Modifications induced by LDL from type 1 diabetic patients on endothelial cells obtained from human umbilical vein.
The aim of the present work was to analyze the effect of LDL obtained from type 1 diabetic patients in good metabolic control on human umbilical vein endothelial cells (HUVECs) after a short incubation period to detect possible atherogenic modifications of endothelial properties. Cultured HUVECs were incubated for 3 h with culture medium alone (control HUVEC), with native LDL from 12 healthy men (control LDL), or with native LDL from 12 type 1 diabetic men (type 1 LDL) (100 pg/ml). After the incubation, the following parameters were evaluated: nitric oxide synthase (NOS) activity, cytoplasmic Ca2+ levels, Na+-K+-ATPase activity, plasma membrane fluidity determined by means of 1,6-diphenyl-1,3,5-hexatriene (DPH) and 1-(4-trimethylaminophenyl)-6-phenyl-1,3,5-hexatriene (TMA-DPH), and plasma membrane conjugated diene (CD) content. The same experiments were repeated after bradykinin stimulation or in the presence of the antioxidant butylated hydroxytoluene (BHT), and nitric oxide (NO) production in intact HUVECs was also evaluated. HUVECs incubated with control LDL in comparison with control HUVECs showed a decreased fluidity of the membrane surface evaluated by TMA-DPH and a higher CD content. These alterations were prevented by the presence of BHT. HUVECs incubated with type 1 LDL in comparison with both control HUVECs and cells incubated with control LDL showed 1) increased NOS and Na+-K+-ATPase activity, cytoplasmic Ca2+ levels, and CD content, and 2) decreased fluidity of the membrane surface evaluated by TMA-DPH. These modifications were blunted--but not abolished--by the presence of BHT. After bradykinin stimulation either in the absence or in the presence of BHT, both cytoplasmic Ca2+ levels and NO production were increased in control HUVECs and in HUVECs incubated with control LDL, while a reduced response was observed in HUVECs incubated with type 1 LDL. The alterations observed in the endothelial function after the cell-LDL interaction might play a central role in the atherogenic process in diabetes.
Rabini
R A
RA
Department of Diabetology, INRCA Hospital, Ancona, Italy.
Cester
N
N
Staffolani
R
R
Salvolini
E
E
Moretti
N
N
Vignini
A
A
Fumelli
D
D
Mazzanti
L
L
D000328:Adult; D002118:Calcium / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism; D002784:Cholesterol / Q000097:blood; D008076:Cholesterol, HDL / Q000097:blood; D008078:Cholesterol, LDL / Q000097:blood; D003922:Diabetes Mellitus, Type 1 / Q000097:blood*; D004161:Diphenylhexatriene / Q000031:analogs & derivatives; D004730:Endothelium, Vascular / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D005456:Fluorescent Dyes; D006801:Humans; D008077:Lipoproteins, LDL / Q000097:blood* / Q000494:pharmacology; D008297:Male; D008560:Membrane Fluidity / Q000502:physiology; D019001:Nitric Oxide Synthase / Q000378:metabolism; D010743:Phospholipids / Q000097:blood; D012016:Reference Values; D000254:Sodium-Potassium-Exchanging ATPase / Q000378:metabolism; D014280:Triglycerides / Q000097:blood; D014471:Umbilical Veins
D008076:Cholesterol, HDL; D008078:Cholesterol, LDL; D005456:Fluorescent Dyes; D008077:Lipoproteins, LDL; D010743:Phospholipids; D014280:Triglycerides; D004161:Diphenylhexatriene; C033628:1-(4-(trimethylamino)phenyl)-6-phenylhexa-1,3,5-triene; D002784:Cholesterol; D019001:Nitric Oxide Synthase; D000254:Sodium-Potassium-Exchanging ATPase; D002118:Calcium