Possible Curative Effects of Boric Acid and Bacillus clausii Treatments on TNBS-Induced Ulcerative Colitis in Rats.

Crohn's disease (CD) and ulcerative colitis (UC) are two chronic relapsing inflammatory bowel diseases (IBD). Although there are several treatment options available to improve the symptoms of IBD patients, there is no effective treatment that provides a definitive solution. In the present study, we aim to investigate the antioxidative/anti-inflammatory effects of oral administration of boric acid and Bacillus clausii in a rat trinitrobenzenesulfonic acid (TNBS)-induced colitis model. The effects of boric acid and B. clausii were examined in serum and colon tissues with the help of some biochemical and histological analyses. Elevated inflammation and oxidative damage were found in the blood and colon tissue samples in the TNBS-induced group according to the complete blood count (CBC), tumor necrosis factor (TNF) alpha, interleukin-35 (IL-35), malondialdehyde (MDA), glutathione peroxidase (GPx), myeloperoxidase (MPO), nitric oxide (NO), and histological findings. Particularly, the highest IL-35 level (70.09 ± 12.62 ng/mL) in the combined treatment group, highest catalase activity (5322 ± 668.1 U/mg protein) in the TNBS-induced group, and lower relative expression of inducible nitric oxide synthase in the TNBS-induced group than the control group were striking findings. According to our results, it can be concluded that boric acid showed more curative effects, even if B. clausii probiotics was partially ameliorative.

Comparative effects of metformin and Cistus laurifolius L. extract in streptozotocin-induced diabetic rat model: oxidative, inflammatory, apoptotic, and histopathological analyzes.

Interest in phytochemical therapy methods in the treatment of diabetes is increasing day by day. Although the antidiabetic and antioxidant effects of Cistus laurifolius L. (CL) have been mentioned, the systemic effects remain unknown. The present study aims at evaluating the antidiabetic effects of the CL aqueous extract via metformin on streptozotocin (STZ)-induced diabetic rats. Forty male Wistar albino rats were divided into five groups of eight animals each: control, diabetic group (55mg/kg STZ), STZ+125mg/kg CL, STZ+250mg/kg CL, and STZ+100mg/kg metformin. The effects of CL and metformin on oxidative, apoptotic, and inflammatory pathways were comparatively investigated. In addition, nuclear factor-κB (NFκB), tumor necrosis factor-alpha (TNF-α), and interleukin (IL)-1ß expressions analysis were carried out. CL treatment resulted in a significant improvement in blood glucose levels, lipid profile, pancreatic markers, and liver and kidney function tests. A 250mg/kg CL treatment decreased by 67.9%, 31.6%, 66.8%, 28.3%, and 31.4% in the total oxidant capacity, NFκB, TNF-α, IL-1ß, caspase3, and cytochrome c levels, respectively, compared to the diabetic group. Additionally, CL treatments showed a dose-dependent reduction in NFκB, TNF-α, and IL-1ß expression levels. A 250mg/kg CL treatment exhibited a greater increase (by 9.6%) in total antioxidant capacity than metformin. CL treatment provided histologically more improvement in the brain, heart, pancreas, spleen, liver, kidney, and testicular tissues compared to the metformin group. Our results suggest that the single treatment of CL aqueous extract at the low doses may have stronger short-term anti-diabetic effects than metformin. Therefore, further studies are needed regarding the long-term hypoglycemic effect or treatment of CL aqueous extract.

Cyproheptadine causes apoptosis and decreases inflammation by disrupting thiol/disulfide balance and enhancing the levels of SIRT1 in C6 glioblastoma cells.

Cyproheptadine is first-generation antihistamine drug, that is, H1 receptor antagonist, with a drug being anesthetic, anti-serotonergic and anti-cholinergic and started to be used clinically in the 1960s. As firstly utilized as an anti-allergic drug, usage of cyproheptadine was expanded to other cases including serotonin syndrome, appetite increasing, migraines and insomnia. However, there are almost few studies seeking to explore the association between cyproheptadine and cancer in general. In the present study, we sought to determine the impact of cyproheptadine on C6 glioblastoma cells by morphological, biochemical and cytotoxic analyzes. We searched the effective doses of cyproheptadine for C6 glioblastoma cells and examined the cells under an inverted microscope. Next, we determined the protein levels of SIRT1, NFκB and IL-6 protein. Then, we measured and calculated the levels of thiols, disulfide bonds and related parameters. After that, we evaluated apoptotic activity by Annexin V and caspase 3 assays. As a result, we detected a dose-dependent increase in apoptosis and SIRT 1 protein levels, and a decrease in inflammatory proteins. Furthermore, we have detected a drop in thiol and disulfide content. Our study suggests that Cyproheptadine causes apoptosis and decreases inflammation by disrupting thiol/disulfide balance and enhancing the levels of SIRT1, offering the potential for being an anti-cancer drug. Therefore, it might be further investigated in future studies.

Concanavalin A induces apoptosis in a dose-dependent manner by modulating thiol/disulfide homeostasis in C6 glioblastoma cells.

Glioma is the most common brain tumor. C6 rat glioblastoma cells provide the possibility to the scientist to study brain cancer. Concanavalin A (Con A) has a lot of antitumoral effects, especially over oxidative stress. In the present study, it was aimed to decide the impacts of various doses of Con A on C6 glioblastoma cells regarding cytotoxicity, thiol/disulfide homeostasis, apoptosis, and inflammation. We detected the cytotoxic activity of Con A (from 7.8 to 500 µg/ml) in C6 cells by utilizing 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) and determined the toxic concentration of Con A. Once the optimal doses were found, the thiol-disulfide homeostasis, levels of total antioxidant and oxidant status (TAS and TOS), malondialdehyde (MDA) and glutathione (GSH), pro-inflammatory cytokines as tumor necrosis factor-alpha (TNF-α) and interleukin-6 (IL-6), apoptotic proteins as cytochrome c (CYCS), and caspase 3 (CASP3) were measured. Apoptotic and morphological changes in the C6 cells were examined with an inverted microscope and flow cytometry technique. Dose-dependent Con A triggered oxidative damage in the C6 cells, affecting the inflammatory pathway, so reducing proliferation with apoptotic proteins and morphological changes. But especially, Con A increased disulfide formation by disrupting the thiol/disulfide balance in C6 cells. This study revealed that Con A, known as carbohydrate-binding protein, generated oxidative damage, inflammation, and apoptosis in a dose-dependent manner by modulating thiol/disulfide homeostasis in C6 glioblastoma cells.

Bexarotene inhibits cell proliferation by inducing oxidative stress, DNA damage and apoptosis via PPARγ/ NF-κB signaling pathway in C6 glioma cells.

Gliomas are one of the most aggressive brain tumors with a poor prognosis in the central nervous system. Bexarotene is a third-generation retinoid X receptor agonist that is promising in the treatment of both cancer and neurodegenerative diseases. In this study, we aimed to investigate the cytotoxic and anti-proliferative effects of bexarotene in C6 glioma cells through the PPARγ/NF-κB pathway. In the study, first cytotoxic bexarotene concentrations for C6 cells were detected, and then apoptosis profile, reactive oxygen species (ROS), total antioxidant (TAS), 8-hydroxy-2'-deoxyguanosine (8-OHdG) and nuclear factor-κB (NF-κB) levels in the cells were determined. In addition, peroxisome proliferator-activated receptor γ (PPARγ) mRNA expression analysis was carried out. As a result, we detected concentration- and time-dependent antiproliferative effects of bexarotene on C6 cells. We found that bexarotene treatment decreased NF-κB and TAS levels and increased PPARγ and 8-OHdG levels in C6 cells. Bexarotene enhanced PPARγ expression in a dose-dependent manner when compared to the control group (P < 0.01). Furthermore, we determined that bexarotene-induced apoptotic C6 cells enhanced through Annexin V-FITC/PI staining and caspase-3/-7 activation analyses since phosphatidylserine level on the outer surface of the cell membrane and caspase-3/-7 activities were increased in the cells treated with bexarotene. In conclusion, bexarotene treatment in C6 glioma cells could modulate apoptosis profile, DNA damage, ROS production, and reduction of TAS levels through inhibition of NF-κB by enhancing PPARγ expression.

Protective Effect of Boric Acid and Omega-3 on Myocardial Infarction in an Experimental Rat Model.

Boric acid and omega-3 are used as essential elements for both animal and human health. Many researchers have shown these beneficial effects on cardiac and inflammatory markers. This study aims to evaluate cardiac protective effect of boric acid and omega-3 against MI (myocardial infarction), probably due to the suppression of pro-inflammatory cytokines of natriuretic peptides in rats. Fifty male Sprague-Dawley rats were randomly divided into five groups: control, MI, MI+boric acid, MI+omega-3, and MI+boric acid+omega-3. Saline solution (2 ml/day), omega-3 (800 mg/kg/day), and boric acid (100 mg/kg/day)+omega-3 (800 mg/kg/day) were orally administered to the relevant groups throughout the 28 days. To constitute the MI model, the rats were exposed to isoproterenol-HCl (ISO) (200 mg/kg, S.C.) on the 27th and 28th. In the MI group, serum levels of CK-MB, BNP, and TNF-α are increased significantly. Also, ST waves and heart rates were higher in the MI than the control. These results demonstrate that biochemical results healed in MI+boric acid, MI+omega-3, and MI+boric acid+omega-3 groups compared MI group. ECG and light microscope results supported the findings as well. The statistical analysis showed that boric acid and/or omega-3 has protective effects on cellular damage in MI.

Concentration-Dependent Effects of Zinc Sulfate on DU-145 Human Prostate Cancer Cell Line: Oxidative, Apoptotic, Inflammatory, and Morphological Analyzes.

Zinc takes part in several of cellular signaling pathways, containing defense against free radicals, apoptosis, and inflammation. However, interaction between zinc and prostate cancer progression is poorly understood. Therefore, zinc treatment in DU-145 human prostate cancer cells was investigated. First, zinc sulfate (ZnSO4) concentrations with antiproliferative effect were determined using MTT assay. Then, ZnSO4-induced oxidative damage was evaluated by malondialdehyde (MDA) levels, glutathione (GSH) levels, total oxidant status (TOS) levels, and total antioxidant status (TAS) levels. Apoptotic effects of ZnSO4 were determined by measuring biochemical and immunohistochemical parameters including caspase 3 (CASP3), cytochrome C (CYC), Bcl-2-associated X protein (Bax), and B cell CLL/lymphoma 2 (Bcl-2) levels. Inflammatory effects of ZnSO4 were investigated by measuring interleukin-6 (IL-6) levels and tumor necrosis factor-alpha (TNF-α) levels. Finally, morphological analysis was performed using hematoxylin-eosin staining. We found that ZnSO4 caused a concentration-dependent increase in oxidative stress, apoptosis, and inflammation pathways. Moreover, there were a number of morphological alterations in treated cells depending on the ZnSO4 concentration. Consequently, our data showed that zinc acts as a regulator of increased oxidative damage and apoptosis through the upregulation of TNF-α and IL-6.

High Concentrations of Boric Acid Trigger Concentration-Dependent Oxidative Stress, Apoptotic Pathways and Morphological Alterations in DU-145 Human Prostate Cancer Cell Line.

Boric acid is known to regulate the proliferation of cancer cells. Prostate cancer is among the types of cancer with high mortality in men. There are a few numbers of studies investigating the effects of boric acid on prostate cancer cells. The objective of the present study was to assess the effects of boric acid at concentrations higher than that can be achieved in blood by dietary intake on DU-145 human prostate cancer cells for 24 h. Firstly, we determined the cytotoxic activity of boric acid (0 to 12.5 mM) on DU-145 human prostate cancer cells by using 3-(4, 5-dimethylthiazol, 2-yl)-2, 5-diphenyl tetrazolium bromide (MTT) and defined the IC50 concentration of boric acid. Then, by employing the doses found in MTT, the levels of antioxidant-oxidant molecules and apoptotic proteins were measured and morphological changes were evaluated. We have concluded that boric acid caused oxidative stress, inhibition of cell growth, apoptosis, and morphological alterations in a concentration-dependent manner in DU-145 cells. Furthermore, treatments with increasing boric acid concentrations decreased the antioxidant levels in cells. We actually revealed that boric acid, known as an antioxidant, may prevent cell proliferation by acting as an oxidant in certain doses. Although the high IC50 concentration of boric acid is perceived to be negative, we think it provides important background for subsequent studies.

Betaine suppresses cell proliferation by increasing oxidative stress-mediated apoptosis and inflammation in DU-145 human prostate cancer cell line.

Prostate cancer is the main cause of cancer-related mortality in men around the world and an important health problem. DU-145 human prostate cancer cells provide an opportunity to investigate prostate cancer. Betaine has a number of anticancer effects, such as inactivation of carcinogens, inhibition of cancer cell proliferation, angiogenesis, and metastasis. However, there is no study investigating the effects of betaine on DU-145 cells. The aim of this study was to evaluate the effects of different concentrations of betaine on the oxidative stress, apoptosis, and inflammation on DU-145 cells. Firstly, we proved the cytotoxic activity of betaine (0 to 150 mg/ml) on DU-145 cells by using 3-(4, 5-dimethylthiazol, 2-yl)-2, 5-diphenyl tetrazolium bromide (MTT) and defined the optimal concentration of betaine. Then, by employing the doses found in MTT, the levels of antioxidant (GSH, SOD, CAT, and TAS) and oxidant (MDA and TOS) molecules, pro-inflammatory cytokines (TNF-a and IL-6), apoptotic proteins (CYCS and CASP3), and DNA fragmentation were measured. Morphological changes and apoptosis were evaluated using H&E technique, Bax and Bcl-2 immunohistochemistry. Results suggested that betaine caused oxidative stress, inflammation, inhibition of cell growth, apoptosis, and morphological alterations in DU-145 cells dose-dependently. Furthermore, treatments with increasing betaine concentrations decreased the antioxidant levels in cells. We actually revealed that betaine, known as an antioxidant, may prevent cell proliferation by acting as an oxidant in certain doses. In conclusion, betaine may act as a biological response modifier in prostate cancer treatment in a dose-dependent manner.

Curcumin Acts as Post-protective Effects on Rat Hippocampal Synaptosomes in a Neuronal Model of Aluminum-Induced Toxicity.

The neurotoxic effects of aluminum are generally associated with reduced antioxidant capacity, increased oxidative stress and apoptosis, which lead to the induction of neurodegenerative processes. Curcumin has a lipophilic polyphenol character and effects of antioxidant and anti-apoptotic. The present study was undertaken to examine possible aluminum exposure in rats brain synaptosomes and to investigate whether protective and therapeutic effects of curcumin on biochemical and morphological changes in both pre- and post-treated groups. Aluminum chloride (AlCl3) at 50 µM concentration and curcumin at 5 and 10 µg/mL doses were applied to hippocampal synaptosomes of rats according to experimental design. Biochemical effects were evaluated by MTT cytotoxicity, malondialdehyde (MDA) levels, nitric oxide (NO) levels, glutathione (GSH) levels, caspase 3 activities, cytochrome c levels, DNA fragmentation values and protein levels. Morphological examinations were done by TEM analysis. AlCI3 exposure in the synaptosomes enhanced oxidative stress, triggered apoptosis and caused ultrastructural alterations which were well reflected in the TEM images. Curcumin pre-treatment slightly ameliorated the MDA levels, NO levels, cytochrome c levels and caspase 3 activities in AlCI3-exposed synaptosomes, but these results were not statistically significant. Furthermore, curcumin post-treatment significantly improved oxidative damage and morphological alterations, and suppressed cytochrome c and caspase 3 activities. Taken together, our data showed that curcumin had more therapeutic effects than protective effects in AlCI3-induced neurotoxicity. Nevertheless, the therapeutic (post-protective) effects of curcumin should be further investigated in in vivo neurodegenerative models involving behavioral tests.

A possible protective role of betain and omega-3 supplementation in traumatic brain injury.

INTRODUCTION: Due to irreversible damage following head trauma, many overlapping pathophysiological events occur including excitotoxicity, acidotoxicity, ionic imbalance, edema, oxidative stress inflammation and apoptosis. MATERIAL AND METHODS: In this this study, after the rats were separated in to groups theserats were fed throughout fourteen days with betaine, omega-3 or betaine+omega-3 combination in physiological limits prior to the trauma. After a closed head trauma, the damaged brain tissues were collected for biochemically and histologically analyses. This examination involved analyses of levels of caspase-3 and cytochrome C and neuron-specific enolase (NSE) levels in brain tissue. RESULTS: These analyses showed that traumatic brain injury (TBI) caused an increase in the levels of caspase-3, cytochrome C and neuron-specific enolase (NED) in the brain tissues examined. DISCUSSION: In this study, apoptotic and/or necrotic cell death via mitochondrial cytochrome C caspase pathway in traumatized cells and neuron-specific enolase (NED) increase indicative of neuronal damage confirmed the research hypothesis. CONCLUSION: Level of the biomarkers induced by brain injury in the groups fed with betaine, omega-3 and betaine+omega-3 combination before the traumatic damage approximated to that of control group values, suggesting that these products may have a neuroprotective role. KEY WORDS: Betain, Caspase-3, Cytochrome C and Neuron-specific enolase, Omega-3, Traumatic brain injury.

Prenatal alcohol-induced neuroapoptosis in rat brain cerebral cortex: protective effect of folic acid and betaine.

PURPOSE: Alcohol consumption in pregnancy may cause fetal alcohol syndrome (FAS) in the infant. This study aims to investigate prenatal alcohol exposure related neuroapoptosis on the cerebral cortex tissues of newborn rats and possible neuroprotective effects of betaine, folic acid, and combined therapy. METHODS: Pregnant rats were divided into five experimental groups: control, ethanol, ethanol + betaine, ethanol + folic acid, and ethanol + betaine + folic acid combined therapy groups. We measured cytochrome c release, caspase-3, calpain and cathepsin B and L. enzyme activities. In order to observe apoptotic cells in the early stages, TUNEL method was chosen together with histologic methods such as assessing the diameters of the apoptotic cells, their distribution in unit volume and volume proportion of cortical intact neuron nuclei. RESULTS: Calpain, caspase-3 activities, and cytochrome c levels were significantly increased in alcohol group while cathepsin B and L. activities were also found to be elevated albeit not statistically significant. These increases were significantly reversed by folic acid and betaine + folic acid treatments. While ethanol increased the number of apoptotic cells, this increase was prevented in ethanol + betaine and ethanol + betaine + folic acid groups. Morphometric examination showed that the mean diameter of apoptotic cells was increased with ethanol administration while this increase was reduced by betaine and betaine + folic acid treatments. CONCLUSION: We observed that ethanol is capable of triggering apoptotic cell death in the newborn rat brains. Furthermore, folic acid, betaine, and combined therapy of these supplements may reduce neuroapoptosis related to prenatal alcohol consumption, and might be effective on preventing fetal alcohol syndrome in infants.

The investigation of the prenatal and postnatal alcohol exposure-induced neurodegeneration in rat brain: protection by betaine and/or omega-3.

PURPOSE: We aim to study the effect of neurodegeneration on the brain of rat pups caused by prenatal and postnatal ethanol exposure with modified liquid diet to elucidate protective effects of betaine and omega-3 supplementation. When ethanol is consumed during prenatal and postnatal periods, it may result in fetal alcohol syndrome (FAS) in the offspring. METHODS: Rats were divided into control, ethanol, ethanol + betaine, ethanol + omega-3, ethanol + omega-3 + betaine groups. The effect of betaine and omega-3 in response to ethanol-induced changes on the brain, by biochemical analyses cytochrome c, caspase-3, calpain, cathepsin B and L, DNA fragmentation, histological and morfometric methods were evaluated. RESULTS: Caspase-3, calpain, cathepsin B, and cytochrome c levels in ethanol group were significantly higher than control. Caspase-3, calpain levels were decreased in ethanol + betaine, ethanol + omega-3, and ethanol + omega-3 + betaine groups compared to ethanol group. Cathepsin B in ethanol + omega-3 + betaine group was decreased compared to ethanol, ethanol + betaine groups. Cathepsin L and DNA fragmentation were found not statistically significant. We found similar results in histological and morfometric parameters. CONCLUSION: We found that pre- and postnatal ethanol exposure is capable of triggering necrotic cell death in rat brains, omega-3, and betaine reduce neurodegeneration. Omega-3 and betaine may prove beneficial for neurodegeneration, particularly in preventing FAS.

Protective effect of calpain inhibitor N-acetyl-L-leucyl-L-leucyl-L-norleucinal on acute alcohol consumption related cardiomyopathy.

Excessive alcohol consumption and alcoholism cause medical problems with high mortality and morbidity rates. In this study we aimed to decrease the alcohol related tissue damage by inhibiting calpain activation which plays an important role in apoptosis and necrosis, in rats with cardiomyopathy induced by acute alcohol consumption. Male Sprague-Dawley rats were separated into four groups (control, vehicle, alcohol and alcohol + inhibitor) with 10 rats in each. Control group received isocaloric maltose while vehicle group received isocaloric maltose with DMSO, and alcohol group received 8 g/kg absolute ethanol by gavage. Inhibitor group received 20 mg/kg calpain inhibitor 1 intraperitonally prior to alcohol administration. Calpain activities, cathepsin L levels and cytochrome c release rates were significantly increased in alcohol group compared to control group (p < 0.05). Serum CK MB and BNP levels of alcohol group were excessively increased compared to control group (respectively p < 0.001 and p < 0.01). Serum BNP levels of alcohol + inhibitor group were significantly (p < 0.05) decreased compared to alcohol group. In addition to these, histological evaluation of light microscope images and the results of DNA fragmentation and immunohistochemical caspase-3 activity results showed significant improvement of these parameters in alcohol + inhibitor group compared to alcohol group. Results of our biochemical and histological evaluation results revealed that the calpain inhibitor N-acetyl-leu-leu-norleucinal may have an ameliorating effect on acute alcohol consumption related cardiac tissue damage due to its effects on cell death pathways.

Effect of carotenoid ß-cryptoxanthin on cellular and humoral immune response in rabbit.

Beta-cryptoxanthin (b-Cr) is a pro-vitamin A and one of the major carotenoids that can be commonly found in mammalian serum and tissues. Foods rich in certain fatty acids are known to be effective to gain a healthy immune system. In the present study, we evaluated the effect of b-Cr on rabbit humoral and cellular immune responses to have a better vision about the mechanism of effect of carotenoids on immune system. Twenty rabbits were randomly divided into five groups (4 per group): Groups consisted of: 1) control group (normal saline; 2) b-Cr (control); 3) vaccine control; 4) 5 mg/kg b-Cr o.p. + vaccine; 5) 10 mg/kg b-Cr o.p. + vaccine. Blood samples were obtained from the marginal ear artery at three time points: days 0, 14 and 21 of the study. Blood CD4+ and CD8+ lymphocytes and Serum Immunoglobulin and Cytokines content were evaluated. Results show that b-Cr administration increased the blood CD4+ lymphocytes count (P > 0.01). Serum IgG, IgM and IgA levels increased (P > 0.05) following b-Cr administration. b-Cr treatment increased serum IL-4 levels (P > 0.05). According to presented results, b-Cr may increase the humoral immunity in mammals. So, it would possible has a potentially beneficial effect on health and on prevention of the immunity related diseases.

Biological assays on the effects of Acra3 peptide from Turkish scorpion Androctonus crassicauda venom on a mouse brain tumor cell line (BC3H1) and production of specific monoclonal antibodies.

Constitutes of the venom scorpion are a rich source of low molecular mass peptides which are toxic to various organisms, including man. Androctonus crassicauda is one of the scorpions from the Southeastern Anatolia of Turkey with public health importance. This work is focused on the investigation of biological effects of Acra3 peptide from Androctonus crassicauda. For this purpose, Acra3 isolated from crude venoms was tested for its cytotoxicity on BC3H1 mouse brain tumor cells using tetrazolium salt cleavage and lactate dehydrogenase activity assays. To determine whether the cytotoxic effects of Acra3 was related to the induction of apoptosis, the morphology of the cells and the nuclear fragmentation was examined by using Acridin Orange staining and DNA fragmentation assay, respectively. Caspase 3 and caspase 9 activities were measured spectrophotometrically and flow cytometric assay was performed using Annexin-V FITC and Propidium Iodide staining. Furthermore toxic peptide Acra3 was used as an antigen for immunological studies. Results showed that Acra3 exerted very strong cytotoxic effect on BC3H1 cells with an IC50 value of 5 µg/ml. Exposure of the cells to 0.1 and 0.5 µg/ml was resulted in very strong appearance of the apoptotic morphology in a dose dependent manner. On the other side, not any DNA fragmentation was observed after treatment of the cells. Caspase 3 and 9 activities were slightly decreased with Acra3. Results from flow cytometry and lactate dehydrogenase activity assays indicate that Acra3 exerts its effects by inducing a stronger necrosis than apoptosis in BC3H1 cells. To evaluate its immunogenicity, monoclonal antibody (MAb) specific for Acra3 antigen (5B9) was developed by hybridoma technology using spleen and lymph nodes of mice and immunoglobulin type of antibody was found to be IgM. We suggest that Acra3 may exert its effects by inducing both necrotic and apoptotic pathway in some way on mouse brain tumor cells. These findings will be useful for understanding the mechanism of cell death caused by venom in vitro. Anti-Acra3 monoclonal antibody can be further used as a bioactive tools for exploring the structure/function relationship and the pharmacological mechanism of scorpion peptide neurotoxins.

Investigation of the possible protective role of gallic acid on paraoxanase and arylesterase activities in livers of rats with acute alcohol intoxication.

Gallic acid, a polyphenyl class natural product from gallnut and green tea, is known to be antioxidant, anti-inflammatory and radical scavenger. In this study, we aimed to investigate the possible protective effects of gallic acid on paraoxonase and arylesterase activities in liver exposed to acute alcohol intoxication. Paraoxonase and arylesterase activities in liver tissue and serum aspartate aminotransferase, alanine aminotransferase and lactate dehydrogenase levels were measured. Histological investigations were also made. In our study, we observed a significant increase of serum alanine aminotransferase, aspartate aminotransferase and lactate dehydrogenase activities, which are indicators of liver damage after acute ethanol consumption. Gallic acid therapy has significantly reduced the increase in these biomarkers, indicating a possible hepatoprotective effect of gallic acid. Ethanol consumption caused a significant decrease in liver paraoxonase activity (P < 0.001). Gallic acid treatment partly restored this decreased paraoxonase activity, which resulted from ethanol administration. A gallic acid dose of 100 mg/kg was observed as highest restoring effect for paraoxonase activity (P < 0.05). The activity of arylesterase was decreased in the ethanol group as compared with the control group, but this was not significant. However, 50 mg/kg of gallic acid treatment restored the loss of this activity due to ethanol exposure (P < 0.001). We observed that gallic acid ameliorates the liver damage caused by excessive alcohol consumption in a dose-dependent way. Our results in this study showed that gallic acid might have a protective effect against alcoholic liver disease.

The neuroprotective effect of acute moderate alcohol consumption on caspase-3 mediated neuroapoptosis in traumatic brain injury: the role of lysosomal cathepsin L and nitric oxide.

Our aim in this study was to investigate the effect of moderate acute alcohol administration on cysteine protease mediated neuronal apoptosis and nitric oxide production in the traumatic brain injury. A total of 29 adult Sprague-Dawley male rats weighing 250-300 g were used. The rats were allocated into four groups. The first group was the control (sham-operated) group in which only a craniotomy was performed, the others were alcohol, trauma and trauma+alcohol groups. Caspase-3 enzyme activity in the trauma group increased significantly in comparison with the control group. The alcohol given group showed a decreased caspase-3 enzyme activity compared to the trauma group. The level of caspase-3 enzyme activity in the alcohol+trauma group decreased in comparison to the trauma group. SF/FEL ratio of cathepsin-L enzyme activity in the trauma group was significantly higher than in the control group. Our results indicate that moderate alcohol consumption may have protective effects on apoptotic cell death after traumatic brain injury. Protective effects of moderate ethanol consumption might be related to inhibition of lysosomal protease release and nitric oxide production.

Preventive role of gallic acid on alcohol dependent and cysteine protease-mediated pancreas injury.

In order to investigate an association between alcohol consumption and lysosomal cysteine protease induced pancreatic injury and preventive effect of gallic acid as dose-dependent, we determined myeloperoxidase and malondialdehyde levels, serum amylase activities and cathepsin B and L activities in the cytosolic and lysosomal fractions of pancreatic tissue in the ethanol (8 g/kg) and ethanol plus gallic acid (at different doses 50, 100 and 200 mg/kg) given rats. Absolute ethanol (8 g/kg) was given by oral gavage. Gallic acid was dissolved in the saline (2 ml/kg) and administered before 30 min the oral administration of ethanol. Pancreatic myeloperoxidase and also malondialdehyde levels and serum amylase activities were measured. Besides, histological investigations were made. Cathepsin B activities in the cytosolic fraction were decreased by gallic acid (200 mg/kg) and increased in ethanol given rats. Cytosolic/lysosomal ratio of cathepsin B and L were found to be low in the all doses of gallic acid as compared to ethanol group. Serum amylase, pancreatic myeloperoxidase activities and malondialdehyde levels in the ethanol group were higher than in the control group. These were not statistically significant for myeloperoxidase and malondialdehyde. Also, our histopathologic results indicated that ethanol administration increased pancreatic tissue injury. Gallic acid especially at 200 mg/kg improved ethanol-mediated pancreatic tissue damage.In conclusion, gallic acid treatments were decreased release of lysosomal cathepsin B and L enzymes into cytoplasmic fraction and prevented alcohol mediated pancreatic tissue injury. Preventive effect of gallic acid might be dose-dependent.

Erythropoietin prevents nitric oxide and cathepsin-mediated neuronal death in focal brain ischemia.

We examined the preventive effect of human recombinant erythropoietin (HrEPO) on nitric oxide (NO)-mediated toxicity to neurons and cysteine protease release into cytoplasm, which is attributed to neuronal death in brain ischemia. Focal cerebral ischemia was induced by permanent occlusion of middle cerebral artery in two sets of rat. The first set was used to monitor NO concentration and cathepsin activity, while the second was used for histological examination with hematoxylin and eosin, and TUNEL staining. A group in both set was administered human recombinant erythropoietin (HrEPO). NO content, cathepsins B and L activity increased significantly in the post-ischemic cerebral tissue (p<0.05). HrEPO treatment reduced NO concentration and cathepsin activity to control level (p>0.05). A significant increase in the number of necrotic and apoptotic neurons was observed in the post-ischemic cerebral cortex (p<0.05). HrEPO treatment was markedly lowered both of these (p<0.05). It is concluded that HrEPO prevents neuronal death by protecting neuronal liposomes from NO-mediated toxicity and suppressing the release of cathepsins.