Investigation on sodium valproate induced germ cell damage, oxidative stress and genotoxicity in male Swiss mice

Sodium valproate (VPA) is the most widely used antiepileptic drug for the treatment of epilepsy, bipolar psychiatric disorders and migraine. However, long-term VPA treatment has several adverse effects on the reproductive system. The present study was aimed to investigate the possible germ cell toxicity of VPA in mice. Animals were treated with VPA intraperitoneally for 10 and 28 days at the doses of 500 mg/kg-d and 100, 200 and 400 mg/kg-d, respectively, and were sacrificed 24h after the last dose. The germ cell toxicity of VPA was assessed using oxidative stress parameters, sperm count, sperm head morphology, sperm comet assay, 8-oxo-dG expression and histology. VPA treatment significantly decreased the sperm count, testes and epididymis weight and significantly increased the sperm head abnormality, sperm DNA damage, oxidative stress and 8-oxo-dG expression in the testes of mice. The present study illustrates that VPA induced germ cell toxicity in mice.     

Responses of testis,epididymis, and sperm of pubertal rats exposed to functionalized multiwalled carbon nanotubes

The present study investigated the response of testes, epididymides and sperm in pubertal Wistar rats following exposure to 0, 0.25, 0.5, 0.75, and 1.0 mg kg^?1 functionalized multi‐walled carbon nanotubes (f‐MWCNTs) for 5 days. The results showed that administration of (f‐MWCNTs) significantly increased the activities of superoxide dismutase, catalase, and glutathione peroxidase in a dose‐dependent manner in both testes and sperm compared with control group. Moreover, the significant decrease in the activity of glutathione‐S‐transferase and glutathione level was accompanied with significant elevation in the levels of hydrogen peroxide and malondialdehyde in both testes and sperm of (f‐MWCNTs)‐treated rats. The spermiogram of (f‐MWCNTs)‐treated rats indicated significant decrease in epididymal sperm number, sperm progressive motility, testicular sperm number and daily sperm production with elevated sperm abnormalities when compared with the control. Exposure to (f‐MWCNTs) decreased plasma testosterone level and produced marked morphological changes including decreased geminal epithelium, edema, congestion, reduced spermatogenic cells and focal areas of tubular degeneration in the testes. The lumen of the epididymides contained reduced sperm cells and there was mild to severe hyperplasia epithelial cells lining the duct of the epididymis. Collectively, pubertal exposure of male rats to (f‐MWCNTs) elicited oxidative stress response resulting in marked testicular and epididymides dysfunction. © 2014 Wiley Periodicals, Inc. Environ Toxicol 31: 543–551, 2016.     

Resveratrol improves sperm motility,prevents lipid peroxidation and enhances antioxidant defences in the testes of hyperthyroid rats

Hyperthyroidism may lead to a loss of sperm motility and an increase in oxidative stress (OS) in testes and may cause male reproductive disorders. Thus, the use of compounds with antioxidant properties may be a strategy for preventing these disorders. The effect of resveratrol (RSV) on sperm motility and on variables of the antioxidant status in the testes of rats with triiodothyronine-induced hyperthyroidism (100 μg/kg) was investigated. Hyperthyroid rats presented lower sperm motility, higher levels of lipid hydroperoxides and thiobarbituric reactive substances, lower catalase and glutathione peroxidase activities and higher glutathione-S-transferase activity in their testes than control animals. RSV treatment (1 mg/kg and 10 mg/kg) was able to prevent these effects in the hyperthyroid rats and had no effect in the control animals. In conclusion, RSV might be a strategy for therapeutic intervention to preserve sperm motility and to prevent OS in testes, preserving testicular function in those with hyperthyroidism.     

Valproic acid induced liver injury: An insight into molecular toxicological mechanism

Valproic acid (VPA) is an anti-seizure drug that causes idiosyncratic liver injury. 2-propyl-4-pentenoic acid (Δ⁴VPA), a metabolite of VPA, has been implicated in VPA-induced hepatotoxicity. This review summarizes the pathogenesis involved in VPA-induced liver injury. The VPA induce liver injury mainly by i) liberation of Δ⁴VPA metabolites; ii) decrease in glutathione stores and antioxidants, resulting in oxidative stress; iii) inhibition of fatty acid β-oxidation, inducing mitochondrial DNA depletion and hypermethylation; a decrease in proton leak; oxidative phosphorylation impairment and ATP synthesis decrease; iv) induction of fatty liver via inhibition of carnitine palmitoyltransferase I, enhancing nuclear receptor peroxisome proliferator-activated receptor-gamma and acyl-CoA thioesterase 1, and inducing long-chain fatty acid uptake and triglyceride synthesis. VPA administration aggravates liver injury in individuals with metabolic syndromes. Therapeutic drug monitoring, routine serum levels of transaminases, ammonia, and lipid parameters during VPA therapy may thus be beneficial in improving the safety profile or preventing the progression of DILI.     

Reproductive toxicity of methomyl insecticide in male rats and protective effect of folic acid

The acute toxicity (LD₅₀) of insecticide methomyl and its effects on male reproduction in rats were carried out. Methomyl was given orally to male rats daily for 65 successive days at two doses (0.5 and 1.0 mg kg⁻¹ b.wt., corresponding to 1/40 and 1/20 LD₅₀) alone and in combination with folic acid (1.1 mg kg⁻¹ b.wt., corresponding to acceptable daily intake, ADI). Fertility index, weight of sexual organs, semen picture, serum testosterone level and histopathology of testes were the parameters used to evaluate the reproductive efficiency of treated rats. The reversibility of methomyl effects was also studied after 65 days post-administration. The oral LD₅₀ of methomyl was 20.0 mg kg⁻¹ b.wt. in male rats. Methomyl significantly decreased the fertility index, weight of testes and accessory male sexual glands, serum testosterone level and sperm motility and count, but increased sperm cell abnormality. It induced testicular lesions characterized by moderate to severe degenerative changes of seminiferous tubules and incomplete arrest of spermatogenesis. These toxic effects were not persistent (reversible). Coadministration of folic acid with methomyl decreased its reproductive toxicity. A great attention should be taken during field application of methomyl to avoid its deleterious effects in farm animals and occupationally exposed humans.     

Effect of nonylphenol on the antioxidant system in epididymal sperm of rats

Nonylphenol, an environmental contaminant, has been shown to induce reproductive abnormalities in male rats. The nature and mechanism of action of nonylphenol on the epididymal sperm has not been elucidated. In the present study we have sought to investigate whether administration of nonylphenol induces oxidative stress in rat epididymal sperm. Nonylphenol was administered orally to male rats at 1, 10 and 100 microg/kg body weight per day for 45 days. Twenty-four hours after the last treatment, rats were weighed and killed using anaesthetic ether. The body weight of the animals treated with nonylphenol did not show any significant change. The weights of the testes and epididymides decreased significantly whereas the weights of seminal vesicles and ventral prostate remained unchanged at all doses of nonylphenol in treated rats. Epididymal sperm were collected by cutting the epididymides into small pieces in Ham's F-12 medium at 32 degrees C. Administration of nonylphenol decreased the epididymal sperm counts in a dose-dependent manner. The activities of antioxidant enzymes superoxide dismutase, catalase, glutathione reductase and glutathione peroxidase decreased significantly while the levels of H(2)O(2) generation and lipid peroxidation increased significantly in the animals treated with nonylphenol when expressed in terms of milligram protein and milligram DNA. The activity of alpha-glucosidase, a negative control against antioxidant enzymes, in the sperm of nonylphenol-treated rats did not show any significant change at any of the doses. The results suggest that graded doses of nonylphenol elicit depletion of antioxidant defence system in sperm, indicating nonylphenol-induced oxidative stress in the epididymal sperm of rats.     

Effect of nonylphenol on male reproduction: analysis of rat epididymal biochemical markers and antioxidant defense enzymes

The mechanism by which nonylphenol (NP) interferes with male reproduction is not fully elucidated. Therefore, the present study was conducted to evaluate the effect of NP on male reproductive organ's weight, sperm characteristics, and to elucidate the nature and mechanism of action of NP on the epididymis. Adult male Wistar rats were gavaged with NP, dissolved in corn oil, at 0, 100, 200 or 300 mg/kg/day for 30 consecutive days. Control rats were gavaged with vehicle (corn oil) alone. Body weight did not show any significant change while, absolute testes and epididymides weights were significantly decreased. Sperm count in cauda and caput/corpus epididymides, and sperm motility was significantly decreased. Daily sperm production was significantly decreased in a dose-related manner. Sperm transit time in cauda epididymis was significantly decreased by 300 mg/kg, while in the caput/corpus epididymis it was significantly decreased by 200 and 300 mg/kg of NP. Plasma LDH was significantly increased while; plasma testosterone was significantly decreased in a dose-related pattern. In the epididymal sperm, NP decreased acrosome integrity, Δψm and 5′-nucleotidase activity. Hydrogen peroxide (H₂O₂) production and LPO were significantly increased in a dose-related pattern. The activities of SOD, CAT and GPx were significantly decreased in the epididymal sperm. In conclusion, this study revealed that NP treatment impairs spermatogenesis and has a cytotoxic effect on epididymal sperm. It disrupts the prooxidant and antioxidant balance. This leads oxidative stress in epididymal sperms of rat. Moreover, the reduction in sperm transit time may affect sperm quality and fertility potential.     

Deciphering the role of ferulic acid against streptozotocin-induced cellular stress in the cardiac tissue of diabetic rats

The cardiomyocytes are one of the major sources of hyperglycemia induced ROS generation. The present study focuses on the ameliorative role of ferulic acid in combating cardiac complications in diabetic rats. Induction of diabetes by STZ in male Wistar rats (at a dose of 50 mg kg⁻¹ body wt, i.p.) reduced body weight and plasma insulin level, enhanced blood glucose, disturbed the intra-cellular antioxidant machineries and disintegrated the normal radiation pattern of cardiac muscle fibers. Induction of ER stress (up-regulation in the levels of CHOP, GRP78, eIF2α signaling, increased calpain-1 expression), caspase-3 activation, PARP cleavage and DNA fragmentation were evidenced from immunoblot analyses and DNA fragmentation assay. However, ferulic acid administration, (at a dose of 50 mg kg⁻¹ body wt, orally for eight weeks) in post-hyperglycemia could reverse such adverse effects. Also, the molecule increased GLUT-4 translocation to the cardiac membrane by enhanced phosphorylation of PI3Kinase, AKT and inactivation of GSK-3β thereby altering the hyperglycemic condition in the cardiac tissue of diabetic rats. Therefore, as a potential therapeutic, ferulic acid, exhibiting antioxidant and hypoglycemic effects, may hold promise in circumventing stress mediated diabetic cardiomyopathy in rats.     

Protection of cisplatin-induced spermatotoxicity,DNA damage and chromatin abnormality by selenium nano-particles

Cisplatin (CIS), an anticancer alkylating agent, induces DNA adducts and effectively cross links the DNA strands and so affects spermatozoa as a male reproductive toxicant. The present study investigated the cellular/biochemical mechanisms underlying possible protective effect of selenium nano-particles (Nano-Se) as an established strong antioxidant with more bioavailability and less toxicity, on reproductive toxicity of CIS by assessment of sperm characteristics, sperm DNA integrity, chromatin quality and spermatogenic disorders. To determine the role of oxidative stress (OS) in the pathogenesis of CIS gonadotoxicity, the level of lipid peroxidation (LPO), antioxidant enzymes including superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GSH-Px) and peroxynitrite (ONOO) as a marker of nitrosative stress (NS) and testosterone (T) concentration as a biomarker of testicular function were measured in the blood and testes. Thirty-two male Wistar rats were equally divided into four groups. A single IP dose of CIS (7 mg/kg) and protective dose of Nano-Se (2 mg/kg/day) were administered alone or in combination. The CIS-exposed rats showed a significant increase in testicular and serum LPO and ONOO level, along with a significant decrease in enzymatic antioxidants levels, diminished serum T concentration and abnormal histologic findings with impaired sperm quality associated with increased DNA damage and decreased chromatin quality. Coadministration of Nano-Se significantly improved the serum T, sperm quality, and spermatogenesis and reduced CIS-induced free radical toxic stress and spermatic DNA damage.In conclusion, the current study demonstrated that Nano-Se may be useful to prevent CIS-induced gonadotoxicity through its antioxidant potential.     

Role of Nrf2 activation and NF-κB inhibition in valproic acid induced hepatotoxicity and in diammonium glycyrrhizinate induced protection in mice

Diammonium glycyrrhizinate (DG), an active compound extracted and purified from liquorices root, has been reported to exhibit antioxidant and anti-inflammatory properties. The aim of this study was to investigate the effect and underlying mechanisms of DG on the hepatotoxicity induced by valproic acid (VPA). DG at the dose of 60 mg/kg was orally administered with VPA (100 mg/kg) to mice once daily for 14 consecutive days. DG treatment attenuated VPA-induced liver dysfunction, structural damage, glutathione depletion and decrease in antioxidant enzymes in BALB/C mice. DG prevented VPA-induced depletion of cytosolic nuclear factor E2-related factor 2 (Nrf2) and suppression of nuclear translocation of Nrf2, which, in turn, up-regulated phase II/antioxidant enzyme activities. The effects of VPA and DG on Nrf2 expression in HepG2 cells were in consistent with that of mice. Furthermore, an increase in the nuclear levels of nuclear factor-kappaB (NF-κB) was observed in the livers of VPA-treated mice that coincided with induction of inflammatory cytokines. In contrast, DG inhibited NF-κB translocation and that subsequently decreased inflammatory cytokines. Taken together, these results demonstrate that DG attenuates VPA-induced liver injury through increasing the expression of Nrf2 mediated phase II/antioxidant enzymes and simultaneously decreasing the expression of inflammatory mediators.     

Protective effects of grape seed procyanidin extract against nickel sulfate-induced apoptosis and oxidative stress in rat testes

This study determined whether nickel sulfate (Ni)-induced reproductive damage occurs via apoptosis and oxidative stress and to examine the expression of Bax and c-kit and their effects on Ni exposure. The study also explored the protective effects of grape seed proanthocyanidin extract (GSPE) against Ni toxicity in the testes. Wistar rats were treated with normal saline, Ni alone (1.25, 2.5, and 5?mg/kg/day), and Ni (2.5?mg/kg/day) plus GSPE (50 and 100 mg/kg/day). After 30 days, Ni significantly decreased sperm motility and the percentage of S-phase cells and enhanced testicular apoptosis in the 2.5 and 5?mg groups. The levels of malondialdehyde (MDA), hydrogen peroxide (H?O?), and nitric oxide (NO) significantly increased. The decreased activity of glutathione peroxidase and catalase in the Ni groups showed that Ni could increase oxidative stress, especially at 2.5 and 5 mg. Western blot analysis showed that the expression of Bax protein and c-kit increased in 2.5 and 5?mg Ni groups compared with controls. Conversely, these changes were partially attenuated in rats simultaneously administered GSPE, especially in the 100?mg group. These results demonstrate the following: (1) Ni exhibits reproductive toxicity in rats by decreasing sperm at concentrations of 2.5 and 5 mg; (2) intratesticular apoptosis, oxidative stress, and c-kit overexpression play pivotal roles in reproductive damage induced by Ni; and (3) GSPE enhances sperm motility by down-regulating c-kit expression and offsetting the apoptosis and oxidative stress induced by Ni by directly decreasing MDA and NO, scavenging H?O?, and down-regulating Bax expression.     

Protective effects of grape seed procyanidin extract against nickel sulfate-induced apoptosis and oxidative stress in rat testes

This study determined whether nickel sulfate (Ni)-induced reproductive damage occurs via apoptosis and oxidative stress and to examine the expression of Bax and c-kit and their effects on Ni exposure. The study also explored the protective effects of grape seed proanthocyanidin extract (GSPE) against Ni toxicity in the testes. Wistar rats were treated with normal saline, Ni alone (1.25, 2.5, and 5?mg/kg/day), and Ni (2.5?mg/kg/day) plus GSPE (50 and 100?mg/kg/day). After 30 days, Ni significantly decreased sperm motility and the percentage of S-phase cells and enhanced testicular apoptosis in the 2.5 and 5?mg groups. The levels of malondialdehyde (MDA), hydrogen peroxide (H₂O₂), and nitric oxide (NO) significantly increased. The decreased activity of glutathione peroxidase and catalase in the Ni groups showed that Ni could increase oxidative stress, especially at 2.5 and 5?mg. Western blot analysis showed that the expression of Bax protein and c-kit increased in 2.5 and 5?mg Ni groups compared with controls. Conversely, these changes were partially attenuated in rats simultaneously administered GSPE, especially in the 100?mg group. These results demonstrate the following: (1) Ni exhibits reproductive toxicity in rats by decreasing sperm at concentrations of 2.5 and 5?mg; (2) intratesticular apoptosis, oxidative stress, and c-kit overexpression play pivotal roles in reproductive damage induced by Ni; and (3) GSPE enhances sperm motility by down-regulating c-kit expression and offsetting the apoptosis and oxidative stress induced by Ni by directly decreasing MDA and NO, scavenging H₂O₂, and down-regulating Bax expression.     

Effects of abamectin exposure on male fertility in rats: Potential role of oxidative stress-mediated poly(ADP-ribose) polymerase (PARP) activation

Despite the known adverse effects of abamectin pesticide, little is known about its action on male fertility. To explore the effects of exposure to abamectin on male fertility and its mechanism, low (1 mg/kg/day) and high dose (4 mg/kg/day) abamectin were applied to male rats by oral gavage for 1 week and for 6 weeks. Weight of testes, serum reproductive hormone levels, sperm dynamics and histopathology of testes were used to evaluate the reproductive efficiency of abamectin-exposed rats. Abamectin level was determined at high concentrations in plasma and testicular tissues of male rats exposed to this pesticide. The testes weights of animals and serum testosterone concentrations did not show any significant changes after abamectin exposure. Abamectin administration was associated with decreased sperm count and motility and increased seminiferous tubule damage. In addition, significant elevations in the 4-hydroxy-2-nonenal (4-HNE)-modified proteins and poly(ADP-ribose) (PAR) expression, as markers for oxidative stress and poly(ADP-ribose) polymerase (PARP) activation, were observed in testes of rats exposed to abamectin. These results showed that abamectin exposure induces testicular damage and affects sperm dynamics. Oxidative stress-mediated PARP activation might be one of the possible mechanism(s) underlying testicular damage induced by abamectin.     

Chronic Exposure to Low Doses of Mercury Impairs Sperm Quality and Induces Oxidative Stress in Rats

Mercury (Hg) is a widespread environmental pollutant that adversely affects the male reproductive system. The precise mechanisms underlying mercuric chloride (HgCl₂)-induced toxicity are not fully understood; however, evidence indicates that oxidative stress may be involved in this process. Although the adverse effects of high levels of inorganic Hg on the male reproductive system have been investigated, the effects of low levels of exposure are unknown. Therefore, the aim of this study was to investigate the effects of chronic exposure to low concentrations of HgCl₂ on sperm parameters, lipid peroxidation, and antioxidant activity of male rats. Three-month-old male Wistar rats were treated for 30 d and divided into groups: control (saline, i.m.) and HgCl₂ group (i.m., first dose 4.6 μg/kg, subsequent doses 0.07 μg/kg/d). Sperm parameters (count, motility and morphology) and biomarkers of oxidative stress in testis, epididymis, prostate, and vas deferens were analyzed. Mercury treatment produced a reduction in sperm quantity (testis and epididymis) and daily sperm production, following by decrease in sperm motility and increase on head and tail morphologic abnormalities. HgCl₂ exposure was correlated with enhanced oxidative stress in reproductive organs, represented not only by augmented lipid peroxidation but also by changes in antioxidant enzymes activity superoxide dismutase (SOD) and catalase (CAT) and nonprotein thiol levels. In conclusion, chronic exposure to low doses of Hg impaired sperm quality and adversely affected male reproductive functions, which may be due, at least in part, to enhanced oxidative stress.     

Reproductive toxicity of chromium in adult bonnet monkeys (Macaca radiata Geoffrey). Reversible oxidative stress in the semen

The present study was designed to test the hypothesis that oxidative stress mediates chromium-induced reproductive toxicity. Monthly semen samples were collected from adult monkeys (Macaca radiata), which were exposed to varying doses (50, 100, 200 and 400 ppm) of chromium (as potassium dichromate) for 6 months through drinking water. Chromium treatment decreased sperm count, sperm forward motility and the specific activities of antioxidant enzymes, superoxide dismutase and catalase, and the concentration of reduced glutathione in both seminal plasma and sperm in a dose- and duration-dependent manner. On the other hand, the quantum of hydrogen peroxide in the seminal plasma/sperm from monkeys exposed to chromium increased with increasing dose and duration of chromium exposure. All these changes were reversed after 6 months of chromium-free exposure period. Simultaneous supplementation of vitamin C (0.5 g/L; 1.0 g/L; 2.0 g/L) prevented the development of chromium-induced oxidative stress. Data support the hypothesis and show that chronic chromium exposure induces a reversible oxidative stress in the seminal plasma and sperm by creating an imbalance between reactive oxygen species and antioxidant system, leading to sperm death and reduced motility of live sperm.     

Toxic effects of Microcystis cell extracts on the reproductive system of male mice

In this paper, the reproductive toxicity of male mice treated with Microcystis aeruginosa cell extracts containing microcystins was examined. In contrast to the control group, male mice exposed intraperitoneally to 3.33 or 6.67 μg microcystins/kg body weight for 14 days had decreased mean body weight, and the mean absolute weight of the testes and epididymides was decreased. However, the mean relative weight of the testes increased compared to the controls. In addition, histological examination of microcystin-treated mice indicated that the testes were damaged and the space between the seminiferous tubules was more pronounced compared to control mice. The quality of mature sperm in the seminiferous tubules was also decreased in treated mice compared with the control group. Further studies showed that motility and viability of the sperm from microcystin-treated mice were reduced, but no significant difference was found in the concentration and abnormality of the sperm from treated mice compared to the control. This study indicated that microcystins had numerous toxic effects on the reproductive system of male mice.     

Assessment of the reproductive toxicity of inhalation exposure to ethyl tertiary butyl ether in male mice with normal,low active and inactive ALDH2

No data are available regarding aldehyde dehydrogenase 2 (ALDH2) polymorphisms related to the reproductive toxicity possibly caused by ethyl tertiary butyl ether (ETBE). In this study, two inhalation experiments were performed in Aldh2 knockout (KO), heterogeneous (HT) and wild type (WT) C57BL/6 male mice exposed to ETBE, and the data about general toxicity, testicular histopathology, sperm head numbers, sperm motility and sperm DNA damage were collected. The results showed that the 13-week exposure to 0, 500, 1,750 and 5,000 ppm ETBE significantly decreased sperm motility and increased levels of sperm DNA strand breaks and 8-hydroxy-deoxyguanosine in both WT and KO mice, the effects were found in 1,750 and 5,000 ppm groups of WT mice, and all of the three exposed groups of KO mice compared to the corresponding control; furthermore, ETBE also caused decrease in the relative weights of testes and epididymides, the slight atrophy of seminiferous tubules of testis and reduction in sperm numbers of KO mice exposed to ≥500 ppm. In the experiment of exposure to lower concentrations of ETBE (0, 50, 200 and 500 ppm) for 9 weeks, the remarkable effects of ETBE on sperm head numbers, sperm motility and sperm DNA damage were further observed in KO and HT mice exposed to 200 ppm ETBE, but not in WT mice. Our findings suggested that only exposure to high concentrations of ETBE might result in reproductive toxicity in mice with normal active ALDH2, while low active and inactive ALDH2 enzyme significantly enhanced the ETBE-induced reproductive toxicity in mice, even exposed to low concentrations of ETBE, mainly due to the accumulation of acetaldehyde as a primary metabolite of ETBE.     

Oxidative stress and DNA damage in the earthworm <Emphasis Type="Italic">Eisenia fetida</Emphasis> induced by toluene,ethylbenzene and xylene

Superoxide dismutase (SOD), guaiacol peroxidase (POD), catalase (CAT), and the comet assay (SCGE) were used as biomarkers to evaluate the oxidative stress and genotoxicity of toluene, ethylbenzene and xylene in earthworms (Eisenia fetida). The results indicated that the exposure of the three pollutants caused a stress response of the three enzymes, an approximate bell-shaped change (a tendency of inducement firstly and then inhibition with increasing concentrations of the pollutants) was mostly found. The three enzymes tested differed in their sensitivity to different pollutants. While the activity of POD was not significantly changed within the concentration range, the concentration thresholds for significant (P < 0.05) responses to toluene based on SOD and CAT were 5 mg kg⁻¹, respectively. Similarly, the concentration thresholds for significant (P < 0.05) responses to ethylbenzene based on CAT and POD were 10 and 5 mg kg⁻¹, respectively, while the activity of SOD was not significantly changed within the concentration range. Significant responses to xylene based on CAT and POD were 5 mg kg⁻¹, respectively, while the activity of SOD was significantly (P < 0.05) induced at 10 mg kg⁻¹. The SCGE assay results showed that these three pollutants could significantly (P < 0.01) induce DNA damage in earthworms and the clear dose-dependent relationships were displayed, indicating potential genotoxic effects of toluene, ethylbenzene, and xylene on E. fetida. The inducement of DNA damage may be attributed to the oxidative attack of toluene, ethylbenzene, and xylene. Toluene seemed to be more genotoxic as it could induce the higher extent of DNA damage than ethylbenzene and xylene. The results suggest that the SCGE assay of earthworms is simple and efficient for diagnosing the genotoxicity of pollutants in terrestrial environment.     

Valproic acid increases formation of reactive oxygen species and induces apoptosis in postimplantation embryos: a role for oxidative stress in valproic acid-induced neural tube defects

Exposure to the anticonvulsant valproic acid (VPA) during the first trimester of pregnancy is associated with an increased risk of congenital malformations including heart defects, craniofacial abnormalities, skeletal and limb defects, and, most frequently, neural tube defects (NTDs). The mechanisms by which VPA induces teratogenic effects are not fully understood, although previous studies support a role for oxidative stress. To investigate the effects of VPA on early development, a whole-embryo culture model was used to evaluate the protective effects of antioxidants, measure intracellular reactive oxygen species (ROS) levels, and assess markers of oxidative damage and apoptosis. Furthermore, in vivo teratological evaluations of antioxidant protection were also completed. VPA (0.60 mM in embryo culture, 400 mg/kg in vivo) induced significant decreases in embryonic growth and increases in NTDs. Of the antioxidants tested, catalase provided partial protection against VPA-mediated reductions in morphological and developmental growth parameters in both whole-embryo culture and in vivo systems. VPA exposure resulted in an increase in ROS staining in the head region, as assessed by whole-mount staining with 5-(and-6)-chloromethyl-2',7'-dichlorodihydrofluorescein diacetate. Markers of embryonic oxidative damage including 8-hydroxyguanosine, 4-hydroxynonenal adducts, and 3-nitrotyrosine were not affected by VPA treatment. Increased ROS levels were correlated with increased staining for apoptotic markers, as assessed by Western blotting and immunohistochemistry. Addition of catalase to the medium attenuated VPA-induced increases in ROS formation and apoptosis. These studies identify regions of the embryo susceptible to ROS and apoptosis induced by VPA, thus establishing a possible molecular pathway by which VPA exerts teratogenicity.     

Benfotiamine attenuates nicotine and uric acid-induced vascular endothelial dysfunction in the rat

The study has been designed to investigate the effect of benfotiamine, a thiamine derivative, in nicotine and uric acid-induced vascular endothelial dysfunction (VED) in rats. Nicotine (2 mg kg⁻¹ day⁻¹, i.p., 4 weeks) and uric acid (150 mg kg⁻¹ day⁻¹, i.p., 3 weeks) were administered to produce VED in rats. The development of VED was assessed by employing isolated aortic ring preparation and estimating serum and aortic concentration of nitrite/nitrate. Further, the integrity of vascular endothelium was assessed using the scanning electron microscopy (SEM) of thoracic aorta. Moreover, the oxidative stress was assessed by estimating serum thiobarbituric acid reactive substances (TBARS) and aortic superoxide anion generation. The administration of nicotine and uric acid produced VED by impairing the integrity of vascular endothelium and subsequently decreasing serum and aortic concentration of nitrite/nitrate and attenuating acetylcholine-induced endothelium dependent relaxation. Further, nicotine and uric acid produced oxidative stress, which was assessed in terms of increase in serum TBARS and aortic superoxide generation. However, treatment with benfotiamine (70 mg kg⁻¹ day⁻¹, p.o.) or atorvastatin (30 mg kg⁻¹ day⁻¹ p.o., a standard agent) markedly prevented nicotine and uric acid-induced VED and oxidative stress by improving the integrity of vascular endothelium, increasing the concentration of serum and aortic nitrite/nitrate, enhancing the acetylcholine-induced endothelium dependent relaxation and decreasing serum TBARS and aortic superoxide anion generation. Thus, it may be concluded that benfotiamine reduces the oxidative stress and consequently improves the integrity of vascular endothelium and enhances the generation of nitric oxide to prevent nicotine and uric acid-induced experimental VED.