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Objective To investigate the effects of ZMS,the active component of Zhimu,on amyloid precursor protein(APP) and ?-site APP cleaving enzyme 1(BACE1) in HEK293sw cells. Methods HEK293sw cells were pretreated with different concentrations of ZMS in routine culture medium for 24 h,and then serum-free medium with ZMS was used for further culture for 48 h.Cells were examined for expression of APP and ?-secretase BACE1 with Western blotting and RT-PCR,and were then examined for BACE1 activity with fluorescence method. Results 1 ?mol/L and 10 ?mol/L ZMS significantly decreased the expression of BACE1 mRNA and protein,while had no effect on the expression of APP mRNA and protein.It was indicated by fluorescence analysis that 10 ?mol/L ZMS significantly decreased the ?-secretase activity.Conclusion ZMS significantly decreases the activity and expression of ?-secretase BACE1,while has no effect on the expression of APP in HEK293sw cells.
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Objective To investigate whether catalpol is the active component responsible for the Yin tonic effect of Radix Rehmanniae.Methods Young NH mice were injected with triiodothyronine to produce the hyperthyroidism model,while old mice were used as the model of natural aging.The single point radioligand binding assay was carried out to determine the ?-adrenergic receptor density and M-cholinergic receptor density.The learning ability(short term memory) was determined by the Y-maze avoidance test. Results In the ?-adrenergic receptor experiment,the densities were(15.7?5.2) and(20.9?7.2) fmol/mg protein in normal control group and in T3 control group(P
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Objective To establish mouse model of dementia by intracranial injection of A?25-35 and small amount of ibotenic acid(IBO) and to explore whether the effects of catalpol can affect the brain M receptor density and the short term memory. Methods The mice were randomly divided into three groups: control group,model group,treat group which were given orally for 2 months with 50 mg?kg-1?d-1 of catalpol.Dementia model was developed by single unilateral injection of 0.3 ?L of a solution of A?(1?L normal saline containing 4 ?g of A?25-35 and 1 ?g of ibotenic acid) into right basal ganglion region according the atlas of mouse brain with the aid of a stereotaxic equipment.The track of injection was observed by HE staining.The learning/memory ability was measured by Y-maze perfor-mance.The brain muscarinic receptor density was analyzed with single-site binding assay using 3H-quinuclidinyl benzilae(QNB).Results Two months after model development,the learning ability as well as the density of muscarinic receptor in brain were significantly decreased in model mice compared with those in control mice.Parallel models treated with daily oral administration of Catalpol for two months improved the learning ability and increased the brain muscarinic receptor density when compared with model mice.The correlation coefficient between total M receptor densities and the learning/memory ability was significant when examined with linear regresion.Conclusion A dementia model was established in mice.Dementia model was developed by single unilateral injection of 0.3 ?L of a solution of A?(1 ?L normal saline containing 4 ?g of A?25-35 and 1 ?g of ibotenic acid) into right basal ganglion region was established in mice.Catalpol can significantly improve the learning and increase the brain muscarinic receptor density of the model.
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Objective To investigate the effect of catalpol from Radix Rehmanniae on A?25-35-induced apoptosis of PC12 cells.Methods PC12 cells were routinely cultivated and treated by A?25-35(final concentration,20 ?mol/L) 24 hours after the addition of catalpol or saline.Forty-eight hours later,cells were examined for viability and apoptosis by MTT method and TUNEL method,respectively,while Bax and Bcl-2 mRNA expression were analyzed by semi-quantitive RT-PCR. Results Catalpol could significantly elevate the viability at 1?10-5 mol/L and 1?10-4 mol/L(P
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Objective To produce a hybridoma secreting stable monoclonal antibodies against Aβ22-35 and to develop a detection method for the assay of Aβ. Methods Spleen cells from Balb/cmice immunized with Aβ22-35-KLH were fused with mouse myeloma cells SP2/0. The techniques of immunoprecipitation and western blotting plus ECL were used to investigate the levels of Aβ in the rat brain. Results Two strains of hybridomas (3A8 and 3B2) secreting stable monoclonal antibodies raised against Aβ22-35 were obtained. The subtypes of Aβ22-35 were IgG3. The levels of Aβ in young and older rat brain were 9.8±2.8 and 13.36±2.65 (pmol/12mg brain tissues, x±s), respectively. Conclusion The Aβ22-35 mAb obtained had high titres and specificity. The levels of Aβ in the older rat brain were significantly increased as compared with the young one (P＜0.05).