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

Melanocortin MC₄ receptor expression sites and local function

The melanocortin MC(4) receptor plays an important role in energy metabolism, but also affects blood pressure, heart rate and erectile function. Localization of the receptors that fulfill these distinct roles is only partially known. Mapping of the melanocortin MC(4) receptor has been stymied by the absence of a functional antibody. Several groups have examined mRNA expression of the melanocortin MC(4) receptor in the rodent brain and transgenic approaches have also been utilized to visualize melanocortin MC(4) receptor expression sites within the brain. Ligand expression and binding studies have provided additional information on the areas of the brain where this elusive receptor is functionally expressed. Finally, microinjection of melanocortin MC(4) receptor ligands in specific nuclei has further served to elucidate the function of melanocortin MC(4) receptors in these nuclei. These combined approaches have helped link the anatomy and function of this receptor, such as the role of paraventricular hypothalamic nucleus melanocortin MC(4) receptor in the regulation of food intake. Intriguingly, however, numerous expression-sites have been identified that have not been linked to a specific receptor function such as those along the optic tract and olfactory tubercle. Further research is needed to clarify the function of the melanocortin MC(4) receptor at these site

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 (fβ-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 fβ-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 fβ-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

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: <p10) showed an adjusted odds ratio (OR) of 5.29 (95%CI 1.10–25.5) for LO-PE. In the Utrecht population, low relaxin levels (<p10) demonstrated adjusted ORs of 1.45 (95%CI 0.54–3.90) and 2.03 (95%CI 1.06–3.88) for EO-PE and LO-PE respectively, the latter increasing to an adjusted OR of 3.18 (95%CI 1.41–7.20) when newborn weight was < 10%. Serum relaxin concentrations slightly improved the detection rate of a previously derived prediction model for LO-PE from 42.5% to 45.1% at a fixed 10% false-positive rate. Conclusion: Relaxin shows little improvement in the performance of first trimester prediction models, which does not support its clinical implementation as a biomarker. Although this study was only correlational, the results point to a possible pathophysiologic role f

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 (fβ-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 fβ-hCG. PAPP-A and fβ-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 fβ-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

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