Janet Sredy

Prevention of urinary albumin excretion in 6 month streptozocin-diabetic rats with the aldose reductase inhibitor tolrestat

Recent clinical data strongly suggest that elevated urinary albumin excretion (UAE) identifies diabetic subjects at risk of developing nephropathy. Elevated UAE is attributed to increased transglomerular pressure, which is associated with poor metabolic control in rats. Because excess glucose in diabetes is metabolized via the polyol pathway, we were interested in whether the diabetes-induced elevation in UAE in rats could be prevented by inhibiting aldose reductase (AR), the first enzyme in the polyol pathway, with the AR inhibitor tolrestat. In fact, in rats made diabetic with streptozocin (35 mg/kg IV), treatment for 6 months with tolrestat (25 mg/kg/day in the diet) prevented both sorbitol accumulation in the kidney and the increase in UAE. Sorbitol accumulation and the increased UAE were not associated with statistically significant mesangial expansion, and the thickening of glomerular basement membranes was not affected by tolrestat treatment. The authors conclude that the 4.7-fold elevation in UAE in chronically diabetic rats is linked to the increased flux of glucose through the polyol pathway since it was prevented by inhibiting aldose reductase with tolrestat.

Structural abnormalities do not explain the early functional abnormalities in the peripheral nerves of the streptozotocin diabetic rat.

The streptozotocin (STZ)‐diabetic rat, the most commonly employed model of experimental diabetic neuropathy, is characterised by a reduction in nerve conduction velocity, pain threshold and blood flow. Whether or not structural abnormalities underlie these functional abnormalities is unclear. 10 adult male Sprague–Dawley STZ‐diabetic rats (diabetes duration 27 d) and 10 age‐matched (23 wk) control animals were studied. Motor nerve conduction velocity (m s−1) was significantly reduced in diabetic (41.31±0.8) compared with control (46.15±1.5) animals (P<0.001). The concentration of sciatic nerve glucose (P<0.001), fructose (P<0.001) and sorbitol (P<0.001) was elevated, and myoinositol (P<0.001) was reduced in diabetic compared with control animals. Detailed morphometric studies demonstrated no significant difference in fascicular area, myelinated fibre density, fibre and axon areas as well as unmyelinated fibre density and diameter. Endoneurial capillary density, basement membrane area and endothelial cell profile number did not differ between diabetic and control animals. However, luminal area (P<0.03) was increased and endothelial cell area (P<0.08) was decreased in the diabetic rats. We conclude there is no detectable structural basis for the reduction in nerve conduction velocity, pain threshold or blood flow, observed in the streptozotocin diabetic rat.

Metabolic Abnormalities of the Hyperglycemic Obese Zucker Rat

In a cross-sectional study, we evaluated the metabolic profiles of lean (Fa/?) and obese (fa/fa) Zucker male rats at 4 to 8 months of age. Although all of the obese rats (N = 108) demonstrated glucose intolerance, most of the obese rats exhibited only mild elevations of fasted and fed plasma glucose. Only 14 of the obese rats were severely hyperglycemic, which resulted in substantial elevations of glycohemoglobin (GHb) levels. The nerve and lens levels of glucose, sorbitol, and fructose were elevated, and the myo-inositol was depleted in all hyperglycemic obese rats, but not in the euglycemic obese rats. With increasing duration of hyperglycemia, the neural myo-inositol level approached normal, while the lenses became cataractous. All obese rats had increased urinary albumin excretion (UAE), which was dependent on age (r = .45, P less than .02) and independent of hyperglycemia, glucosuria, and polyuria. In conclusion, although the euglycemic obese rats exhibited some diabetic abnormalities, the hyperglycemic obese Zucker rat more closely resembled the altered metabolic profile associated with type II diabetes mellitus.

Antihyperglycemic activity of new 1,2,4-oxadiazolidine-3,5-diones.

A series of 1,2,4-oxadiazolidine-3,5-diones was synthesized and evaluated as oral antihyperglycemic agents in the obese insulin resistant db/db and ob/ob mouse - the two models for Type 2 diabetes mellitus. The majority of the prepared methoxy- and ethoxy-linked oxazole 1,2,4-oxadiazolidine-3,5-diones normalized plasma glucose levels at the 100 mg kg(-1) oral dose in the db/db diabetic mouse model, and several amongst them reduced the glucose levels at the 20 mg kg(-1) oral dose. The most potent compounds in the db/db mouse model were also active in the ob/ob mouse model normalizing the plasma glucose levels at the 20 mg kg(-1) oral dose. The trifluoromethoxy analog 32 was the most active compound of the series, reducing significantly the plasma glucose levels at the 5 mg kg(-1) oral dose. Oxadiazole-tailed 1,2,4-oxadiazolidine-3,5-diones were also active in both the db/db and ob/ob diabetic mouse models normalizing plasma glucose levels at the 100 mg kg(-1) oral dose.

Polyol pathway activity in nervous tissues of diabetic and galactose-fed rats: Effect of dietary galactose withdrawal or tolrestat intervention therapy

Enhanced polyol pathway activity resulting in an accumulation of sorbitol and a depletion of myoinositol in nervous tissues has been proposed to be important in development of diabetic neuropathies. This investigation demonstrated that in two models of diabetic complications, streptozocin (STZ)-induced diabetic rats and galactose-fed rats, 5 weeks of disease led to an accumulation of sorbitol or galactitol, respectively, in three cranial nerves (the optic (II), trigeminal (V), and vagus (X) nerves), as well as the sciatic nerve, cerebral cortex, and retina. In both models, the cranial nerves and cerebral cortex contained lower levels of accumulated polyol than the sciatic nerve. In addition, myoinositol depletion was observed in the sciatic nerve only. In a second galactose-fed rat study, returning 5-week galactose-fed rats to a normal diet for 6 weeks led to complete elimination of galactitol from the optic nerve, sciatic nerve, and retina and normalization of the sciatic nerve myoinositol concentration. Similarly, continuing the galactose diet for 6 more weeks (ie, a total of 11 weeks) as well as administration of the aldose reductase inhibitor (ARI) tolrestat (20 and 40 mg/kg/day), caused the sciatic nerve to contain a normal myoinositol concentration and a galactitol concentration that was 95% below the level observed in galactose-fed controls. In the optic nerve and retina, tolrestat was less effective, resulting in 69-78% lower galactitol levels. In conclusion, these findings indicate that sorbitol and galactitol accumulate in cranial nerves, brain, and retina without a concomitant decrease in myoinositol. Either withdrawal of the galactose diet or intervention with tolrestat normalized polyol levels in the sciatic nerve.(ABSTRACT TRUNCATED AT 250 WORDS)

Insulin resistance is associated with abnormal dephosphorylation of a synthetic phosphopeptide corresponding to the major autophosphorylation sites of the insulin receptor

Insulin resistance in the ob/ob mouse model is associated with a reduction in insulin-induced protein-tyrosine phosphorylation in tissues such as liver. To ascertain whether this decrease in phosphorylation may be due to increased phosphatase activity, protein-tyrosine phosphatase (PTPase) activity was determined in particulate and soluble fractions from livers of 5- to 23-week-old ob/ob mice and age-matched lean littermates. PTPase activity was measured using a synthetic phosphopeptide, TRDIY(P)ETDY(P)Y(P)RK, as the substrate, corresponding to residues 1142 to 1153 of the insulin receptor and containing the major autophosphorylation sites of the regulatory domain. The ob/ob mice were hyperinsulinemic across all age groups, but only the youngest mice (aged 5 to 7 weeks) were hyperglycemic. Most PTPase activity was present in the liver particulate fraction and was 19% to 114% greater in ob/ob mice as compared with controls. PTPase activity in the liver soluble fraction was 26% less than control values in the youngest ob/ob mice (5 to 7 weeks), but increased with age and was 41% and 131% above control values at 21 to 23 and 25 to 27 weeks of age, respectively. Oral administration of the PTPase inhibitor sodium orthovanadate (0.6 mg/mL in drinking water for 2 weeks) to young ob/ob mice caused a significant reduction in the elevated particulate PTPase activity, with concomitant decreases in plasma insulin and plasma glucose. Assessment of PTPase activity with a monophosphate form of the same synthetic peptide, TRDIY(P)ETDYYRK, showed lower PTPase activities as compared with the triphosphate form and no significant differences between ob/ob and control preparations.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenosine Triphosphatase Activity in Sciatic Nerve Tissue of Streptozocin-Induced Diabetic Rats with and without High Dietary Sucrose: Effect of Aldose Reductase Inhibitors

Abstract The ability of aldose reductase inhibitors to prevent the decline in neural Na+,K+-ATPase activity in diabetic rats has not been confirmed by all laboratories. In this study, the efficacy of two structurally different aldose reductase inhibitors was evaluated under different experimental conditions. Na+,K+-ATPase activity was measured in sciatic nerves from streptozocin-induced diabetic rats fed normal rodent chow or a chow supplemented with 68% sucrose. Nerve homogenates from chow-fed rats were prepared with a Dounce tissue grinder, whereas homogenates from the sucrose-fed rats were prepared with an Ultra-Turrax disperser. In the chow-fed rats, 4 weeks of untreated diabetes resulted in an increase in neural sorbitol and fructose, a decrease in myoinositol, and a 54% decline in Na+,K+-ATPase activity. Sorbinil administration (20 mg/kg/day) completely prevented the rise in sorbitol and fructose and the depletion of myoinositol, but did not prevent the decline in Na+,K+-ATPase activity. In diabetic rats fed the sucrose diet for 4, 6, and 8 weeks, the neural sorbitol and fructose levels were elevated, the myoinositol concentration declined, and the Na+,K+-ATPase activity was 26 to 28% below the control. Prevention or intervention treatment with sorbinil (20 mg/kg/day) or tolrestat (50 mg/kg/day) for 4 to 6 weeks prevented the alterations in sorbitol, fructose, and myoinositol, and also prevented the decline in Na+,K+-ATPase activity. In conclusion, prevention and intervention therapy with aldose reductase inhibitors prevented the decline in Na+,K+-ATPase in sciatic nerves of sucrose-fed streptozocin-diabetic rats that were homogenized with an Ultra-Turrax disperser, but not in sciatic nerves from streptozocin-diabetic rats fed normal rodent chow that were homogenized with a Dounce tissue grinder. These findings indicate that the assessment of aldose reductase inhibitor efficacy is dramatically affected by the type of nerve preparation assayed and/or the diet.

Discovery of [3-(4,5,7-trifluoro-benzothiazol-2-ylmethyl)-pyrrolo[2,3-b]pyridin-1-yl]acetic acids as highly potent and selective inhibitors of aldose reductase for treatment of chronic diabetic complications.

Efforts to identify treatments for chronic diabetic complications have resulted in the discovery of a novel series of highly potent and selective [3-(4,5,7-trifluoro-benzothiazol-2-ylmethyl)-pyrrolo[2,3-b]pyridin-1-yl]acetic acid aldose reductase inhibitors. The lead candidate, [6-methyl-3-(4,5,7-trifluoro-benzothiazol-2-ylmethyl)-pyrrolo[2,3-b]pyridin-1-yl]acetic acid example 16, inhibits aldose reductase with an IC50 of 8 nM, while being inactive against aldehyde reductase (IC50>100 microM), a related enzyme involved in the detoxification of reactive aldehydes.

Synthesis and PTP1B Inhibition of Novel 4-Aryl-1-Oxa-9-Thiacyclopenta[b]fluorenes

Novel 4-aryl-1-oxa-9-thiacyclopenta[b]fluorenes were designed, synthesized, and evaluated as inhibitors of the protein tyrosine phosphatase, PTP1B. Compounds 3 (IC50 = 284 nM) and 4 (IC50 = 74 nM), showed nanomolar potency against PTP1B (TRDI(P)YETD(P)Y(P)YRK as substrate). Compound 4 also lowered insulin in the diabetic ob/ob mouse at a dose of 10 mg/kg/day, p.o.

Vanadate as an oral antidiabetic agent

Todays need for effective oral glucose lowering agents and the recent advances in the understanding of the molecular mechanisms of insulin action have increased attention to the use of vanadate as an oral euglycaemic agent. Vanadate has metabolic actions like insulin. It is efficacious in rodent models of insulin-dependent diabetes (IDDM) and non-insulin-dependent diabetes (NIDDM). Moreover, recent clinical trials of two to three week duration in human NIDDM and IDDM subjects demonstrated that oral vanadate improved insulin sensitivity in all NIDDM subjects and some IDDM subjects. No major side effects were observed. Although vanadate is a potent protein-tyrosine phosphatase (PTPase) inhibitor, the precise glucose lowering mechanism is debatable since the metal is unstable and it is metabolised by cells. Recently chemists have taken advantage of the complex chemical properties of vanadate and have synthesised a variety of vanadium derivatives that are much more potent than vanadate. Some show in vitro en...