placental growth factor (PlGF). Excessive sFlt-1 causes hypertension and proteinuria in rats. The purpose of this study was to determine whether sFlt-1 levels correlate with the severity of preeclampsia. STUDY DESIGN: After IRB approval, blood samples from normotensive controls and from patients with preeclampsia were collected. Patients were classified with mild or severe disease based on ACOG criteria. Patients with chronic hypertension, collagen vascular disease, diabetes, and multiple gestations were excluded. Serum was stored at ÿ708C. sFlt-1 concentration (pg/mL) was measured by ELISA. Statistical analysis was performed with ANOVA and Student-t test. RESULTS: 32 controls and 47 patients with preeclampsia were enrolled. 23 patients had mild preeclampsia and 24 had severe. Mean gestational age (GA) (weeks) was similar for both controls and mildly affected patients (37.2 ± 2.0 vs 35.8 ± 2.7, P = NS) but GA was more premature for severely affected patients (31.6 ± 3.0 P < 0.03) When compared to controls, sFlt-1 levels were increased in preeclamptic patients (100.3 ± 142.0 vs 507.7 ± 441.5, P < 0.05). sFlt-1 was significantly higher in patients with severe preeclampsia compared to patients with mild disease (644.9 ± 457.9 vs 344.3 ± 367.9, P < 0.05). sFlt-1 was also significantly higher in patients with more than 1 gm of proteinuria than less than 1 gm (320.4 ± 408 vs 631.7 ± 406.7, P < 0.04) CONCLUSION: sFLt-1 levels are increased in patients with preeclampsia. sFlt-1 levels correlate with the severity of disease. s-Flt-1 may antagonize the normal effects of PlGF and VEGF.
normal (left) and PE (right)