Ketosis (acetoacetate) can generate oxygen radicals and cause increased lipid peroxidation and growth inhibition in human endothelial cells.

Elevated level of cellular lipid peroxidation can increase the incidence of vascular disease. The mechanism by which ketosis causes accelerated cellular damage and vascular disease in diabetes is not known. This study was undertaken to test the hypothesis that elevated levels of ketone bodies increase lipid peroxidation in endothelial cells. Human umbilical venous endothelial cells (HUVEC) were cultured for 24 h at 37 degrees C with ketone bodies (acetoacetate, beta-hydroxybutyrate). Acetoacetate, but not beta-hydroxybutyrate, caused an increase in lipid peroxidation and growth inhibition in cultured HUVEC. To determine whether ketone bodies generate oxygen radicals, studies using cell-free buffered solution were performed. They showed a significant superoxide dismutase (SOD) inhibitable reduction of cytochrome C by acetoacetate, but not by beta-hydroxybutyrate, suggesting the generation of superoxide anion radicals by acetoacetate. Additional studies show that Fe2+ potentiates oxygen radical generation by acetoacetate. Thus, elevated levels of ketone body acetoacetate can generate oxygen radicals and cause lipid peroxidation in endothelial cells, providing a possible mechanism for the increased incidence of vascular disease in diabetes.

[1]  H. Heinzl,et al.  Endothelial eicosanoid metabolism and signal transduction during exposure to oxygen radicals injury. , 1997, Thrombosis research.

[2]  T. Lüscher.  The endothelium as a target and mediator of cardiovascular disease , 1993, European journal of clinical investigation.

[3]  V. Monnier,et al.  Immunohistochemical colocalization of glycoxidation products and lipid peroxidation products in diabetic renal glomerular lesions. Implication for glycoxidative stress in the pathogenesis of diabetic nephropathy. , 1997, The Journal of clinical investigation.

[4]  K. Audus,et al.  21-aminosteroid and 2-(aminomethyl)chromans inhibition of arachidonic acid-induced lipid peroxidation and permeability enhancement in bovine brain microvessel endothelial cell monolayers. , 1995, Free radical biology & medicine.

[5]  G. Czapski.  [24] Reaction of ·OH , 1984 .

[6]  V. Kalra,et al.  Diabetic RBC-induced oxidant stress leads to transendothelial migration of monocyte-like HL-60 cells. , 1997, The American journal of physiology.

[7]  S. Jain,et al.  Hyperglycemia can cause membrane lipid peroxidation and osmotic fragility in human red blood cells. , 1989, The Journal of biological chemistry.

[8]  G. Paolisso,et al.  Oxidative Stress and Diabetic Vascular Complications , 1996, Diabetes Care.

[9]  R. Cohen,et al.  Free radicals mediate endothelial cell dysfunction caused by elevated glucose. , 1992, The American journal of physiology.

[10]  S. Ahmed,et al.  A new rapid and simple non-radioactive assay to monitor and determine the proliferation of lymphocytes: an alternative to [3H]thymidine incorporation assay. , 1994, Journal of immunological methods.

[11]  P. Guillausseau,et al.  Rapid Publication , 1971, Nature.

[12]  S. Muller,et al.  Decreased Erythrocyte Membrane Fluidity in Poorly Controlled IDDM: Influence of ketone bodies , 1995, Diabetes Care.

[13]  V. Kalra,et al.  Increased adherence of oxidant‐treated human and bovine erythrocytes to cultured endothelial cells , 1987, Journal of cellular physiology.

[14]  S. Jain,et al.  Elevated lipid peroxidation levels in red blood cells of streptozotocin-treated diabetic rats. , 1991, Metabolism: clinical and experimental.

[15]  M. Limburg,et al.  PREDISPOSITION FOR ATOPY OR ALLERGY LINKED TO HL-A , 1975, The Lancet.

[16]  S. Jain,et al.  The effect of malonyldialdehyde on viscosity of normal and sickle red blood cells. , 1990, Biochemical medicine and metabolic biology.

[17]  S. Jain,et al.  In vivo externalization of phosphatidylserine and phosphatidylethanolamine in the membrane bilayer and hypercoagulability by the lipid peroxidation of erythrocytes in rats. , 1985, The Journal of clinical investigation.

[18]  J. Baynes.  Role of Oxidative Stress in Development of Complications in Diabetes , 1991, Diabetes.

[19]  S. Jain,et al.  Reduced Vitamin E and Increased Lipofuscin Products in Erythrocytes of Diabetic Rats , 1991, Diabetes.

[20]  R. Sundaram,et al.  Antioxidant status and lipid peroxidation in type II diabetes mellitus with and without complications. , 1996, Clinical science.

[21]  E. Kitzmüller,et al.  Arginine reduces kidney collagen accumulation, cross-linking, lipid peroxidation, glycoxidation, kidney weight and albuminuria in the diabetic kk mouse. , 1997, Nephron.

[22]  N. Mohandas,et al.  The effect of malonyldialdehyde, a product of lipid peroxidation, on the deformability, dehydration and 51Cr‐survival of erythrocytes , 1983, British journal of haematology.

[23]  B. Feldt-Rasmussen,et al.  FEATURES OF ENDOTHELIAL DYSFUNCTION IN EARLY DIABETIC NEPHROPATHY , 1989, The Lancet.

[24]  E. Joslin,et al.  Joslin's Diabetes Mellitus , 1971 .

[25]  D. Collen,et al.  Malondialdehyde-modified low density lipoproteins in patients with atherosclerotic disease. , 1995, The Journal of clinical investigation.

[26]  J. Tooke.  Peripheral microvascular disease in diabetes. , 1996, Diabetes research and clinical practice.

[27]  H. Esterbauer,et al.  [38] Detection of malonaldehyde by high-performance liquid chromatography , 1984 .

[28]  D. Trunkey,et al.  Mechanism of peroxide‐induced cellular injury in cultured adult cardiac myocytes , 1991, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[29]  E. Krane.  Diabetic Ketoacidosis: Biochemistry, Physiology, Treatment, and Prevention , 1987 .

[30]  H. Temme,et al.  Endothelial Cells in Culture , 1983 .

[31]  P. Davies,et al.  Mechanical stress mechanisms and the cell. An endothelial paradigm. , 1993, Circulation research.

[32]  J. Curnutte,et al.  Kinetic microplate assay for superoxide production by neutrophils and other phagocytic cells. , 1990, Methods in enzymology.

[33]  J. Satoh,et al.  N-acetylcysteine inhibits loss of diaphragm function in streptozotocin-treated rats. , 1996, American journal of respiratory and critical care medicine.

[34]  S. Jain,et al.  Erythrocyte Membrane Lipid Peroxidation and Glycosylated Hemoglobin in Diabetes , 1989, Diabetes.

[35]  R. Dean,et al.  Macrophages require both iron and copper to oxidize low-density lipoprotein in Hanks' balanced salt solution. , 1995, Archives of biochemistry and biophysics.

[36]  M. Runge,et al.  Induction of vascular endothelial growth factor in balloon-injured baboon arteries. A novel role for reactive oxygen species in atherosclerosis. , 1997, Circulation research.

[37]  Sushil K. Jain,et al.  Generation of superoxide radicals by hydrazine: Its role in phenylhydrazine-induced hemolytic anemia , 1979 .

[38]  A. J. Valente,et al.  Pathogenesis of the Atherosclerotic Lesion: Implications for diabetes mellitus , 1992, Diabetes Care.

[39]  S. Jain.  The accumulation of malonyldialdehyde, a product of fatty acid peroxidation, can disturb aminophospholipid organization in the membrane bilayer of human erythrocytes. , 1984, The Journal of biological chemistry.

[40]  N. Khaper,et al.  Probucol improves antioxidant activity and modulates development of diabetic cardiomyopathy. , 1995, Nutrition.

[41]  D. Morel,et al.  Antioxidant treatment of diabetic rats inhibits lipoprotein oxidation and cytotoxicity. , 1989, Journal of lipid research.

[42]  A. Veves,et al.  Rheological and microvascular parameters in diabetic peripheral neuropathy. , 1996, Clinical science.

[43]  R. Natarajan,et al.  Glucose induces lipid peroxidation and inactivation of membrane‐associated ion‐transport enzymes in human erythrocytes in vivo and in vitro , 1991, Journal of cellular physiology.

[44]  D. Erdogan,et al.  Antioxidant and triglyceride-lowering effects of vitamin E associated with the prevention of abnormalities in the reactivity and morphology of aorta from streptozotocin-diabetic rats. Antioxidants in Diabetes-Induced Complications (ADIC) Study Group. , 1997, Metabolism: clinical and experimental.

[45]  R. Micciolo,et al.  Mechanisms involved in the in vitro modification of low density lipoprotein by human umbilical vein endothelial cells and copper ions. , 1996, Journal of lipid mediators and cell signalling.