Human & Experimental Toxicology

Environmental chemicals and drugs can induce cardiotoxicity, mainly by generating free radicals. Reactive oxygen species play a critical role in the pathogenesis of cardiac tissue injury. This highlights a need for prevention of cardiotoxicity by scavenging free radicals. Melatonin has been shown to act as a protector against various conditions in which free radicals cause molecular and tissue injury. Some of the mechanisms by which melatonin operates as a free radical scavenger and antioxidant have been identified. The importance of endogenous melatonin in cardiovascular health and the benefits of melatonin supplementation in different cardiac pathophysiological disorders have been shown in a variety of model systems. Melatonin continues to attract attention for its potential therapeutic value for cardiovascular toxicity. The therapeutic potential of melatonin in treatment of cardiotoxicities caused by various chemicals along with suggested molecular mechanisms of action for melatonin is reviewed.

Nonylphenol (NP) is an environmental contaminant, which adversely affects the male fertility due to endocrine disruption and generation of oxidative stress. The current research was planned to assess the effects of nobiletin (NOB), a polymethoxyflavone, on NP-induced testicular damages. Twenty-four male rats were divided into 4 groups: control (0.1% DMSO), NP group (50 mg/kg), NP+NOB group (50 mg/kg + 25 mg/kg), and NOB group (25 mg/kg). Our results revealed that NP brought down the activities of catalase (CAT), superoxide dismutase (SOD), glutathione peroxidase (GPx) and glutathione reductase (GSR), while elevated the level of thiobarbituric acid reactive substances (TBARS). Additionally, NP decreased the level of follicle-stimulating hormone (FSH), luteinizing hormone (LH), plasma testosterone, daily sperm production (DSP), epididymal sperm count, viability, motility, gene expression of testicular steroidogenic enzymes (StAR, 3β-HSD and 17β-HSD) and anti-apoptotic protein (Bcl-2), as well as number of spermatogenic cells belonging to various stages. Whereas, sperm (head, mid-piece/neck and tail) abnormalities, expression of apoptotic proteins (Bax and caspase-3), and histopathological damages were increased. However, NOB remarkably reversed all the damages caused by NP. Therefore, it is deduced that NOB could be used as a potential therapeutic to counter the NP-prompted oxidative stress and apoptotic damages in testes.

Butylated hydroxyanisole (BHA) has been widely used in the cosmetics, pharmaceutical, and food industries due to its antioxidant activity. Despite the antioxidant effects, reported adverse effects of BHA at the cellular level have made its use controversial. In this regard, this study was performed to elucidate the potential toxicity mechanism caused by BHA at the molecular level in zebrafish embryos. For this purpose, zebrafish embryos were exposed to BHA at levels of 0.5, 1, 5, 7.5 and 10 ppm and monitored at 24, 48, 72 and 96 hours. Survival rate, hatching rate and malformations were evaluated. We examined the potential for reactive oxygen species (ROS) production and apoptosis signalling accumulation in the whole body. Moreover, we evaluated histopathological and immunohistochemical (8-OHDG) characterization of the brain in zebrafish embryos at the 96th hour. We also examined apoptosis, histopathological and immunohistochemical (8-OHDG) characteristics in 96 hpf zebrafish larvae exposed to tertiary butylhydroquinone (TBHQ), one of the major metabolites of BHA, at doses of 0.5, 2.5, 3.75 and 5 ppm. Consequently, it has been considered that increased embryonic and larval malformations in this study may have been caused by ROS-induced apoptosis. After 96 h of exposure, positive 8-OHdG immunofluorescence, degenerative changes, and necrosis were observed in the brain of BHA and TBHQ-treated zebrafish larvae in a dose-dependent manner. BHA and TBHQ exposure could lead to an increase in 8-OHdG activities by resulting oxidative DNA damage. In particular, the obtained data indicate that the induction of ROS formation, occurring during exposure to BHA and/or multiple hydroxyl groups, could be responsible for apoptosis.

Over the last decade, electrolyzed water (EW) produced by salt and tap water has gained importance due to its antimicrobial effects. Regarding to chlorine-based compounds, EW also used in post-harvest safety of food processing and sterilization of surfaces. The latest studies suggested that EW might act as wound healing agent due to anti-infective and cell proliferative properties. In this study, we evaluated acute contact cytotoxicity in L929 mice fibroblast cells and wound healing activity of EWs in vitro. In addition, mutagenic activity was evaluated by Ames test with and without metabolic activation by S9 fraction and the stability profile of freshly prepared EWs has been followed up. According to the results, strong acid (StAEW) and mixed EW (MEW) showed dose-dependent cytotoxicity due to possible high HOCl concentration, while slightly acidic and catholyte EW (CEW) were not cytotoxic even applied directly for 30 sec. Further, StAEW and CEW showed a significant increase in L929 cell migration in scratch assay. Likewise, with/ without metabolic activation, neither of EWs had shown mutagenic profile in TA 98 and TA100 strains of Salmonella typhimurium. Follow-up of ORP (oxidation-reduction potential), pH and FCC (free chlorine concentration) showed that temperature and light were important storage conditions to maintain a stable profile particularly for ORP and FCC, which are the most important indicators for biological activity of EW. According to the present findings, it can be suggested that particularly StAEW, may represent a valuable wound healing agent with an achievable, economical and easy production system when stored under proper conditions.

Bupivacaine is frequently used for conducting regional anesthesia. When accidentally injected or excessively absorbed into circulation, bupivacaine can induce severe arrhythmia and potentially lead to cardiac arrest. The specific mechanisms underlying this cardiotoxicity, however, remain to be clarified. We transfected HEK-293 cells to express the small conductance calcium-activated potassium type-2 channel (SK₂), and used a whole-cell patch clamp method in order to explore how bupivacaine affected these channels. We subsequently used SK₂ knockout mice to explore the relevance of SK₂ channels in bupivacaine-induced cardiotoxicity in isolating mouse hearts, mounting them on a Langendorff apparatus, and perfusing them with bupivacaine. Using this system, arrhythmia, asystole, and cardiac functions were monitored. We observed dose-dependent inhibition of SK₂ channels by bupivacaine: half-maximal inhibitory concentration (IC50) value = 18.6 μM (95% CI 10.8–32.1). When SK₂ knockout (SK₂ ^−/−) or wild-type (WT) mice were perfused with Krebs-Henseleit buffer (KHB), we did not observe any instances of arrhythmia. When SK₂ ^−/− mice or WT were perfused with KHB containing bupivacaine (40 μM), the time to arrhythmia (T_arrhythmia) and time to asystole (T_asystole) were both significantly longer in SK₂ ^−/− mice relative to WT mice (P < 0.001). Similarly, SK₂ ^−/− mice exhibited a significantly longer time to 25%, 50%, and 75% reductions in heart rate (HR) and rate-pressure product (RPP) relative to WT mice following bupivacaine perfusion (P < 0.001). These results reveal that bupivacaine was able to mediate a dose-dependent inhibition of SK₂ channels in HEK-293 cells, and deletion of SK₂ channels can delay bupivacaine-induced cardiotoxicity in isolated mouse hearts.

Senescence marker protein 30 (SMP30) is a senescence marker molecule and identified as a calcium regulatory protein. Currently, SMP30 has emerged as a cytoprotective protein in a wide range of cell types. However, the role of SMP30 in regulating neuronal survival during cerebral ischemia/reperfusion injury remains unclear. In the present study, we aimed to investigate the biological function and regulatory mechanism of SMP30 on neuronal survival using a cellular model induced by oxygen-glucose deprivation/reoxygenation (OGD/R). The results showed that SMP30 expression was significantly decreased by OGD/R exposure in neurons. Functional experiments demonstrated that SMP30 overexpression significantly rescued the decreased cell viability and attenuated the apoptosis and reactive oxygen species generation in OGD/R-exposed neurons. By contrast, SMP30 knockdown exhibited the opposite effect. Mechanism research revealed that SMP30 overexpression contributed to the activation of nuclear factor erythroid 2-related factor (Nrf2)/antioxidant response element (ARE) signaling associated with downregulation of Kelch-like ECH-associated protein (Keap1). Keap1 overexpression or Nrf2 silencing significantly reversed SMP30-mediated neuroprotection against OGD/R-induced injury. Overall, these findings demonstrate that SMP30 overexpression protects neurons from OGD/R-induced apoptosis and oxidative stress by enhancing Nrf2/ARE antioxidant signaling via inhibition of Keap1. These data highlight the importance of the SMP30/Keap1/Nrf2/ARE signaling axis in regulating neuronal survival during cerebral ischemia/reperfusion injury.

Endemic goitre and associated iodine deficiency disorders (IDDs) are a major concern in public health even in the period of post salt iodization in many regions. Among others the consumption of cyanogenic plants found responsible for the persistence of such diseases. Bamboo shoots (BS) is one such cyanogenic plant food that caused disruption of certain thyroid hormone synthesizing regulatory element as has already been reported in our earlier study. In this investigation the possible mechanism of thyrocytes disruption along with interruption of thyroid hormone biosynthesis by BS has been worked out. Commonly consumed BS, Bambusa Balcooa Roxb (BBR) water extract was analysed by GC MS; three doses below IC₅₀ were administered to thyrocytes in culture with and without iodine. Expressions of thyroglobulin (Tg), pendrin (PDS) and monocarboxylate transporter 8 (MCT8) were evaluated in thyrocytes with cell cycle analysis, reactive oxygen species (ROS) generation, DNA oxidation and apoptotic regulation through Bax, Bcl-2 and p53. Phytochemical analysis of BBR extract revealed the presence of precursors and metabolic end products of cyanogenic glycosides. Dose dependent decrease in expression of Tg and PDS with concomitant decrease in gene expression of these with MCT8 were observed. Increased ROS, DNA oxidation and associated imbalance were found through increased Bax and p53 with decreased Bcl-2 that perturbed thyrocytes cell cycle. Cyanogenic constituents of BBR generates ROS associated oxidative changes in thyrocytes with DNA damage and oxidation and cell cycle disruption followed by inhibition of thyroid hormone synthesizing regulatory elements; addition of extra iodine showed partial prevention.

The high incidence of alcoholic poisoning in Indonesia is caused by the use of methanol as a surrogate ingredient of nonconforming alcohol. The product of methanol metabolism is toxic to the liver. Ranitidine has been studied as an antidote to reduce the effect of methanol toxicity. The present study aimed to assess the effect of ranitidine administration on the liver damage of Wistar rats with acute methanol intoxication. This research was an experimental study with randomized and posttest-only control group design. A total of 24 male Wistar rats divided into four groups, each consisting of six Rats. The control group (K0) served as a reference normal value. The control group (K1) was intoxicated with methanol 7 g/kg body weight (b.w.) through rat gavage tube. Treatment groups were intoxicated with methanol and 15 minutes after that, Ranitidine (30 mg/kg b.w. (P1) or 60 mg/kg b.w. (P2)) was given intraperitoneally. Ranitidine administration was proven to protect the liver tissue from damage due to methanol intoxication as was indicated from the histopathological examination. Ranitidine with dose 60 mg/kg b.w. is better in reducing the degree of liver tissue damage in acute methanol intoxication rats.

The organic compound di(2-ethylhexyl) phthalate (DEHP) is widely used as a plasticizer in many products. Exposure to DEHP has been reported to lead to adverse pregnancy outcomes by suppressing placenta growth and development. The aim of this study was to determine the gene expression profiles of rat placenta exposed to (DEHP) and identify genes crucial for the DEHP response. Three groups of Wistar rats were administered an intragastric dose of 1,000 mg/kg DEHP, 500 mg/kg DEHP, or corn oil, RNA was isolated from placenta tissue, and hybridization was performed. Gene expression profiles were analyzed by identifying functional enrichment, differentially expressed genes (DEGs), protein–protein interaction (PPI) networks and modules, and transcription factor (TF)-miRNA-target regulatory networks. We obtained 2,032 DEGs, including cytochrome P450, family 2, subfamily R, polypeptide 1 (CYP2R1), sterol O-acyltransferase 2 (SOAT2), and 24-dehydrocholesterol reductase (DHCR24) from the steroid biosynthesis pathway and somatostatin receptor 4 (SSTR4) and somatostatin receptor 2 (SSTR2) in the neuroactive ligand-receptor interaction pathway. The PPI network included 476 nodes, 2,682 interaction pairs, and three sub-network modules. Moreover, eight miRNAs, three TFs, and 176 regulatory pairs were obtained from the TF-miRNA-target regulatory network. CYP2R1, SOAT2, DHCR24, SSTR4, and SSTR2 may affect DEHP influence on rat placenta development.

Mercury is a widespread pollutant. Mercuric ions uptake into tubular cells is supported by the Organic anion transporter 1 (Oat1) and 3 (Oat3) and its elimination into urine is through the Multidrug resistance-associated protein 2 (Mrp2). We investigated the effect of recombinant human erythropoietin (Epo) on renal function and on renal expression of Oat1, Oat3, and Mrp2 in a model of mercuric chloride (HgCl₂)-induced renal damage. Four experimental groups of adult male Wistar rats were used: Control, Epo, HgCl₂, and Epo + HgCl₂. Epo (3000 IU/kg, b.w., i.p.) was administered 24 h before HgCl₂ (4 mg/kg, b.w., i.p.). Experiments were performed 18 h after the HgCl₂ dose. Parameters of renal function and structure were evaluated. The protein expression of Oat1, Oat3 and Mrp2 in renal tissue was assessed by immunoblotting techniques. Mercury levels were determined by cold vapor atomic absorption spectrometry. Pretreatment with Epo ameliorated the HgCl₂-induced tubular injury as assessed by histopathology and urinary biomarkers. Immunoblotting showed that pretreatment with Epo regulated the renal expression of mercury transporters in a way to decrease mercury content in the kidney. Epo pretreatment ameliorates HgCl₂-induced renal tubular injury by modulation of mercury transporters expression in the kidneys.

Acute paracetamol over dose-induced hepatotoxicity is considered an important medical hazard especially among women. Omega-3 long-chain polyunsaturated fatty acids (Omega-3 PUFAs) daily doses are nowadays recommended for their antioxidant and anti-inflammatory potentials. Fourier transform infrared (FTIR) spectroscopy is considered a reliable method in analyzing cellular alterations and is now efficiently used to diagnose several diseases and the efficacy of drugs even in the early stages. The aim of our study was to evaluate the hepatoprotective effect of Omega-3 PUFAs against paracetamol-induced hepatotoxicity in rats confirmed through measuring protein alterations in hepatocytes by FTIR. Rats were pretreated with Omega-3 PUFAs (50 and 100 mg/kg) for 21 days prior to oral ingestion of paracetamol. FTIR results revealed that Omega-3 PUFAs (50 mg/kg) limited the toxic effects of paracetamol by restoring the hepatic amide I to amide II ratio. In addition; biochemical analyses demonstrated that serum ALT, AST, Cholesterol, LDL-cholesterol and Il-6 levels as well as hepatic TNF-α, MDA, NOx levels were decreased. Besides; serum HDL-cholesterol level and hepatic GSH level were increased. Histopathological examinations of hepatic sections validated the hepatoprotective potential. The overall effect of this dose was comparable to those of the usual recommended hepatoprotective supplement; silymarin. In conclusion; it would be recommended to use Omega-3 PUFAs in low doses on daily bases as a hepatoprotective agent.

Bisphenol A (BPA), an imperative environmental contaminant used in polycarbonate plastics. Due to limited information concerning the effect of BPA on lungs, this study design to assess whether BPA cause alterations in histopathology and trace metal content in lungs of rats. They were divided into five groups with five rats per group. Group I was named as control group. Group L6 and L12 were received BPA (10 mg/kg body weight/day) for 6 weeks and 12 weeks respectively. Group H6 and L12 were given BPA (25 mg/kg body weight/day) for 6 weeks and 12 weeks respectively. Considerable alteration in Cu, Zn and Fe was detected in experimental groups. BPA also caused significant increase in the expression of tumor necrosis factor α that mediate the pulmonary inflammatory response. Comparative study of resolved proteins i.e. 72 KDa (matrix metalloproteinase 2 fragment) and 109 KDa (nucleolin) on SDS-PAGE showed their altered expression in experimental groups. Histopathology of experimental groups revealed altered architecture of lungs. Special staining of BPA treated groups showed significant number of mast cells in alveoli and bronchioles. Prolonged administration of BPA causes deleterious aggravating lung damage even at extremely low dose, so the use of BPA should be prohibited in plastic synthesizing industries.