Assessment of In Vitro Tests as Predictors of the Antioxidant Effects of Insulin, Metformin, and Taurine in the Brain of Diabetic Rats.

Hyperglycemia-induced oxidative stress is an intrinsic feature of diabetes mellitus and a recognized causative factor of complications associated with the disease. As a result, compounds possessing antioxidant properties are commonly investigated as possible ways of minimizing and even preventing diabetes-related oxidative stress. On these premises, the present study was carried out to investigate the antioxidant properties of metformin (MET), a common oral hypoglycemic agent, of taurine (TAU), a sulfonic acid compound with known antioxidant benefits in diabetes, and of insulin (INS), a standard antidiabetic serving as a reference compound, by using in vitro and in vivo tests. A battery of seven in vitro tests was used to assess antioxidant/antiradical activity. The addition of a treatment compound led to a mean percentage decrease of values for free radical/lipid peroxidation (LPO) that ranged from very high (82%) with INS to moderate (43%) with MET) and to low (31%) with TAU. Combining MET with TAU leads to an improvement of the effect seen with MET alone (46%). By contrast, under the same conditions, N-acetylcysteine, a known antioxidant, was more potent (92%) than any of the test compounds. In vivo studies were conducted using rats made diabetic with streptozotocin and treated with daily doses of INS, MET, TAU, and MET-TAU for 6 weeks. Among the test compounds, the greatest hypoglycemic effect was attained with INS (>90% decrease), followed by MET (~70% decrease), with TAU providing only a modest effect (-30% decrease). Unexpectedly, however, all three compounds reduced the diabetic values for brain LPO, nitric oxide, antioxidant enzymes, glutathione, and glutathione-related enzymes to values that varied in extent within a narrow range (<12% from one another). On the other hand, pairing MET with TAU led to a small enhancement (<10%) of the effects seen with MET alone. In short, while in vitro tests for antioxidant/antiradical activity suggest marked differences in potency for INS, MET, and TAU as a result of different structures, changes in the values of indices of oxidative stress affected by these compounds in the brain of diabetic rats varied within a rather narrow range. Also, the present results suggest that although hyperglycemia is an important determinant of the oxidative stress of diabetes, other factors may be involved since a weak hypoglycemic like TAU demonstrated in vivo antioxidant actions that were comparable to those of more potent hypoglycemic agents like INS and MET.

Taurine Enhances the Protective Actions of Fish Oil Against D-Galactosamine-Induced Metabolic Changes and Hepatic Lipid Accumulation and Injury in the Rat.

This study has evaluated the effects of a supplementation with taurine (TAU) on the actions of fish oil (FO) against the hypoglycemia, hypoproteinemia, and hepatic accumulation of lipids and liver damage caused by D-galactosamine (GAL) in the rat. To this end, male Sprague-Dawley rats (200-225 g), in groups of 6, were orally treated with physiological saline (2.5 mL, control group), FO (60 mg/kg), TAU (2.4 mmol/kg) or FO-TAU for three consecutive days and before a single oral dose of GAL (400 mg/kg) given on day 3. In parallel, rats receiving only GAL on day 3 or N-acetylcysteine (NAC, 2.4 mmol/kg) for 3 days before GAL served as controls. On day 4 blood samples were collected by cardiac puncture and used to either measure glucose (GLC) or to obtain plasma fractions. Immediately thereafter, the livers were excised, made into a homogenate in phosphate buffered saline pH 7.4, and centrifuged to obtain clear supernatant. Plasma samples were assayed for their total protein (TP), triglycerides (TG), cholesterol (CHOL), phospholipids (PLP), free fatty acids (FFA) and total bilirubin (TB) and direct bilirubin (DB) contents, and for the activities of alanine transaminase (ALT), aspartate transaminase (AST) and alkaline phosphatase (ALP). The liver homogenates were used to measure TG, CHOL, PLP and total lipids (TL) contents. Without exceptions, GAL was found to markedly affect (p < 0.001) all of the experimental parameters examined, with increases occurring in all instances except for the values of the plasma GLC, TP and PLP which were decreased. A pretreatment with either FO or TAU led to significant attenuation of the effects of GAL and which, in most cases, were of similar magnitude. On the other hand, a combined pretreatment with FO plus TAU usually resulted in a greater protection than with either agent alone (p ≤ 0.05). NAC, serving as a reference treatment, was, in most instances, equipotent with FO alone and. in addition, was the only agent that significantly attenuated the increases in both liver weight and liver weight to body weight ratio caused by GAL.

Taurine Improves the Actions of Metformin and Lovastatin on Plasma Markers of Carbohydrate and Lipid Dysfunction of Diabetic Rats.

The present study has investigated the effect of adding taurine (TAU) to a treatment of diabetes with metformin (MET), a hypoglycemic, and lovastatin (LOV), an antihyperlipidemic. To this end, male Sprague-Dawley rats, agent, 250-275 g in weight, were made diabetic with a single 60 mg/kg intraperitoneal (i.p.) dose of streptozocin (STZ) in 10 mM citrate buffer pH 4.5, and, after 14 days, treated daily with oral doses of MET (2.4 mM/kg), LOV (0.075 mM/kg) or TAU (2.4 mM/kg), and with binary and ternary combinations of these agents. Rats receiving only 10 mM citrate buffer pH 4.5 or only STZ served as negative and positive controls, respectively. In addition, rats receiving insulin (INS, 4 units/kg) by the subcutaneous route served as a reference treatment. All the rats were sacrificed on day 57 and their bloods collected into heparinized tubes. The corresponding plasma samples were analyzed for their glucose (GLC), insulin (INS), glycated hemoglobin (HbA1c), cholesterol (CHOL) and triglycerides (TG) contents. In comparison to normal rats, diabetic ones showed marked increases in GLC (+313%), HbA1c (+207%), CHOL (+66%) and TG (+188) and a profound decrease of INS levels (-76%) (p < 0.001 vs. control values). Among the various treatments, one with INS produced the greatest lowering effect on the plasm a GLC (+23%, p < 0.05), INS (+23%, p < 0.05) and TG (+3%), with the remaining changes being similar to those seen with MET. A treatment with MET reduced all the diabetic changes by at least threefold; and one with LOV had a significant (p < 0.001) lowering effect on the plasma CHOL and TG but was without an effect on the plasma GLC, INS and HbA1c. In common with LOV, TAU reduced the diabetic levels of both CHOL and TG and, in addition, reduced the diabetic plasma GLC and raised the corresponding INS level. Among binary combinations, one with LOV-MET provided a greater effect than MET alone only in terms of the plasma CHOL and TG; and one with LOV-TAU was only significantly better than TAU alone in lowering the TG levels. However, a treatment with LOV-MET-TAU led to reductions in all the plasma parameters examined that were much greater than those achieved with any of the individual agents or with their binary combinations (at p ≤ 0.05).

Modification by Ethanol and Taurine, Singly and in Combination, of Changes in Indices of Renal Dysfunction Caused by Diabetes in Rats.

The present study was carried out in diabetic rats to examine the effects of ethanol (EtOH) and taurine (TAU), singly and in combination, in reducing the changes of laboratory test values indicating renal dysfunction. For this purpose, male Sprague-Dawley rats, 250-280 g in weight and in groups of 6, were made diabetic with a single, 60 mg/kg intraperitoneal dose of streptozotocin in 10 mM citrate buffer pH 4.5. On day 15 and for the remaining 14 days of the study, the diabetic rats (a) started to drink 5% EtOH in place of water, (b) received a single daily 2.4 mM/kg oral dose of TAU or (c) were allowed to drink 5% EtOH after receiving a dose of TAU. Starting from day 28 and ending on day 29, a 24 h urine sample was collected, its volume was measured, and then used to measure glucose (GLC), total protein (TP) and electrolytes (Na+, K+, Ca++, Mg++). Blood samples collected immediately thereafter via cardiac puncture were processed for the plasma fractions which were analyzed for their creatinine (CRT) and urea nitrogen (UN) contents. In comparison to normal (control) rats, diabetic ones showed a higher output of urine (+5.6-fold), a massive increase in plasma GLC (+473%), passed more GLC (+73.8-fold) and TP (+8.2-fold) in the urine, showed higher plasma CRT (+241%) and UN (+74%) levels, a lower plasma UN/CRT ratio (-47%) and a greater output of electrolytes in the urine (by at least twofold). By themselves both EtOH and TAU were found to markedly lower the effects of diabetes, with EtOH generally appearing more effective than TAU. However, the concurrent availability of EtOH and TAU was found to be more protective than either treatment alone.

The Effect of Taurine and Its Immediate Homologs on Diabetes-Induced Oxidative Stress in the Brain and Spinal Cord of Rats.

This study has examined the acute effects of taurine (TAU) and of its two immediate homologs aminomethanesulfonic acid (AMSA) and homotaurine (HTAU) on the oxidative stress that develops in the brain of rats as a result of type 2 diabetes mellitus. Male Sprague-Dawley rats, 220-225 g in weight, were divided into groups of 6 each, and treated with a single intraperitoneal (i.p.) dose of streptozotocin (STZ) in 10 mM citrate buffer pH 4.5 (60 mg/kg). The treatment compound (AMSA, HTAU or TAU) was administered by the i.p. route in two equal doses (1.2 mM/kg each) at 75 and 45 min before STZ. Control rats received only 10 mM citrate buffer pH 4.5 or only STZ by the i.p. route. The rats were sacrificed at 24 h after a dosing with STZ under general anesthesia, and their brains and spinal cords collected by the freeze clamp technique. A portion of brain, of a brain area (cerebellum, cortex, brain stem) or of spinal cord from each animal was extracted into 0.1 M PBS pH 7.4, and the extract was used for the assay of malondialdehyde (MDA), nitric oxide (NO), catalase (CAT), glutathione peroxidase (GPx) and superoxide dismutase (SOD). An extract for the assay of the reduced (GSH) and disulfide (GGSG) forms of glutathione was prepared in similar manner but using 2% metaphosphoric acid plus 0.1 M PBS pH 8.0 as the extracting medium. Diabetes was found to markedly increase the formation of MDA (by 160-202%), NO (by 138-313%) and GSSG (by 103-241%), and to lower the values of GSH (by 57-65%), GSH/GSSG ratio (79-89%) and activities of CAT (by 61-69%), GPx (by 52-66%) and SOD (by 55-68%) in the brain, brain areas and spinal cord relative to corresponding control values (all at p < 0.001). These effects were reduced to values that were generally at least one-half of those seen in untreated diabetic rats, with TAU providing a greater attenuation of the formation of MDA and NO, an about similar action on the depletion of GSH, and a lower action on the decrease in the GSH/GSSG ratio caused by diabetes than either AMSA or HMTAU. In contrast AMSA and HMTAU were about equipotent with each other and more potent than TAU in preventing the loss of antioxidant enzyme activities associated with diabetes. In short, pretreating diabetic rats with AMSA, HMTAU or TAU is found to protect the brain against changes in biochemical parameters indicative of oxidative stress, with potency differences among the test compounds varying within a narrow range.

Investigation of the Role of a Supplementation with Taurine on the Effects of Hypoglycemic-Hypotensive Therapy Against Diabetes-Induced Nephrotoxicity in Rats.

This study has examined the role of supplementing a treatment of diabetic rats with captopril (CAP), metformin (MET) or CAP-MET with the antioxidant amino acid taurine (TAU) on biochemical indices of diabetes-induced metabolic changes, oxidative stress and nephropathy. To this end, groups of 6 male Sprague-Dawley rats (250-375 g) were made diabetic with a single, 60 mg/kg, intraperitoneal dose of streptozotocin (STZ) in 10 mM citrate buffer pH 4.5 and, after 14 days, treated daily for up to 42 days with either a single oral dose of CAP (0.15 mM/kg), MET (2.4 mM/kg) or TAU (2.4 mM/kg), or with a binary or tertiary combination of these agents. Rats receiving only 10 mM citrate buffer pH 4.5 or only STZ served as negative and positive controls, respectively. All rats were sacrificed by decapitation on day 57 and their blood and kidneys collected. In addition, a 24 h urine sample was collected starting on day 56. Compared to normal rats, untreated diabetic ones exhibited frank hyperglycemia (+313%), hypoinsulinemia (-76%) and elevation of the glycated hemoglobin value (HbA1c, +207%). Also they showed increased plasma levels of Na+ (+35%), K+ (+56%), creatinine (+232%), urea nitrogen (+158%), total protein (-53%) and transforming growth factor-ß1 (TGF-ß1, 12.4-fold) values. These changes were accompanied by increases in the renal levels of malondialdehyde (MDA, +42%), by decreases in the renal glutathione redox state (-71%), and activities of catalase (-70%), glutathione peroxidase (-71%) and superoxide dismutase (-85%), and by moderate decreases of the urine Na+ (-33%) and K+ (-39%) values. Following monotherapy, MET generally showed a greater attenuating effect than CAP or TAU on the changes in circulating glucose, insulin and HbA1c levels, urine total protein, and renal SOD activity; and CAP appeared more potent than TAU and MET, in that order, in antagonizing the changes in plasma creatinine and urea nitrogen levels. On the other hand, TAU generally provided a greater protection against changes in glutathione redox state and in CAT and GPx activities, with other actions falling in potency between those of CAP and MET. Adding TAU to a treatment with CAP, but not to one with MET, led to an increase in protective action relative to a treatment with drug alone. On the other hand, the actions of CAP-MET, which were about equipotent with those of MET, became enhanced in the presence of TAU, particularly against the changes of the glutathione redox state and activities of antioxidant enzymes. In short, the present results suggest that the addition of TAU to a treatment of diabetes with CAP or CAP-MET, and sometimes to one with MET, will lead to a gain in protective potency against changes in indices of glucose metabolism and of renal functional impairment and oxidative stress.

The Effect of Metformin and Taurine, Alone and in Combination, on the Oxidative Stress Caused by Diabetes in the Rat Brain.

This study has compared the effects of metformin (MET) and taurine (TAU), singly and in combination, on the oxidative stress caused by diabetes in the rat brain. For this purpose, male Sprague-Dawley rats, 200-225 g in weight, assigned to groups of 6, were intraperitoneally (i.p.) treated with the diabetogen streptozotocin (STZ, 60 mg/kg, in citrate buffer pH 4.5) on day 1 and, after 14 days, orally (p.o.) with either MET, TAU or MET-TAU (each at 2.4 mM/kg, in water). Control rats received only citrate buffer pH 4.5 (2 mL) or only STZ on day 1 by the i.p. route. All the animals were sacrificed by decapitation on day 57 and their brains collected by the freeze clamp technique. Blood samples were placed in heparinized tubes and used for the assay of the plasma glucose (GLC) and blood insulin (INS) levels. Immediately thereafter, the brains were surgically removed and a portion was used to prepare a homogenate in 0.1 M PBS pH 7.4, which was used for the assay of indices of oxidative stress. Diabetes raised the plasma GLC level (+313%) but lowered that of the blood INS (-76%) compared to corresponding values from nondiabetic rats. In addition it raised the brain malondialdehyde level (+59%) but lowered the reduced/disulfide glutathione ratio (-46%), and activities of catalase (-43%), glutathione peroxidase (-48%), superoxide dismutase (-65%), glutathione reductase (-50%) and glutathione S-transferase (-51%) significantly (all at p < 0.001). Except for the greater decrease in GLC (+90% vs. +22%) and increase in INS (-26% vs. -52%) levels seen in rats receiving MET than in rats receiving TAU, these compounds protected the brain against oxidative stress to significant (p ≤ 0.05%) and rather similar extents. Furthermore, the concurrent administration of MET and TAU to the diabetic rats led to brain values of indices of oxidative stress that were lower than those attained with MET alone, although generally not to a statistically significant degree.

Comparative evaluation of the effects of taurine and thiotaurine on alterations of the cellular redox status and activities of antioxidant and glutathione-related enzymes by acetaminophen in the rat.

The present study was carried out to ascertain the impact of replacing the sulfonate group of TAU with thiosulfonate, as present in thiotaurine (TTAU), on the protective actions of TAU against hepatocellular damage and biochemical alterations related to oxidative stress and glutathione redox cycling, synthesis, and utilization caused by a high dose of acetaminophen (APAP). To this end, male Sprague-Dawley rats, 225-250 g, were intraperitoneally treated with a 2.4 mmol/kg dose of TAU (or TTAU), followed 30 min later by 800 mg/kg of APAP. A reference group received 2.4 mmol/kg of N-acetylcysteine (NAC) prior to APAP. Naive rats served as controls. The animals were sacrificed 6 h after receiving APAP and their blood and livers were collected. Plasma and liver homogenates were analyzed for indices of cell damage (plasma transaminases, plasma lactate dehydrogenase), -oxidative stress (malondialdehyde = MDA, reduced glutathione = GSH, glutathione disulfide = GSSG, catalase, glutathione peroxidase, superoxide dismutase), glutathione cycling (glutathione reductase), utilization (glutathione S-transferase), and synthesis (γ-glutamylcysteine synthetase) activities. APAP increased MDA formation and lowered the GSH/GSSG ratio and all enzyme activities, especially those of antioxidant enzymes. In general, TTAU was equipotent with NAC and more potent than TAU in protecting the liver. Taken into account the results of a previous study comparing the actions of TAU and hypotaurine (HTAU), the sulfinate analog of TAU, it appears that the sulfinate and thiosulfonate analogs are somewhat more effective than the parent sulfonate TAU in counteracting APAP-induced hepatic alterations in the liver and plasma.

Protection by taurine and thiotaurine against biochemical and cellular alterations induced by diabetes in a rat model.

In this study, the actions of taurine (TAU), a sulfonate, and thiotaurine (TTAU), a thiosulfonate, on diabetes-mediated biochemical alterations in red blood cells (RBCs) and plasma and on the RBC membrane, morphology and spectrin distribution were examined in rats. Diabetes was induced in male Sprague-Dawley rats with streptozotocin (60 mg/kg i.p.) and allowed to progress for 14 days. From days to 56, the rats received a daily, 2.4 mmol/kg, oral dose of TAU or TTAU, 2 mL oral dose of physiological saline or 4 U/kg subcutaneous dose of isophane insulin (INS). Naive rats served as the control group. The rats were sacrificed on day 57 and their blood was collected to measure HbA(1c), to isolate intact RBCs, and to obtain plasma. A 6-weeks treatment with INS effectively lowered the elevations in plasma glucose, cholesterol, triglycerides, and plasma and RBC malondialdehyde and glutathione disulfide while effectively counteracting the decreases in plasma INS, plasma and RBC glutathione redox status, and plasma and RBC activities of antioxidant enzymes caused by diabetes. Also, INS returned the echynocytic appearance and peripheral location of spectrin seen in RBCs from diabetic rats to the normal discocytic shape and uniform distribution. TAU and TTAU were as effective as INS in inhibiting malondialdehyde formation, changes in redox status and oxidative stress in both the plasma and RBC, but were much less effective in controlling hyperglycemia and hypoinsulinemia. Furthermore TTAU was more effective than INS or TAU in lowering the increase in cholesterol to phospholipids ratio in the RBC membrane and, unlike TAU, it was able to normalize the RBC morphology and spectrin distribution.

The effects of taurine and thiotaurine on oxidative stress in the aorta and heart of diabetic rats.

This study has compared the actions of the sulfur-containing compounds taurine (TAU) and thiotaurine (TTAU) with those of insulin (INS) on the oxidative stress that develops in the aorta and heart as a result of diabetes. Diabetes was induced in male Sprague-Dawley rats with streptozotocin (60 mg/kg, i.p.). Starting on day 15, and continuing for the next 41 days, the diabetic rats received each day 2 mL of physiological saline or 2.4 mmol/kg/2 mL of TAU (or TTAU) p.o. or 4 U/kg of isophane INS s.c. Normal rats served as controls. The rats were sacrificed on day 57 to collect blood, heart and thoracic aorta samples. Untreated diabetic rats exhibited a lower body weight gain (by 34%), higher than normal plasma glucose (by ∼4-fold), cholesterol (by 66%) and triglycerides (by 188%) levels, and lower INS levels (by 76%). Also there was a marked increase in catalase activity (≥90%); and clear decreases in nitrite (≥40%), glutathione redox status (≥67%), and glutathione peroxidase (≥66%) and superoxide dismutase (≥51%) activities in both the aorta and heart. With only a few isolated instances (plasma lipids), TTAU was either markedly more effective (plasma glucose, plasma INS, aorta and heart glutathione, aorta redox status, and antioxidant enzymes) or marginally more effective (heart redox status) than TAU in attenuating the alterations brought about by diabetes. These results suggest that replacing the sulfonic acid group of TAU by thiosulfonic acid can lead to a greater potency against diabetes-related biochemical changes in the plasma, heart and aorta. However, except for effects on plasma lipids, these sulfur-containing compounds were less effective than INS in counteracting diabetes-related changes.

Comparative evaluation of taurine and thiotaurine as protectants against diabetes-induced nephropathy in a rat model.

Taking into account the proven effectiveness of antioxidants in preventing experimentally induced diabetes in laboratory animals, this study was carried out with the specific purpose of comparing the effectiveness of two known antioxidants, the ß-aminosulfonate taurine (TAU) and ß-aminothiosulfonate thiotaurine (TTAU), in preventing biochemical, functional and histological alterations indicative of -diabetic nephropathy. In the study, streptozotocin (60 mg/kg, orally) was used to induce type 2 diabetes mellitus in Sprague-Dawley rats. Starting on day 15 and continuing up to day 56, the rats received a daily single 2.4 mmol/kg oral dose of a sulfur-containing compound (TAU or TTAU) or 4 U/kg subcutaneous dose of isophane insulin (INS). Rats not receiving any treatment served as controls. After obtaining a 24 h urine sample, the animals were sacrificed by decapitation on day 57, and their blood and kidneys immediately collected. Diabetic rats exhibited marked hyperglycemia, hypoinsulinemia, hypoproteinemia, hyponatremia, hyperkalemia, azotemia, hypercreatinemia, increased plasma TGF ß(1), lipid peroxidation, plasma and kidney nitrite, and urine output; decreased glutathione redox status in plasma and kidney, decreased urine Na(+) and K(+), proteinuria and hypocreatinuria. Without exceptions, all the treatment compounds were found to markedly and variously attenuate these changes. Confirmation of protection by INS, TAU and TTAU was provided by the results of histological examination of kidney sections and which showed a more normal appearance than sections from diabetic animals. In most instances protection by TTAU was about equal to that by INS but greater than that by TAU.

Simple reversed-phase HPLC method with spectrophotometric detection for measuring acetaminophen-protein adducts in rat liver samples.

A simple reversed-phase HPLC method for measuring hepatic levels of acetaminophen- (APAP-) protein adduct following an overdose of APAP was developed. An aliquot of liver homogenate in phosphate-buffered saline pH 7.4 (PBS) was placed on a Nanosep centrifugal device, which was centrifuged to obtain a protein residue. This residue was incubated with a solution of p-aminobenzoic acid (PABA), the internal standard, and bacterial protease in PBS, transferred to a Nanosep centrifugal device, and centrifuged. A 100 µL portion of the filtrate was analyzed on a YMC-Pack ODS-AMQ C18 column, using 100 mM potassium dihydrogen phosphate-methanol-acetic acid (100 : 0.6 : 0.1) as the mobile phase, a flow rate of 1 mL/min, and photometric detection at 254 nm. PABA and APAP-cystein-S-yl (APAP-Cys) eluted at ~14.7 min and 22.7 min, respectively. Method linearity, based on on-column concentrations of APAP-Cys, was observed over the range 0.078-40 µg. Recoveries of APAP-Cys from spiked blank liver homogenates ranged from ~83% to 91%. Limits of detection and of quantification of APAP-Cys, based on column concentrations, were 0.06 µg and 0.14 µg, respectively. RSD values for interday and intraday analyses of a blank liver homogenate spiked with APAP-Cyst at three levels were, in all cases, ≤1.0% and <1.5%, respectively. The proposed method was found appropriate for comparing the antidotal properties of N-acetylcysteine and taurine in a rat model of APAP poisoning.

Comparison of the protective actions of N-acetylcysteine, hypotaurine and taurine against acetaminophen-induced hepatotoxicity in the rat.

When used in overdoses, acetaminophen (APAP) is a common cause of morbidity and mortality in humans. At present, N-acetylcysteine (NAC) is the antidote of choice for acetaminophen overdoses. Prompt administration of NAC can prevent the deleterious actions of APAP in the liver. In view of the similarities in antioxidant effects demonstrated by NAC, hypotaurine (HYTAU) and taurine (TAU) in this and other our laboratories, the present study was undertaken to compare these compounds for the ability to attenuate plasma and liver biochemical changes associated with a toxic dose of APAP. For this purpose, fasted male Sprague-Dawley rats, 225-250 g in weight, were intraperitoneally treated with APAP (800 mg/kg), NAC, HYTAU or TAU (2.4 mM/kg) followed 30 min later by APAP, or 50% PEG 400 (the vehicle for APAP). At 6 hr after APAP administration, all animals were sacrificed by decapitation and their blood and livers collected. The plasma fractions were analyzed for indices of liver damage (alanine transaminase, aspartate transaminase, lactate dehydrogenase), levels of malondialdehyde (MDA), reduced (GSH) and oxidized (GSSG) glutathione, and activities of glutathione reductase (GR), glutathione S-transferase (GST) and gamma-glutamylcisteinyl synthetase (GCS). Suitable liver homogenates were analyzed for the same biochemical parameters as the plasma but indices of liver damage. By itself, APAP increased MDA formation and had a significant lowering influence on the levels of GSH and GSSG, the GSH/GSSH ratio, and the activities of GR, GST and GCS both in the plasma and liver. In addition, APAP promoted the leakage of transaminases and lactate dehydrogenase from the liver into the plasma. Without exceptions, a pretreatment with a sulfur-containing compound led to a significant attenuation of the liver injury and the biochemical changes induced by APAP. Within a narrow range of potency differences, HYTAU appeared to be the most protective and TAU the least. The present results suggest that, irrespective of the differences in structural features and in vitro antioxidant properties that may exist among NAC, TAU and HYTAU, these compounds demonstrate equivalent patterns of protection and, to a certain extent, equipotent protective actions against the toxic actions of APAP in the liver when tested in equimolar doses and under the same conditions in an animal model.

Comparative study of the binding characteristics to and inhibitory potencies towards PARP and in vivo antidiabetogenic potencies of taurine, 3-aminobenzamide and nicotinamide.

BACKGROUND: Poly(ADP-ribose) is a NAD+-requiring, DNA-repairing, enzyme playing a central role in pancreatic beta-cell death and in the development of endothelial dysfunction in humans and experimental animals. PARP activation is also relevant to the development of complications of diabetes. Hence, agents capable of inhibiting PARP may be useful in preventing the development of diabetes and in slowing down complications of diabetes. METHODS: PARP inhibition was assessed with a colorimetric assay kit. Molecular docking studies on the active site of PARP were conducted using the crystalline structure of the enzyme available as Protein Data Bank Identification No. 1UK1. Type 2 diabetes was induced in male Sprague-Dawley rats with streptozotocin (STZ, 60 mg/kg, i.p.). The test compounds (3-aminobenzamide = 3-AB, nicotinamide = NIC, taurine = TAU) were given by the i.p. route 45 min before STZ at 2.4 mM/kg (all three compounds) or 1.2 and 3.6 mM/kg (only NIC and TAU). Blood samples were collected at 24 hr after STZ and processed for their plasma. The plasma samples were used to measure glucose, insulin, cholesterol, triglycerides, malondialdehyde, nitric oxide, and glutathione levels using reported methods. RESULTS: 3-AB, NIC and TAU were able to inhibit PARP, with the inhibitory potency order being 3-AB>NIC> or =TAU. Molecular docking studies at the active site of PARP showed 3-AB and NIC to interact with the binding site for the nicotinamide moiety of NAD+ and TAU to interact with the binding site for the adenine moiety of NAD+. While STZ-induced diabetes elevated all the experimental parameters examined and lowered the insulin output, a pretreatment with 3-AB, NIC or TAU reversed these trends to a significant extent. At a dose of 2.4 mm/kg, the protective effect decreased in the approximate order 3-AB>NIC> or =TAU. The attenuating actions of both NIC and TAU were dose-related except for the plasma lipids since NIC was without a significant effect at all doses tested. CONCLUSIONS: At equal molar doses, 3-AB was generally more potent than either TAU or NIC as an antidiabetogenic agent, but the differences were not as dramatic as would have been predicted from their differences in PARP inhibitory potencies. NIC and TAU demonstrated dose-related effects, which in the case of TAU were only evident at doses > or =2.4 mM/kg. The present results also suggest that in the case of NIC and TAU an increase in dose will enhance the magnitude of their attenuating actions on diabetes-related biochemical alterations to that achieved with a stronger PARP inhibitor such as 3-AB. Hence, dosing will play a critical role in clinical studies assessing the merits of NIC and TAU as diabetes-preventing agents.

Protective action of taurine, given as a pretreatment or as a posttreatment, against endotoxin-induced acute lung inflammation in hamsters.

To assess the effect of taurine on lipopolysaccharide (LPS)-induced lung inflammation, oxidative stress and apoptosis, female Golden Syrian hamsters were intratracheally instilled with bacterial LPS (0.02 mg in phosphate buffered saline (PBS) pH 7.4), before or after a 3-day intraperitoneal treatment with a single dose of taurine (50 mg/kg/day in PBS pH 7.4), and bronchoalveolar lavage fluid (BALF) and lung tissue samples were collected at 24 hr after the last treatment. In comparison to BALF samples from animals receiving only PBS pH 7.4, and serving as controls, those of LPS-stimulated animals exhibited a higher count of both total leukocytes and neutrophils and increased expression of tumor necrosis factor receptor 1. In comparison to lungs from control animals, those from LPS-treated animals showed increased cellular apoptosis, lipid peroxidation, decreased glutathione levels, altered activities of antioxidant enzymes (catalase, glutathione peroxidase, superoxide dismutase) and focal inflammation confined to the parenchyma. A treatment with taurine was found to significantly attenuate all these alterations, with the protection being, in all instances, greater when given before rather than after LPS. The present results suggest that taurine is endowed with antiinflammatory and antioxidant properties that are protective in the lung against the deleterious actions of Gram negative bacterial endotoxin.

Attenuating effect of taurine on lipopolysaccharide-induced acute lung injury in hamsters.

This study has evaluated the ability of the semiessential amino acid taurine to attenuate lipopolysaccharide (LPS)-induced lung inflammation, oxidative stress and apoptosis in a small animal model. For this purpose, bacterial LPS (0.02mg in phosphate buffered saline (PBS) pH 7.4) was instilled intratracheally into female Golden Syrian hamsters, before or after a 3-day intraperitoneal treatment with a single dose (50mg/kg in PBS pH 7.4) of taurine. At 24h after the last treatment, lung tissue and bronchoalveolar lavage fluid (BALF) samples were collected. In comparison to samples from animals receiving only PBS pH 7.4, serving as controls, those of LPS-stimulated animals exhibited a higher count of both total leukocytes and neutrophils in the BALF, and increased incidence of apoptosis, depletion of intracellular glutathione and evidence of inflammation confined to the parenchyma in the lung. In addition, LPS caused cells in the BALF to exhibit a higher expression of tumor necrosis factor-1, a higher activity of caspase-3, marked lipid peroxidation, and altered activities of catalase, glutathione peroxidase and superoxide dismutase relative to control samples. In contrast, a treatment with taurine was found to significantly attenuate all of the cellular and biochemical alterations induced by LPS, more so when given before rather than after the endotoxin. The present results suggest that taurine possesses intrinsic antiinflammatory and antioxidant properties that may be of benefit against the deleterious actions of LPS in the lung.