Circulating pregnancy hormone relaxin as a first trimester biomarker for preeclampsia.

OBJECTIVE: Preeclampsia, a multi-system hypertensive disorder, is associated with perturbations in the maternal cardiovascular system during early pregnancy. The corpus luteal hormone relaxin, a potent vasodilator, may contribute to physiological circulatory changes especially in early gestation when circulating levels are highest. This study investigated whether first trimester circulating relaxin may be a suitable biomarker for the early prediction of preeclampsia. METHODS: Relaxin was initially measured in first-trimester samples of women who developed late-onset preeclamptic (LO-PE; delivery ≥ 34 weeks; n = 33) and uncomplicated pregnancies (n = 25) in Pittsburgh, USA. Subsequently, to expand the group numbers, relaxin was measured in women who developed LO-PE (n = 95), early-onset preeclamptic (EO-PE; delivery < 34 weeks; n = 57), and uncomplicated pregnancies (n = 469) in Utrecht, the Netherlands. RESULTS: In the Pittsburgh subjects, low relaxin levels (lowest centile:

Subcellular localization of MC4R with ADCY3 at neuronal primary cilia underlies a common pathway for genetic predisposition to obesity.

Most monogenic cases of obesity in humans have been linked to mutations in genes encoding members of the leptin-melanocortin pathway. Specifically, mutations in MC4R, the melanocortin-4 receptor gene, account for 3-5% of all severe obesity cases in humans1-3. Recently, ADCY3 (adenylyl cyclase 3) gene mutations have been implicated in obesity4,5. ADCY3 localizes to the primary cilia of neurons 6 , organelles that function as hubs for select signaling pathways. Mutations that disrupt the functions of primary cilia cause ciliopathies, rare recessive pleiotropic diseases in which obesity is a cardinal manifestation 7 . We demonstrate that MC4R colocalizes with ADCY3 at the primary cilia of a subset of hypothalamic neurons, that obesity-associated MC4R mutations impair ciliary localization and that inhibition of adenylyl cyclase signaling at the primary cilia of these neurons increases body weight. These data suggest that impaired signaling from the primary cilia of MC4R neurons is a common pathway underlying genetic causes of obesity in humans.

External validation of prognostic models to predict risk of gestational diabetes mellitus in one Dutch cohort: prospective multicentre cohort study.

OBJECTIVE:  To perform an external validation and direct comparison of published prognostic models for early prediction of the risk of gestational diabetes mellitus, including predictors applicable in the first trimester of pregnancy. DESIGN:  External validation of all published prognostic models in large scale, prospective, multicentre cohort study. SETTING:  31 independent midwifery practices and six hospitals in the Netherlands. PARTICIPANTS:  Women recruited in their first trimester (<14 weeks) of pregnancy between December 2012 and January 2014, at their initial prenatal visit. Women with pre-existing diabetes mellitus of any type were excluded. MAIN OUTCOME MEASURES:  Discrimination of the prognostic models was assessed by the C statistic, and calibration assessed by calibration plots. RESULTS:  3723 women were included for analysis, of whom 181 (4.9%) developed gestational diabetes mellitus in pregnancy. 12 prognostic models for the disorder could be validated in the cohort. C statistics ranged from 0.67 to 0.78. Calibration plots showed that eight of the 12 models were well calibrated. The four models with the highest C statistics included almost all of the following predictors: maternal age, maternal body mass index, history of gestational diabetes mellitus, ethnicity, and family history of diabetes. Prognostic models had a similar performance in a subgroup of nulliparous women only. Decision curve analysis showed that the use of these four models always had a positive net benefit. CONCLUSIONS:  In this external validation study, most of the published prognostic models for gestational diabetes mellitus show acceptable discrimination and calibration. The four models with the highest discriminative abilities in this study cohort, which also perform well in a subgroup of nulliparous women, are easy models to apply in clinical practice and therefore deserve further evaluation regarding their clinical impact.

Predictive performance of a seven-plex antibody array in prenatal screening for Down Syndrome.

We evaluated the use of multiplex antibody array methodology for simultaneous measurement of serum protein markers for first trimester screening of Down Syndrome (DS) and other pregnancy outcomes such as preeclampsia. For this purpose, we constructed an antibody array for indirect ("sandwich") measurement of seven serum proteins: pregnancy-associated plasma protein-A (PAPP-A), free beta subunit of human chorionic gonadotropin (fß-hCG), alpha-fetoprotein (AFP), angiopoietin-like 3 (ANGPTL3), epidermal growth factor (EGF), insulin-like growth factor 2 (IGFII), and superoxide dismutase 1 (SOD1). This array was tested using 170 DS cases and 510 matched controls drawn during the 8th-13th weeks of pregnancy. Data were used for prediction modelling and compared to previously obtained AutoDELFIA immunoassay data for PAPP-A and fß-hCG. PAPP-A and fß-hCG serum concentrations obtained using antibody arrays were highly correlated with AutoDELFIA data. Moreover, DS prediction modeling using (log-MoMmed) antibody array and AutoDELFIA data gave comparable results. Of the other markers, AFP and IGFII showed significant changes in concentration, although adding these markers to a prediction model based on prior risk, PAPP-A and fß-hCG did not improve the predictive performance. We conclude that implementation of antibody arrays in a prenatal screening setting is feasible but will require additional first trimester screening markers.

Comparison of different blood collection, sample matrix, and immunoassay methods in a prenatal screening setting.

We compared how measurements of pregnancy-associated plasma protein A (PAPP-A) and the free beta subunit of human chorionic gonadotropin (fß-hCG) in maternal blood are influenced by different methods for blood collection, sample matrix, and immunoassay platform. Serum and dried blood spots (DBS) were obtained by venipuncture and by finger prick of 19 pregnant women. PAPP-A and fß-hCG from serum and from DBS were measured by conventional indirect immunoassay on an AutoDELFIA platform and by antibody microarray. We compared methods based on the recoveries for both markers as well as marker levels correlations across samples. All method comparisons showed high correlations for both marker concentrations. Recovery levels of PAPP-A from DBS were 30% lower, while those of fß-hCG from DBS were 50% higher compared to conventional venipuncture serum. The recoveries were not affected by blood collection or immunoassay method. The high correlation coefficients for both markers indicate that DBS from finger prick can be used reliably in a prenatal screening setting, as a less costly and minimally invasive alternative for venipuncture serum, with great logistical advantages. Additionally, the use of antibody arrays will allow for extending the number of first trimester screening markers on maternal and fetal health.

Positive predictive values for detection of trisomies 21, 18 and 13 and termination of pregnancy rates after referral for advanced maternal age, first trimester combined test or ultrasound abnormalities in a national screening programme (2007-2009).

OBJECTIVE: The objective of this article is to analyse the positive predictive value (PPV) of trisomies 21, 18 and 13 after referral for advanced maternal age (AMA), first trimester combined test or ultrasound findings to suggest improvements for clinical practice. METHODS: Data (48 457 combined tests, 134 000 fetal anomaly scans and 24 379 invasive prenatal tests) were combined to calculate PPV and termination of pregnancy rates. RESULTS: For referral for AMA, the PPV for T21 was 1.0% and 1.8% for amniocentesis and chorionic villus biopsy, respectively; for the combined test at a maternal age ≥36 years, these percentages were 4.9% and 12.5%, respectively and for maternal age <36 years, 4.4% and 8.1%, respectively. For ultrasound findings, the PPV was 5.3% and 14.8%, respectively. Termination of pregnancy rate upon trisomy 21 diagnosis was >90% unless detected after referral for ultrasound findings (71.5-85.9%). About 50% of pregnant women with a high combined test risk chose not to have invasive testing. CONCLUSIONS: Advanced MA is still a large contributor to invasive testing but should be abandoned (low PPV, high fetal loss rate) and be replaced by reimbursable combined test screening for all women. Patient information on second trimester ultrasound screening should indicate that abnormal ultrasound findings are associated with high trisomy rate.

Identification of interleukin-1 beta, but no other inflammatory proteins, as an early onset pre-eclampsia biomarker in first trimester serum by bead-based multiplexed immunoassays.

OBJECTIVE: This study aimed to determine the predictive value of growth factors, cardiovascular, and immunological markers for first trimester identification of early onset pre-eclampsia (PE). METHODS: In a retrospective case-control study, maternal serum samples of 35 early onset PE cases and 35 controls were analysed by multiplexed immunoassays, to determine serum concentrations of 41 proteins whose functionality can be associated with PE pathogenesis. All levels were converted into multiples of the gestation-specific normal median. For prediction modelling, proteins that were found to be significant were combined with previously obtained values of three established PE markers, that is, placental growth factor, placental protein 13, and pregnancy-associated plasma protein A. Prediction modelling was used to determine predicted detection rates for 5% and 10% false-positive rates. RESULTS: Three of the proteins examined in this study, interleukin-1 beta (IL-1ß), fibrinogen, and carcinoembryonic antigen, showed significantly different serum levels at p < 0.05. In prediction modelling, only IL-1ß added predictive value to the three previously established biomarkers, by increasing detection from 38.2% to 44.1% at a 5% false-positive rate. CONCLUSIONS: This study indicates that IL-1ß has potential to improve first trimester prediction of pre-eclampsia. Studies on larger cohorts will be needed to validate these findings.

Arginine vasopressin immunoreactivity is decreased in the hypothalamic suprachiasmatic nucleus of subjects with suprasellar tumors.

Suprasellar tumors with compression of the optic chiasm are associated with an impaired sleep-wake rhythm. We hypothesized that this reflects a disorder of the biological clock of the human brain, the suprachiasmatic nucleus (SCN), which is located just above the optic chiasm. In order to test this hypothesis, we investigated the expression of two key neuropeptides of the SCN, that is, arginine vasopressin (AVP) and vasoactive intestinal peptide (VIP), as assessed by quantitative immunocytochemistry in post-mortem hypothalamic tissue of patients with a suprasellar tumor inducing permanent visual field defects. Post-mortem hypothalamic tissue of 5 patients with a suprasellar tumor inducing permanent visual field defects (acromegaly n = 2, nonfunctioning macro-adenoma n = 1, macroprolactinoma n = 1, infundibular metastasis of a colorectal adenocarcinoma n = 1) and 15 age- and gender-matched controls was obtained from the Netherlands Brain Bank. Total AVP immunoreactivity in the SCN was lower in patients with a suprasellar tumor than in controls (P = 0.03). By contrast, total VIP immunoreactivity was not different between patients and controls (P = 0.44). Suprasellar tumors leading to permanent visual field defects are associated with reduced AVP, but not VIP immunoreactivity, in the SCN. These findings raise the possibility that selective impairment of the SCN contributes to sleep-wake disturbances in these patients.

Melanocortin 4 receptor distribution in the human hypothalamus.

OBJECTIVE: The melanocortin 4 receptor (MC4R) is an essential regulator of energy homeostasis and metabolism, and MC4R mutations represent the most prevalent monogenetic cause of obesity in humans known to date. Hypothalamic MC4Rs in rodents are well characterized in neuroanatomical and functional terms, but their expression pattern in the human hypothalamus is unknown. DESIGN AND METHODS: To determine the topographic distribution and identity of cells expressing MC4R mRNA in the human hypothalamus, locked nucleic acid in situ hybridization was performed on nine human postmortem hypothalami. In addition, co-expression of MC4R with glial fibrillary acidic protein (GFAP), vasopressin/oxytocin (AVP/OXT), corticotropin-releasing hormone (CRH), neuropeptide Y (NPY), agouti-related protein (AgRP), and α-melanocyte stimulating hormone (α-MSH) was examined. RESULTS: Most intense MC4R mRNA expression was present in the paraventricular nucleus (PVN), the supraoptic nucleus (SON), and the nucleus basalis of Meynert. Most MC4R-positive cells in the SON also expressed AVP/OXT. Co-expression with AVP/OXT in the PVN was less abundant. We did not observe co-expression of MC4R mRNA and GFAP, CRH, NPY, AgRP, or α-MSH. However, fiber-like staining of NPY, AgRP, and α-MSH was found adjacent to MC4R-positive cells in the PVN. CONCLUSION: Expression of MC4R mRNA in the human hypothalamus is widespread and in close approximation to endogenous MC4R binding partners AgRP and α-MSH.