Endothelial cell injury by high glucose and heparanase is prevented by insulin, heparin and basic fibroblast growth factor.
BACKGROUND
Uncontrolled hyperglycemia is the main risk factor in the development of diabetic vascular complications. The endothelial cells are the first cells targeted by hyperglycemia. The mechanism of endothelial injury by high glucose is still poorly understood. Heparanase production, induced by hyperglycemia, and subsequent degradation of heparan sulfate may contribute to endothelial injury. Little is known about endothelial injury by heparanase and possible means of preventing this injury.


OBJECTIVE
To determine if high glucose as well as heparanase cause endothelial cell injury and if insulin, heparin and bFGF protect cells from this injury.


METHODS
Cultured porcine aortic endothelial cells were treated with high glucose (30 mM) and/or insulin (1 U/ml) and/or heparin (0.5 microg/ml) and /or basic fibroblast growth factor (bFGF) (1 ng/ml) for seven days. Cells were also treated with heparinase I (0.3 U/ml, the in vitro surrogate heparanase), plus insulin, heparin and bFGF for two days in serum free medium. Endothelial cell injury was evaluated by determining the number of live cells per culture and lactate dehydrogenase (LDH) release into medium expressed as percentage of control.


RESULTS
A significant decrease in live cell number and increase in LDH release was found in endothelial cells treated with high glucose or heparinase I. Insulin and/or heparin and/or bFGF prevented these changes and thus protected cells from injury by high glucose or heparinase I. The protective ability of heparin and bFGF alone or in combination was more evident in cells damaged with heparinase I than high glucose.


CONCLUSIONS
Endothelial cells injured by high glucose or heparinase I are protected by a combination of insulin, heparin and bFGF, although protection by heparin and/or bFGF was variable.
Han
Juying
J
Department of Veterinary Biomedical Sciences, University of Saskatchewan, Saskatoon, Saskatchewan, S7N 5B4, Canada. juyinghan@yahoo.ca
Mandal
Anil K
AK
Hiebert
Linda M
LM
D000818:Animals; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D003925:Diabetic Angiopathies / Q000473:pathology / Q000503:physiopathology; D004730:Endothelium, Vascular / Q000187:drug effects* / Q000473:pathology / Q000503:physiopathology; D016222:Fibroblast Growth Factor 2 / Q000494:pharmacology* / Q000502:physiology; D005947:Glucose / Q000494:pharmacology* / Q000502:physiology; D005966:Glucuronidase / Q000494:pharmacology* / Q000502:physiology; D006493:Heparin / Q000494:pharmacology* / Q000502:physiology; D006943:Hyperglycemia / Q000201:enzymology / Q000473:pathology / Q000503:physiopathology; D007328:Insulin / Q000494:pharmacology* / Q000502:physiology; D007770:L-Lactate Dehydrogenase / Q000032:analysis; D013552:Swine; D000099175:Heparanase
D016222:Fibroblast Growth Factor 2; D005947:Glucose; D005966:Glucuronidase; D006493:Heparin; D007328:Insulin; D007770:L-Lactate Dehydrogenase; D000099175:Heparanase