| BackgroundAlzheimer’s disease (AD), accounts for up to two-thirds of cases of dementia in the elderly individuals and affects more than 35 million people in the world, and is now estimated to be the third-leading cause of death after heart disease and cancer. The global prevalence of dementia is predicted to double every 20 years through to 2040.AD is a chronic neurodegenerative disease. The most common symptoms are gradual memory loss and deterioration of higher cognitive functions. Other characteristics include Language problems, difficulty in daily living, behavioural and neuropsychiatric problems.The pathological features are chiefly characterized by extracellular senile plaques and intracellular neurofibrillary tangles and loss of neurons. Extracellular senile plaques, also named Beta-Amyloid Plaques, which are made up of fragments of a protein called beta-amyloid peptide mixed with a collection of additional proteins, remnants of neurons, and bits and pieces of other nerve cells. Although amyloid plaque deposition does not correlate to cognitive decline, more and more evidence indicates that soluble Aβ oligomers may play a important role in AD pathogenesis and associated cognitive impairment. Neurofibrillary tangles, found inside neurons, are composed of highly phosphorylated forms of the microtubule-associated protein tau. Normal tau is required for healthy neurons. However, in AD, phosphorylated tau proteins accumulate early in neurons, even before formation of neurofibrillary tangles. Neurons fail to function normally and eventually die. The presence and extent of neurofibrillary tangles tightly linked to neuronal dysfunction and the degree of dementia. Neuronal loss is the main pathological substrate of cortical atrophy. Multiple factors initiate and promote neuronal degeneration in AD.Glutamate is the major excitatory neurotransmitter in the central nervous system and involved in all aspects of learning and memory. Glutamate is vesicularly released from pre-synaptic neurons into the synaptic cleft and binds glutamate receptors located on post-synaptic neurons to terminate the signal and prepare for another cycle of glutamate release. Glutamate is toxic in high concentrations and triggers the death of neurons. Extracellular concentrations of glutamate should be kept low to prevent exitotoxicity. Glutamate transporters are important in preventing glutamate neurotoxicity. central nervous system of mammals have five subtypes of glutamate transporters, named EAAT1 (GLAST), EAAT2 (GLT-1), EAAT3 (EAAC1), EAAT4, and EAAT5.. EAAT1 and EAAT2 are more often expressed in astrocytes. EAAT3, EAAT 4, and EAAT 5 are are neuronal glutamate transporters. Recent studies show that impairment in glutamate transporters is also plays an important role in the progression of AD.In 2005, Rothstein and his colleagues found that the β-lactam antibiotic ceftriaxone can increase GLT-1 expression to offer neuroprotection in animal model of amyotrophic lateral sclerosis. Ceftriaxone delayed loss of neurons and muscle strength, and finally increased mouse lifespan. Subsequently, in multiple in vivo and in vitro experiments confirmed that ceftriaxone effectivelyIn this study, APP/PS1 transgenic mice were treated with ceftriaxone. The aim is to investigate whether ceftriaxone would increase glutamate transporters in transgenic mouse model of AD. MethodsAPP/PS1 double-transgenic male mice and normal wild male C57BL/6 mice were obtained from Animal Research Institute of Chinese Academy of Medical Sciences. These mice were fed in a controlled environment (12:12h light-dark cycle,60% humidity,24℃), with standard diet and water. Their weight are 25-29g. Four-month-old APP/PS1 transgenic mice and normal mice assigned into four group. Each group includes 8 mices. WT+NS group mice were not treated any drug; WT+Cef group mice were intraperitoneally injected with ceftriaxone,200 mg/kg qd for seven days; APP/PS1+NS group mice were intraperitoneally injected with normal saline for seven days; APP/PS1+Cef group mice were intraperitoneally injected with ceftriaxone, 200 mg/kg qd for seven days; Three groups continued to be keep for seven days after above mentioned treatment.Mice were anaesthetized intraperitoneally with sodium pentobarbital. The brains were removed, bilateral hippocampus were save in EP tubes and preserved in Liquid nitrogen. Morphology and the number of neurons in CA1 region were observed by the method of Cresyl Violet staining. Senile plaques in the hippocampus were observed by the method of Immunohistochemical staining. GLAST and GLT-1 protein were determined by western blotting analysis. Real-time-PCR quantification of GLAST and GLT-1 mRNA was performed in order to compare their changements.Results1. Cresyl violet staining (Nissl staining):Neurons in hippocampus of mice in WT groupShowed a normal morphology and neuronal nuclear was regular. In APP/PS1 group, the Nissl body decreased and dissolved, cell body shrinkage, neuronal nuclear staing was irregular.2. The number of amyloid plaques of APP/PS1 group was significantly more than the number of WT group. After injection of ceftriaxone, APP/PS1 group did not significantly increase.3.Western blotting:GLAST and GLT-1 expression in APP/PS1 group were significantly reduced than in WT group(p<0.05). After injection of ceftriaxone, GLT-1 expression increased, but GLAST expression no significantly change.4. Realtime quantitative PCR analysis demonstrated that GLAST mRNA was no obviously difference in WT group and APP/PS1 group. GLT-1 mRNA was significantly reduced in APP/PS1 group than WT group. After injection of ceftriaxone, only GLT-1 mRNA expression increasedConclusion1. APP/PS1 transgenic mice demonstrated the number of neuron reduced, senile plaques in the brain was significantly increased.2. GLAST and GLT-1 protein expression decreased in APP/PS1 transgenic mice. GLT-1mRNA also expression decreased.3. Ceftriaxone display over-expression of GLT-1 in APP/PS1 transgenic mice. |
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