The Effects Of Native And Oxidized High Density Lipoprotein On Some Secretive Functions Of Endothelial Cells

Many factors are involved in the development ofatherosclerosis(AS), of which the injury and disfunction of endothelial cells is the initial step. Studies indicated that high density lipoprotein(HDL) help maintain endothelial integrity, facilitate vascular relaxation, inhibit blood cell adhesion to vascular endothelium. Therefore, it can prevent and eventually correct endothelial dysfunction. Studies in our laboratory and abroad revealed that HDL can be oxiditively modified by endothelial cells , macrophages and smooth muscle cells in vivo and by Cu²⁺ and HClO in vitro. Once HDL is oxiditively modified, it will not only lose its above functions, but becomes toxic to endothelial cells and promotes the development of AS. To further understand the role of OX-HDL in the development of AS, the present study is to explore the effects of native and oxidized high density lipoprotein on some secretive functions of endothelial cells.The native high density lipoprotein was isolated by the density gradient ultra-centrifugation and oxiditively modified by Cu²⁺. The oxidation of HDL was identified by agarose gel relative electrophoretic mobility(REM), absorbance at 234nm (A234,conjugated diene,CD) and thiobarbituric acidsubstance(TBARS) contents. Using 0.01mol/L phosphate bufferedsaline(PBS) as the negative control, OX-HDL of 5mg/L, lOmg/L, 50mg/L were co-incubated with human umbilical vein endothelial cells(HUVECs) for 24h in vitro. The activity of nitric oxide synthase(NOS) and the levels of nitric oxide(NO) in the medium were detected after the incubation. The protein content of each poll was measured to correct the difference induced by different quantity of endothelial cells. Compared with the negative control, the activity of NOS(p<0.01) and the levels of NO(p<0.05) were significantly decreased in a dose-dependent way. And a concentration of 50mg/L OX-HDL protein has the most significant effect.To explore the effects of native HDL and oxidized HDL on some secretive functions of endothelial cells, with O.Olmol/L of PBS and 50mg/L of N-HDL as the negative and positive control, OX-HDL of 50mg/L was co-incubated with HUVECs for 24h in vitro. The activiry of NOS, the levels of NO, and the levels of tissue plasminogen activator(t-PA) and plasminogen activator inhibitor-l(PAI-l) in the medium were detected after the incubation. The protein content of each poll was also measured to correct the difference induced by different quantity of endothelial cells. Compared with the negative control, the activity of NOS(><0.01) and the levels of NO(><0.05) were significantly increased in the N-HDL group,but the changes of tPA and PAI-1 had no statistical significance(p>0.05); In the OX-HDL group, the activity of NOS and the levels of NO were significantly decreased, but the PAI-1 activity(p<0.05) was increased. The levels of tPA(/?>0.05) were not affected. These results suggest that N-HDLsignificantly increased the activity of NOS and the levels of NO secreted by HUVECs, but had no influence on the levels of tPA and PAI-1. It may be one of the mechanisms of endothelial protective functions of N-HDL. On the contrary, OX-HDL significantly decreased the activity of NOS and the synthesis of NO, and increased PAI-1 activity, but had no effect on the secretion of tPA. This may be one of the mechanisms that OX-HDL promotes the development of AS.