Jacques Massé

Quantification of all fetal nucleated cells in maternal blood between the 18th and 22nd weeks of pregnancy using molecular cytogenetic techniques

Different types of nucleated fetal cells (trophoblasts, erythroblasts, lymphocytes, and granulocytes) have been recovered in maternal peripheral blood. In spite of many attempts to estimate the number of fetal cells in maternal circulation, there is still much controversy concerning this aspect. The numbers obtained vary widely, ranging from 1 nucleated cell per 104 to 1 per 109 nucleated maternal cells. The purpose of our project was to determine the absolute number of all different types of male fetal nucleated cells per unit volume of peripheral maternal blood. Peripheral blood samples were obtained from 12 normal pregnant women known to carry a male fetus between 18 and 22 weeks of pregnancy. Three milliliters (3 ml) of maternal blood has been processed without any enrichment procedures. Fluorescence in situ hybridization (FISH) and primed in situ labeling (PRINS) were performed, and fetal XY cells were identified (among maternal XX cells) and scored by fluorescent microscopy screening. The total number of male fetal nucleated cells per milliliter of maternal blood was consistent in each woman studied and varied from 2 to 6 cells per milliliter within the group of normal pregnancies. The number of fetal cells in maternal blood, at a given period, is reproducible and can therefore be assessed by cytogenetic methods. This confirms the possibility of developing a non‐invasive prenatal diagnosis test for aneuploidies. Furthermore, we demonstrate that it is possible to repeatedly identify an extremely small number of fetal cells among millions of maternal cells.

Early occurrence of metabolic syndrome after hypertension in pregnancy.

OBJECTIVE: The objective of the present study was to evaluate the cardiovascular risk profile and the prevalence of metabolic syndrome among women with a history of pregnancy-induced hypertension (PIH). METHODS: From a cohort of 3,799 nulliparous women prospectively recruited between 1989 and 1997, we performed an observational study on 168 case-control pairs 7.8 years after delivery. Participants were scheduled for a visit with a research nurse to evaluate their cardiovascular risk profile using a questionnaire, anthropometric measurements and blood specimen analysis. RESULTS: One hundred sixty-eight women with prior PIH (105 with gestational hypertension and 63 with preeclampsia) and 168 controls matched for age and year of index delivery were evaluated. The women with PIH (34.6 ± 4.4 years) were more obese and had higher systolic (115 mm Hg versus 108 mm Hg) and diastolic (75 mm Hg versus 70 mm Hg) blood pressures (P < .001) than the 168 controls (35.1 ± 4.5 years). They had lower high-density lipoprotein cholesterol level (1.30 mmol/L versus 1.42 mmol/L; P < .001), increased fasting blood glucose concentration (5.2 mmol/L versus 5.0 mmol/L; P = .002), and higher insulin levels (119 versus 91 pmol/L; P < .001). The prevalence of the metabolic syndrome was higher in the PIH group (unadjusted odds ratio = 4.9; 95% confidence interval 2.1–10.9) compared with controls, even after adjustment for confounders (adjusted odds ratio = 3.6; 95% confidence interval 1.4 –9.0). CONCLUSION: In white women in their mid-30s, the prevalence of the metabolic syndrome is 3- to 5-fold increased in those with a history of PIH in their first pregnancy. This emphasizes the importance of long-term follow-up assessment for cardiovascular risk factors in these women. LEVEL OF EVIDENCE: II-2

Prediction of pre‐eclampsia, low birthweight for gestation and prematurity by uterine artery blood flow velocity waveforms analysis in low risk nulliparous women

Objective To assess the performance of four previously reported Doppler abnormalities of uterine artery velocity waveforms (presence of a protodiastolic notch, peak systolic over protodiastolic velocities (A:C ratio) > 2.5, peak systolic over end diastolic velocities (A:B ratio) > 90th centile, resistance index (RI) ([A‐B]/A) ≥0.58) in predicting pre‐eclampsia, low birthweight and prematurity.

Trophoblastic remodeling in normal and preeclamptic pregnancies: implication of cytokines.

OBJECTIVE To summarize the recent knowledge on the implications of placenta and cytokines in normal and preeclamptic pregnancies. DATA SOURCES A literature search was conducted of applicable articles related to interactions between trophoblast and cytokines in generating preeclampsia. CONCLUSIONS The initiating event in preeclampsia has been postulated to be the reduced uteroplacental perfusion as a result of abnormal extravillous cytotrophoblast invasion and remodeling of the uterine spiral arteries. Focal ischemia and hypoxia, deportation of hypoxemic trophoblast cells and abnormal expression of various placental biologic molecules, particularly the cytokines, are thought to lead to widespread dysfunction of the maternal vascular endothelium resulting in overproduction of endothelin and thromboxane, enhanced vascular sensitivity to angiotensin II, and reduced secretion of vasodilators such as nitric oxide and prostacyclin. These alterations, in turn, cause hypertension, proteinuria and edema, and pathologies in many organ systems (kidney, lung, liver, brain).

A prospective study of several potential biologic markers for early prediction of the development of preeclampsia.

OBJECTIVE The purpose of this study was to prospectively evaluate the predictive performance of several potential biologic markers of preeclampsia used alone or in combination. STUDY DESIGN A prospective cohort of 1366 nulliparous women was followed up longitudinally on three occasions during pregnancy. The predictive performance of the tests, used either alone or in combination (stepwise multiple logistic regression), was assessed and compared with that of the mean arterial pressure. RESULTS Preeclampsia occurred in 109 of the pregnant women. At a specificity of 80% the sensitivity and the positive and negative predictive values for mean arterial pressure (at a threshold of 87 mm Hg) were 46.6%, 23.5%, and 92.0%, respectively, and the corresponding values for a multiple logistic model at 15 to 24 weeks that included some biologic markers, as well as the mean arterial pressure, were 57.1%, 26.9%, and 93.7%, respectively. CONCLUSION Preeclampsia can be predicted by a combination of simple biologic tests with a performance similar to second-trimester mean arterial pressure. However, this procedure is insufficient in terms of clinical usefulness.

Evaluation of the analytical performance of the Boehringer Mannheim Elecsys 2010 immunoanalyzer.

OBJECTIVES We evaluated the analytical performance of the Elecsys 2010 immunoanalyzer (Boehringer Mannheim Canada), which is based on a new detection technology, electrochemical luminescence. DESIGN AND METHODS We used six representative assays from the initial launch menu of the instrument: thyroid stimulating hormone (TSH), free thyroxine (FT4), troponin T, human chorionic gonadotropin (hCG), carcinoembryonic antigen (CEA), and prostate specific antigen (PSA). Within-run and between-day imprecision were evaluated using pools of human specimens at low, mid and high concentrations. Linearity was evaluated by diluting specimens with high analyte concentrations with specimens that had a low level of this analyte. Carry over and hook effect were investigated using specimens with high concentrations of hCG. Functional sensitivity was studied by running low TSH specimens over 21 daily runs, and by comparing the scatterplot of FT4 as a function of TSH. Over 100 specimens distributed across the analytical range were analyzed with two comparison methods: ES 300 (Boehringer Mannheim Canada) and AxSYM (Abbott Laboratories). RESULTS Within-run and between-day imprecisions were less than 4% and 10%, respectively, for most assays. All assays were linear over the whole analytical range. Carry over was minimal (< 0.0002%). A hook effect was present for hCG levels greater than 560,000 U/L. The functional sensitivity of the TSH assay was lower than 0.02 mlU/L. Correlation coefficients were all > 0.94. Small proportional errors were observed in comparison studies for the CEA and PSA assays. CONCLUSIONS The Elecsys 2010 system was shown to have an acceptable analytical performance for the rapid analysis of a wide variety of analytes. The hook effect observed with hCG assay would imply that the laboratory informs the clinicians of the possibility of falsely low values in trophoblastic diseases or that all specimens with values greater than 3000 U/L are reassayed after dilution.

Candidate biochemical markers for screening of pre-eclampsia in early pregnancy

Abstract Pre-eclampsia (PE) and other hypertensive disorders of pregnancy (HDP) are a leading cause of adverse outcomes. Their pathophysiology remains elusive, hampering the development of efficient prevention. The onset of HDP and PE and the severity of their clinical manifestations are heterogeneous. The advent of preventive measures, such as low-dose aspirin that targets high-risk women, emphasizes the need of better prediction. Until recently, only environmental information and maternal risk factors were considered, with equivocal predictive value. No validated screening procedures were available to identify at-risk women despite the emergence of Doppler ultrasonography parameters for the uterine artery (e.g., pulsatility index and bilateral notching) and pathophysiological biochemical markers (e.g., angiogenesis, inflammation, and endothelial dysfunction). Owing to its heterogeneity and lack of specific, sensitive markers among those studied so far (>200), PE is unlikely to be detected early by a single predictive parameter. Systematic reviews have concluded that no single test fulfilling World Health Organization criteria for biomarker selection can diagnose/predict a disease. However, by combining antenatal risk factors, clinical parameters, as well as biophysical and biochemical markers into multivariate algorithms, the risk of PE can be estimated with performance levels that could reach clinical utility. Performance characteristics of selected algorithms will be presented and discussed with respect to transferability to different geographic and healthcare environments.

SCREENING FOR DOWN SYNDROME DURING THE FIRST AND SECOND TRIMESTERS : IMPACT OF RISK ESTIMATION PARAMETERS

OBJECTIVES To evaluate impact of risk estimation parameters for screening for Down Syndrome during the first and second trimesters. METHODS We prospectively examined for their performance in the prenatal prediction of trisomy 21, alphafetoprotein (AFP), unconjugated estriol (uE3), total human chorionic gonadotropin (hCG), and its free subunits (free alpha-hCG, free beta-hCG) at both the first and second trimesters, and the impact of three sets of published risk estimation parameters. A total of 14,612 pregnancies were studied. All Down syndrome specimens (12 and 11 cases for first and second trimesters, respectively) and a sample of the unaffected pregnancies were analyzed. RESULTS The median multiple of median (MoM) for total hCG was lower in the first trimester (1.83 vs. 2.01 in the second trimester) but no loss in discriminative power was observed if the lower variability of the results in the first trimester is taken into account (interquartile range of 0.251 vs. 0.338). The choice of distribution parameters did not alter significantly the detection rates for the various combinations of markers (p > 0.05). False positive rates were affected significantly however and for the combination AFP-uE3-free beta-hCG they varied from 14.6% to 22.6% (p < 0.001). CONCLUSIONS Our results suggest that specific distribution parameters would be necessary to account for the lower variability of the markers in the first trimester and the peculiarity of the total hCG assay we used.

Peak systolic over protodiastolic ratio as an objective substitute for the uterine artery notch

Objective To measure the inter‐rater agreement for the identification of a uterine artery notch, as well as the association between an observed notch and the peak systolic over protodiastolic (A/C) ratio.

Pathophysiology and maternal biologic markers of preeclampsia.

Preeclampsia—increased blood pressure and proteinuria appearing after the twentieth week of pregnancy—is a major cause of materal and neonatal morbidity, leading to iatrogenic prematurity. Several lines of evidence suggest that the disorder is owing to diminished invasion of spiral arteries by trophoblastic cells, followed by reduced perfusion of the fetoplacental unit and oxidative stress. These alterations, in the presence of maternal predisposition, lead to endothelial dysfunction and occurrence of the clinical syndrome of preeclampsia (multisystemic lesions). Although the pathophysiology of preeclampsia is still unknown, progress has been made during the past 10 yr, and the early identification of at-risk women with the use of biochemical; ultrasonographic; and, more recently, genetic susceptibility markers has been the subject of intense research. In the present review, markers of maternal predisposition, placental implantation, oxidative stress, vasomotor regulation, and endothelial dysfunction are investigated as candidate markers in the early prediction of preeclampsia. Unfortunately, at the present time, no marker has been proven to have a clinically useful predictive performance in the general pregnant population, and, therefore, more research in that area is warranted.