الفهرس 
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Abstract
Biochemical influence of alpha-lipoic acid on lipid peroxidation and antioxidant enzymes in blood and gastric tissue of ethanol induced gastric mucosal erosions in rats were investigated. Moreover, the possible protective and therapeutic effects of alpha lipoic acid on serum nitric oxide, sialic acid, tumor necrosis factor alpha and interlukin-6 in addition to enzymatic and non-enzymatic antioxidants, biomarkers of oxidative stress and DNA-fragmentation in gastric tissue in rat model of ethanol-induced gastric ulcer were also evaluated. A total number of 60 male albino rats, 12-16 weeks old and average body weight 200-250 gm were used in the experimental investigation of this study. Rats were housed in separated metal cages and kept at constant environmental and nutritional conditions throughout the period of experiment. The animals were fed on constant ration and water was supplied ad- libitum. The animals were left 14 days for acclimatization before the beginning of the experiment. Ethanol-induced gastric mucosal erosions: Rats were fasted for 18 hours and allowed free access of water prior to the administration of ethanol for gastric ulcer induction. All the rats except those of the control group orally administrated with absolute ethanol at a dose of 1ml/rat. Experimental design: Rats were randomly divided into four main equal groups, 15 rats each, placed in individual cages and classified as follow: Group I (normal control group): received no drugs. This group was divided into 2 subgroups: Subgroup (a): Included 7 rats sacrificed at the 8th day of the experiment, served as the normal control rats for early ulcer group. Subgroup (b): Included 8 rats sacrificed at the 18th day of the experiment, served as the normal control rats for non-treated late ulcer group. Group II (ulcerated non-treated group): Included 15 rats, administrated once orally with 1ml/rat absolute ethanol for induction of gastric ulcer. This group was divided into 2 subgroups: Subgroup (a): Consisted of 7 rats served as, early ulcerated non-treated group, for comparison with alpha lipoic acid protected group. This group received absolute ethanol at a dose of (1ml/rat) on empty stomach and the rats were sacrificed after one hour later of ethanol administration. Subgroup (b): Consisted of 8 rats served as, late ulcerated non-treated group, for comparison with alpha lipoic acid treated group. This group received absolute ethanol at a dose of (1ml/rat) on empty stomach and the rats were left free and sacrificed ten days later of ethanol administration. Group III (alpha lipoic acid protected group): Comprised 15 male rats received alpha lipoic acid (100 mg/kg body weight/day) orally for seven days prior absolute ethanol administration. One hour after the administration of ethanol the animals were sacrificed. Group IV (alpha lipoic acid treated group): Included 15 male rats received alpha lipoic acid orally(100 mg/kg body weight) for seven days before ethanol administration and the treatment were continued with ALA for ten days later. Sampling: Blood samples and tissue specimens (gastric tissues) were collected one hour after administration of ethanol in normal control group (subgroup a), ulcerated non-treated group (subgroup a) and alpha lipoic acid protected ulcer group at 8days from the onset of treatment with α-lipoic acid. Also, blood samples and tissue specimens (gastric tissues) were collected after 17days from the onset of treatment with α-lipoic acid in normal control group (subgroup b), ulcerated non-treated group subgroup (b) and alpha lipoic acid treated ulcer group. Blood samples: Blood samples for serum separation were collected by ocular vein puncture at the end of each experimental period in dry, clean, and screw capped tubes and serum were separated by centrifugation at 2500 r.p.m for 15 minutes .The clean, clear serum was separated by automatic pipette and received in dry sterile samples tube and kept in a deep freeze at -20ْC until used for subsequent biochemical analysis. All serum samples were analyzed for NO, SA, TNF-α, IL-6 and L-MDA. Tissue samples (gastric tissue): a- For biochemical analysis: After seven and seventeen days of treatment with alpha-lipoic acid the rats were sacrificed by cervical decapitation. The stomach was quickly removed, and opened along the greater curvature using a scrapper, cleaned by rinsing with cold saline and stored at -20°C for subsequent biochemical analyses. Stomach tissue preparation: Briefly, gastric tissues were cut, weighed and minced into small pieces, homogenized with a glass homogenizer in 9 volume of ice-cold 0.05 mM potassium phosphate buffer (pH7.4) to make 10% homogenates. The homogenates were centrifuged at 6000 r.p.m for 15 minutes at 4°C then the resultant supernatant were used for the determination of the following parameters: superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx), glutathione reductase (GR), L-malondialdehyde (L-MDA), reduced glutathione (GSH), myeloperoxidase (MPO), vitamin C and DNA fragmentation. b- Histopathological examination: Gastric tissue specimens were taken from different parts of the stomach. The specimens were preserved in 10% buffered neutral formalin and subjected for microscopical examination. The obtained results summarized the followings: 1- Serum nitric oxide: A significant decrease in serum nitric oxide concentration was observed in ethanol-induced gastric mucosal erosion (early ulcer) in rats after one hour compared with the normal control group. Pretreatment with α-Lipoic acid to ethanol- induced gastric ulcer (early ulcer) in rats exhibited a non-significant increase in serum nitric oxide concentration compared with control one hour early ulcer group. A non-significant decrease in serum nitric oxide concentration was observed in ethanol-induced gastric erosion (late ulcer) in rats after 10 days compared with the normal control group. Alpha-Lipoic acid treatment in ethanol- induced gastric erosion (late ulcer) in rats exhibited a non-significant decrease in serum nitric oxide concentration compared with control late ulcer group. 2- Serum L-malondialdehyde (L-MDA): A significant increase in serum L-malondialdehyde concentration is observed in ethanol-induced gastric mucosal erosion (early ulcer) in rats when compared with the normal control group. Pretreatment with α-Lipoic acid to ethanol- induced gastric mucosa erosion (early ulcer) in rats exhibited a non-significant increase in serum L-malondialdehyde concentration when compared with early ulcer non-treated group. A significant increase in serum L-malondialdehyde concentration is observed in ethanol-induced gastric mucosal erosion (late ulcer) in rats when compared with normal control group. Treatment with α-Lipoic acid to ethanol- induced gastric mucosal erosion (late ulcer) in rats exhibited a non-significant increase in serum L-malondialdehyde concentration when compared with late ulcer non-treated group. 3- Gastric tissue L-malondialdehyde (L- MDA): A significant increase in gastric tissue L-malondialdehyde concentration is observed in ethanol-induced gastric mucosal erosion (early ulcer) in rats when compared with the normal control group. Pretreatment with α-Lipoic acid to ethanol- induced gastric mucosal erosion (early ulcer) in rats exhibited a non-significant decreased in gastric tissue L-malondialdehyde concentration when compared with early ulcer non-treated group. A significant increase in gastric tissue L-malondialdehyde concentration is observed in ethanol-induced gastric mucosal erosion (late ulcer) in rats when compared with normal control group. Alpha-Lipoic acid treatment in ethanol- induced gastric mucosal erosion (late ulcer) in rats exhibited a non-significant decrease in gastric tissue L-malondialdehyde concentration when compared with late ulcer non-treated group. 4- Gastric tissue Glutathione peroxidase (GPx): A significant decrease in gastric tissue Glutathione peroxidase activity was observed in ethanol–induced gastric mucosal erosion (early ulcer) in rats when compared with the normal control group. Pretreatment with α-Lipoic acid to ethanol- induced gastric mucosal erosion (early ulcer) in rats exhibited a non-significant decrease in gastric tissue Glutathione peroxidase activity when compared with early ulcer non-treated group. A significant decrease in gastric tissue Glutathione peroxidase activity was observed in ethanol–induced gastric mucosal erosion (late ulcer) in rats when compared with the normal control group. Treatment with α-Lipoic acid to ethanol- induced gastric mucosal erosion (late ulcer) in rats exhibited a significant decrease in gastric tissue Glutathione peroxidase activity when compared with late ulcer non-treated group. 5-Gastric tissue superoxide dismutase (SOD): A significant decrease in gastric tissue superoxide dismutase activity was observed in ethanol–induced gastric mucosal erosion (early ulcer) in rats when compared with the normal control group. Pretreatment with α-Lipoic acid to ethanol- induced gastric mucosal erosion (early ulcer) in rats exhibited a significant increase in gastric tissue superoxide dismutase activity when compared with early ulcer non-treated group. A significant decrease in gastric tissue superoxide dismutase activity was observed in ethanol–induced gastric mucosal erosion (late ulcer) in rats when compared with the normal control group. Alpha-Lipoic acid treatment in ethanol- induced gastric mucosal erosion in rats exhibited a non-significant increase in gastric tissue superoxide dismutase activity when compared with late ulcer non-treated group. 6-Gastric tissue catalase (CAT): A significant decrease in gastric tissue catalase (CAT) activity was observed in ethanol–induced gastric mucosal erosion (early ulcer) in rats when compared with the normal control group. Pretreatment with α-Lipoic acid to ethanol-induced gastric mucosal erosion (early ulcer) in rats exhibited a non-significant increase in gastric tissue catalase activity when compared with early ulcer non-treated group. A significant decrease in gastric tissue catalase (CAT) activity was observed in ethanol–induced gastric mucosal erosion (late ulcer) in rats when compared with the normal control group. Treatment with α-Lipoic acid to ethanol- induced gastric mucosal erosion (late ulcer) in rats exhibited a non-significant increase in gastric tissue catalase activity when compared with late ulcer non-treated group. 7- Gastric tissue glutathione reductase (GR): A significant increase in gastric tissue glutathione reductase activity was observed in ethanol–induced gastric mucosal erosion (early ulcer) in rats when compared with the normal control group. Pretreatment with α-Lipoic acid to ethanol- induced gastric mucosal erosion (early ulcer) in rats exhibited a significant decrease in gastric tissue glutathione reductase activity when compared with early ulcer non-treated group. A significant increase in gastric tissue glutathione reductase activity was observed in ethanol–induced gastric mucosal erosion (late ulcer) in rats when compared with the normal control group. Alpha-Lipoic acid treatment in ethanol- induced gastric mucosal erosion (late ulcer) in rats exhibited a non-significant increase in gastric tissue glutathione reductase activity when compared with late ulcer non-treated group. 8-Gastric tissue reduced glutathione (GSH): A non-significantly increases in gastric tissue reduced glutathione (GSH) concentration was observed in ethanol-induced gastric mucosal erosion (early ulcer) in rats when compared with the normal control group. Pretreatment with α-Lipoic acid to ethanol- induced gastric mucosal erosion (early ulcer) in rats exhibited a non-significant increase in gastric tissue reduced glutathione when compared with early ulcer non-treated group. A significant increase in gastric tissue reduced glutathione (GSH) concentration was observed in ethanol-induced gastric mucosal erosion (late ulcer) in rats when compared with the normal control group. Alpha-Lipoic acid treatment in ethanol- induced gastric mucosal erosion (late ulcer) in rats exhibited a significant increase in gastric tissue reduced glutathione when compared with late ulcer non-treated group. 9-Gastric tissue vitamin C: A significant decrease in gastric tissue vitamin C concentration was observed in ethanol–induced gastric mucosal erosion (early ulcer) in rats when compared with the normal control group. Pretreatment with α-Lipoic acid to ethanol- induced gastric mucosal erosion (early ulcer) in rats exhibited a significant increase in gastric tissue vitamin C when compared with early ulcer non-treated group. A non-significantly increase in gastric tissue vitamin C concentration was observed in ethanol–induced gastric mucosal erosion (late ulcer) in rats when compared with the normal control group. Treatment with α-Lipoic acid to ethanol- induced gastric mucosal erosion (late ulcer) in rats exhibited a significant increase in gastric tissue vitamin C concentration when compared with late ulcer non-treated group. 10- Serum tumor necrosis factor-alpha (TNF-α): A significant increase in serum tumor necrosis factor-alpha concentration was observed in ethanol – induced gastric mucosal erosion (early ulcer) in rats compared with the normal control group. Pretreatment with α-Lipoic acid to ethanol- induced gastric mucosal erosion (early ulcer) in rats exhibited a significant increase in serum tumor necrosis factor-alpha level in comparison with early ulcer non-treated group. A significant increase in serum tumor necrosis factor-alpha concentration was observed in ethanol – induced gastric mucosal erosion (late ulcer) in rats when compared with the normal control group. Alpha-Lipoic acid treatment in ethanol- induced gastric mucosal erosion (late ulcer) in rats exhibited a non-significant increase in serum tumor necrosis factor-alpha level when compared with late ulcer non-treated group. 11- Serum interleukin-6 (IL-6):- A significant increase in serum interleukin-6 concentration was observed in ethanol–induced gastric mucosal erosion (early ulcer) in rats when compared with the normal control group. Pretreatment with α-Lipoic acid a non-significant reduced elevated serum interleukin-6 concentrations in ethanol–induced gastric mucosal erosion (early ulcer) in rats when compared with early ulcer non-treated group A significant increase in serum interleukin-6 concentration was observed in ethanol–induced gastric mucosal erosion (late ulcer) in rats when compared with the normal control group. Treatment with α-Lipoic acid resulted in a non-significant reduced elevated serum interleukin-6 concentrations in ethanol–induced gastric mucosal erosion (late ulcer) in rats when compared with late ulcer non-treated group. 12- Gastric tissue myeloperoxidase (MPO): A significant increase in gastric tissue myeloperoxidase activity was observed in ethanol-induced gastric mucosal erosion (early ulcer) in rats when compared with the normal control group. Pretreatment with α-Lipoic acid to ethanol- induced gastric mucosal erosion (early ulcer) in rats exhibited a significant decrease in gastric tissue myeloperoxidase activity when compared with early ulcer non-treated group. A significant increase in gastric tissue myeloperoxidase activity was observed in ethanol-induced gastric mucosal erosion (late ulcer) in rats when compared with the normal control group. Alpha-Lipoic acid treatment in ethanol- induced gastric mucosal erosion (late ulcer) in rats exhibited a non-significant decrease in gastric tissue myeloperoxidase activity when compared with late ulcer non-treated group. 13-Serum sialic acid: A significant decrease in serum sialic acid concentration was observed in ethanol-induced gastric mucosal erosion (early ulcer) in rats when compared with the normal control group. Pretreatment with α-Lipoic acid to ethanol- induced gastric mucosal erosion (early ulcer) in rats exhibited a significant increase in serum sialic acid concentration when compared with early ulcer non-treated group. A non-significant decrease in serum sialic acid concentration was observed in ethanol-induced gastric mucosal erosion (late ulcer) in rats when compared with the normal control group. Treatment with α-Lipoic acid to ethanol- induced gastric mucosal erosion (late ulcer) in rats exhibited a significant increase in serum sialic acid concentration when compared with late ulcer non-treated group. 14-Gastric tissue DNA fragmentation: A significant increase in gastric tissue DNA-fragmentation was observed in ethanol-induced gastric mucosal erosion (early ulcer) in rats when compared with the normal control group. Pretreatment with α-Lipoic acid significantly reduced elevated gastric tissue DNA-fragmentation in ethanol-induced gastric mucosal erosion (early ulcer) in rats when compared with early ulcer non-treated group. A significant increase in gastric tissue DNA-fragmentation was observed in ethanol-induced gastric mucosal erosion (late ulcer) in rats when compared with the normal control group. Treatment with α-Lipoic acid significantly reduced elevated gastric tissue DNA fragmentation in ethanol-induced gastric mucosal erosion (late ulcer) in rats when compared with late ulcer non-treated group. Histopathological Results Group (1): Control normal group showing normal histological structure of the glandular epithelium of the gastric mucosa of the rat. Group (2): ulcerated non-treated group. Subgroup (a): Showing severe desquamation of the glandular epithelium lining of the gastric mucosa, the desquamated epithelium mixed with eosinophilic debris and erythrocytes were seen in the gastric lumen. Moreover, focal areas of hemorrhage and necrosis were also noticed among the affected glandular epithelium. Also, the main noticed lesions in this group were represented by the presence of multiple ulcertic areas scattered all over the gastric mucosa and the ulcerated areas showing complete destruction of the glandular tissue and the muscular layer. Moreover, the muscular layer showing leukocytic infiltration particularly of mononuclear cells. Subgroup (b): Showing severe congestion of gastric blood vessels with the presence of focal areas of hemorrhage and the desquamation of the epithelial cell lining of the gastric mucosa were also seen. Moreover, Small areas of ulceration were also noticed. Group (3): ulcerated + alpha lipoic acid protective groups showing mild desquamation of the glandular epithelium. Moreover, some of the glandular epithelium of the stomach of this group showing mild degenerative changes represented by vacuolar degeneration in the cytoplasm of the epithelial cells. Also, a focal mononuclear infiltration in the deep layer of the sub mucosa and muscular were also noticed. Group (4): ulcerated + alpha lipoic acid treated groups showing mild congestion of the gastric mucosal blood vessels with mild degenerative changes of the lining epithelial. CONCLUSION AND RECOMMENDATION from the obtained results, it could be concluded that, ethanol- induced gastric mucosal erosions in male rats caused significant decreased in serum NO and SA concentrations and in gastric tissue vitamin C level, GPX, SOD, and CAT activities. On the other hand, a marked increase in TNF-α, IL-6, MPO, L-MDA, GR and DNA-fragmentation were observed in ethanol induced gastric damage. Pretreatment of ALA was able to mitigate gastric mucosal damage induced by ethanol through increasing of SA, vitamin C, SOD, CAT, GSH in addition to decreasing DNA-fragmentation and MPO in gastric tissue. The findings of the present study demonstrated that, alpha-lipoic acid possesses significantly gastro protection and treatment effects against gastric ulcer and oxidative damage in gastric tissue induced by ethanol in rats. Since, α-lipoic acid was able to ameliorate serum biochemical parameters, enzymatic and non-enzymatic antioxidant defense system, mucus secretion, decrease the lipid peroxidation and prevent DNA fragmentation and myeloperoxidase activity in gastric tissue. Based on the data of the current study, the effect of alpha-lipoic acid against ethanol-induced gastric lesions can be attributed to the inhibitory effects on neutrophil infiltration, and its reduction of pro-inflammatory cytokines as well as its antiapoptotic effect. Because, ALA may be effective in enhances the healing of gastric ulcers by its radical scavenging and antiapoptotic activity, adjusting the pro-inflammatory cytokine, inhibited neutrophil accumulation and regenerating endogenous antioxidant mechanisms. So we recommended that, administration of diet rich in the antioxidant alpha lipoic acid is very important for protection of different body tissue, especially gastric tissue, against oxidative stress or even inflammation or erosion. Also, we strongly support the use of alpha lipoic acid as pure active ingredients in pharmacological industry for production of new drugs used as therapeutics for treatment of gastric diseases like ulcer and inflammation.