Extract from Aronia melanocarpa, Lonicera caerulea, and Vaccinium myrtillus Improves near Visual Acuity in People with Presbyopia.

Presbyopia is a global problem with an estimated 1.3 billion patients worldwide. In the area of functional food applications, dietary supplements or herbs, there are very few reports describing the positive effects of their use. In the available literature, there is a lack of studies in humans as well as on an animal model of extracts containing, simultaneously, compounds from the polyphenol group (in particular, anthocyanins) and iridoids, so we undertook a study of the effects of a preparation composed of these compounds on a condition of the organ of vision. Our previous experience on a rabbit model proved the positive effect of taking an oral extract of Cornus mas in stabilizing the intraocular pressure of the eye. The purpose of this study was to evaluate the effect of an orally administered ternary compound preparation on the status of physiological parameters of the ocular organ. The preparation contained an extract of the chokeberry Aronia melanocarpa, the honeysuckle berry Lonicera caerulea L., and the bilberry Vaccinium myrtillus (hereafter AKB) standardized for anthocyanins and iridoids, as bioactive compounds known from the literature. A randomized, double-blind, cross-over study lasting with a "wash-out" period of 17 weeks evaluated a group of 23 people over the age of 50, who were subjects with presbyopia and burdened by prolonged work in front of screen monitors. The group of volunteers was recruited from people who perform white-collar jobs on a daily basis. The effects of the test substances contained in the preparation on visual acuity for distance and near, sense of contrast for distance and near, intraocular pressure, and conjunctival lubrication, tested by Schirmer test, LIPCOF index and TBUT test, and visual field test were evaluated. Anthocyanins (including cyanidin 3-O-galactoside, delphinidin 3-O-arabinoside, cyanidin 3-O-glucoside, cyanidin 3-O-rutinoside, cyanidin 3-O-arabinoside) and iridoids (including loganin, sweroside, loganic acid) were identified as substances present in the extract obtained by HPLC-MS. The preliminary results showed that the composition of AKB applied orally does not change visual acuity in the first 6 weeks of administration. Only in the next cycle of the study was an improvement in near visual acuity observed in 92.3% of the patients. This may indicate potential to correct near vision in presbyopic patients. On the other hand, an improvement in conjunctival wetting was observed in the Schirmer test at the beginning of week 6 of administration in 80% of patients. This effect was weakened in subsequent weeks of conducting the experiment to 61.5%. The improvement in conjunctival hydration in the Schirmer test shows the potential beneficial effect of the AKB formulation in a group of patients with dry eye syndrome. This is the first study of a preparation based on natural, standardized extracts of chokeberry, honeysuckle berry, and bilberry. Preliminary studies show an improvement in near visual acuity and conjunctival hydration on the Schirmer test, but this needs to be confirmed in further studies.

Cornelian Cherry (Cornus mas L.) Iridoid and Anthocyanin Extract Enhances PPAR-α, PPAR-γ Expression and Reduces I/M Ratio in Aorta, Increases LXR-α Expression and Alters Adipokines and Triglycerides Levels in Cholesterol-Rich Diet Rabbit Model.

Cornelian cherry (Cornus mas L.) fruits possess potential cardiovascular, lipid-lowering and hypoglycemic bioactivities. The aim of this study is to evaluate the influence of resin-purified cornelian cherry extract rich in iridoids and anthocyanins on several transcription factors, intima/media ratio in aorta and serum parameters, which determine or are valuable indicators of the adverse changes observed in the course of atherosclerosis, cardiovascular disease, and metabolic syndrome. For this purpose, male New Zealand rabbits were fed a diet enriched in 1% cholesterol for 60 days. Additionally, one group received 10 mg/kg b.w. of cornelian cherry extract and the second group 50 mg/kg b.w. of cornelian cherry extract. PPAR-α and PPAR-γ expression in the aorta, LXR-α expression in the liver; cholesterol, triglycerides, adipokines, apolipoproteins, glucose and insulin levels in serum; the intima and media diameter in the thoracic and abdominal aorta were determined. Administration of cornelian cherry extract resulted in an enhancement in the expression of all tested transcription factors, a decrease in triglycerides, leptin and resistin, and an increase in adiponectin levels. In addition, a significant reduction in the I/M ratio was observed for both the thoracic and abdominal aorta. The results we have obtained confirm the potential contribution of cornelian cherry extract to mitigation of the risk of developing and the intensity of symptoms of obesity-related cardiovascular diseases and metabolic disorders such as atherosclerosis or metabolic syndrome.

Acute intranasal intoxication with mercuric chloride taken accidently instead of cocaine - A case report.

CONTEXT: Mercuric chloride (mercury (II) chloride) belongs to inorganic mercury compounds characterized by good water solubility and associated high toxicity. The paper describes an unusual case of intranasal intoxication with corrosive sublimate confused with cocaine by a young male. CASE REPORT: Intranasal administration of corrosive sublimate caused severe local symptoms of chemical burn within the nasal cavity. From the 2nd day the patient developed symptoms of renal dysfunction with transient polyuria and serum retention of nitrogen metabolites. The patient was undergoing chelation therapy with DMPS, N-acetylcysteine and d-penicyllamine. Four procedures of haemodialysis were performed with simultaneous DMPS and N-acetylcysteine treatment. The urine mercury level on the first day of hospitalization was 1989 µg/L, and after 26 days of treatment returned to the physiological level. During treatment renal function was normalized, the patient was discharged in general good condition. DISCUSSION: Mercuric chloride is readily absorbed from the nasal cavity. Its administration may cause intoxication manifested by both chemical burn at the exposure site and systemic symptoms, particularly renal impairment. Even in case of renal dysfunction the use of DMPS seems safe, if haemodialysis is performed at the same time. Simultaneous haemodialysis and chelation therapy may accelerate elimination of mercury from the organism.

The iridoid loganic acid and anthocyanins from the cornelian cherry (Cornus mas L.) fruit increase the plasma l-arginine/ADMA ratio and decrease levels of ADMA in rabbits fed a high-cholesterol diet.

BACKGROUND: Although fruit and vegetable-rich diets have beneficial effects on cardiovascular diseases, we have little knowledge of the impact of fruits and their constituents, iridoids and anthocyanins, on the l-arginine-ADMA-DDAH pathway. Our previous study demonstrated the modulation of those factors by the oral administration of the cornelian cherry fruit. HYPOTHESIS/PURPOSE: We have assessed the effects of the oral administration of two main constituents isolated from the cornelian cherry fruit, iridoid loganic acid and anthocyanins, on l-arginine, its derivatives (ADMA, SDMA), metabolites (DMA, l-citrulline), and the hepatic DDAH activity and its isoform expression in rabbits fed a high-cholesterol diet. We have also analyzed eNOS expression in the thoracic aorta as well as the redox status in blood. STUDY DESIGN: In the present study, we used an animal model of diet induced atherosclerosis. For 60 days, white New Zealand rabbits were fed a standard diet, a 1% cholesterol enriched diet, or concomitantly with the investigated substances. l-arginine, ADMA, SDMA, DMA, and l-citrulline were assessed using the LC-MS/MS method. DDAH activity and redox parameters were analyzed spectrophotometrically. DDAH1 and DDAH2 isoform expressions were assessed by western blotting, mRNA expression of eNOS was quantified by real-time PCR. RESULTS: We demonstrated that the administration of loganic acid (20 mg/kg b.w.), and to a lesser extent of anthocyanins (10 mg/kg b.w.), caused an increase in the l-arginine level and the l-arginine/ADMA ratio. Also, both substances decreased ADMA, DMA, and l-citrulline, but not SDMA levels. Anthocyanins, but not loganic acid, enhanced the activity of DDAH in the liver. Anthocyanins also significantly enhanced both DDAH1 and DDAH2 expression, while loganic acid to a lesser extent enhanced DDAH1 but not DDAH2 expression. Both loganic acid and anthocyanins pronouncedly increased mRNA expression of eNOS in thoracic aortas. Both loganic acid and anthocyanins reversed the blood glutathione level depleted by dietary cholesterol. Cholesterol feeding decreased the blood GPx level, and the change was not reversed by anthocyanins or loganic acid. We did not observe any significant differences in the blood levels of MDA or SOD among the groups. CONCLUSION: Iridoids and anthocyanins may modulate the l-arginine-ADMA pathway in subjects fed a high-cholesterol diet.

Fatal iatrogenic vinorelbine poisoning: a case report.

The paper describes the case of a 69-year-old man with non-small-cell lung cancer who, owing to a mistake, received intravenously 500 mg of vinorelbine. Within 3 days of intoxication, the bone marrow of the patient was damaged with subsequent pancytopenia that did not respond to treatment. On the fifth day after the poisoning, features of intestinal obstruction appeared. The patient died on the sixth day after the drug overdose. The case presented by us constitutes the first description of a fatal iatrogenic poisoning with this drug.

Loganic acid and anthocyanins from cornelian cherry (Cornus mas L.) fruits modulate diet-induced atherosclerosis and redox status in rabbits.

BACKGROUND: Cornelian cherry (Cornus mas L.) is a plant growing in southeast Europe, in the past used in folk medicine. There are many previous publications showing the preventive effects of (poly)phenolic compounds, especially anthocyanins, on cardiovascular diseases, but there is a lack of studies comparing the effects of (poly)phenolics and other constituents of fruits. OBJECTIVES: We have attempted to determine if iridoids and anthocyanins from cornelian cherry fruits may affect the formation of atherosclerotic plaques in the aorta as well as lipid peroxidation and oxidative stress in the livers of cholesterol-fed rabbits. MATERIAL AND METHODS: Fractions of iridoids and anthocyanins were analyzed using the high-performance liquid chromatography (HPLC) and liquid chromatography-mass spectrometry (LC-MS) methods. Loganic acid (20 mg/kg b.w.) and a mixture of anthocyanins (10 mg/kg b.w.) were administered orally for 60 days to rabbits fed with 1% cholesterol. Histopathological samples of the aortas and the livers were stained with hematoxylin and eosin. Lipid peroxidation (malondialdehyde - MDA) and redox status (glutathione - GSH, glutathione peroxidase - Gpx and superoxide dismutase - SOD) were analyzed using spectrophotometrical methods. RESULTS: Both loganic acid (an iridoid) and a mixture of anthocyanins diminished the formation of atherosclerotic plaques in the aorta. Both substances also diminished lipid peroxidation, measured as a decrease of MDA, and attenuated oxidative stress, measured as an increase of GSH in the livers depleted by cholesterol feeding. Unexpectedly, cholesterol feeding decreased the Gpx activity in the liver, which was reversed by both investigated substances. CONCLUSIONS: We have shown that both iridoids and anthocyanins help prevent fed-induced atherosclerosis, and the consumption of fruits rich in these substances may elicit beneficial effects on the cardiovascular system.

Nonfatal and fatal intoxications with pure caffeine - report of three different cases.

Caffeine is not usually perceived as a drug by most people because it is found in many foods and drinks, including caffeinated energy drinks, as well as in over the counter analgesics and cold preparations. Recently in Poland it has become increasingly common to take pure caffeine, bought through online stores, as a psychoanaleptic. This creates a much higher risk of severe and even fatal poisoning in comparison with the risk associated with the abuse of food products and non-prescription medicines containing low doses of caffeine. This paper presents three different cases of poisoning that occurred when pure caffeine was taken as psychostimulant; in cases 1 and 2 poisoning was the result of a single overdose, while in the case 3 poisoning resulted from a cumulative overdose. In the case 1 there was a severe intoxication (persistent vomiting, hypotension, tremor), and the concentration of caffeine in the blood was found to be 80.16 µg/mL. The patient was treated using hemodialysis, which caused a rapid decrease in blood levels of caffeine and relief of the clinical symptoms of poisoning. Cases 2 and 3 were fatal poisonings, and recorded levels of caffeine in post mortem blood samples were 140.64 µg/mL and 613.0 µg/mL. In case 2 the patient died 10 min after admission to hospital as a result of sudden cardiac arrest, which was preceded by an attack of convulsions, and in case 3 death occurred in home and was also sudden in nature. Taking pure caffeine as a stimulant is associated with a high risk of overdose and the development of serious and even fatal poisoning, and those using pure caffeine are generally completely unaware of these risks. In such cases, death is usually sudden due to functional mechanisms.

Iridoid-loganic acid versus anthocyanins from the Cornus mas fruits (cornelian cherry): Common and different effects on diet-induced atherosclerosis, PPARs expression and inflammation.

BACKGROUND AND AIMS: Cardiovascular benefits of fruits are attributed mainly to their (poly)phenolic constituents, especially anthocyanins. The main aim of our study is to compare effects of iridoids and anthocyanins from one fruit on diet-induced atherosclerosis. The cornelian cherry is a native or cultivated plant that grows in many European countries, used in cuisine and folk medicine. In our previous study, we showed its constituents and proved that oral administration of lyophilized fruits to hypercholesterolemic rabbits had preventive effects on atherosclerosis through the activation of PPARα expression. In this study, we have compared the effects of the main constituents of the cornelian cherry:iridoid loganic acid and anthocyanins. METHODS: Our experiment followed the model used in our previous study, in which rabbits were fed 1% cholesterol. RESULTS: We showed that both loganic acid (20 mg/kg b.w.) and a mixture of anthocyanins (10 mg/kg b.w.) administered orally for 60 days had a positive impact on dyslipidemia caused by cholesterol-rich diet, although the effects of anthocyanins were more pronounced. Anthocyanins decreased total and LDL-cholesterol and triglycerides and increased HDL-cholesterol. Loganic acid showed similar effects, but only the triglycerides and HDL-cholesterol changes achieved statistical significance. Anthocyanins, and to a lesser extent loganic acid, significantly decreased intima thickness and intima/media ratio in the thoracic aorta. Both substances decrease ox-LDL in the plasma. Anthocyanins significantly increased expression of PPARγ and α in the liver. Loganic acid also increased their expression, but to a lesser extent. Conversely, loganic acid showed pronounced anti-inflammatory effects, decreasing TNF-α and IL-6 activity. CONCLUSIONS: Our results imply that both substances have a positive effect on factors contributing to the development of diet-induced atherosclerosis. Our results also indicate the potential health benefits of fruits containing anthocyanins and iridoids, and support the idea of creating composed phytopharmaceuticals containing both groups of substances.

Impact of morin-5'-sulfonic acid sodium salt on cyclophosphamide-induced gastrointestinal toxicity in rats.

BACKGROUND: The aim of this study was to evaluate the effect of morin-5'-sulfonic acid sodium salt (NaMSA) on cyclophosphamide-induced gastrointestinal changes in rats. METHODS: Rats received intragastrically 0.9% saline (group C), cyclophosphamide (15 mg/kg) (group CX), NaMSA (100 mg/kg) (group M) or cyclophosphamide (15 mg/kg) with NaMSA (100 mg/kg) (group M-CX), respectively, for 10 days. RESULTS: No histological lesions were observed in the liver and the large intestine in the control group and group receiving NaMSA. In the cyclophosphamide-treated group, a generalized blurred trabecular structure, hepatocyte apoptosis, focal and diffuse necrosis were noticed in the liver and atypia of epithelial cells or adenoma were noticed in the large intestine. In the group receiving both cyclophosphamide and NaMSA, hepatocyte apoptosis in the liver was observed less frequently. Histological examination of the small intestine revealed: low-grade dysplasia adenoma in the C, M, CX and M-CX group (in 44%, 0%, 100%, and 55.6% of specimens, respectively) with adenocarcinoma in 55.6% of specimens in the cyclophosphamide-receiving group only. Adenoma with high-grade dysplasia was observed in the control and NaMSA-receiving group with a similar frequency (22%). In addition to the histological evaluation, blood cell count parameters, as well as total protein concentration, blood glucose level, amylase, ALT, AST and GGTP activities were evaluated. Cyclophosphamide impaired weight gain, decreased blood cell count parameters and total protein concentration, and increased the GGTP activity. Those changes were not reversed by NaMSA. CONCLUSIONS: Summing up, NaMSA may protect against some cyclophosphamide-induced histological abnormalities in the gastrointestinal tract, including intestinal neoplasia in rats.

Application of Cornelian Cherry Iridoid-Polyphenolic Fraction and Loganic Acid to Reduce Intraocular Pressure.

One of the most common diseases of old age in modern societies is glaucoma. It is strongly connected with increased intraocular pressure (IOP) and could permanently damage vision in the affected eye. As there are only a limited number of chemical compounds that can decrease IOP as well as blood flow in eye vessels, the up-to-date investigation of new molecules is important. The chemical composition of the dried Cornelian cherry (Cornus mas L.) polar, iridoid-polyphenol-rich fraction was investigated. Loganic acid (50%) and pelargonidin-3-galactoside (7%) were found as the main components. Among the other constituents, iridoid compound cornuside and the anthocyans cyanidin 3-O-galactoside, cyanidin 3-O-robinobioside, and pelargonidin 3-O-robinobioside were quantified in the fraction. In an animal model (New Zealand rabbits), the influence of loganic acid and the polyphenolic fraction isolated from Cornelian cherry fruit was investigated. We found a strong IOP-hypotensive effect for a 0.7% solution of loganic acid, which could be compared with the widely ophthalmologically used timolol. About a 25% decrease in IOP was observed within the first 3 hours of use.

The impact of morin, a natural flavonoid, on cyclophosphamide-induced changes in the oxidative stress parameters in rat livers.

BACKGROUND: Cyclophosphamide (CPX) has many adverse effects, partly due to oxidative stress induction in various tissues. Morin is one of the natural flavonoids with strong antioxidant properties. OBJECTIVES: The aim of the current research was to estimate the influence of morin on changes in antioxidant parameters in rat livers after cyclophosphamide administration. MATERIAL AND METHODS: The study was performed on Wistar rats. The rats in Group C received 0.9% saline; those in Group CX received cyclophosphamide (CPX); and those in Group M-CX received CPX with morin. Cyclophosphamide and morin were given by gastric gavage for 10 consecutive days at doses of 15 mg/kg and 100 mg/kg, respectively. Malondialdehyde (MDA) and glutathione (GSH) concentrations, superoxide dismutase (SOD) activity and catalase (CAT) activity were determined in liver tissue homogenates. RESULTS: CPX caused a significant decrease in SOD activity and GSH levels, but only the latter was fully restored by morin. There were no significant differences in CAT activity in the various groups. CPX also insignificantly decreased MDA levels, which was aggravated by co-administration of morin. CONCLUSIONS: The results obtained indicate that morin may exert some protective action on CPX-induced changes in the antioxidant state in rat livers.

Suicidal overdose with relapsing clomipramine concentrations due to a large gastric pharmacobezoar.

The paper presents a case of fatal intoxication after massive sustained-release clomipramine overdosage with prolonged toxicity related to a large gastric pharmacobezoar. 42-year-old female was admitted to the toxicology unit 14 h after drugs ingestion. At admission patient was deeply unconscious, required controlled mechanical ventilation. Serum total level of TCAs was 1955 ng/mL. Gastric lavage revealed no pills. Within the next 12h the patient's clinical condition improved. TCAs level decreased to 999 ng/mL. However, after another 10h the clinical condition started deteriorating again and the patient went into a deep coma requiring controlled mechanical ventilation. TCAs level increased to 2011 ng/mL. X-ray and computed tomography revealed large pharmacobezoar consisted from radio-opaque pills. In the 28th h of hospitalization gastrotomy was performed, confirming presence of pharmacobezoar formed from Anafranil SR tablets. After surgery TCAs level was gradually decreasing. However, the patient's condition did not improve, she died 32 h after gastrotomy. Post-mortem analyses revealed drug and its metabolite toxic levels in blood (clomipramine - 1729 ng/mL, norclomipramine - 431 ng/mL) and toxic levels in internal organs: myocardium (clomipramine - 14,420 ng/g, norclomipramine - 35,930 ng/g), vitreous humor (clomipramine - 1000 ng/mL, norclomipramine - 3110 ng/mL). Described case report indicates that sustained release clomipramine tablets may form pharmacobezoar. X-ray and computed tomography examinations should be considered in cases of massive abuse of sustained release clomipramine, particularly if symptoms of intoxication are recurrent or persistent.

Effect of cyclophosphamide and morin-5'-sulfonic acid sodium salt, alone or in combination, on ADMA/DDAH pathway in rats.

BACKGROUND: The aim of the study was to evaluate the effect of cyclophosphamide (CPX) and morin-5'-sulfonic acid sodium salt (NaMSA) on plasma asymmetric dimethylarginine (ADMA) level and dimethylarginine dimethylaminohydrolase (DDAH) activity in rat liver. METHODS: The study was performed on Wistar rats receiving normal saline, CPX (15 mg/kg/day), NaMSA (100 mg/kg/day) or both CPX and NaMSA for 10 consecutive days. RESULTS: Significant decrease in ADMA level was found in all groups when compared to the control. DDAH activity in the liver was significantly higher in CX group compared to the control group. CONCLUSION: Obtained results of ADMA/DDAH pathway parameters require further research.

Effect of quercetin-5'-sulfonic acid sodium salt on SOD activity and ADMA/DDAH pathway in extracorporeal liver perfusion in rats.

BACKGROUND: Quercetin-5'-sulfonic acid sodium salt (NaQSA) exerts good aqueous solubility, strong antioxidant activity and low toxicity. OBJECTIVES: The aims of this study were to investigate the effect of NaQSA on superoxide dismutase (SOD) activity and ADMA/DDAH pathway during extracorporeal liver perfusion (ELP). MATERIAL AND METHODS: The study was carried out on male Wistar rats. Isolated livers were perfused with Krebs-Henseleit bicarbonate buffer (KHB) + 1 microM ADMA (group C), or with KHB + 1 microM ADMA and either 10 microM NaQSA (Q10) or 50 microM NaQSA (Q50). In group 0 (sham) livers were perfused with KHB alone. Levels of ADMA, alanine (ALT) and aspartate (AST) aminotransferases activities were measured during perfusion. After 90 min. of perfusion superoxide dismutase (SOD) and dimethylarginine dimethylaminohydrolase (DDAH) activities were estimated in liver homogenates. RESULTS: DDAH activity in Q10 group was significantly higher as compared to control and Q50 groups. No significant differences were observed between Q50 and control group. The decrease in ADMA concentration during perfusion was observed in all groups, but the most pronounced in the group Q10 and the least in group Q50. During perfusion AST activities were the lowest in Q50 group. No significant difference in SOD activity in groups perfused with NaQSA as compared to control group was noted. CONCLUSIONS: The impact of NaQSA on ADMA/DDAH system depends on its concentration. In lower concentration NaQSA exerted some beneficial properties which vanished in higher concentration. No increase in SOD activity during perfusion with NaQSA was observed.

Omeprazole does not change the oral bioavailability or pharmacokinetics of vinpocetine in rats.

Previous studies proved that food strongly enhanced the bioavailability of vinpocetine. Food may change the pharmacokinetics of a drug by affecting various factors, including gastrointestinal pH. However, the influence of proton pump inhibitor-induced pH alterations on vinpocetine pharmacokinetics is not known. The aim was to evaluate the influence of omeprazole on the pharmacokinetics of oral vinpocetine. One group of male Wistar rats received single oral doses of vinpocetine (2 mg/kg - regimen V). In the second group, omeprazole (10 mg/kg) was administered intraperitoneally for 5 days before vinpocetine administration (regimen OV). For analysis of vinpocetine pharmacokinetics, blood samples were obtained before and 0.25, 0.5, 1, 1.5, 2, 3, 4, 5, 6, 8, 10 and 12 h after vinpocetine administration. Vinpocetine concentrations were measured by high performance liquid chromatography (HPLC). The mean values of AUC(0-t), AUC(0-inf) and C(max) in regimen V were very similar to respective values in regimen OV. The mean T(max) in both regimens was estimated for 1.5 h. There were no statistically significant differences between both regimens. In conclusion, omeprazole did not affect the pharmacokinetic profile of vinpocetine.

Fulminant hepatic failure in woman with iron and non-steroidal anti-inflammatory drug intoxication.

A 17-year-old, previously healthy female ingested 16,000 mg iron sulphate (96.15 mg of iron ions per kg of b.wt.) with a suicidal intent. The patient was admitted to a toxicology unit 10 hours after the drug ingestion. Serum iron concentration at admission was 2351 µg% (421.0 µmol/L). In the course of the intoxication, hemorrhagic gastritis, renal insufficiency and increasing signs of fulminant hepatic failure complicated with coagulopathy and encephalopathy were observed. Treatment with deferoxamine was started immediately after admission to the hospital and continued for 15 hours until the serum concentration of iron decreased to 145 µg% (25.9 µmol/L). Patient was qualified for liver transplant, therefore albumin dialysis as a bridge to liver transplantation was performed. In spite of two procedures of albumin dialysis using the Prometheus system, deep coma, shock and respiratory insufficiency developed. The patient died 80 hours after iron ingestion. In the presented case, the ingestion of a very high dose of iron and late introduction of deferoxamine treatment contributed to fulminant liver failure and fatal outcome of the intoxication.

Influence of commonly used clinical antidotes on antioxidant systems in human hepatocyte culture intoxicated with alpha-amanitin.

α-Amanitin (α-AMA) is the main toxin of Amanita phalloides and its subspecies (A. virosa and A. verna). The primary mechanism of α-AMA toxicity is associated with protein synthesis blocking in hepatocytes. Additionally, α-AMA exhibits prooxidant properties that may contribute to its severe hepatotoxicity. The aim of the present study was to assess the effect of α-AMA on lipid peroxidation and the activities of superoxide dismutase (SOD) and catalase (CAT) in human hepatocyte culture. The effects of benzylpenicillin (BPCN), N-acetyl-L-cysteine (ACC), and silibinin (SIL) on SOD and CAT activities and on lipid peroxidation in human hepatocyte culture intoxicated with α-AMA were also examined. In human hepatocyte culture, 48-hour exposure to α-AMA at a 2-µM concentration caused an increase in SOD activity, a reduction of CAT activity, and a significant increase in lipid peroxidation. Changes in SOD and CAT activity caused by α-AMA could probably enhance lipid peroxidation by increased generation of hydrogen peroxide combined with reduced detoxification of that oxygen radical. The addition of antidotes (ACC or SIL) to the culture medium provided more effective protection against lipid peroxidation in human hepatocytes intoxicated with α-AMA than the addition of BPCN, possessing no antioxidant properties.

Effects of melatonin on lipid peroxidation and antioxidative enzyme activities in the liver, kidneys and brain of rats administered with benzo(a)pyrene.

Benzo(a)pyrene [B(a)P] is a widespread pollutant with a mutagenic, carcinogenic and strong prooxidative properties. The present study evaluated the melatonin effects on lipid peroxidation products levels and on activity of antioxidative enzymes in the course of B(a)P intoxication. Control rats were treated with 0.9% NaCl; another group was given 10mg melatonin/kg bw; a third group was injected twice a week with B(a)P at the dose of 10mg/kg bw; the fourth group received both B(a)P and melatonin at the dose as mentioned above. The experiment continued for 3 months. In homogenates of brain, liver and kidneys lipid peroxidation was appraised by evaluation of malonyldialdehyde and 4-hydroxyalkenal (MDA+4HDA) levels. Activities of glutathione peroxidase (GPx), superoxide dysmutase (SOD) and catalase (CAT) and concentration of reduced glutathione (GSH) were also estimated. In animals receiving both B(a)P and melatonin, lower levels of MDA+4HDA were observed in all organs as compared to the group treated with B(a)P only. Following administration of B(a)P, GSH level decreased in brain and kidney. Melatonin in combination with B(a)P induced rises in the GSH level in liver and brain, as compared to the receiving B(a)P alone. The activity of SOD increased in the rats treated with melatonin alone but the highest activity was observed in rats treated with B(a)P plus melatonin. CAT activity in the melatonin-treated group increased in brain and liver. Similar to SOD, activity of the enzyme significantly increased in the group treated in combination with B(a)P and melatonin, as compared to the remaining groups in all tested tissues. The results suggest that melatonin protects cells from the damaging action of B(a)P. According to our knowledge, there are no studies describing the effects of melatonin on lipid peroxidation markers and antioxidative enzymes during intoxication of B(a)P in the brain, liver and kidneys. The results of present study give a perspective for further studies of its free radical scavenger properties in prevention of oxidative stress dependent diseases, among others cancers caused by carcinogens such as B(a)P.

Benzylpenicyllin and acetylcysteine protection from α-amanitin-induced apoptosis in human hepatocyte cultures.

High mortality rate in Amanita phalloides (death cap) intoxications is a result of the acute liver failure following hepatocyte damage due to hepatocellular uptake of amatoxins. α-Amanitin (α-AMA), the major amatoxin, blocks a RNA polymerase II, which results in inhibition of transcription of DNA and protein synthesis processes and leads to hepatocyte death. α-AMA is also a strong apoptosis inductor and may play a significant role in pathogenesis of hepatic damage in course of amanitin intoxication. The aim of this study was to examine mechanisms of α-AMA-induced apoptosis in human hepatocytes, as well as in determining if commonly clinically used antidotes benzylpenicillin (BPCN) and N-acetylcysteine (ACC) are able to protect human hepatocytes against α-AMA-induced apoptosis. The experiment was performed on cultured human hepatocytes. Viability of cultured hepatocytes was assessed using the MTT assay, whereas apoptosis processes were evaluated by the electron microscopy, detection of DNA laddering, determination of caspase-3 activity, and measuring annexin V, p53 and Bcl-2 protein concentration. Cytotoxicity and apoptosis evaluation were performed after 24 h of exposure to α-AMA and/or tested antidotes.Both ACC and BPCN were well tolerated by human hepatocyte cultures, and exposure to those substances did not reduce cell viability nor induce apoptosis. Exposure of hepatocytes to α-AMA at concentration 2µM resulted in derangement of cell cultures, apoptosis and significant reduction in cell viability. α-AMA-induced apoptosis in human heptocyte cultures is p53- and caspase-3-dependent. Human hepatocyte cultures are exposed simultaneously to α-AMA and tested antidotes (BPCN or ACC) showed significantly higher cell viability and significantly lower values of apoptosis markers compared to the cultures exposed to α-AMA only.