Nuovi approcci molecolari nello studio della fisiopatologia del diabete gestazionale

Abstract research study 1 Cross-talk between foetal membranes and visceral adipose tissue involves HMGB1-RAGE and VIP-VPAC2 pathways in human gestational diabetes mellitus Introduction: Gestational diabetes mellitus (GDM) is defined as glucose intolerance that is first diagnosed during pregnancy. Foetal membranes (FMs) and maternal visceral adipose tissue (VAT) secrete various molecules that are relevant players in the pathogenesis of GDM. Aim: This pilot study aimed to comparatively evaluate the expression of high mobility group box 1 protein (HMGB1) and its receptor for advanced glycation end products (RAGE), and vasoactive intestinal peptide (VIP) and its receptors (VPAC1, VPAC2) in FMs and VAT in GDM and in healthy pregnant women. Patients and Methods: FMs, omental VAT explants, and serum samples were obtained from twelve patients with GDM and twelve pregnant women with normal glucose tolerance (NGT) at delivery. The expression of HMGB1, RAGE and VIP, VPAC1 and VPAC2 was detected by Western Blotting in explants; circulating levels and in vitro release of HMGB1 and VIP were measured by ELISA tests. Results: HMGB1 tissue expression was higher in FMs obtained from GDM patients (p=0.02) than in FMs from NGT women. VPAC2 (p=0.03) and RAGE (p=0.03) tissue expressions were significantly increased in VAT from GDM patients compared to NGT. Only FMs of NGT released detectable levels of HMGB1, which was not observed in samples obtained from GDM. VAT of GDM released lower levels of VIP (p=0.05) than NGT samples. Conclusions: This study suggests that a fine tuned regulation exists between FMs and VAT throughout pregnancy to maintain immune metabolic homeostasis. In GDM a balance between pro-inflammatory and anti-inflammatory mediators has been observed. Further studies are needed to establish their exact role on foetal and maternal outcomes in GDM. Abstract research study 2 MicroRNA expression profile in circulating exosomes and plasma of patients with GDM and healthy pregnant women Introduction: MicroRNAs are small non-coding RNAs, playing critical roles in modulating gene expression. The deregulation of microRNAs has been observed in GDM, highlighting their crucial involvement both in the pathogenic mechanisms of this condition and in the development of its complications. Circulating microRNAs can be packaged into exosomes, and exosome signalling has emerged as a novel mechanism of cell-to-cell communication. Through exosomes, microRNAs are delivered in distant target cells and are able to affect gene expression. Aim: The aim of this study was to explore microRNA expression in circulating exosomes and in plasma obtained from patients with GDM and healthy control subjects in the third trimester of gestation, to potentially elucidate some relevant aspects of GDM pathophysiology and individuate novel potential candidate biomarkers for GDM. Patients and Methods: A profiling cohort of plasma samples collected from GDM (n=3, age: 34.7 ± 4.9 years; BMI 27.0 ± 3.7 Kg/m2) and NGT women (n=3, age: 34.3 ± 3.1 years; BMI 26.4 ± 1.1 Kg/m2) was recruited. In addition, a profiling cohort of healthy non-pregnant age- and BMI-matched women (NP, n=5) was used as negative control. The microRNA patterns of expression in exosomes and plasma have been assessed with the innovative technology NanoString nCounter microRNA expression (NanoString Technologies inc., Seattle, WA, USA). Target gene identification and bioinformatics analysis of the differentially expressed microRNAs have been performed with Ingenuity Pathway Analysis (IPA, QIAGEN Redwood City, USA). Results: A specific set of microRNAs resulted to be differentially expressed in exosomes and plasma from GDM patients compared to NGT. Specifically, five exosomal microRNAs were significantly upregulated, while 23 were downregulated in GDM compared to NGT. As for plasma, 4 microRNA were upregulated, while 9 were downregulated in GDM compared to NGT. In addition, two microRNAs, miR-196a-5p and miR-652, resulted to be significantly downregulated in GDM compared both to NGT and NP in exosomes and plasma, respectively, suggesting that their deregulation might hallmark GDM pregnancy. In bioinformatics analysis the major predicted target genes and biological processes of the deregulated microRNAs were associated with insulin resistance, abnormal glucose and lipid metabolism, consistently linked to GDM pathophysiology. Conclusions: GDM might markedly alter microRNA profile in exosomes and plasma, conceivably mirroring the metabolic alterations described in GDM pregnancy. In light of this, exploring circulating microRNA expression might help unravel the molecular events leading to the metabolic alterations observed in GDM.