Sorbitol Dehydrogenase Levels Research Articles

Paeonol attenuates retinopathy in streptozotocin-induced diabetes in rats by regulating the oxidative stress and polyol pathway.

The current research work was planned to study the effects of paeonol in the management of diabetic retinopathy. Diabetes was induced in male Sprague Dawley rats using Streptozotocin (55 mg/kg, i.p.). After 4 weeks, the diabetic animals were treated with paeonol at a dose of 50, 100, and 200 mg/kg body weight daily for the next 4 weeks. At the end of treatment, retinal physiology was studied by recording an electroretinogram (ERG); biochemical parameters and oxidative stress were estimated. The histopathology of the retina was also carried out at the end of the study. The ERG of paeonol-treated animals showed a significant improvement in a-wave amplitude, b-wave amplitude, a-wave latency, and b-wave latency (p < 0.001) at 15 cd s/m2 when compared with the diabetic control animals. The paeonol treatment (200 mg/kg) in diabetic animals showed a significant decrease in the plasma glucose level (p < 0.001) when compared to the animals in diabetic control group. Paeonol also significantly decreased the lactate dehydrogenase, aldose reductase, and sorbitol dehydrogenase levels when compared with the diabetic control animals. The oxidative stress in the eye was significantly reduced after the paeonol treatment in the diabetic rats. The histopathology showed a significant reduction (p < 0.05) in the retinal thickness after the paeonol treatment. The results of the study indicate that paeonol can be considered an effective management option for diabetic retinopathy.

Antifertility effects of 60-day oral gavage of ethanol extract of Spondias mombin leaves in guinea pigs

Objective: To investigate the impact of ethanol extract of Spondias (S.) mombin leaves on the biochemical parameters of testicular function, hormonal profile, sperm profile and reproductive outcomes of guinea pigs. Methods: Sexually matured male [(1000.40±8.12) g] and female [(810.00±7.09) g] guinea pigs were used. In testicular function study, a total of 24 male guinea pigs were randomised into 4 groups of 6 guinea pigs each. Group A (control) were orally administered 1 mL of physiological saline, once daily for 60 days while groups B, C and D were treated like the control group except that they were orally administered 100, 250 and 500 mg/kg body weight of ethanol extract of S. mombin leaves. For the fertility study, the same animal groupings and treatments in the testicular function study were adopted. The male guinea pigs were paired with the females (1:1) and afterwards examined for pregnancy outcomes. Results: The ethanol extract of S. mombin leaves contained saponins, alkaloids, flavonoids, tannins, steroids, phenolics, phlobatannins, cardiac glycosides, cardenolides and dienolides with saponins (4.80 mg/mL) occurring the most whilst cardenolides and dienolides (0.08 mg/mL) were the least. The ethanol extract of S. mombin leaves significantly and dose dependently reduced the activities of alkaline phosphatase, glutamate dehydrogenase, 3-hydroxy-3-methylglutaryl coenzyme A reductase, malic enzyme, 17-β-hydroxysteroid dehydrogenase, lactate dehydrogenase, catalase and superoxide dismutase as well as levels of testosterone, glycogen, total protein and ascorbic acid in the testes when compared with the control group (P&lt;0.05). All the doses of ethanol extract of S. mombin leaves also reduced the levels of sorbitol dehydrogenase, 3-β-hydroxysteroid dehydrogenase, glucose-6-phosphate dehydrogenase and sialic acid whereas the levels of testicular acid phosphatase, gamma glutamyl transferase and cholesterol increased dose dependently (P&lt;0.05). The serum luteinising hormone, testosterone and estradiol were reduced after the administration of ethanol extract of S. mombin leaves whereas levels of serum follicle stimulating hormone increased significantly. The 100 and 200 mg/kg body weight of ethanol extract of S. mombin leaves increased the testosterone/estradiol ratios whilst the 500 mg/kg body weight of ethanol extract of S. mombin leaves decreased it. The sperm motility, sperm count, normal sperm morphology, sperm density, sperm viability and semen viscosity were significantly reduced in the ethanol extract of S. mombin leaves-treated guinea pigs (P&lt;0.05) whereas the head-, tail- and neck-defects increased significantly when compared with the control group (P&lt;0.05). In contrast, the semen volume and pH were not significantly altered by the ethanol extract of S. mombin leaves (P&gt;0.05). The ethanol extract of S. mombin leaves at both 100 and 250 mg/kg body weight significantly reduced the total number, circumference, weight and length of the pups whereas the 500 mg/kg body weight of ethanol extract of S. mombin leaves-treated rats did not produce any pup. The 250 and 500 mg/kg body weight of ethanol extract of S. mombin leaves induced degenerative and necrotic changes in the seminiferous tubules with vacuoles in the germinal epithelium and a few to complete absence of spermatozoa. In all of these, the 500 mg/kg body weight of ethanol extract of S. mombin leaves produced the most pronounced alteration on the parameters. Conclusions: S. mombin leaves have induced infertility in the male guinea pigs via endocrine dysregulation, anti-spermatogenic activity, testicular dysfunction and oxidative stress and made possible by the presence of saponins, alkaloids, flavonoids, tannins, steroids, phenolics, and cardiac glycosides.

Reduction of Glut1 in the Neural Retina But Not the RPE Alleviates Polyol Accumulation and Normalizes Early Characteristics of Diabetic Retinopathy

Hyperglycemia is a key determinant for development of diabetic retinopathy (DR). Inadequate glycemic control exacerbates retinopathy, while normalization of glucose levels delays its progression. In hyperglycemia, hexokinase is saturated and excess glucose is metabolized to sorbitol by aldose reductase via the polyol pathway. Therapies to reduce retinal polyol accumulation for the prevention of DR have been elusive due to low sorbitol dehydrogenase levels in the retina and inadequate inhibition of aldose reductase. Using systemic and conditional genetic inactivation, we targeted the primary facilitative glucose transporter in the retina, Glut1, as a preventative therapeutic in diabetic male and female mice. Unlike wildtype diabetics, diabetic Glut1+/- mice did not display elevated Glut1 levels in the retina. Furthermore, diabetic Glut1+/- mice exhibited ameliorated ERG defects, inflammation and oxidative stress, which was correlated with a significant reduction in retinal sorbitol accumulation. RPE-specific reduction of Glut1 did not prevent an increase in retinal sorbitol content or early hallmarks of DR. However, like diabetic Glut1+/- mice, reduction of Glut1 specifically in the retina mitigated polyol accumulation and diminished retinal dysfunction and the elevation of markers for oxidative stress and inflammation associated with diabetes. These results suggest that modulation of retinal polyol accumulation via Glut1 in photoreceptors can circumvent the difficulties in regulating systemic glucose metabolism and be exploited to prevent DR.SIGNIFICANCE:Diabetic retinopathy (DR) affects one third of diabetic patients and is the primary cause of vision loss in adults aged 20-74. While anti-VEGF and photocoagulation treatments for the late-stage vision threatening complications can prevent vision loss, a significant proportion of patients do not respond to anti-VEGF therapies and mechanisms to stop progression of early-stage symptoms remain elusive. Glut1 is the primary facilitative glucose transporter for the retina. We determined that a moderate reduction in Glut1 levels, specifically in the retina, but not the RPE, was sufficient to prevent retinal polyol accumulation and the earliest functional defects to be identified in the diabetic retina. Our study defines modulation of Glut1 in retinal neurons as a targetable molecule for prevention of DR.

The Beneficial Radioprotective Effect of Tomato Seed Oil Against Gamma Radiation–Induced Damage in Male Rats

ABSTRACTRadiation protection research receives intense focus due to its significant impact on human health. The present study was undertaken to investigate the protective effect of pretreatment with tomato seed oil (TSO) against gamma radiation–induced damage in rats. Male Wistar rats were divided into four groups: (1) untreated control; (2) TSO-supplemented; (3) gamma-irradiated; (4) TSO-pretreated and gamma-irradiated. Acute exposure of animals to a single gamma radiation dose (6 Gy) induced oxidative stress in major body organs, altered serum lipid homeostasis, significantly increased serum testosterone and sorbitol dehydrogenase levels, and elicited a systemic inflammation as manifested by the induction of serum vascular cell adhesion molecule-1. Oral pretreatment with TSO (1 ml/kg; 3 times/week for 8 weeks) before exposure to gamma radiation protected rats against ionizing radiation-induced oxidative stress, restored lipid homeostasis, and suppressed systemic inflammation. Histological findings of target tissues verified biochemical data. The radioprotective ability of TSO was attributed to its content of phytosterols, policosanol, and antioxidants, including lycopene, β-carotene, lutein, and tocopherols. TSO is considered a promising radioprotective agent that can be effectively used to protect the body from the damaging effects of harmful radiation.

Effects of S-Allylcysteine on Biomarkers of the Polyol Pathway in Rats with Type 2 Diabetes.

We evaluated the effects of S-allylcysteine (SAC) on biomarkers of the polyol pathway in streptozotocin-nicotinamide (STZ-NA)-induced diabetes in rats. Diabetes was induced in male albino Wistar rats by intraperitoneal administration of STZ (55 mg kg-1 bw-1) and NA (110 mg kg-1 bw-1). SAC (150 mg kg-1 bw-1) was orally administered to the rats with diabetes for 45 days to assess its effects on blood glucose, insulin, insulin resistance, glycated hemoglobin, aldose reductase (AR), sorbitol dehydrogenase (SDH), sorbitol, fructose, thiobarbituric acid-reactive substances (TBARS), hydroperoxide, hemoglobin and glutathione (GSH). On SAC administration in the rats with diabetes, the levels of blood glucose, insulin resistance, glycated hemoglobin, AR, SDH, sorbitol, fructose, TBARS and hydroperoxide increased significantly (p<0.05), whereas those of insulin, hemoglobin and GSH decreased. SAC showed therapeutic effects similar to those of gliclazide in decreasing blood glucose, AR, SDH, sorbitol, fructose, glycosylated hemoglobin, TBARS and hydroperoxides levels and significant increases in insulin, hemoglobin and GSH activity in rats with diabetes. Moreover, histopathologic studies also revealed the protective effect of SAC on pancreatic beta cells. The results indicate that SAC prevents complications of diabetes by reducing the influx of glucose in the polyol pathway, thereby elevating the GSH level and reducing the activities of AR and SDH. Therefore, SAC may have imperative implications for the deterrence and early treatment of type 2 diabetes.

Effects of hot air and methyl jasmonate treatment on the metabolism of soluble sugars in peach fruit during cold storage

Abstract Soluble sugar metabolism affects the quality and chilling resistance of postharvest peach fruit. Although hot air (HA) and methyl jasmonate (MeJA) treatments are often effective in reducing chilling injury (CI), little is known about the relationship between sugar metabolism and HA or MeJA treatments in peach fruit. In this study, peach fruit was treated with hot air at 37 °C for 3 days or MeJA vapor at 10 μmol/L for 24 h before storage at 5 °C. Soluble sugar content, gene expression and enzyme activities associated with sugar metabolism were measured. Both treatments resulted in an initial increase, then a decrease in sucrose content over the course of storage time. Sucrose levels at every time point, but one, throughout the experiment were significantly higher than in control fruit, paralleled by higher gene expression and activity of SPS (sucrose phosphate synthase) and lower expression and activity of AI (acid invertase). HA-treated fruit had the highest sucrose content at the end of storage and the mildest CI symptoms. All treated fruit had higher sorbitol content and lower levels of SDH (sorbitol dehydrogenase) gene expression than control fruit. After 21 days in cold storage, sucrose content had decreased sharply in the control group, hexose content was not markedly affected, perhaps due to the increased expression of PFK (phosphofructokinase), resulting in more glucose entering the Embden–Meyerhof–Parnas pathway (EMP). These results suggest that the increase in sucrose observed during cold storage, associated with higher SPS and lower AI levels, enhances the chilling tolerance observed in HA- and MeJA-treated fruit.

D-penicillamine-induced granulomatous hepatitis in brown Norway rats.

The mechanism of idiosyncratic drug reactions (IDRs) remains poorly understood. D-penicillamine treatment is associated with a wide range of autoimmune reactions including liver injury. An animal model which utilizes brown Norway (BN) rats has been used to investigate the mechanism of D-penicillamine-induced IDRs because it mimics the autoimmune reactions that occur in humans. The purpose of this study was to investigate the type of liver injury that results from D-penicillamine treatment in BN rats. We had previously noted that D-penicillamine caused histological changes in the liver, but there was no increase in alanine transaminase (ALT), and we assumed that there was no significant injury. However, we subsequently discovered that D-penicillamine inhibits the ALT assay. In the present study, we found that treatment of BN rats with a low doses of D-penicillamine (10 or 15 mg/day) resulted in a mild increases in glutamate dehydrogenase (GLDH) and sorbitol dehydrogenase (SDH) activities; however, this was not associated with histological changes. A higher dose of D-penicillamine (20 mg/day) resulted in 63% of the rats developing a skin rash, and these rats had elevated serum GLDH and SDH levels with histopathological changes characteristic of granulomatous hepatitis. This included large clusters of leukocytes in the form of granulomas that contained neutrophils, macrophages, and CD8 T cells. These changes did not occur in the rats that did not get sick. This model may be a good model to investigate the characteristics of drug-induced granulomatous hepatitis.

Expression patterns of genes involved in sugar metabolism and accumulation during apple fruit development.

Both sorbitol and sucrose are imported into apple fruit from leaves. The metabolism of sorbitol and sucrose fuels fruit growth and development, and accumulation of sugars in fruit is central to the edible quality of apple. However, our understanding of the mechanisms controlling sugar metabolism and accumulation in apple remains quite limited. We identified members of various gene families encoding key enzymes or transporters involved in sugar metabolism and accumulation in apple fruit using homology searches and comparison of their expression patterns in different tissues, and analyzed the relationship of their transcripts with enzyme activities and sugar accumulation during fruit development. At the early stage of fruit development, the transcript levels of sorbitol dehydrogenase, cell wall invertase, neutral invertase, sucrose synthase, fructokinase and hexokinase are high, and the resulting high enzyme activities are responsible for the rapid utilization of the imported sorbitol and sucrose for fruit growth, with low levels of sugar accumulation. As the fruit continues to grow due to cell expansion, the transcript levels and activities of these enzymes are down-regulated, with concomitant accumulation of fructose and elevated transcript levels of tonoplast monosaccharide transporters (TMTs), MdTMT1 and MdTMT2; the excess carbon is converted into starch. At the late stage of fruit development, sucrose accumulation is enhanced, consistent with the elevated expression of sucrose-phosphate synthase (SPS), MdSPS5 and MdSPS6, and an increase in its total activity. Our data indicate that sugar metabolism and accumulation in apple fruit is developmentally regulated. This represents a comprehensive analysis of the genes involved in sugar metabolism and accumulation in apple, which will serve as a platform for further studies on the functions of these genes and subsequent manipulation of sugar metabolism and fruit quality traits related to carbohydrates.

Effects of Synthetic Androgens on Liver Function Using the Rabbit as a Model

The objective of this study was to determine whether the rabbit was a suitable model to test new synthetic androgens for potential liver toxicity within a short dosing interval. Adult male rabbits were dosed orally daily on days 0-13 with 17α-methyltestosterone (MT) as a positive control and testosterone (T) as a negative control to validate this model. Synthetic androgens tested were: 7α-methyl-19-nortestosterone (MENT), dimethandrolone-undecanoate (DMAU), and 11β-methyl-19-nortestosterone-17β-dodecylcarbonate (11β-MNTDC). Serum levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), gamma glutamyl transpeptidase (GGT), and sorbitol dehydrogenase (SDH), as well as clearance of intravenous injected bromsulfonphthalein (BSP) from serum on days 0, 7, and 14, were determined. As expected, T (10 mg/kg/d) did not adversely affect BSP retention or serum liver enzymes. MT (10 mg/kg/d) increased BSP retention, and AST, ALT, GGT, and SDH levels, indicating that this model could detect androgens known to be hepatotoxic. DMAU and MENT (10 mg/kg/d) increased BSP retention and all 4 serum liver enzymes as well, but the effects were less than those observed with MT at the same dose. All parameters returned to baseline 2 weeks after cessation of dosing. 11β-MNTDC at 10 mg/kg/d did not have an effect on BSP retention or liver enzymes, but a slight increase in serum GGT levels was observed in rabbits treated with 25 mg/kg/d. For the androgens that exhibited liver toxicity at 10 mg/kg/d (MT, DMAU, and MENT), a no-observed-effect level of 1 mg/kg/d was established. Overall ranking of the synthetic androgens from most to least hepatotoxic on the basis of percent BSP retention was: MT & DMAU > MENT > 11β-MNTDC. Hence, the rabbit appears to be a promising model for detection of potential liver toxicity by synthetic androgens using BSP clearance and serum liver enzyme levels as early indicators of injury.

ZAC1 Is Up-regulated by Hypertonicity and Decreases Sorbitol Dehydrogenase Expression, Allowing Accumulation of Sorbitol in Kidney Cells

Affymetrix GeneChip technology was employed to detect differentially expressed genes in inner medullary collecting duct (IMCD3) cells grown under isotonic and hypertonic conditions. A marked up-regulation was found for the zinc-finger protein ZAC1 under hypertonic stress (219-fold, p < 0.001). Changes in expression for ZAC1 were verified by quantitative PCR for message and Western blotting for protein. In mouse and human kidney tissues, ZAC1 expression was substantial in the papilla and was absent in the cortex. Furthermore, ZAC1 expression significantly increased in the papilla of mice following 36 h of fluid restriction and decreased in polyuric mice consuming sucrose in water. Because ZAC1 has been described to be a potential negative regulator of sorbitol dehydrogenase (SDH) in hippocampal cells, we examined whether this relationship also occurs in kidney cells under hypertonic stress. We found that stable IMCD3 clones silenced for ZAC1 to varying levels demonstrated an inverse effect on SDH expression. ZAC1 binds to a consensus repression site within the promoter of SDH, pointing to a mechanism whereby ZAC1 acts by repressing SDH transcriptional activity during hypertonic conditions. Taken together, these data strongly suggest that ZAC1 is up-regulated under hypertonic stress and negatively regulates expression of SDH, allowing for accumulation of sorbitol as a compatible organic osmolyte.

Requirement of Aldose Reductase for the Hyperglycemic Activation of Protein Kinase C and Formation of Diacylglycerol in Vascular Smooth Muscle Cells

Activation of protein kinase C (PKC) has been linked to the development of secondary diabetes complications. However, the underlying molecular mechanisms remain unclear. We examined the contribution of aldose reductase, which catalyzes the first, and the rate-limiting, step of the polyol pathway of glucose metabolism, to PKC activation in vascular smooth muscle cells (VSMCs) isolated from rat aorta and exposed to high glucose in culture. Exposure of VSMCs to high glucose (25 mmol/l), but not iso-osmotic mannitol, led to an increase in total membrane-associated PKC activity, which was prevented by the aldose reductase inhibitors tolrestat or sorbinil or by the ablation of aldose reductase by small interfering RNA (siRNA). The VSMCs were found to express low levels of sorbitol dehydrogenase, and treatment with the sorbitol dehydrogenase inhibitor CP-166572 did not prevent high-glucose-induced PKC activation. Stimulation with high glucose caused membrane translocation of conventional (alpha, beta1, beta2, and gamma) and novel (delta and epsilon) isoforms of PKC. Inhibition of aldose reductase prevented membrane translocation of PKC-beta2 and -delta and delayed the activation of PKC-beta1 and -epsilon, whereas membrane translocation of PKC-alpha and -gamma was not affected. Treatment with tolrestat prevented phosphorylation of PKC-beta2 and -delta. High glucose increased the formation of diacylglycerol (DAG) and enhanced phosphorylation of phospholipase C-gamma1 (PLC-gamma1). Inhibition of aldose reductase prevented high glucose-induced DAG formation and phosphorylation of PLC-gamma1 and PLC-beta2 and -delta. Inhibition of phospholipid hydrolysis by D609, but not by the synthetic alkyl-1-lysophospholipid 1-O-octadecyl-2-O-methyl-rac-glycerophosphocholine, or edelfosine, prevented DAG formation. Treatment with sorbinil decreased the levels of reactive oxygen species in high-glucose-stimulated VSMCs. Hence, inhibition of aldose reductase, independent of sorbitol dehydrogenase, appears to be effective in diminishing oxidative stress and hyperglycemic changes in signaling events upstream to the activation of multiple PKC isoforms and PLC-gamma1 and may represent a useful approach for preventing the development of secondary vascular complications of diabetes.

Colocalization of polyol-metabolizing enzymes and immunological detection of fructated proteins in the female reproductive system of the rat.

The expression of aldose reductase (AR) and sorbitol dehydrogenase (SDH), which, in concert, catalyze the conversion of glucose to fructose via sorbitol, in the rat ovary, oviduct, and uterus, was investigated by immunohistochemical and biochemical analyses. The activities and protein levels of AR and SDH were higher in the ovary than in the oviduct and uterus. A strong immunoreactivity to the anti-AR antibody was observed in granulosa cells and epithelia of the oviduct, endometrium, and endometrial glands, and virtually the same tissues were strongly stained with the anti-SDH antibody. The application of an anti-fructated lysine antibody, which detects an adduct of fructose with the epsilon-amino group of lysine in proteins, in this study detected marked staining mainly in the egg and luminal surface of the oviductal epithelia. Collectively, these data indicate that fructose is produced by coordinately expressed AR and SDH in the egg and epithelia of the oviduct and suggest that the resulting sorbitol and fructose can be used as energy sources for spermatozoa motility during the fertilization process. The abundance of AR compared with SDH suggests that it also plays an additional role in the reproductive system, which might include a source of reducing power and protection against toxic carbonyl compounds.

Effects of high copper supplements on performance, health, plasma copper and enzymes in goats

Six growing female Nubian goats (average BW=34.8+/-0.55kg, 7-8 months of age) were randomly assigned to either a basal diet (BD, 10-15ppm Cu/DM), or to medium Cu (MC, BD+50mgCu), or to high Cu (HC, BD+100mgCu) diets for 9 weeks. This level would cause Cu toxicity in sheep, but none occurred in the goats. Therefore, Cu supplementation was then increased to 150 and 300mg per head per day, for the following 14 weeks; to 300 and 600mg per head per day, for the next 8 weeks; and to 600 and 1200mg per head per day, for an additional 4 weeks, in the MC and HC group, respectively. Body weight and vital signs were recorded and blood samples collected at different time intervals. Hematological parameters, plasma Cu, sorbitol dehydrogenase (SDH), glutamic oxaloacetic transaminase (GOT), and gamma-glutamyl transferase (GGT) were determined. At the termination of the study, tissue Cu concentration in different organs was also determined. During first 23 weeks (<300mgCu per day) of the study there were no apparent signs of Cu toxicity. Cu supplementation at 600mg per head per day in young Nubian does, had no effect on respiration rate (RR), heart rate (HR), and decreased (P<0.05) rectal temperature (RT) in the HC group only. Leukocyte counts were positively correlated with Cu supplementation (r=+0.296, P<0.02) and negatively correlated (r=-0.254, P<0.05) with RT in the HC group. Plasma SDH increased (P<0.05) when Cu supplementation was >/=300mg per head per day, thus, SDH may serve as an early indicator of Cu toxicosis in goats. Increases (P<0.05) in GOT were noted when Cu intake was >/=600mg per head per day. Contrary to the results observed for SDH and GOT, feeding goats 50mgCu per day or more, resulted in an increased plasma GGT as compared to BD goats. Levels of SDH, GOT and GGT of the BD goats were within normal range. Plasma Cu was not indicative of Cu status of animals. Copper improved ADG by 28% at the 100-150ppm level in diet. No relationship between Cu intake and hair Cu was found in the present study. Highest concentration of Cu was found in liver, followed by duodenum, rumen and brain. Results of this study indicate that goats are more resistant to Cu toxicity than sheep. This is one of the first reports documenting significant differences in Cu requirements and tolerance between goats and sheep.

Bosentan, an endothelin antagonist, augments hepatic graft function by reducing graft circulatory impairment following ischemia/reperfusion injury

Endothelin is a potent hepatic vasoconstrictor. We evaluated the role of an endothelin antagonist in hepatic ischemia/reperfusion injury. Bosentan, a novel endothelin receptor antagonist, was infused directly into the portal vein prior to cold ischemia and immediately on reperfusion, in five porcine livers. Five other pigs underwent routine liver harvest and reperfusion without bosentan treatment. Hepatic vascular resistance and liver tissue blood flow, as measured by thermistor flow probes, were determined following reperfusion. Hepatocellular damage was assessed through hepatic venous levels of sorbitol dehydrogenase and lactate dehydrogenase. Endothelial cell damage was determined in sections immuno-stained for factor VIII. Graft function was determined through oxygen consumption, bile production, and response to bile acid challenge. Organs treated with bosentan demonstrated lower vascular resistance and enhanced tissue blood flow (P < 0.05) as compared to untreated organs. Portal vein inflow to hepatic tissue was significantly enhanced (4.4-fold) in the bosentan-treated organs (P < 0.05). No difference was observed in hepatocellular damage. Pathology scores for factor VIII immunohistochemical staining were 2.3-fold higher in the bosentan-treated livers as compared to untreated livers (P < 0.05). The bosentan-treated livers also demonstrated enhanced oxygen consumption, increased bile production, and augmented biliary response to a bile acid challenge (P < 0.05). These results indicate that administration of bosentan before and after ischemia/reperfusion reduces hepatic circulatory disturbances, diminishes endothelial cell damage, and augments hepatic graft function.

Studies on some serum enzyme levels in various liver diseases.

Serum alkaline phosphatase (ALP) (EC 3.1.3.1), 5'nucleotidase (5'NT) (EC 3.1.3.5), aldolase (ALD) (EC 4.1.2.13) and sorbitol dehydrogenase (SDH) (EC 1.1.1.14) were estimated in infective hepatitis, alcoholic hepatitis, chronic active hepatitis, obstructive jaundice, cirrhosis of liver and amoebic liver abscess. It was observed that serum ALP and 5'NT were significantly increased in all cases of chronic active hepatitis and obstructive hepatic disease. However, the elevation observed in the latter was much higher than the former. Serum SDH and ALD levels were elevated in all cases of infective hepatitis, studied though increase in the former was much higher than the latter, suggesting its significance in the diagnostic confirmation of this disease. Results presented suggest 5'NT and SDH as more reliable diagnostic test compared to ALP and ALD for obstructive jaundice and infective hepatitis respectively.

Further evaluation of the incorporation of an immunotoxicological functional assay for assessing humoral immunity for hazard identification purposes in rats in a standard toxicology study

A previous study (Ladics et al., 1995) conducted in our laboratory using the known immunosuppressant agent, cyclophosphamide, indicated that a functional assay for assessment of humoral immunity may be conducted in rats in a standard toxicology study. The objective of this study was to further examine the feasibility of conducting an immunotoxicological assay for assessing humoral immunity in rats in a standard toxicology study using a chemical, carbon tetrachloride (CCl4), whose principal target organ of toxicity is not the immune system. Specifically, the previous study and this study were done to determine whether the conduct of an assay for assessing humoral immune function would affect standard toxicological endpoints. Male CD rats were untreated or dosed orally for 30 or 90 days, excluding weekends, with vehicle or 12.5 or 25 mg/kg CCl4. Six days prior to sacrifice, selected rats were injected intravenously with sheep red blood cells (SRBC) for assessment of humoral immune function. One day prior to necropsy, blood for hematological and clinical chemical measurements was collected from each rat. On the day of necropsy standard protocol tissues were collected, weighed, processed to slides, and later examined microscopically. One-half of each spleen was used to assess spleen cell numbers and quantitate lymphocyte subsets (Thelper; Tcyt/sup; total T- and B-cells) by flow cytometry. Serum was analyzed for anti-SRBC IgM antibody by using an enzyme-linked immunosorbent assay. Administration of 12.5 and 25 mg/kg CCl4 for 30 days decreased SRBC-specific serum IgM levels 42 and 45%, respectively, while 25 mg/kg CCl4 for 90 days increased SRBC-specific IgM levels by 50%. CCl4 did not alter splenic lymphocyte subset numbers nor the weight nor morphology of lymphoid organs. Exposure to 25 mg/kg CCl4 did increase liver weight and serum sorbitol dehydrogenase levels, as well as produce centrilobular fatty change. SRBC administration did not alter any hematological or clinical chemistry parameters, nor lymphocyte subset numbers. With the expected exception of the spleen (slight increase in number and size of germinal centers), administration of SRBC did not significantly alter the weights or morphology of routine protocol tissues. Furthermore, administration of SRBC did not mask the rather mild hepatotoxic effects of CCL4 exposure observed in this study. Based on these and previous findings, it appears that a functional assay for assessing humoral immunity may be conducted in animals on standard toxicology study without altering standard toxicological endpoints.

Antihepatotoxic zeaxanthins from the fruits ofLycium chinense

A CHCl(3): MeOH extract of the fruit ofLycium chinense Mill. (Solanaceae) was found to afford significant protection against carbon tetrachloride-induced toxicity in primary cultures of rat hepatocytes. Subsequent activity-guided fractionation resulted in the isolation of zeaxanthin and zeaxanthin dipalmitate as antihepatotoxic components. Incubation of injured hepatocytes with zeaxanthin dipalmitate reduced the levels of glutamic pyruvic transaminase (GPT) and sorbitol dehydrogenase (SDH) released from damaged cells to 60.5% and 76.3% of those released from untreated controls, respectively. Zeaxanthin also reduced the levels of GPT and SDH to 68.5% and 61.3% of the levels of those released from the untreated control. The results confirm the hepatoprotective activities of zeaxanthins. Antihepatotoxic activities of zeaxanthins are comparable to that of silybin.

COMPARISON OF MOUSE STRAINS FOR SUSCEPTIBILITY TO STYRENE-INDUCED HEPATOTOXICITY AND PNEUMOTOXICITY

Styrene is known to cause both hepatotoxicity and pneumotoxicity in mice. Strain differ ences have been reported by other investigators suggesting that Swiss mice are less suscep tible than non-Swiss mice to styrene-induced liver damage. In this study, A/I and C57BL/6 mice were found to be similar to non-Swiss albino (NSA) mice in susceptibility whereas CD-1 (Swiss) mice were more resistant to hepatotoxicity as assessed by serum sorbitol de hydrogenase levels and pneumotoxicity as determined by gamma-glutamyltranspeptidase and lactate dehydrogenase measurements in bronchoalveolar lavage fluid. Styrene was hepatotoxic in CD-I mice treated with pyridine to induce CYP2E1. CYP2EI apoprotein levels and p-nitrophenol hydroxylase activities in control and pyridine-induced mice were similar in the two strains. Hepatic and pulmonary microsomal preparations from both strains metabolized styrene to styrene oxide at similar rates. CD-1 mice were as susceptible as the NSA mice to the effects of styrene oxide. The data suggest that there are no differ ences in the bioactivation of styrene to styrene oxide or innate susceptibility to the active metabolite that would account for the differences between the CD-1 and NSA mice.

Physiologically Based Pharmacokinetic Estimated Metabolic Constants and Hepatotoxicity of Carbon Tetrachloride after Methanol Pretreatment in Rats

A single 6-hr exposure to inhaled methanol (MeOH) has been shown to enhance carbon tetrachloride (CCl4) hepatotoxicity. The objective of the present study was to use gas uptake data and the development of a physiologically based pharmacokinetic model (PBPK) to determine in vivo changes in CCl4 metabolism resulting from MeOH pretreatment. Adult male F344 rats (167-197 g) were exposed to 10,000 ppm MeOH (constant concentration) via inhalation for 6 hr. Individual rats were exposed using gas uptake techniques to CCl4 alone or to CCl4 either 24 or 48 hr after initiation of MeOH pretreatment. The following initial concentrations were used for CCl4: 0, 25, 100, 250, and 1000 ppm with exposures lasting 6 hr. Vmax (metabolic rate) was estimated from gas uptake data and Km (Michaelis constant) was assumed constant after methanol pretreatment. For CCl4 alone, Vmax was 0.11 mg/hr (Vmaxc = 0.37 mg/hr/kg) and Km was 1.3 mg/liter. Vmax was 0.48 mg/hr (Vmaxc = 1.6 mg/hr/kg) for the 24-hr MeOH + CCl4 group and Vmax was 0.18 mg/hr (Vmaxc = 0.6 mg/hr/kg) for the 48-hr MeOH + CCl4 group. For CCl4 alone, serum markers of hepatotoxicity alanine aminotransferase (ALT) and sorbitol dehydrogenase (SDH) were increased significantly only at 1000 ppm CCl4. Both serum markers of hepatotoxicity in the 24-hr MeOH + CCl4 group increased as a function of CCl4 concentration when compared with 0 ppm CCl4 controls. The maximum increase occurred at 1000 ppm CCl4, where ALT and SDH increased by 392- and 286-fold, respectively. At 100, 250, and 1000 ppm CCl4, ALT and SDH values for the 24-hr MeOH + CCl4 groups were significantly increased relative to control (0 ppm CCl4), CCl4 alone, and 48-hr MeOH + CCl4. ALT and SDH levels in the 48-hr MeOH + CCl4 groups were not statistically different from the respective CCl4 alone groups.

Comparison of Styrene Hepatotoxicity in B6C3F1 and Swiss Mice

Inhalation exposure to styrene at concentrations that cause metabolic saturation results in significantly greater hepatotoxicity in B6C3F1 mice than in Swiss mice; females of both strains are more susceptible than males. These studies were conducted to investigate the mouse strain and gender differences in susceptibility to hepatotoxicity caused by repeated exposure to styrene at concentrations that do not cause metabolic saturation. Male and female B6C3F1 and Swiss mice (8 weeks old) were exposed to 0, 150, or 200 ppm styrene for 6 hr/day, 5 days/week, for up to 2 weeks. Changes in body and liver weights, serum alanine aminotransferase (ALT) and sorbitol dehydrogenase (SDH) levels, liver histopathology, and total liver glutathione (GSH) were evaluated after 2, 3, 5, and 10 exposures (six mice/sex/strain/time point/concentration). Blood levels of styrene and styrene-7,8-oxide (SO) were measured in mice exposed to 200 ppm styrene for 2,3, or 5 days (six mice/sex/strain/time point/concentration). Serum ALT and SDH levels were significantly elevated only in female B6C3F1 mice after 3 exposures to 200 ppm styrene; enzyme levels had returned to control levels when measured after 5 and 10 exposures. Degeneration and coagulative necrosis of centrilobular hepatocytes were observed in female B6C3F1 mice exposed 2, 3, and 5 days to 150 or 200 ppm styrene; incidences of these lesions were greater in the 200 ppm than in the 150 ppm dose group. After 10 days of exposure to 150 or 200 ppm styrene, hepatocellular lesions had resolved, although a residual chronic inflammation was present in livers of most female B6C3F1 mice.(ABSTRACT TRUNCATED AT 250 WORDS)