Biopathological Effects Of Dicarbony1 Compounds On The Human Vascular Endothelial Cells

Accumulation of dicarbonyl compounds in circulation has been demonstrated in the patients with chronic renal failure and diabetes. Some of these compounds, including 3-deoxyglucosone(3-DG) and methylglyoxal (MOO), are important intermediates in Maillard reaction, in which advanced glycation end products(AGEs) are produced ultimately. Cytotoxicity of dicarbonyl compounds has been found in some kinds of cells, such as macrophage, neuron and pancreatic cell. This study was conducted to elucidate the effect of 3-DG and MGO on viability and expression of adhesion molecules of human vascular endothelial cells (VECs). Methods: VECs were isolated from human umbilical vein and cultured with 3-DO or MOO in vitro. Apoptosis was observed by light microscope and transmission electric microscope and confirmed by fluorescent TUNEL assay. The number of apoptosis and death cells was quantitated by flow cytometer using FITC- Annexin-V and propidium iodid (P1) staining. Expression of intercellular adhesion molecule-i (ICAM-1) and vascular cell adhesion molecule-i (VCAM-1) on the surface of VECs was detected by flow cytometer. Intracellular oxidative levels were measured by flow cytometric assay using an oxidant sensitive dye, 2,7-dichlorefluoresin (DCFH). Immune fluorescence assay was applied to observe the translocation of nuclear factor- K B(NF- K B) p65. Results: Apoptosis occurred when VECs were incubated with 3-DG or MOO evidenced by morphological changes and increased proportion of TUNEL positive cells. Apoptosis and death in VECs induced by 3-DO and MOO were both in a time- and dose-dependent manner. Up- regulation of expression of ICAM-l and VCAM-1 was induced by 3-DO or MOO. The highest expression of ICAM-1 appeared at 12 hours after stimulation, and VCAM-1 expression reached the top at 6 hours. Furthermore, markedly increasing in the levels of intracellular oxidation was observed, which started 3 bours after 3-DO and MOO were added into the cultures. In the unstimulated VECs, NF- K B p65 was located in cytoplasma and transferred into nucleus immediately after the cells were incubated with 3- DO and MGO. All of these effects of 3-DG and MOO, including induction of apoptosis and oxidative stress, up-regulation of adhesion molecules and activation of NF- K B were inhibited when a dicarbonyl scavenger, aminoguanidine(AO), was included in the culture, as well as preincubation of VECs with oxidative inhibitors, N-acetylcysteine(NAC) or probucol. Conclusion: Dicarbonyl compounds induce apoptosis and up-regulate expression of adhesion molecules of VECs by increasing intracellular oxidative level and activating NF- IC B, which could be partially blocked by both dicarbonyl scavenger and antioxidants. These results indicate that the accumulation of dicarbonyl compounds in circulation might be involved in the pathogeneses of vascular diseases seen in chronic renal failure and diabetes.