|Year : 2015 | Volume
| Issue : 1 | Page : 41-48
Protective effect of Nigella sativa against cerebral ischemia and sodium valproate-induced hepatotoxicity
Elsayed A Abd El Latif, Rezk A Sanad, Omaima M Abdallah, Yasmin M Ismail MSc
Department of Pharmacology and Therapeutics, Faculty of Medicine, Benha University, Benha, Egypt
|Date of Submission||13-May-2015|
|Date of Acceptance||15-May-2015|
|Date of Web Publication||26-Nov-2015|
Yasmin M Ismail
Department of Pharmacology, Faculty of Medicine, Benha University, 3112 Benha
Source of Support: None, Conflict of Interest: None
Nigella Sativa (NS) is one of the traditionally used herb well known for its healing properties. The most of the therapeutic properties of this plant is due to the presence of Thymoquinone (TQ), the major bioactive component of the essential oil. TQ is also a promising dietary chemopreventive agent for the treatment of number of diseases. Very low level of toxicity has been reported through acute and chronic toxicity studies.
The present study aimed to investigate the anti-ischemic effect of NS using carotid artery occlusion model in rats.This study also aimed to investigate the effect of NS on SVP induced hepatotoxicity.
0Animals given NS and TMZ for 21 days then subjected to 45 min for brain ischemia then reperfusion.Animals administered NS, SVP for 21 days then subjected to serum measurement of serum ALT & AST and liver histopathological examination.
NS significantly reduced the percent of necrosis and reduced the size of cerebral infarction compared to control and enhanced the effect of TMZ on cerebral ischemia.NS produced significant decrease in serum ALT and AST and improvement of histopathlogical picture compared to SVP group.
These results indicate that the NS could have a therapeutic effect against cerebral ischemia.NS has protective effect against SVP induced hepatotoxicity.
Keywords: Cerebral ischemia, hepatotoxicity, Nigella sativa
|How to cite this article:|
Abd El Latif EA, Sanad RA, Abdallah OM, Ismail YM. Protective effect of Nigella sativa against cerebral ischemia and sodium valproate-induced hepatotoxicity. Benha Med J 2015;32:41-8
|How to cite this URL:|
Abd El Latif EA, Sanad RA, Abdallah OM, Ismail YM. Protective effect of Nigella sativa against cerebral ischemia and sodium valproate-induced hepatotoxicity. Benha Med J [serial online] 2015 [cited 2018 Jan 23];32:41-8. Available from: http://www.bmfj.eg.net/text.asp?2015/32/1/41/170558
| Introduction|| |
Nigella sativa (of the family Ranunculaceae) is a plant that is synonymous with Nigella cretica and is commonly called black cumin. Other names include Kalonji seeds, Ajaji, black caraway seed, and Habbatu Sawda  . The medicinal usage of these seeds mostly encompasses diarrhea and abdominal pain, dyslipidemia, asthma and cough, headache, dysentery, renal calculi, infections, obesity, back pain, hypertension, and dermatological problems  . The essential oil of the seeds contains a variety of molecules, of which thymoquinone (TQ) is considered as the main component of the entire seed  . In folk medicine, it has been traditionally used for a variety of applications, including treatments related to respiratory health, stomach, and intestinal diseases, kidney and liver function, circulatory and immune system support, and for general overall well-being  . N. sativa and TQ inhibit nonenzymatic lipid peroxidation  . TQ can attenuate picrotoxin-induced seizures, as well as potentiate the protective effects of sodium valproate (SVP) when coadministered  . Isolated TQ appears to be relatively safe, with an oral LD 50 of 794.3 mg/kg in rats and 870.9 mg/kg in mice, which is ~100-150 times higher than its therapeutic level. There have been some isolated cases of topical usage of N. sativa causing contact dermatitis, suggesting that it is possible to be allergic to the seed oil  . SVP is the sodium salt of valproic acid and is an anticonvulsant used in the treatment of epilepsy, anorexia nervosa, panic attack, anxiety disorder, and bipolar disorder, as well as other psychiatric conditions requiring the administration of a mood stabilizer  . SVP is a weak blocker of sodium ion channels; it is also a weak inhibitor of enzymes that deactivate GABA, such as GABA transaminase. It may also stimulate the synthesis of GABA, but the direct mechanism is not known  . The present study aimed to investigate the anti-ischemic effect of N. sativa using carotid artery occlusion model in rats. This study also aimed to investigate the effect of N. sativa on SVP-induced hepatotoxicity.
| Materials and methods|| |
N. sativa was supplied as gelatinous capsules of 450 mg (Mepaco Company, Cairo, Egypt). SVP was supplied as tablets, 200 mg (Sanofi Aventis Company, Paris, France). Trimetazidine (TMZ) was supplied as tablets, 20 mg (Global Napi Pharmaceutical Company, Cairo, Egypt).
N. sativa was dissolved in corn oil  . SVP and TMZ were dissolved in distilled water , .
Experimental animals (60 rats weighing 250-275 g) were kept six per cage under complete healthy conditions throughout the experiment, including a clean environment, good ventilation, and good nutrition. The animals were housed in groups to acclimatize to laboratory conditions for 3 days before the experiment, such as diet, water, and temperature (22°C). Food and water were freely accessible. The experimental rats were under the care of professional technicians and qualified researchers. The work was approved by the ethical committee (Faculty of Medicine, Benha University).
Part I: Effect of N. sativa on cerebral ischemia
This procedure was carried out to investigate the effect of N. sativa either singly or in combination with the anti-ischemic agent TMZ. A total of 36 adult male albino rats were used in this study. They were divided into six groups of six animals each: group 1 (normal control animals) received no drugs; group 2 (the corn oil group) received corn oil in volumes comparable to that of tested drugs; group 3 (the saline group) received saline in volumes comparable to that of tested drugs; group 4 (N. sativa-treated cerebral ischemic rats) received N. sativa at 4 mg/kg intraperitoneally for 3 weeks  and then was exposed to cerebral ischemia by ligation of carotid artery; group 5 (TMZ-treated cerebral ischemic rats) received TMZ at 2.5 mg/kg  for 3 weeks and then was exposed to cerebral ischemia by ligation of carotid artery; and group 6 (TMZ and N. sativa-treated ischemic rats) received both TMZ and N. sativa at the same doses as groups 3 and 4 and then was exposed to cerebral ischemia by ligation of carotid artery.
Method of induction of cerebral ischemia: Cerebral ischemia was induced using the four-vessel-occlusion technique described by Pulsinelli and Briely  . Anesthesia was induced with an intraperitoneal injection of ketamine/xylazine (60 and 6 mg/kg, respectively). Following a dorsal neck incision, the first cervical vertebra and alar foraminae were exposed. Vertebral arteries were electrocauterized permanently. On the next day, under brief anesthesia, the common carotid arteries were dissected from the surrounding tissues and temporarily ligated using the microvascular clamps for 20 min. At the end of the ischemic period, the clamps were released and reperfusion restored. Same procedures were applied to the control group, and, instead of TMZ, an equal amount of intraperitoneal saline was administered daily. After 3 days of reperfusion, the animal was killed with a high-dose anesthetic and cervical dislocation. Craniotomy was performed to remove the brain and preserve it in formalin.
Brain samples were removed and placed in 10% formalin. Initially, they were preserved for 72 h, and then 0.5-cm-thick sections were cut and placed in formalin for another 24 h. Using a rat-brain Atlas More Details  , sections including both hippocampuses were cut and placed in paraffin blocks. From these blocks 4-μm-thick sections were cut and stained with hematoxylin and eosin  .
Cells that showed karyopyknosis, hyperchromasia, or contour roughness were regarded as delayed ischemic damage, and the proportion of damaged cells were expressed as the percentage of total cells counted.
Part II: Assessment of the hepatoprotective activity of N. sativa against sodium valproate-induced hepatotoxicity
A total of 24 male albino rats were used. They were divided into four groups of six rats each: group 1 (the control group) was not administered any drug; group 2 was administered SVP at a dose of 500 mg/kg; group 3 was administered N. sativa at a dose of 0.2 mg/kg; and group 4 was administered N. sativa at a dose of 0.2 mg/kg and SVP at a dose of 500 mg/kg. All drugs were administered intraperitoneally for 3 weeks.
SVP at a dose of 500 mg/kg/day has been described and proven to be hepatotoxic , . For the hepatoprotective effect of N. sativa, the daily dose was calculated to be 0.2 mg/kg/day (intraperitoneal), as already reported by Aniya et al.  . The treatment continued for the same duration as for valproate. Rats from the treated and control groups were killed after 21 days. Blood samples were drawn by means of direct cardiac puncture into nonheparinized capillary tubes. Serum was separated by centrifugation for 5 min and stored at −20°C until biochemical assessment of serum alanine transaminase (ALT) and aspartate aminotransferase (AST) was carried out.
Data were presented as mean ± SD and analyzed with nonparametric statistics using the Kruskal-Wallis test, followed by the Dunn-post test using GraphPad Prism (California, USA) 5 software at P less than 0.05 (N = 6). For other parameters, data were presented as mean ± SD and analyzed using the one-way analysis of variance test, followed by the Tukey Kramer-post test using GraphPad Prism 5 software at P less than 0.05 (N = 6).
| Results|| |
Effect on cerebral infarction
N. sativa showed significant neuroprotective effect on the size of cerebral infarction (2540.17 ± 26 μm 2 ) in comparison with controls (20968.5 ± 13 μm 2 ). No significant difference was found between the control, corn oil (20166 ± 63 μm 2 ), and saline-treated groups (20279 ± 64 μm 2 ) (P > 0.05). Therefore, other groups were compared with the control group. No significant difference was found in the size of cerebral infarction between the N. sativa-treated (2540.17 ± 26 μm 2 ) and the TMZ-treated group (2384.2 ± 31 μm 2 ) (P > 0.05). N. sativa caused significant potentiation of the neuroprotective effect of TMZ (P < 0.05). This has been observed on comparison of the size of cerebral infarction between TMZ+N. sativa-treated group (1039.1 ± 83 μm 2 ) and the TMZ-treated group ([Table 1]).
|Table 1 Effect of Nigella sativa on size of cerebral infarction in comparison with trimetazidine|
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The brain of control rats showed necrotic changes in about 50% of neurons. Infarction size was 20968.5 ± 13 μm 2 ([Figure 1]). The corn oil and saline groups showed necrotic changes in about 50% of neurons, and infarction size was 20166.4 ± 63 and 20279.5 ± 64 μm 2 , respectively ([Figure 2]). The brain of N. sativa group rats showed necrotic changes in 10-50% of neurons, and infarction size was 2540.17 ± 26 μm 2 ([Figure 3]). The brain of TMZ group rats showed necrotic changes in 10-50% of neurons, and infarction size was 2384.2 ± 31 μm 2 ([Figure 4]). The brain of N. sativa+TMZ group rats showed necrotic changes in less than 10% of neurons, and infarction size was 2039.1 ± 83 μm 2 ([Figure 5]).
|Figure 1 A section of the brain from a control group rat showing necrotic changes in more than 50% of neurons. Area of cerebral infarction. H&E, ×40.|
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|Figure 2 A section of the brain from corn oil and water group rats showing necrotic changes in more than 50% of neurons.|
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|Figure 3 A section of the brain from a Nigella sativa-treated rat showing necrotic changes in about 10– 50% of neurons. Arrows are pointing to infarction tissue. H&E, ×40.|
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|Figure 4 A section of the brain from a trimetazidine-treated rat showing necrotic changes in about 10– 50% of neurons. Arrows are pointing to infarction tissue. H&E, ×40.|
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|Figure 5 A section of the brain from a Nigella sativa plus trimetazidine-treated rat showing necrotic changes in less than 10% of neurons. Arrows are pointing to infarction tissue. No part lables need to be inserted. H&E, ×40.|
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Protective effect of N. sativa against valproate-induced hepatotoxicity
Administration of toxic dose of SVP caused statistically significant (P < 0.05) increase in liver enzymes (ALT 142.22 ± 14 and AST 355.5 ± 30) in comparison with the control (ALT 35.5 ± 8 and AST 100.1 ± 21). Elevated liver enzymes decreased significantly (P < 0.05) in the group that received both SVP and N. sativa (ALT 39.5 ± 10 and AST 106.17 ± 19).
The effect of N. sativa on liver function enzymes compared with valproate-induced hepatotoxicity is shown in [Table 2].
|Table 2 The effect of Nigella sativa on liver function enzymes compared with valproate-induced hepatotoxicity|
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The liver of control rats showed normal histological architecture. The hepatic lobules showed central veins from which the hepatocytes radiated in the form of cords. These cords were separated by blood sinusoids ([Figure 6]). The liver of rats from the SVP intoxication group showed focal areas of mononuclear cellular infiltrations. The hepatocytes of the affected area appeared as cords without clear cell boundaries ([Figure 7]).
|Figure 6 A section showing normal histological architecture in control group rats. The hepatic lobules show central veins from which the hepatocytes radiate in the form of cords. These cords are separated by blood sinusoids. The arrow is pointing to central vein. H&E, ×40.|
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|Figure 7 A section showing congestion and dilation of blood sinusoids and central vein in the valproate-intoxicated group. The liver shows focal areas of mononuclear cellular infiltrations. No hepatocyte boundaries. Arrow is pointing to congestion and dilation of blood sinosoids and central vein. H&E, ×40.|
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The liver of rats that received N. sativa revealed more or less normal hepatic architecture ([Figure 8]). On combination of N. sativa with SVP, the liver showed many hepatocytes with small well-circumscribed vacuoles (microvesicular steatosis). Kupffer cells appeared swollen and hypertrophied. Hepatocytes of some rats showed strong acidophilic cytoplasm and small, darkly stained nuclei ([Figure 9]).
|Figure 8 A section showing the effect of Nigella sativa oil on the liver of rats, revealing more or less normal hepatic architecture. Arrow is pointing to normal central vein. H&E, ×40.|
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|Figure 9 A section showing the effect of combination of Nigella sativa oil with sodium valproate, revealing many hepatocytes with small well-circumscribed vacuoles (microvesicular steatosis). Kupffer cells appear swollen and hypertrophied. H&E, ×100.|
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Hepatocytes of some rats showed strong acidophilic cytoplasm and small, darkly stained nuclei.
| Discussion|| |
Many pharmacological investigations have proved the potential therapeutic effects of N. sativa seed, as well as its oil, through various studies  . Generation of free radicals may be the basis of many neurological and neurodegenerative disorders, such as ischemia-reperfusion and seizures  . N. sativa is a herbal drug that phytochemically confirms to have antioxidant properties, which suppress reactive oxygen and nitrogen species formation. This scavenging reactive oxygen and nitrogen species play a major role in protecting the antioxidant defense system  . The pathogenesis of neuronal damage in response to brain ischemia has been widely studied. A number of observations indicate that a series of events occur in response to a transient ischemic insult and attendant reperfusion, which precede and is necessary for eventual neuronal death. Among these are a decrease in the production of ATP, which results in energy failure  . In cerebral ischemia, particularly after reperfusion, free radical production is elevated, thus disrupting the endogenous antioxidant system  . Free-radical-induced lipid peroxidation produces cytotoxic aldehydes, including malondialdehyde (MDA), 4-hydroxynonenal, and acrolein  .
It was already reported that N. sativa decreased lipid peroxidation and increased the antioxidant defense system activity  . The results obtained in the present investigation suggest that N. sativa decreases cerebral ischemia-reperfusion injury-induced pathological stress in the rat model. The size of cerebral infarction and percentage of necrotic changes had significantly decreased in the N. sativa group in comparison with the control group. These findings confirm the results of previous studies that proved that the active constituent of N. sativa has an anti-ischemic effect  . The exact mechanism of action of anti-ischemic activities is not clear. N. sativa seed has a variety of activities, including antioxidant , , lipoxygenase and cyclooxygenase inhibitory activities  , and a decreasing effect on intracellular calcium in mast cells  . The efficacy of TMZ has recently been investigated in reducing reperfusion damage in the myocardium  . In our findings, pretreatment with TMZ for 21 days in animals subjected to 15 min of global cerebral ischemia has exhibited better neurologic outcomes. As regards infarction size and necrotic changes, there was insignificant difference between the N. sativa-pretreated and TMZ-pretreated groups. A combination of N. sativa with TMZ produced reduction in the size of cerebral infarction and decreased neurological damage to 10%. Thus, N. sativa produced enhancement in the anti-ischemic effect of TMZ. To our knowledge, no previous studies have been conducted comparing N. sativa with TMZ. TMZ significantly reduces free radical production  .
Several studies have shown that the antioxidant compounds and free radical scavengers inhibit lipid peroxidation caused by free radicals and excitatory amino acid-induced neuronal injury following ischemia  . Gupta et al. 2004  showed that rats pretreated with N. sativa for 7 days showed improvement in motor performance tests. These findings suggested that the improvement in the motor performance test could be due to reduction in the ischemic lesion. Our finding confirmed the results of a previous study by Al-Omar et al.  , in which chloroform and petroleum extracts of N. sativa significantly reduced the infarct size when compared with middle cerebral artery (MCA) occluded rats. Thus, pretreatment with both extracts of N. sativa showed neuroprotection. Reports from previous in-vivo studies indicated that N. sativa can protect the brain, through its antioxidant activities, from oxidative stress resulting from lipid peroxidation in transient global ischemia to the brain  . El-Abhar et al.  showed that N. sativa has a marked protective effect against ischemia reperfusion injury in gastric mucosa. It has been reported that N. sativa seeds exhibit an inhibitory effect on nitric oxide production  . Excessive production of oxygen free radicals has been reported in ischemic reperfused liver leading to tissue damage  . It has been shown in many studies that supplementation of free radical scavengers is helpful in reducing hepatic ischemia-reperfusion-induced tissue damage  . N. sativa has been identified as a potent antioxidant acting as a free radical scavenger  . Therefore, it should not be surprising that N. sativa pretreatment has a protective effect on hepatic ischemia reperfusion injury in rats.
SVP is a simple fatty acid largely used as an anticonvulsant. The structure facilitates the interaction of SVP with cell membranes, which may participate in both its therapeutic and adverse effects. SVP is extensively metabolized by the liver through glucuronic acid conjugation, mitochondrial β-oxidation and cytosolic ω-oxidation to produce multiple metabolites. As the clinical usage of SVP increased, reports of its hepatotoxicity began to appear. This hepatotoxicity ranged from mild increase in aminotransferase enzyme in 15-30% of patients to liver cell failure and death in some patients. The mechanism by which SVP caused hepatotoxicity was poorly understood , . This study reinforced the previous results , . El-Gharieb et al. 2010  reported that administration of SVP caused significant elevation in liver enzymes. In contrast, Lee et al.  reported that ALT and AST did not change with oral administration of SVP. This might be because of different route and/or dose of drug administration. N. sativa has protective effect against SVP-induced hepatotoxicity, which has been demonstrated by the prevention of elevation in AST and ALT in the group that received both N. sativa and SVP compared with the group that received SVP alone, in which there was marked elevation in both ALT and AST. Our findings confirm the results of a previous study, which demonstrated the protective effect of N. sativa against hepatotoxicity induced by anticancer drug cyclophosphamide (CTX). Toxicity related to anticancer drugs is usually associated with significant hepatotoxicity due to the alteration in ALT and lipid peroxidation. The data presented here show that CTX treatment is associated with disregulation of liver function, as shown by increases in AST and ALT levels. However, these effects can be ameliorated after treatment with N. sativa or TQ, indicating their protective effects against CTX-induced toxicity  .
The current study revealed that intraperitoneal administration of a toxic dose of SVP for 21 days caused focal areas of liver cell degeneration, which were accompanied by occasional eosinophilia of hepatocytes, mononuclear cellular infiltration, prominent microvesicular steatosis (which referred to a variant form of hepatic fatty infiltration), remark cell lines of hepatocytes, and Kupffer cell enlargement. These findings confirmed the results of others  . There was an elevation in serum ALT as well. This is parallel to the results of Loscher and Nau  . Addition of N. sativa to SVP protected against SVP-induced hepatotoxicity indicated by inhibition of serum ALT and improvement of the histopathological picture compared with SVP alone.
As the mechanism by which SVP causes hepatic damage is uncertain, hepatotoxicity was suspected to result from the formation of toxic SVP metabolites  . A possible mechanism of SVP-induced hepatotoxicity was that it caused depression of free radical scavenging enzyme activities  . The cause of liver cell injury was due to decreased plasma and tissue carnitine  . Another cause for liver cell injury might be due to decreased activity of complex IV of the respiratory chain and/or depletion of hepatic pool of glutathione. N. sativa protects against toxic damage caused by free radical generation. One of the first noticeable morphological changes in SVP-treated livers was the accumulation of cytoplasmic fat  . This might be due to the accumulation of esterified SVP and failure of lipid secretion, which were due to interference with vesicular movement, which might lead to fat accumulation. A second study mentioned that SVP interfered with the mitochondrial inner membrane. The loss of that membrane integrity might interfere with the release of calcium from internal stores. This calcium could be a contributor to the secretory problems caused by SVP, either directly affecting the movement of the vesicles along microtubules or indirectly through a second messenger system; thus, lipid accumulation was not only due to overproduction but also due to inability to be secreted at a rate to match the production  . Microvesicular steatosis, which is marked in SVP-associated liver injury, possibly occurs through reactive intermediates that interfere with the process of fatty acid β-oxidation, which inhibit key enzymes in β-oxidation cycle, or through idiosyncrasy for production of toxic SVP metabolites  . Oda et al.  demonstrated acidophilic degeneration observed with light microscopy. Kupffer cell hypertrophy might be due to lipid deposition in their cytoplasm  . Administration of SVP and N. sativa caused some improvement in the histological changes at the light microscopic level. At the light microscopic level, hepatocytes revealed less microvesicular steatosis. Hepatocytes did not return completely to the control pattern. This might be due to the short duration of N. sativa. The findings of the present study indicate that N. sativa has hepatoprotective activity. Daba and Abdel-Rahman  tested TQ in isolated rat hepatocyte as a protective agent against butylhydroperoxide toxicity. Moreover, our findings confirmed the results of previous study by Al-Gharably et al.  , in which pretreatment with N. sativa protected against hepatotoxicity induced by CCL 4 . This hepatoprotective effect is demonstrated through significant prevention of any increase in ALT and AST. Therefore, it can be mentioned that N. sativa behaves as an antioxidant and protects liver against the detrimental effects of SVP. The antioxidant action of N. sativa and TQ may explain the protective effect of these agents against hepatotoxic models in vivo and in vitro  , as well as against liver fibrosis and cirrhosis  .
| Conclusion|| |
The result of this study revealed that N. sativa has a protective effect against cerebral ischemia, which is proved by reduction in the percentage of necrosis and decrease in infarction size in the N. sativa-treated group in comparison with the control group. Moreover, N. sativa enhanced the anti-ischemic effect of TMZ. N. sativa also has a protective effect against SVP-induced hepatotoxicity, which is proved by the reduction in elevated serum ALT and AST and improvement in histopathological picture in the N. sativa-treated group in comparison with the control group.
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Conflicts of interest
There are no conflicts of interest.
| References|| |
Ahmad A, Husain A, Mujeeb M, Khan SA, Najmi AK, Siddique NA, et al
.. A review on therapeutic potential of Nigella sativa
: a miracle herb. Asian Pac J Trop Biomed 2013; 3
Ali BH, Blunden G. Pharmacological and toxicological properties of Nigella sativa
. Phytother Res 2003; 17
Nickavar B, Mojab F, Javidnia K, Amoli MA. Chemical composition of the fixed and volatile oils of i L. from Iran. Z Naturforsch C 2003; 58
Anwar MA. Nigella sativa
. Malaysia J Lib Infor Sci 2005; 10
Houghton PI, Zarka R, De Las Heras B, Hoult JRS. Fixed oil of Nigella sativa
and derived thymoquinone inhibit eicosanoid generation in leukocytes and membrane lipid perioxidation. Plasma Med 1995; 61
Raza M, Alghasham AA, Alorainy MS, El-Hadiyah TM. Potentiation of valproate-induced anticonvulsant response by Nigella sativa
seed constituents: the role of GABA receptors. Int J Health Sci (Qassim) 2008; 2
Ali Z, Ferreira D, Carvalho P, Avery MA, Khan IA. Nigellidine-4-O-sulfite, the first sulfated indazole-type alkaloid from the seeds of Nigella sativa
. J Nat Prod 2008; 71
De Gelder B, Hadjikhani N. Non-conscious recognition of emotional body language. Neuroreport 2006; 17
de Gelder B, Morris JS, Dolan RJ. Unconscious fear influences emotional awareness of faces and voices. Proc Natl Acad Sci USA 2005; 102
Kruse HJ, Kuch H. Etifoxine: evaluation of its convulsant profile in comparison with sodium valproate, phenytoin and clonazepam. Arzneimittelforschung 1985; 35
Ganesh M, Jeraldmaria Antony G, Saravankumar A, Rajesh R, Rajasekar K. A new validated spectrophotometric method for determination of trimetazidine in formulation and comparison with UV method. Der Pharma Chemica 2009; 1
Burits M, Bucar F. Antioxidant activity of Nigella sativa
essential oil. Phytother Res 2000; 14
Harpey C, Clanser P, Labrid C, Freyria JL, Poirier JP. Trimetazidine, a cellular anti-ischemic agent. Cardiovasc Drug Rev 1989; 6
Pulsinelli WA, Briely JB. Anew model of bilateral hemispheric ischemia in unanaesthetized rat. Stroke 1979; 10
Paxinos G, Watson C. The rat in stereotaxic coordinates
. New York, NY: Academic Press; 1982.
Loscher W, Nau H. Valproic acid: metabolic concentrations in plasma and brain, anticonvulsant activity and effects on GABA metabolism during subacute treatment in mice. Arch Int Pharmacodyn Ther 1982; 257
Raza M, Al-Bekairi M, Ageel AM, Qureshi S. Biochemical basis of sodium valproate hepatotoxicity and renal tubular disorder. Time dependence of perioxidative injury. Pharmacol Res 1997; 35
Aniya Y, Shimabukuro H, Shimoji M, et al.
Antioxidant and hepatoprotective action of the medical herb Artemisia campestries from the Okinawa islands. Boil Pharm Bull 2000; 23
Sharma PC, Yelne MB, Dennis TJ. Database on Medicinal Plants used in Ayurvedic Plants used in Ayurveda. Vol. 6
. CCRAS, New Delhi 2005; 6:420-440.
Gilgun-Sherki Y, Melamed E, Offen D. Oxidative stress induced-neurodegenerative diseases: the need for antioxidants that penetrate the blood brain barrier. Neuropharmacology 2001; 40
Moyer CA, Rounds J, Hannum JW. A meta-analysis of massage therapy research. Psychol Bull 2004; 130
Hansen AJ. Effect of anoxia on ion distribution in the brain. Physiol Rev 1985; 65
Gargaril P, Vahideh EA, Rafraf M, Gorbani A. Effect of dietary supplementation with Nigella sativa L. on serum lipid profile, lipid peroxidation and antioxidant defense system in hyperlipidemic rabbits. J Med Plants Res 2009; 3
Rao MA, Hatcher JF, Dempsey RJ. Lipid metabolism in ischemic neuronal death. Rec Res Dev Neurochem 1999; 2
Padhye S, Banerjee S, Ahmad A, Mohammad R, Sarkar FH. From here to eternity - the secret of Pharaohs: therapeutic potential of black cumin seeds and beyond. Cancer Ther 2008; 6
Araki T, Kato H, Kogure K. Postischemic alteration of muscarinic acetylcholine and adenosine a-1 binding sites in gerbil brain protective effects of a novel vinca alkaloid derivative vinconate and pentobarbital using an autoradiographic study. Res Exp Med 1992; 192
Turkdogan MK, Agaoglu Z, Yener Z, Sekeroglu R, Akkan HA, Avci ME. The role of antioxidant vitamins (C and E), selenium and Nigella sativa
in the prevention of liver fibrosis and cirrhosis in rabbits: new hopes. Dtsch Tierarztl Wochenschr 2001; 108
Araki S, Makino S, Mukai R, Nishikawa T, Saruwatari H, "Fundamental limitation of frequency domain blind source separation for convolved mixture of speech," in Proc. ICA2001 (International Conference on Independent Component Analysis and Blind Signal Separation), 2001; p. 132-137.
Chakravarty N. Inhibition of histamine release from mast cells by nigellone. Ann Allergy 1993; 70
Libersa C, Honor E, Adamantidis M, Rouet E, Dupuis B. Antiischemic effect of trimetazidiue: enzymatic and electric response in a model of in-vitro
myocardial ischemia. Cardiovasc Drugs Ther 1990; 4
Maupil V, Rochette L, Tabard A, Clauser P, Harpey C. Evaluation of the free radical formation during low flow ischemia and reperfusion in isolated rat heart. Cardiovasc Drugs Ther 1990; 4
Wang Q, Xu J, Rottinghaus GE, Simonyi A, Lubahn D, Sun GY, Sun AY. Resveratrol protects against global cerebral ischemic injury in gerbils. Brain Res 2002; 958
Gupta PC, Gupta R, Pednekar MS, Gupta PC, Gupta R, Pednekar MS, Gupta R. Hypertension prevalence and blood pressure trends in 88 653 subjects in Mumbai, India. J Hum Hypertens 2004; 18
Al-Omar FA, Nagi MN, Abdulgadir MM, Al Joni KS, Al-Majed AA. Immediate and delayed treatments with curcumin prevent forebrain ischemia-induced neuronal damage and oxidative insult in the rat hippocampus. Neurochem Res 2006; 31
Hosseinzadeh H, Parvardeh S, Asl MN, Sadeghnia HR, Ziaee T. Effect of thymoquinone and Nigella sativa
seeds oil on lipid peroxidation level during global cerebral ischemia-reperfusion injury in rat hippocampus. Phytomedicine 2007; 14
El-Abhar HS, DM Abdallah, S Saleh. Gastroprotective activity of Nigella sativa
oil and its constituent, thymoquinone, against gastric mucosal injury induced by ischaemia/reperfusion in rats. J Ethnopharmacol 2003; 84
Mahmood S, Ali S, Bhatti MH, Mazhar M, Iqbal R. Synthesis, characterization and biological applications of organotin (IV) derivatives of 2-(2-Fluoro-4-biphenyl) propanoic Acid, Turkish Journal of Chemistry, 2003; 27
Hassan-Khabbar S, Cottart CH, Wendum D, Vibert F, Clot JP, Savouret JF, et al.
Postischemic treatment by trans-resveratrol in rat liver ischemia-reperfusion: a possible strategy in liver surgery. Liver Transpl 2008; 14
Polat KY, Aydinli B, Polat O, Aydin U, Yazici P, Ozturk G, et al.
The protective effect of aprotinin and alpha-tocopherol on ischemia-reperfusion injury of the rat liver. Transplant Proc 2008; 40
Khalife KH, Lupidi G. Nonenzymatic reduction of thymoquinone in physiological conditions. Free Radic Res 2007; 41
Baran O, Yildirim A, Akkus M. The protective role of folic acid and vitamin E against toxical effects of valproic acid on liver tissue during period of gestation. Dicle Tup Dergisi 2004; 31
Lheureux PE, Hantson P. Carnitine in the treatment of valproic acid-induced toxicity. Clin Toxicol (Phila) 2009; 47
Kimura A, Yoshida I, Yamashita F, Kuriya N, Yamamoto M, Nagayama K. The occurrence of intramitochondrial Ca2+ granules in valproate-induced liver injury. J Pediatr Gastroenterol Nutr 1989; 8
El-Gharieb M, El-Masry T, Emara A, Hashem M. Potential hepatoprotective effects of vitamin E and Nigella sativa
oil on hepatotoxicity induced by chronic exposure to malathion in human and male albino rats. Toxicol Environ Chem J 2010; 92
Lee M, Lee Y, Mim B, Shin K, Chung B, Beak D, Jung B. The relationship between glucoronic levels and hepatotoxicity after oral administration of valproic acid. Arch Pharm Res 2009; 32
Nagi MN, Alam K, Badary OA, Al-Shabanah OA, Al-Sawaf HA, Al-Bekairi AM. Thymoquinone protects against carbon tetrachloride hepatotoxicity in mice via an antioxidant mechanism. Biochem Mol Biol Int 1999; 47
Khan S, Shakoor K, Jan M, Khattak A, Shah S. Study of histopathologic changes in the liver of albino rats, induced by toxic dose of valproic acid. Gomal J Med Sci 2005; 3
Siemes H, Nau H, Schultze K, Wittfoht W, Drews E, Penzien J, Seidel U. Valproate (VPA) metabolites in various clinical conditions of probable VPA-associated hepatotoxicity. Epilepsia 1993; 34
Hamza AA, Amin A. Apium graveolens modulates sodium valproate-induced reproductive toxicity in rats. J Exp Zool A Ecol Genet Physiol 2007; 307
Knapp AC, Todesco L, Beier K, Terracciano L, Sägesser H, Reichen J, Krähenbühl S. Toxicity of valproic acid in mice with decreased plasma and tissue carnitine stores. J Pharmacol Exp Ther 2008; 324
Olson M, Handler J, Thurman R. Valproic acid & nonalcoholic fatty liver disease: A possible association?. Mol Pharmacol 1986; 30
Bellringer M, Rahman K, Coleman R. Sodium valproate inhibits the movement of secretory vesicles in rat hepatocytes. Biochem J 1988; 243
Hautekeete ML, Degott C, Benhamou JP. Microvesicular steatosis of the liver. Acta Clin Belg 1990; 45
Oda M, Funatsu K, Okazaki I, Kamegaya K, Tsuchiya M, Sambe K. Morphogenesis of acidophilic bodies in experimental D-galactoisamine induced hepatitis. Gastroenterol Japonica 1973; 8
Roth B, Fkelund M, Fan B, Hägerstrand I, Nilsson-Ehle P. Lipid deposition in Kupffer cells after parenteral fat nutrition in rats: a biochemical and ultrastructural study. Intensive Care Med 1996; 22
Daba MH, Abdel-Rahman MS. Hepatoprotective activity of thymoquinone in isolated rat hepatocytes. Toxicol Lett 1998; 95
Al-Gharably NM, Badary O, Nagi M. Protective effect of thymoquinone against carbon tetrachloride-induced hepatotoxicity in mice. Res Comm Pharmacol Toxicol 1997; 2
Mansour MA, Nagi MN, EL-Khatib AS, AL-Bekairi AM. Effects of thymoquinone on antioxidant enzyme activities, lipid perioxidation and DT-diaphorase in different tissues in mice: a possible mechanism of action. Cell Biochem Funct 2002; 20
[Figure 1], [Figure 2], [Figure 3], [Figure 4], [Figure 5], [Figure 6], [Figure 7], [Figure 8], [Figure 9]
[Table 1], [Table 2]