Cardiovascular disease is a major cause of morbidity and premature mortality in diabetes. HDL plays an important role in limiting vascular damage by removing cholesterol and cholesteryl ester hydroperoxides from oxidized low density lipoprotein and foam cells. Methionine (Met) residues in apolipoprotein A-I (apoA-I), the major apolipoprotein of HDL, reduce peroxides in HDL lipids, forming methionine sulfoxide [Met(O)]. We examined the extent and sites of Met(O) formation in apoA-I of HDL isolated from plasma of healthy control and type 1 diabetic subjects to assess apoA-I exposure to lipid peroxides and the status of oxidative stress in the vascular compartment in diabetes. Three tryptic peptides of apoA-I contain Met residues: Q84-M86-K88, W108-M112-R116, and L144-M148-R149. These peptides and their Met(O) analogs were identified and quantified by mass spectrometry. Relative to controls, Met(O) formation was significantly increased at all three locations (Met86, Met112, and Met148) in diabetic patients. The increase in Met(O) in the diabetic group did not correlate with other biomarkers of oxidative stress, such as Nϵ-malondialdehyde-lysine or Nϵ-(carboxymethyl)lysine, in plasma or lipoproteins. The higher Met(O) content in apoA-I from diabetic patients is consistent with increased levels of lipid peroxidation products in plasma in diabetes. Using the methods developed here, future studies can address the relationship between Met(O) in apoA-I and the risk, development, or progression of the vascular complications of diabetes.

Chemical modification of proteins by advanced glycation and lipoxidation end products is implicated in the pathogenesis of macrovascular disease in aging and diabetes. To identify biomarkers of the lipoxidative modification of protein, we studied the oxidation of phospholipids in the presence of the model protein RNase A and compared protein-bound products formed in these reactions with those formed during oxidation of plasma proteins. Metal-catalyzed oxidation of 1-palmitoyl-2-arachidonoyl-phosphatidylcholine or 1-palmitoyl-2-linoleoyl-phosphatidylcholine in the presence of RNase led to the loss of amino groups in RNase and the incorporation of phosphate, hexanoate, pentanedioate, nonanedioate, and palmitate into protein. Protein-bound palmitate and phosphate correlated strongly with one another, and protein-bound pentanedioate and nonanedioate, derived from arachidonate and linoleate, respectively, accounted for ∼20% of the cross-linking of lipid phosphorus to protein. Similar results were obtained on oxidation of total plasma or isolated LDL. We conclude that alkanedioic acids are quantitatively important linkers of oxidized phospholipids to proteins and that measurement of protein-bound phosphate and long-chain fatty acids may be useful for assessing long-term lipid peroxidative damage to proteins in vivo. Analyses of plasma proteins from control and diabetic patients indicated significant increases in lipoxidative modification of protein in diabetic compared with control subjects.

Prolonged hyperglycemia, dyslipidemia and oxidative stress in diabetes result in the production and accumulation of AGEs. It is now clear that AGEs contribute to the development and progression of cardiovascular disease in diabetes, as well as other complications. AGEs are thought to act through receptor-independent and dependent mechanisms to promote vascular damage, fibrosis and inflammation associated with accelerated atherogenesis. As a result, novel therapeutic agents to reduce the accumulation of AGEs in diabetes have gained interest as potential cardioprotective approaches. A variety of agents have been developed which are examined in detail in this review. These include aminoguanidine, ALT-946, pyridoxamine, benfotiamine, OPB-9195, alagebrium chloride, N-phenacylthiazolium bromide and LR-90. In addition, it has been demonstrated that a number of established therapies have the ability to reduce the accumulation of AGEs in diabetes including ACE inhibitors, angiotensin receptor antagonists, metformin, peroxisome proliferators receptor agonists, metal chelators and some antioxidants. The fact that many of these inhibitors of AGEs are effective in experimental models, despite their disparate mechanisms of action, supports the keystone role of AGEs in diabetic vascular damage. Nonetheless, the clinical utility of AGE inhibition remains to be firmly established. Optimal metabolic and blood pressure control, that is achieved early and sustained indefinitely, remains the best recourse for inhibition of AGEs until more specific interventions become a clinical reality.

Des comparaisons ont ete effectuees entre des estimes d'indice de surface foliaire (SAI non projete du feuillage), calcules a partir des relations allometriques et du pourcentage d'ouverture du couvert, et les caracteristiques de reflectance spectrale de peuplements de qualites variables, âges de 8 ans, de Pinus elliottii Engelm. var. elliottii × Pinus caribaea var. hondurensis Barr. et Golf. Les estimes allometriques de SAI du feuillage, effectues en ayant recours a des relations derivees empiriquement entre la surface terriere (1,3 m) de l'aubier et la surface foliaire (R 2 = 0,88, n = 11), variaient entre 4,46 et 9,77. Les estimes de SAI du feuillage calcules en utilisant le pourcentage d'ouverture du couvert (LI-COR LAI-2000 plant canopy analyser, PCA), apres correction pour la distribution non aleatoire des elements du feuillage, etaient bien correles (R 2 = 0,90) avec les estimes directs, tout en etant en moyenne inferieurs de 11%. Les relations entre le SAI du feuillage (allometrique) et les caracteristiques de reflectance spectrale ont ete analysees. Des relations inverses et non lineaires etroites entre le SAI du feuillage et les bandes de capteur thematique (TM) 5 (1 550 - 1 750 nm, R 2 = 0,91) et 3 (630-690 nm, R 2 = 0,71), la composante principale (CP) 1 des bandes 3, 4 et 5 (CP1, R 2 = 0,86) et le compose des bandes 3, 4 et 5 (R 2 = 0,92) ont ete observees. De plus, des correlations lineaires positives entre l'indice de vegetation par difference normalisee (NDVI, R 2 = 0,81) et le ratio simple de bande TM 4 a bande TM 3 (SR, R 2 = 0,78) ont aussi ete observees. Cependant, le SAI du feuillage demontrait une relation faible (R 2 = 0,02) avec la bande TM 4 (760-900 nm). Les relations etroites observees entre les diverses caracteristiques de reflectance spectrale et les parametres structuraux le plus souvent mesures en foret, tels la surface terriere et la hauteur predominante, suggerent que ces methodes de teledetection pourraient s'averer prometteuses pour utilisation dans des systemes d'inventaire en bas âge.

We examined the effect of two endogenous antioxidant agents, taurine and vitamin E, on renal function in experimental diabetes. Male Sprague-Dawley rats, rendered diabetic with streptozocin (STZ), were assigned to one of the following groups: 1) untreated; 2) insulin treatment with 6 U Ultralente insulin/day in two doses; 3) taurine supplementation by 1% taurine in drinking water; and 4) vitamin E supplementation at 100 IU vitamin E/kg chow. Animals were kept for 52 wk. The survival rate was similar (70-90%) in all groups except vitamin E-treated animals, of which 84% died by 6 mo. At 52 wk, glomerular filtration rate was elevated in untreated and taurine-treated STZ rats compared with normal or insulin-treated diabetic rats. Taurine supplementation reduced total proteinuria and albuminuria by nearly 50%. This treatment also prevented glomerular hypertrophy, preserved immunohistochemical staining for type IV collagen in glomeruli, and diminished glomerulosclerosis and tubulointerstitial fibrosis in diabetic animals. The changes in renal function and structure in taurine-treated diabetic rats were associated with normalization of renal cortical malondialdehyde content, lowering of serum free Fe2+ concentration, and decreased formation of the advanced glycooxidation products, pentosidine, and fluorescence in skin collagen. Administration of the vitamin E-enriched diet exacerbated the nephropathy in STZ-diabetic rats. In addition, vitamin E supplementation increased serum free Fe2+ concentration, enhanced renal lipid peroxidation, and accelerated the accumulation of advanced glycosylation end products (AGEs) in skin collagen. We conclude that administration of taurine, but not vitamin E, to rats with STZ-diabetes ameliorates diabetic nephropathy. The beneficial effect of taurine is related to reduced renal oxidant injury with decreased lipid peroxidation and less accumulation of AGEs within the kidney.

Residualizing labels are radioactive or fluorescent tracers used for identifying the tissue and cellular sites in which circulating proteins are catabolized in the body. When attached to protein the labels do not affect normal mechanisms of protein catabolism, but remain at the cellular site of protein uptake, after the carrier protein itself is degraded to diffusible catabolites. Until recently these labels consisted of biologically indigestible carbohydrates attached to a radioactive reporter molecule. In this report we describe the synthesis and purification of a new fluorescent residualizing label, N,N-dilactitol-N'-fluoresceinyl-ethylenediamine. The label is prepared by first derivatizing ethylenediamine 1:1 with fluorescein isothiocyanate and then coupling lactose to the remaining primary amino group by reductive amination. A rapid one step purification of this and other glycoconjugate labels by reversed-phase high-pressure liquid chromatography is described.

The kinetics of plasma clearance of highly purified human placental glucocerebrosidase in rats were biphasic with 75% of the infused dose showing a rapid clearance (t1/2 = 11 min) and the remaining 25% a considerably lower rate (t1/2 = 60 min). The majority of the enzyme (60%) was taken up by the liver. Although saturation kinetics for the clearance or uptake were not observed, the very high hepatic endocytic index (217 microliter/min) of glucocerebrosidase uptake indicated that liver uptake was mediated by an adsorptive endocytic process. Analysis of the cellular distribution of recovered glucocerebrosidase revealed predominantly parenchymal cell uptake with 38% of the exogenous enzyme in hepatocytes and only 2% in sinusoidal cells. High-mannose glycoproteins blocked hepatocyte and sinusoidal cell uptake of glucocerebrosidase equally. Kinetic experiments failed to demonstrate a transfer or shuttle of exogenous glucocerebrosidase from sinusoidal cells to hepatocytes. The possibility was raised that uptake of enzyme by the liver may be mediated by a common receptor that functions in both hepatocytes and sinusoidal cells. The catabolic turnover of exogenous glucocerebrosidase in rat liver was biphasic and the rate of decline was similar in hepatocytes and sinusoidal cells.

Prekeratin has been isolated from human epidermis using the method of Matoltsy for bovine nose epidermis. Analysis by sedimentation equilibrium indicates that human prekeratin is heterogeneous in solution. The molecular weight range is from about 200 X 10(3) to at least 2 X 10(6). Polyacrylamide gel electrophoresis shows three polypeptide chains of molecular weights 65,000 60,000 and 58,000. A lower than average ratio of polar/apolar amino acid residues suggests that hydrophobic interactions are important for stability.