Myo‐inositol in Down syndrome amniotic fluid. A case‐control study

Down syndrome (DS) is caused by the trisomy of chromosome 21. In DS, gene products of the extra chromosome 21 may cause abnormal levels of certain metabolites directly or by affecting gene expression of other chromosomes. Myo-inositol (MI) is one of these; it is a polyalcohol deriving from glucose, which is synthesized by a few organs in the body, such as the testes, kidneys, and brain. MI concentrations are significantly increased in DS cerebrospinal fluid, and its levels are directly correlated with the plasmatic ones. Thus, it is plausible to hypothesize that DS could be associated with MI accumulation not only in cerebrospinal fluid but also in other fluids, in which it might be detected very early in life. The aim of our study was to evaluate midtrimester amniotic fluid (AF) in DS pregnancies, compared with normal pregnancies. A retrospective study was carried out on 22 AF samples of DS pregnancies stored at 80 °C, collected over the last 6 years, after amniocentesis, performed between 15 and 18weeks gestation at our Prenatal Diagnosis Center. A group of 25 samples of normal pregnancy AF, matched for maternal age, body mass index (BMI) and gestational age, were used as a control group. The protocol was consistent with the principles of the Declaration of Helsinki, and all the participants gave their written informed consent for the utilization of 1mL of their AF for experimental purposes. MI quantification was performed by Chelab Pharma Division using gas chromatography–mass spectrometry analysis after extraction with organic solvents and derivation. Injection (1.0μL) was performed in a split-less mode at 270 °C and a capillary column Agilent 122-5532 DB-5 ms (0.25mm×30m×0.25 ×μm) was used. The total run-time was 15min: oven at 70 °C from 0 to 1min; 20 °C/min to 150 °C; 10 °C/min to 240 °C; 4 min at 320 °C post run. The flow rate was fixed at 1.2mL/min, and results were analyzed by an MS 5973 Network Series detector in sim mode. Statistical analysis was carried out with an SPSS statistical package version 17 (SPSS, Chicago, IL). Data are expressed as medians and interquartile range (IQR). The Mann–Whitney test was used to compare AF MI values between groups, and Spearman’s test was used to correlate normal pregnancy AF values with maternal age, BMI, and parity. The general characteristics of the two groups are reported in Table 1. There were no statistical differences between them in BMI, gestational age, parity, and maternal age. The indications for amniocentesis were maternal age in 17 and positive biochemical genetic screening test in four. In all DS pregnancies, an induced abortion was requested and then performed. A significant difference (p< 0.001) was found inMI concentrations between groups (Figure 1): DS 135. 2μmol/L (IQR 119.2–147.8); control group 84. 0μmol/L (IQR 63.1–112.8) (Figure 1). In the control group, there was no correlation between AF MI levels andmaternal age (r=0.15, p=0.46); BMI (r=0.11, p=0.57); parity (r=0.2, p=0.32); gestational age (r=0.21, p=0.3) or neonatal weight (r= 0.003, p=0.9). Inositol is found in the cellular membrane as inositol phosphoglycans (IPGs). The placenta plays a fundamental role in the complex metabolism of inositol. In fact, IPGs are released into microvilli caveolae by the action of the enzyme glycosylphosphatidylinositol phospholipase D (GPI-PLD) on their lipid precursors, the membrane-associated glycosylphosphatidylinositol (GPI). Placental GPI-PLD, necessary for the production of IPG, is not produced by the placenta but is taken up from maternal plasma. Placental microvilli have therefore a potential source for placenta-derived IPG molecules which can be released into the maternal blood stream, where they could act as systemic (paracrine) factors, in addition to their role as autocrine mediators of a large number of growth factors and hormones. The lack of correlation between AF MI concentrations and maternal parameters confirms a possible fetal origin of this substance. An increased production of MI AF in subjects affected by DS is in accordance with an accumulation of MI in other DS tissues. In particular, fibroblasts of non-DS patients with