The Protection Of Taurine Combined With DFP Against Aluminum-induced Neural Oxidative Injuries And Apoptosis

ObjectiveAluminum (Al) is the most abundant metal elements in earth’s crust which widely exists in air, soil, and water. Because of its specific physico-chemical properties, Al was also widely used for the materials of purifying agent, tableware, food packaging, food additives and medicine. People are more frequently exposed to aluminum, which causes Al intake. Al can accumulate in human body and cause toxic effects on many kinds of organs and tissue. The neurotoxicity induced by aluminum has been paid close attention in recent years. There is circumstantial evidence demonstrated that aluminum neurotoxicity links with Alzheimer’s disease(AD), Parkinson’s Disease (PD) and Dialysis Encephalopathy (DE). Al can produce toxic effect in nerve cells by enhancing lipid peroxidation, which caused oxidative damage, and ultimately lead to apoptosis. Early studies have confirmed that 1,2-dimethyl-3-hydroxypyrid-4-one(DFP) is used as a chelator for Al overload, with characteristic of cheap, orally active and low-toxic. Taurine (Tau), an abundant amino acid, plays a wide range of biological effects in the body. Very little is known yet about Tau combined with DFP protection against Al-induced oxidative damage and apoptosis. Wistar rats were administered with Aluminum Chloride Hexahydrate (AlC13·6H2O) intragastrically to build animal models, and then were treated with Tau or DFP alone or Tau combined with DFP in aspect of treatment. We determined, antioxidant enzyme activities, oxidative damage, adenosine triphosphatase (ATPase) activity, and expression of apoptosis-related gene and protein, to study and discuss the protective role of Tau combined with DFP against Al-induced neural apoptosis and oxidative injuries in rats.Methods1. Animal experiment84 male Wistar rats (180-220g) were radomized into seven groups of 12 animals each based on their body weights. The tested chemicals were given by gavage, once a day. Rats in control group received 1.0ml/d saline for 8 weeks. Al exposure group, Tau does group, DFP low dose group, DFP high dose group, Tau+DFP low dose group, Tau+ DFP high dose group received 281.4mg/kg daily Aluminum Chloride Hexahydrate (AICl3·6H2O) for the first 4 weeks. Then for the last 4 weeks, they were given 1.0ml/d saline,400mg/kg Tau,13.82mg/kg DFP,27.44mg/kg DFP,400mg/kg Tau,400mg/kg Tau, respectively; Tau+DFP low dose group and Tau+DFP high dose group were given 13.82,27.44 mg/kg DFP 6h right after the taurine treatment, respectively.Two rats selected from each group were perfused by 4% paraformaldehyde for apoptosis of hippocampus assay. The other rats were fasted for 24h after the treatments and then were sacrificed, and blood was collected. Their brains were quickly removed and isolated cortex and hippocampus. All the samples were stored for the use of bioassays.2. Effect of Tau and DFP on antioxidant system in Al-exposed rats’cortexAfter the rats were sacrificed, cortexes were separated and made into tissue homogenate with normal saline. The Malondialdehyde (MDA) content, the activities of Glutathione peroxidase (GSH-Px) and Superoxide dismutase (SOD) were determined, to investigate the protective role of Tau combined with DFP against Al-induced oxidative injuries in cortex of rats.3. Effect of Tau and DFP on antioxidant system in serum of rats exposed to AlAll the rats were sacrificed after the treatments. Blood was immediately collected in centrifuge tubes and centrifuged to collect serum. The MDA content, the activities of GSH-Px and SOD were determined, to study the effect of Tau combined with DFP on improving the antioxidant indices in serum of Al-exposed rats.4. Effect of Tau and DFP on the activity of ATPase in Al-exposed rats’ cortexAfter the rats were sacrificed, cortexes were separated and made into tissue homogenate. The activities of Na+K+-ATPase. Ca2+-ATPase and Mg2+-ATPase were tested, to investigate antagonistic effect of Tau combined with DFP on Al-induced reduction of ATPase activities in cortex of rats.5. Effect of Tau and DFP on the activity of ATPase in blood of rats exposed to AlThe blood of was immediately collected after they were sacrificed and a part of it was made into dissolving blood. The activities of Na+K+-ATPase, Ca2+-ATPase and Mg2+-ATPase in blood were tested, to study the protective role of Tau combined with DFP on ATPase in blood of rats exposed to Al.6. Effect of Tau and DFP on apoptosis in Al-exposed rats’ hippocampusAfter all the treatments, the rats were administered with cardiac perfusion by 4% paraformaldehyde to fixate brain tissues and make paraffin sections for Terminal dexynucleotidyl transferase(TdT)-mediated dUTP nick end labeling(TUNEL) test. The expression of Bcl-2 and Bax mRNA were analyzed by RT-PCR, and the expression of Bcl-2, Bax and cleaved Caspase-3 were analyzed by western blotting, to investigate antagonistic effect of Tau combined with DFP on Al-induced apoptosis in hippocampus of rats.Results1. Effect of Tau and DFP on antioxidant system in Al-exposed rats’ cortexExposure of AlCl3·6H2O significantly raised level of MDA, and decreased activities of GSH-Px and SOD in cortex, as compared to control rats (P<0.05). As compared to Al-exposure rats, treatments with Tau, high does DFP, Tau+low-dose DFP and Tau+high-dose DFP significantly reversed the increase of MDA level and decrement activities of antioxidant enzymes (P<0.05). Treatments with Tau+ low-dose DFP induced lower MDA level and higher activities of GSH-Px and SOD, when compared with either drug alone (P<0.05). Administration of Tau+high-dose DFP significantly improved activities of GSH-Px and SOD, and reduced MDA level, as compared with Tau only (P<0.05).2. Effect of Tau and DFP on antioxidant system in serum of rats exposed to AlExposure of AlCl3·6H2O significantly impaired activity of GSH-Px and raised level of MDA in serum, as compared to control rats (P<0.05). When compared to Al-exposure rats, treatment with Tau, high dose DFP, Tau+ low-dose DFP and Tau+ high-dose DFP significantly reduced level of MDA and improved activity of GSH-Px (P<0.05). Treatments with Tau+ low-dose DFP induced lower MDA level and higher activity of GSH-Px, when compared with either drug alone (P<0.05). Administration of Tau+ high-dose DFP significantly improved GSH-Px activity, and reduced MDA level, as compared with Tau only.3. Effect of Tau and DFP on the activity of ATPase in Al-exposed rats’cortexExposure of AlC13·6H2O significantly impaired activity of Na+K+-ATPase Mg2+-ATPase and Ca2+-ATPase in cortex, as compared to control rats (P<0.05). As compared to Al-exposure rats, treatments with Tau, high dose DFP, Tau+low-dose DFP and Tau+high-dose DFP significantly improved activities of ATPase (P<0.05). Treatments with Tau+low-dose DFP induced higher activities of Na+K+-ATPase, Mg2+-ATPase and Ca2+-ATPase, when compared with either drug alone (P<0.05). Administration of Tau+high-dose DFP significantly improved Na+K+-ATPase, Mg2+-ATPase and Ca2+-ATPase, as compared with Tau only (P<0.05).4. Effect of Tau and DFP on the activity of ATPase in blood of rats exposed to AlExposure of A1C13-6H2O significantly impaired activity of Na+K+-ATPase, Mg2+-ATPase and Ca2+-ATPase in blood, as compared to control rats (P<0.05). As compared to Al-exposure rats, treatments with high dose DFP, Tau+low-dose DFP and Tau+high-dose DFP significantly improved activities of ATPase (P<0.05). Treatments with Tau+low-dose DFP induced higher activities of Na+K+-ATPase, Mg2+-ATPase and Ca2+-ATPase, when compared with either drug alone (P<0.05). Administration of Tau+ high-dose DFP significantly improved Na+K+-ATPase, Mg2+-ATPase and Ca2+-ATPase, as compared with Tau only (P<0.05).5. Effect of Tau and DFP on apoptosis in Al-exposed rats’ hippocampusNo TUNEL-positive cells were shown in control rats, but Al-treated led to numerous TUNEL-positive cells (brown colored). Further, treatment with taurine or DFP alone as well as combination of DFP and taurine could decrease the TUNEL positivity. There were no TUNEL-positive cells in Tau+ DFP high dose group’s rats.Exposure of AlCl3·6H2O significantly raised mRNA level of Bax, and decreased mRNA level of Bcl-2 in hippocampus, as compared to control rats (.P<0.05). As compared to Al-exposure rats, treatments with taurine, DFP, and Tau+DFP significantly reversed the increase of mRNA level of Bax and decrement mRNA level of Bcl-2 (P<0.05). Besides, Treatments with Tau+DFP induced lower mRNA level of Bax and higher mRNA level of Bcl-2, when compared with either drug alone (P<0.05).Exposure of AICl3·6H2O significantly reduced level of Bcl-2 and raised level of Bax and cleaved Caspase-3 in hippocampus, as compared to control rats (P<0.05). When compared to Al-exposure rats, treatment with taurine, DFP, and Tau+ DFP significantly decreased levels of Bax and cleaved caspase-3 and improved level of Bcl-2 (P<0.05). When compared with either drug alone, treatments with Tau+ DFP induced lower levels of Bax and cleaved caspase-3 and higher Bcl-2 level, except Tau +low-dose DFP compared with low-dose DFP only.Conclusions1. Tau combined with DFP was effective in protecting rats’cortex and serum from Al-induced oxidative injuries.2. Tau combined with DFP could effectively inhibit the significant reduction of ATPase activity induced by Al in rats’cortex and blood.3. Tau could effectively regulate the expression of neural apoptosis related gene and protein, and thus effectively alleviate Al-induced hippocampus cells apoptosis.In conclusion, Tau combined with DFP can effectively antagonize neural oxidative injuries and apoptosis induced by Al and thus play a role in protecting rats’nervous system, and it is prior to DFP or taurine only.