ABSTRACT
In order to study the expression of melatonin receptors Mel1a and RORβ in the human brain,and to furthe investigate the melatonin's function in Alzheimer's disease,two melatonin receptor cDNAs were cloned and introduced into pBluescript Ⅱ (KS+)vector.The recombinant plasmids were identified by using restriction enzymes and then sequenced.The results indicated that the cloned DNA sequences and the Genbandk Sequences were inentical.The cRNA probes made from them can be used in the in situ hybridization immunohistochemistry to detect the mRNA expression level in the future study.
ABSTRACT
<p><b>OBJECTIVES</b>To explore the mechanisms of neuronal loss and apoptosis in the brains of Alzheimer's disease (AD) patients, through studying the expression of proteins related to signal transduction pathways, which are important for neuron survival.</p><p><b>METHODS</b>(1) Immunohistochemistry: Sections were double stained with Tunel and NSE antibodies. (2) The hippocampal tissue taken from 6 cases of AD and 6 cases of non-AD brains was homogenized. Protein estimation was done by Lowry method. Equal amounts of protein were taken from each specimen and immunoprecipitation was performed and analyzed by Western blot; color development was done by alkaline phosphatase method or luminol reagent.</p><p><b>RESULTS</b>(1) Tunel positive neurons were found in both AD and non-AD brains, but the number in the former was more than the latter. (2) The AD hippocampal tissue showed diminished expression of Akt/PKB, CREB, P-CREB, increased expression of apoptosis-related protein apoptosis-inducing factor, and diminished expression of apoptosis-related protein bcl-2. The expression of bax did not change.</p><p><b>CONCLUSIONS</b>Diminished expression of CREB, P-CREB, bcl-2 in AD hippocampus indicates that the neuron survival signal transduction pathway in AD brains is impaired. Neurons are in apoptotic or pro-apoptotic state. In addition, increased expression of apoptosis-inducing factor, diminished expression of bcl-2, which is an anti-apoptotic factor, promotes further neuron apoptosis.</p>
Subject(s)
Aged , Aged, 80 and over , Humans , Male , Alzheimer Disease , Metabolism , Pathology , Apoptosis , Cyclic AMP Response Element-Binding Protein , Metabolism , Neurons , Metabolism , Pathology , Proto-Oncogene Proteins c-bcl-2 , Metabolism , Signal Transduction , PhysiologyABSTRACT
Objective To investigate p75 NTR expressing neurons and hyperphosphorylated tau protein containing neurons in the hippocampal CA1 subfield of Alzheimer's disease. Methods Samples of hippocampus of 10 female AD patients and 10 non-demented female controls matched with age and postmortem delay were obtained from the Netherlands Brain Bank. The main body of hippocampus was dissected, dehydrated and embedded in paraffin. Serial 6-?m coronal sections were cut, and 3 successive sections were selected. The first section was stained with 0.5% thionin, the second was processed for p75 NTR immunocytochemistry and the third was processed for p75 NTR double-labeling immunocytochemistry with Alz-50. For quantitative analysis of the total number of neurons, p75 NTR expressing neurons and neurons colocalizing p75 NTR and Alz-50 in the CA1 subfields, a MetaMorph image acquisition and processing software (Universal Imaging Corp, USA) was used.Results The total number of neurons, p75 NTR immunoreactive neurons and ratio of the latter to the former in 1 mm2 of the CA1 subfield of AD patients were 293.2?37.0, 116.0?20.4 and 39.7%?5.3%, respectively, of controls were 473.6?63.1, 136.7?24.4 and 28.9%?3.7%, respectively. There was significant decrease in the total number of neurons of the AD patients in comparison with controls (P=0.000 2). However, the ratio of p75 NTR neurons to total number of neurons was significant increase in AD patients compared with controls (P=0.001). The number of Alz-50 neurons, Alz-50 and p75 NTR double-labeling neurons and ratio of the latter to the former in 1 mm2 of the CA1 subfield of AD patients were 87.5?29.2, 76.4?26.6 and 86.6%?5.0%, respectively. Furthermore, the number of p75 NTR containing neurons was significantly correlated to the number of Alz-50 expressing neurons (r=0.79, P=0.006). Conclusion The p75 NTR may interact with hyperphosphorylated tau protein and involve in the pathogenesis of Alzheimer′s disease.
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AIM To investigate the relationships between the alterations in circadian rhythms of cortisol, melatonin and nocturnal asthma METHODS Circadian rhythms of salivary free cortisol and melatonin levels were investigated in 15 control subjects and 8 exacerbation and 7 remission patients with bronchial asthma The serial salivary samples were collected at 12 time points during a 24 hour period The intensity of illumination was restricted to 50 lux during the night Salivary cortisol and melatonin levels were measured by radioimmunoassay RESULTS The 24 hour mean levels (mesor) of salivary cortisol was significantly different by one way ANOVA among the three groups ( P