A rise in postprandial serum triglycerides (PP-sTG) can potentiate inflammatory responses in vascular endothelial cells (ECs) and thus serves as an independent risk factor for predicting increased cardiovascular morbidity. We examined postprandial triglyceride-rich lipoproteins (PP-TGRLs) in subjects ranging from normal to hypertriglyceridemic for their capacity to alter EC acute inflammatory responses. Cultured human aortic ECs (HAECs) were conditioned with PP-TGRLs isolated from human serum at the peak after a moderately high-fat meal. VLDL particle size increased postprandially and varied directly with the subject's PP-sTG level and waist circumference. PP-TGRL particles bound to HAECs and were internalized via LDL receptor-mediated endocytosis. PP-TGRL alone did not induce an inflammatory response over the range of individuals studied. However, combined with low-dose TNF-α stimulation (0.3 ng/ml), it elicited a net 10-15% increase above cytokine alone in the membrane expression of VCAM-1, ICAM-1, and E-selectin, which was not observed with fasting TGRLs. In contrast to upregulation of ICAM-1 and E-selectin, VCAM-1 transcription and expression varied in direct proportion with individual PP-sTG and waist circumference. The extent of monocyte arrest on inflamed HAECs under shear stress also correlated closely with VCAM-1 expression induced by conditioning with PP-TGRL and TNF-α stimulation. This ex vivo approach provides a quantitative means to assess an individual's inflammatory potential, revealing a greater propensity for endothelial inflammation in hypertriglyceridemic individuals with abdominal obesity.

BackgroundMaintaining proper adhesion between neighboring cells depends on the ability of cells to mechanically respond to tension at cell-cell junctions through the actin cytoskeleton. Thus, identifying the molecules involved in responding to cell tension would provide insight into the maintenance, regulation, and breakdown of cell-cell junctions during various biological processes. Vinculin, an actin-binding protein that associates with the cadherin complex, is recruited to cell-cell contacts under increased tension in a myosin II-dependent manner. However, the precise role of vinculin at force-bearing cell-cell junctions and how myosin II activity alters the recruitment of vinculin at quiescent cell-cell contacts have not been demonstrated.ResultsWe generated vinculin knockdown cells using shRNA specific to vinculin and MDCK epithelial cells. These vinculin-deficient MDCK cells form smaller cell clusters in a suspension than wild-type cells. In wound healing assays, GFP-vinculin accumulated at cell-cell junctions along the wound edge while vinculin-deficient cells displayed a slower wound closure rate compared to vinculin-expressing cells. In the presence of blebbistatin (myosin II inhibitor), vinculin localization at quiescent cell-cell contacts was unaffected while in the presence of jasplakinolide (F-actin stabilizer), vinculin recruitment increased in mature MDCK cell monolayers.ConclusionThese results demonstrate that vinculin plays an active role at adherens junctions under increased tension at cell-cell contacts where vinculin recruitment occurs in a myosin II activity-dependent manner, whereas vinculin recruitment to the quiescent cell-cell junctions depends on F-actin stabilization.