Monica A. Valentovic

Comparison of S-Adenosyl-l-methionine and N-Acetylcysteine Protective Effects on Acetaminophen Hepatic Toxicity

Nutraceuticals are widely used by the general public, but very little information is available regarding the effects of nutritional agents on drug toxicity. Excessive doses of acetaminophen (APAP, 4-hydroxyacetanilide) induce hepatic centrilobular necrosis. The naturally occurring substance S-adenosyl-l-methionine (SAMe) has been reported to reduce the hepatic toxicity of APAP. The present study was designed to investigate the hepatoprotective effects of SAMe in comparison to the clinically used antidote N-acetylcysteine (NAC). Male C57BL/6 mice were injected intraperitoneally (i.p.) with an equimolar dose (1.25 mmol/kg) of either SAMe or NAC just before APAP, and the groups were denoted SAMe+APAP and NAC+APAP, respectively. Mice were immediately injected i.p. with 300 mg/kg APAP, and hepatotoxicity was evaluated after 4 h. SAMe was more hepatoprotective than NAC at a dose of 1.25 mmol/kg as liver weight was unchanged by APAP injection in the SAMe+APAP group, whereas liver weight was increased in the NAC+APAP group. SAMe was more hepatoprotective for APAP toxicity than NAC, because alanine aminotransferase levels were lower in the SAMe+APAP. Pretreatment with SAMe maintained total hepatic glutathione (GSH) levels higher than NAC pretreatment before APAP, although total hepatic GSH levels were lower in the SAMe+APAP and NAC+APAP groups than the vehicle control values. Oxidative stress was less extensive in the SAMe+APAP group compared with the APAP-treated mice as indicated by Western blots for protein carbonyls and 4-hydroxynonenal-adducted proteins. In summary, SAMe reduced APAP toxicity and was more potent than NAC in reducing APAP hepatotoxicity.

Pediatric all-terrain vehicle trauma: a 5-year statewide experience.

BACKGROUND/PURPOSE This is a retrospective review of the pediatric all-terrain vehicle trauma victims who presented to the five major trauma centers serving the state of West Virginia during the 5-year period from January 1991 to December 1995. The purpose of this research is to characterize the nature of the injuries and the individuals injured to better appreciate the magnitude of the problem of ATV-related injuries in the pediatric population. METHODS This study is a retrospective review of these 218 consecutive pediatric patients from trauma registry data and their medical records. RESULTS Two hundred eighteen patients between the ages of 2 years and 16 years presented during the study period. Boys outnumbered girls three to one. The average Injury Severity Score (ISS) was 8.76, the average Glasgow Coma Score (GCS) was 14.4, and the average Trauma Score (TS) was 15.2. The most common injuries were orthopedic followed by head and facial injuries. The majority of the children did not wear helmets, and their injuries resulted in an average hospital length of stay of 4.3 days. Thirty-eight percent of the children required surgery. There were a total of four deaths for a mortality rate of 1.8%. The estimated total hospitalization cost for the 218 patients was

Resveratrol attenuates cisplatin renal cortical cytotoxicity by modifying oxidative stress

1,918,400.00. CONCLUSIONS All-terrain vehicle-related trauma remains an ongoing safety concern facing society today. Every physician who cares for children should address this important issue when talking to children and parents about safety issues and injury prevention.

Novel Protective Mechanisms for S-Adenosyl-L-methionine against Acetaminophen Hepatotoxicity: Improvement of Key Antioxidant Enzymatic Function

Cisplatin, a cancer chemotherapy drug, is nephrotoxic. The aim of this study was to investigate whether resveratrol (RES) reduced cisplatin cytotoxicity and oxidative stress. Rat renal cortical slices were pre-incubated 30min with 0 (VEH, ethanol) or 30μg/ml RES followed by 60, 90 or 120min co-incubation with 0, 75, or 150μg/ml cisplatin. Lactate dehydrogenase (LDH) leakage was unchanged at 60 and 90min by cisplatin. Cisplatin increased (p<0.05) LDH leakage at 120min which was protected by RES. Cisplatin induced oxidative stress prior to LDH leakage as cisplatin depressed glutathione peroxidase and superoxide dismutase (SOD) activity, increased lipid peroxidation, protein carbonyls and 4-hydroxynonenal (4-HNE) adducted proteins within 60min. RES failed to reverse glutathione (GSH) depression by cisplatin. In order to eliminated an extracellular interaction between RES and cisplatin, additional studies (RINSE studies) allowed a 30min RES uptake into slices, transfer of slices to buffer lacking RES, followed by 120min cisplatin incubation. RES in the RINSE studies prevented LDH leakage by cisplatin indicating that RES protection was not via a physical interaction with cisplatin in the media. These findings indicate that RES diminished cisplatin in vitro renal toxicity and prevented the development of oxidative stress.

Attenuation of Cisplatin Nephrotoxicity by Streptozotocin-Induced Diabetes

Acetaminophen (APAP) overdose leads to severe hepatotoxicity, increased oxidative stress and mitochondrial dysfunction. S-adenosyl-L-methionine (SAMe) protects against APAP toxicity at a mmol/kg equivalent dose to N-acetylcysteine (NAC). SAMe acts as a principle biological methyl donor and participates in polyamine synthesis which increase cell growth and has a role in mitochondrial protection. The purpose of the current study tested the hypothesis that SAMe protects against APAP toxicity by maintaining critical antioxidant enzymes and markers of oxidative stress. Male C57Bl/6 mice were treated with vehicle (Veh; water 15 ml/kg, ip), SAMe (1.25 mmol/kg, ip), APAP (250 mg/kg, ip), and SAMe+APAP (SAMe given 1 h following APAP). Liver was collected 2 and 4 h following APAP administration; mitochondrial swelling as well as hepatic catalase, glutathione peroxidase (GPx), glutathione reductase, and both Mn- and Cu/Zn-superoxide dismutase (SOD) enzyme activity were evaluated. Mitochondrial protein carbonyl, 3-nitrotyrosine cytochrome c leakage were analyzed by Western blot. SAMe significantly increased SOD, GPx, and glutathione reductase activity at 4 h following APAP overdose. SAMe greatly reduced markers of oxidative stress and cytochrome C leakage following APAP overdose. Our studies also demonstrate that a 1.25 mmol/kg dose of SAMe does not inhibit CYP 2E1 enzyme activity. The current study identifies a plausible mechanism for the decreased oxidative stress observed when SAMe is given following APAP.

Comparison of the in vitro toxicity of dichloroaniline structural isomers

The therapeutic use of cisplatin is associated with acute renal failure. The purpose of this study was to determine (a) if streptozotocin (STZ) was toxic to renal proximal tubules and (b) the nephrotoxicity of cisplatin in STZ-diabetic rats. Male Sprague-Dawley rats were injected with STZ (55 mg/kg, ip) to induce a diabetic state. BUN and renal cortical slice uptake of p-aminohippurate (PAH) and tetraethylammonium (TEA) were not altered, relative to normoglycemic rats, 3, 16, and 28 days following STZ treatment. These results indicate that STZ is not toxic to renal proximal tubules. Cisplatin nephrotoxicity studies were then conducted in STZ-diabetic and normoglycemic rats. Cisplatin nephrotoxicity was also evaluated in diabetic rats pretreated for 8 days with insulin. Diabetic and normoglycemic rats were administered 5 mg/kg cisplatin or water (ip). Increased kidney weight, BUN levels, glucosuria, and proteinuria were measured in normoglycemic rats 4 days after cisplatin administration. Renal cortical TEA and lactate-stimulated PAH uptake (p less than 0.05) were diminished in the normoglycemic rats 4 days after cisplatin injection. No change in kidney weight, BUN levels, or renal cortical slice accumulation of PAH and TEA was observed in diabetic rats treated with cisplatin. However, cisplatin administration to diabetic rats pretreated with insulin resulted in increased mortality, proteinuria, glucosuria and elevated kidney weight. These results indicate that the diabetic state attenuates cisplatin nephrotoxicity. Additionally, these results indicate that diabetes attenuation of cisplatin nephrotoxicity is dependent on the severity of the diabetic state.

3,4-Dicholoroaniline acute toxicity in male Fischer 344 rats

Acute exposure to certain dichloroaniline (DCA) isomers results in renal and hepatic toxicity in vivo. In the present study we examined whether dichloroaniline structural isomers were cytotoxic to liver and kidney slices in vitro and compared the toxicities of the different structural isomers. These studies were necessary in order to validate the use of an in vitro slice system for examination of the cellular mechanisms for toxicity. Renal cortical and hepatic slices were incubated for 90 min with 2,3-DCA, 2,4-DCA, 2,5-DCA, 2,6-DCA, 3,4-DCA or 3,5-DCA at a final concentration of 0-1 mm. Pyruvate-directed gluconeogenesis was measured following an additional 30-min incubation with 10 mM pyruvate. Cytotoxicity was also determined by measurement of lactate dehydrogenase (LDH) release 120 min after the addition of dichloroaniline isomers at a final concentration of 0, 0.5, 1 or 2 mM. Gluconeogenesis in renal cortical slices was inhibited by all of the isomers beginning at a concentration of 0.5 mM. Renal slice LDH leakage was elevated above control levels by 1-2 mM 3,4-DCA or 3,5-DCA. A final concentration of 2 mM was needed for 2,3-DCA, 2,4-DCA, 2,5-DCA or 2,6-DCA in order to detect a significant (P < 0.05) increase in renal slice LDH leakage. Hepatic slices incubated with 0.5-2 mM 2,3-DCA or 2 mM 2,5-DCA exhibited diminished pyruvate-directed gluconeogenesis. After exposure to 2,4-DCA, 2,6-DCA, 3,4-DCA or 3,5-DCA, pyruvate-directed gluconeogenesis was similar to that in the controls. LDH leakage was increased significantly (P < 0.05) above control values by exposure to 2 mM 3,4-DCA or 3,5-DCA. In conclusion, DCA structural isomers were toxic in vitro to liver and kidney slices. These results indicated that the kidney was more sensitive than the liver to DCA isomers, and that the most toxic isomer was 3,5-DCA. These results are similar to those previously observed in vivo.

Acute renal and hepatic toxicity of 2-haloanilines in Fischer 344 rats

The aromatic amine, 3,4-dichloroaniline (DCA) is an important intermediate in the chemical production of agricultural chemicals. A previous study had shown that nephrotoxicity was apparent 48 h after injection of 3,4-DCA. The purpose of this study was to examine the potential for 3,4-DCA to be toxic to the kidney, liver and urinary bladder 24 h after acute administration. Male Fischer 344 (F344) rats were injected (intraperitoneal (i.p.)) with 0.4, 0.8 or 1.0 mmol/kg 3,4-DCA hydrochloride (HCl) salt (2.5 ml/kg, 25% ethanol). Nephrotoxicity was apparent within 24 h in the 0.8 and 1.0 mmol/kg 3,4-DCA treated group and was characterized by elevated (P < 0.05) blood urea nitrogen (BUN) and kidney weight. Renal cortical slice accumulation ofp-aminohippurate (PAH) was also decreased in the 0.8 and 1.0 mmol/kg 3,4-DCA treated group relative to pair fed controls (PFC). Cellular changes were noted in the liver and bladder 24 h after 3,4-DCA administration. Plasma alanine transaminase (ALT) activity was elevated (P < 0.05) above PFC values 24 h after treatment with 0.8 or 1.0 mmol/kg indicating liver damage was apparent within 24 h. Morphological damage was apparent along the centrilobular region. Hematuria was observed in the 0.8 and 1.0 mmol/kg 3,4-DCA treated groups. Infiltration of erythrocytes and polymorphonuclear leukocytes was apparent within the urinary bladder upon examination by light microscopy. These results indicated that 3,4-DCA was toxic within 24 h and that the target tissues were the kidney, liver and urinary bladder. In vitro studies were conducted to compare the toxicity of two forms of 3,4-DCA, the free base and hydrochloride salt to determine whether chemical form contributes to renal cortical slice toxicity. Lactate dehydrogenase (LDH) release was elevated above control by 120 min exposure to 2 mM 3,4-DCA free base or hydrochloride salt. Pyruvate directed gluconeogenesis in renal slices was decreased relative to control by 0.5 mM 3,4-DCA free base and hydrochloride salt. The results from the in vitro studies indicates that the chemical form did not modify in vitro renal cortical slice toxicity.

Characterization of myoglobin toxicity in renal cortical slices from Fischer 344 rats.

Aniline and its halogenated derivatives are widely used as chemical intermediates. The purpose of this study was to determine the hepatotoxic and nephrotoxic potential of the 2-haloanilines. Male Fischer 344 rats (n > or = 4) were injected (i.p.) with 1.0 or 1.25 mmol/kg of: aniline (A), 2-fluoroaniline (2-FA), 2-chloroaniline (2-ClA), 2-bromoaniline (2-BrA), 2-iodoaniline (2-IA) or vehicle (0.9% saline, 2.5 ml/kg). All compounds were injected as hydrochloride salts. Renal and hepatic function was monitored 24 h after treatment. All of the 2-haloanilines induced oliguria, diminished kidney weight, tubular casts and decreased renal cortical slice accumulation of organic anions. Blood urea nitrogen (BUN) levels were increased (P < 0.05) by treatment with 1.0 or 1.25 mmol/kg of 2-FA, 2-ClA or 2-BrA. Hepatic alterations were also observed and characterized by elevated plasma ALT/GPT activity and altered morphology in the centrilobular region. The nephrotoxic and hepatotoxic potentials were similar among the 2-haloanilines but aniline was less toxic than its 2-halo derivatives. These results demonstrated that halogen substitution at the 2-position of aniline increased hepatic and renal toxicity. However, the severity of toxicity was not influenced by the nature of the halogen substituent.

Potentiation of 5-hydroxytryptamine-induced contraction in rat aorta by chlorpheniramine, citalopram and fluoxetine

Rhabdomyolysis is associated with acute renal failure. The following study first characterized myoglobin in vitro toxicity using renal cortical slices isolated from male Fischer 344 rats. This model provided interaction between various cells within the nephron and provides myoglobin access predominantly through the basolateral membrane. Second, this study examined the effect of deferoxamine (DFX) and glutathione on myoglobin toxicity to determine the role of radicals and iron. Renal cortical slices were incubated for 30-120 min with 0, 4, 10 or 12 mg/ml myoglobin. Myoglobin was pretreated with 4 mM ascorbic acid prior to addition to the slices to ensure that myoglobin was in its reduced state. In other experiments tissues were pretreated for 15 min with 0.1 mM of the iron chelator DFX or 30 min with 1 mM glutathione prior to co-incubation with myoglobin. Finally, slices were pretreated with 1 mM glutathione for 30 min, rinsed and incubated only with myoglobin. Early event changes occurred within a 60 min exposure and included a decline in pyruvate-stimulated gluconeogenesis, increased lipid peroxidation levels and decreased glutathione levels. Loss of ATP levels and increased lactate dehydrogenase (LDH) release required a 120 min exposure to myoglobin. DFX reduced myoglobin induced effects on LDH leakage but had no effect on gluconeogenesis suggesting that myoglobin toxicity had an iron dependent (LDH) and independent (gluconeogenesis) pathway. Pretreatment with glutathione provided complete protection and was mediated by intracellular events.