Abstract 547: Excess Glucose Dowregulates Factors for Invasion and Induces an Anti-angiogenic Milieu in First Trimester Cytotrophoblast Cell

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Abstract

Objective: Diabetes mellitus is a known risk factor for the development of preeclampsia (preE). First trimester cytotrophoblastic (CTB) cells invade the decidua using the serine protease plasmin. The enzymatic conversion from the inactive plasminogen to the active form plasmin, is performed by urokinase plasminogen activator (uPA), which is regulated by plasminogen activator inhibitor 1 (PAI-1) in CTBs. Disruption of placental development, setting the stage for preE development is believed, in part, to be caused by a dysfunction of the invasive nature of CTBs. The aim of this study is to determine whether excess glucose induces anti-invasive and anti-angiogenic effects in CTBs in vitro.

Methods: Human CTB cells (Sw. 71) were derived from first trimester placenta. The CTB cells were treated with 45, 135, 225, 495 or 945 mg/dL glucose for 72h. Thereafter, cell lysates were utilized to measure uPA and PAI-1 protein expression and phosphorylation of p38 MAPK by western blot. The mRNA expression of uPA and PAI-1 in glucose-induced CTB cell lysates was measure by qPCR. Levels of angiogenic and anti-angiogenic factors (sFLT-1, sENG, VEGF165, PlGF) and the level of inflammatory cytokine IL-6 were measured in the culture media by the commercially available kits. Statistical comparisons were performed using analysis of variance with Dunnett's post hoc tests.

Results: Both uPA and PAI-1 protein and mRNA expression were downregulated (p<0.05) in CTB cells treated with >135 mg/dL glucose as compared to those in basal (45 mg/dL). The anti-angiogenic factors (sENG and sFLT-1) were upregulated, while the angiogenic factors (VEGF165 and PlGF) were downregulated in the presence of >135 mg/dL glucose. The level of IL-6 was higher (p<0.05) in the presence of glucose >135 mg/dL compared to basal (45 mg/dL) conditions. The p38 MAPK phosphorylation was upregulated (p<0.05) in excess glucose-treated CTBs.

Conclusions: Exposure of CTB cells to glucose at levels associated with diabetes causes a disruption in the invasive profile of CTB cells by decreasing the synthesis of uPA and PAI-1 (both protein and mRNA expression); by downregulating expression of VEGF165 and PlGF; and by upregulating expression of sENG, sFLT-1, and IL-6. Moreover, glucose induced a stress signaling in CTBs.

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