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Human Milk Oligosaccharides in Cord Blood Are Altered in Gestational Diabetes and Stimulate Feto-Placental Angiogenesis In Vitro.

ABSTRACT: (1) Background: Human milk oligosaccharides (HMOs) are present in maternal serum during pregnancy and their composition is altered in gestational diabetes (GDM). HMOs are also in fetal cord blood and in contact with the feto-placental endothelium, potentially affecting its functions, such as angiogenesis. We hypothesized that cord blood HMOs are changed in GDM and contribute to increased feto-placental angiogenesis, hallmark of GDM. (2) Methods: Using HPLC, we quantified HMOs in cord blood of women with normal glucose tolerance (NGT, n = 25) or GDM (n = 26). We investigated in vitro angiogenesis using primary feto-placental endothelial cells (fpECs) from term placentas after healthy pregnancy (n = 10), in presence or absence of HMOs (100 µg/mL) isolated from human milk, 3'-sialyllactose (3'SL, 30 µg/mL) and lactose (glycan control) and determined network formation (Matrigel assay), proliferation (MTT assays), actin organization (F-actin staining), tube formation (fibrin tube formation assay) and sprouting (spheroid sprouting assay). (3) Results: 3'SL was higher in GDM cord blood. HMOs increased network formation, HMOs and 3'SL increased proliferation and F-actin staining. In fibrin assays, HMOs and 3'SL increased total tube length by 24% and 25% (p < 0.05), in spheroid assays, by 32% (p < 0.05) and 21% (p = 0.056), respectively. Lactose had no effect. (4) Conclusions: Our study suggests a novel role of HMOs in feto-placental angiogenesis and indicates a contribution of HMO composition to altered feto-placental vascularization in GDM.

SUBMITTER: Hoch D

PROVIDER: S-EPMC8705424 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:ObjectiveGestational diabetes mellitus (GDM) is a serious complication in pregnancy. Despite controlling the plasma glucose levels with dietary intervention (GDM-D) or insulin therapy (GDM-I), children born of diabetic mothers suffer more long-term complications from childhood to early adulthood. Placental circulation and nutrient exchange play a vital role in fetal development. Additionally, placental endothelial function is an indicator of vascular health, and plays an important role in maintaining placental circulation for nutrient exchange. This study was conducted to assess changes in fetal endothelial dysfunction in GDM under different interventions during pregnancy.MethodsThe primary human umbilical vein endothelial cells (HUVECs) were obtained from normal pregnant women (n = 11), GDM-D (n = 14), and GDM-I (n = 12) patients. LC-MS/MS was used to identify differentially expressed proteins in primary HUVECs among the three groups, after which Bioinformatics analysis was performed. Glucose uptake, ATP level, apoptosis, and differentially expressed proteins were assessed to investigate changes in energy metabolism.ResultsA total of 8174 quantifiable proteins were detected, and 142 differentially expressed proteins were identified after comparing patients with GDM-D/GDM-I and healthy controls. Of the 142, 64 proteins were upregulated while 77 were downregulated. Bioinformatics analysis revealed that the differentially expressed proteins were involved in multiple biological processes and signaling pathways related to cellular processes, biological regulation, and metabolic processes. According to the results from KEGG analysis, there were changes in the PI3K/AKT signaling pathway after comparing the three groups. In addition, there was a decrease in glucose uptake in the GDM-I (P < 0.01) group. In GDM-I, there was a significant decrease in the levels of glucose transporter 1 (GLUT1) and glucose transporter 3 (GLUT3). Moreover, glucose uptake was significantly decreased in GDM-I, although in GDM-D, there was only a decrease in the levels of GLUT1. ATP levels decreased in GDM-I (P < 0.05) and apoptosis occurred in both the GDM-D and GDM-I groups. Compared to the normal controls, the levels of phosphate AKT and phosphate AMPK over total AKT and AMPK were reduced in the GDM-I group.ConclusionIn summary, endothelial dysfunction occurred in pregnancies with GDM even though the plasma glucose levels were controlled, and this dysfunction might be related to the degree of glucose tolerance. The energy dysfunction might be related to the regulation of the AKT/AMPK/mTOR signaling pathway.

Project description:Aims/hypothesisReduced occupancy of junctional occludin is a feature of human placental vessels in the diabetic milieu. However, the functional consequence of this and whether this loss is due to differential expression of occludin splice variants is not known. Our study aimed to investigate the effects of gestational diabetes mellitus (GDM), and its treatment, on endothelial junctional integrity, gene and protein expression of occludin splice variants, and potential regulation of expression by microRNAs (miRNAs).MethodsTerm placentas were obtained from normal pregnancies (n = 21), and pregnancies complicated by GDM where glucose levels were controlled by diet (n = 11) or metformin (n = 6). Gene and microRNA (miRNA) expression were determined by quantitative real-time PCR; protein expression by immunoblotting; endothelial junctional occupancy by fluorescence microscopy and systematic sampling; and paracellular leakage by perfusion of placental microvascular beds with 76 Mr dextran. Transfection studies of miRNAs that target OCLN were performed in HUVECs, and the trans-endothelial electrical resistance and tracer permeability of the HUVECs were measured.ResultsAll three predicted OCLN gene splice variants and two occludin protein isoforms were found in human placental samples. In placental samples from diet-controlled GDM (d-GDM) pregnancies we found a lower percentage of conduit vessels showing occludin immunoreactivity (12%, p < 0.01), decreased levels of the fully functional occludin isoform-A protein (29%), and differential gene expression of OCLN variant 2 (33% decrease), variant 3 (3.3-fold increase). These changes were not seen in samples from the group with metformin-controlled GDM. In d-GDM placentas, increased numbers of conduit microvessels demonstrated extravasation of 76 Mr dextran (2.0-fold). In d-GDM expression of one of the five potential miRNAs targeting OCLN, miR-181a-5p, expression was 2.1-fold that in normal pregnancies. Experimental overexpression of miR-181a-5p in HUVECs from normal pregnancies resulted in a highly significant downregulation of OCLN variant 1 (69%) and variant 2 (46%) gene expression, with decreased trans-endothelial resistance (78%) and increase in tracer permeability (1.3-fold).Conclusions/interpretationDownregulation of expression of OCLN variant 2 and the fully functional occludin isoform-A protein are a feature of placentas in d-GDM pregnancies. These may be behind the loss of junctional occludin and the increased extravasation of exogenous dextran observed. miR-181a-5p was in part responsible for the downregulation of occludin in placentas from d-GDM pregnancies. Induced overexpression of miR-181a-5p compromised the integrity of the endothelial barrier. Our data suggest that, despite good glucose control, the adoption of lifestyle changes alone during a GDM pregnancy may not be enough to prevent an alteration in the expression of occludin and the subsequent functional consequences in placentas and impaired vascular barrier function in offspring. Graphical abstract.

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Human Milk Oligosaccharides in Cord Blood Are Altered in Gestational Diabetes and Stimulate Feto-Placental Angiogenesis In Vitro.

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