RESUMEN
AIMS/HYPOTHESIS: The aim of this study was to quantify protein damage by glycation, oxidation and nitration in a rat model of diabetes at the sites of development of microvascular complications, including the effects of thiamine and benfotiamine therapy. METHODS: Diabetes was induced in male Sprague-Dawley rats by 55 mg/kg streptozotocin and moderated by insulin (2 U twice daily). Diabetic and control rats were given thiamine or benfotiamine (7 or 70 mg kg(-1) day(-1)) over 24 weeks. Plasma, urine and tissues were collected and analysed for protein damage by stable isotopic dilution analysis MS. RESULTS: There were two- to fourfold increases in fructosyl-lysine and AGE content of glomerular, retinal, sciatic nerve and plasma protein in diabetes. Increases in AGEs were reversed by thiamine and benfotiamine therapy but increases in fructosyl-lysine were not. Methionine sulfoxide content of plasma protein and 3-nitrotyrosine content of sciatic nerve protein were increased in diabetes. Plasma glycation free adducts were increased up to twofold in diabetes; the increases were reversed by thiamine. Urinary excretion of glycation, oxidation and nitration free adducts was increased by seven- to 27-fold in diabetes. These increases were reversed by thiamine and benfotiamine therapy. CONCLUSIONS/INTERPRETATION: AGEs, particularly arginine-derived hydroimidazolones, accumulate at sites of microvascular complication development and have markedly increased urinary excretion rates in experimental diabetes. Thiamine and benfotiamine supplementation prevented tissue accumulation and increased urinary excretion of protein glycation, oxidation and nitration adducts. Similar effects may contribute to the reversal of early-stage clinical diabetic nephropathy by thiamine.
Asunto(s)
Diabetes Mellitus Experimental/metabolismo , Glomérulos Renales/metabolismo , Proteínas/metabolismo , Retina/efectos de los fármacos , Retina/metabolismo , Tiamina/análogos & derivados , Tiamina/farmacología , Adyuvantes Inmunológicos/farmacología , Animales , Diabetes Mellitus Experimental/sangre , Diabetes Mellitus Experimental/tratamiento farmacológico , Diabetes Mellitus Experimental/orina , Modelos Animales de Enfermedad , Glomérulos Renales/efectos de los fármacos , Masculino , Espectrometría de Masas , Modelos Biológicos , Estrés Oxidativo/efectos de los fármacos , Ratas , Ratas Sprague-Dawley , Tiamina/uso terapéutico , Complejo Vitamínico B/farmacologíaRESUMEN
AIMS/HYPOTHESIS: Cardiovascular disease in diabetes is linked to increased risk of atherosclerosis, increased levels of triglyceride-rich lipoproteins and enhanced hepatic lipogenesis. The hepatic hexosamine pathway has been implicated in signalling for de novo lipogenesis by the liver. In this study, we assessed if decrease of flux through the hexosamine pathway induced by high-dose thiamine therapy counters diabetic dyslipidaemia. METHODS: The model of diabetes used was the streptozotocin-induced diabetic rat with maintenance insulin therapy. Normal control and diabetic rats were studied for 24 weeks with and without oral high-dose therapy (7 and 70 mg/kg) with thiamine and benfotiamine. Plasma total cholesterol, HDL cholesterol and triglycerides were determined at 6-week intervals and hepatic metabolites and transketolase activity after death of the rats at 24 weeks. RESULTS: We found that thiamine therapy (70 mg/kg) prevented diabetes-induced increases in plasma cholesterol and triglycerides in diabetic rats but did not reverse the diabetes-induced decrease of HDL. This was achieved by prevention of thiamine depletion and decreased transketolase activity in the liver of diabetic rats. There was a concomitant decrease in hepatic UDP-N-acetylglucosamine and fatty acid synthase activity. Thiamine also normalised food intake of diabetic rats. A lower dose of thiamine (7 mg/kg) and the thiamine monophosphate prodrug benfotiamine (7 and 70 mg/kg) were ineffective. CONCLUSIONS/INTERPRETATION: High-dose thiamine therapy prevented diabetic dyslipidaemia in experimental diabetes probably by suppression of food intake and hexosamine pathway signalling but other factors may also be involved. Benfotiamine was ineffective.
Asunto(s)
Diabetes Mellitus Experimental/complicaciones , Hiperlipidemias/prevención & control , Tiamina/uso terapéutico , Animales , Colesterol/sangre , HDL-Colesterol/sangre , Diabetes Mellitus Experimental/sangre , Diabetes Mellitus Experimental/tratamiento farmacológico , Insulina/uso terapéutico , Ratas , Triglicéridos/sangreRESUMEN
The accumulation of AGEs (advanced glycation end products) in diabetes mellitus has been implicated in the biochemical dysfunction associated with the chronic development of microvascular complications of diabetes--nephropathy, retinopathy and peripheral neuropathy. We investigated the concentrations of fructosyl-lysine and AGE residues in protein extracts of renal glomeruli, retina, peripheral nerve and plasma protein of streptozotocin-induced diabetic rats and normal healthy controls. Glycation adducts were determined by LC with tandem MS detection. In diabetic rats, the fructosyl-lysine concentration was increased markedly in glomeruli, retina, sciatic nerve and plasma protein. The concentrations of N (epsilon)-carboxymethyl-lysine and N (epsilon)-carboxyethyl-lysine were increased in glomeruli, sciatic nerve and plasma protein, and N(epsilon)-carboxymethyl-lysine also in the retina. Hydroimidazolone AGEs derived from glyoxal, methylglyoxal and 3-deoxylglucosone were major AGEs quantitatively. They were increased in the retina, nerve, glomeruli and plasma protein. AGE accumulation in renal glomeruli, retina, peripheral nerve and plasma proteins is consistent with a role for AGEs in the development of nephropathy, retinopathy and peripheral neuropathy in diabetes. High-dose therapy with thiamine and Benfotiamine suppressed the accumulation of AGEs, and is a novel approach to preventing the development of diabetic complications.