Beta-Ecdysone Protects Mouse Osteoblasts from Glucocorticoid-Induced Apoptosis In Vitro.

Glucocorticoid-induced osteoporosis is a common form of secondary osteoporosis. Glucocorticoids affect both bone formation and resorption, and prolonged glucocorticoid exposure can suppress osteoblast activities. beta-Ecdysone, found in many plants, is involved in protein synthesis, carbohydrate and lipid metabolism, and immunologic modulation. Here, we evaluated the effects of beta-ecdysone on osteoblast viability by assessing apoptosis following treatment with excess glucocorticoids. Mouse bone marrow stromal cells were induced to differentiate and grow into osteoblasts, and then treated with 10 µM glucocorticoid and 10, 1, or 0.1 µM beta-ecdysone. The expression levels of osteoblast growth and differentiation factors (runt-related transcription factor 2, osteogenic protein-1, and alkaline phosphatase), apoptosis-related genes (transformation-related protein 53, ataxia telangiectasia mutated protein, caspase-3, and caspase-8), and Akt1 and phospho-Akt (Thr308) were then assessed via alkaline phosphatase staining, acridine orange-propidium iodide staining, annexin V/PI apoptosis assay, real-time RT-PCR, and Western blot analyses. Notably, treatment with 10 µM glucocorticoid resulted in reduced osteoblast viability and the specific activity of alkaline phosphatase as well as reduced runt-related transcription factor 2, osteogenic protein-1, and alkaline phosphatase mRNA expression in vitro, indicating that glucocorticoid inhibited osteogenic differentiation. Moreover, glucocorticoid treatment yielded increased transformation-related protein 53, ataxia telangiectasia mutated protein, caspase-3, and caspase-8 expression and decreased Akt1 and phospho-Akt levels, indicating glucocorticoid-induced apoptosis. Meanwhile, beta-ecdysone inhibited glucocorticoid function, preserving the expression of Akt1 and phospho-Akt and reducing the expression of transformation-related protein 53, ataxia telangiectasia mutated protein, caspase-3, and caspase-8. Thus, beta-ecdysone prevented glucocorticoid-induced osteoblast apoptosis in vitro. These data highlight the potential for beta-ecdysone as a treatment for preventing the effects of glucocorticoid on bone growth.

Effects of 5-hydroxymethyl furfural extracted from Rehmannia glutinosa Libosch on the expression of signaling molecules relevant to learning and memory among hippocampal neurons exposed to high concentration of corticosterone.

OBJECTIVE: To determine the effects of 5-hydroxymethyl furfural (5-HMF), an extract of Rehmannia glutinosa Libosch, on several down-regulated signaling molecules involved in learning and memory in hippocampal neurons. METHODS: After cultured for 7 days, primary hippocampal neurons were divided into 5 groups: normal, corticosterone model, RU38486, 5-HMF, and donepezil group. Neuron survival rates were calculated 24 h later using SYTO13-PI double-fluorescence staining and an 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. ß-galactosidase activity was also assayed. Protein expressed by the glucocorticoid receptor (GCR), brainderived neurotrophic factor (BDNF), and N-methyl-D-aspartate receptor 2B (NR2B), as well as phosphorylationcyclic adenosine monophosphate (cAMP) response element binding protein (p-CREB), phosphorylation-extracellular signal-regulated kinase (p-ERK), and phosphorylation-synapsin (p-synapsin) were quantified with Western blot. RESULTS: Hippocampal neuron survival rates and the above-mentioned proteins were dramatically decreased (P<0.05), ß-galactosidase activity was significantly increased in the model group. but the effect was reversed by 5-HMF, RU38486, and to a lesser extent by donepezil (P<0.05). CONCLUSION: 5-HMF extracts from the Chinese herb Rehmannia glutinosa Libosch could protect hippocampal neurons from glucocorticoid injury and from down-regulated signaling molecules in the GCR-BDNF-NR2B-p-ERK-p-CREB-p-synapsin signal transduction pathway.

[Analysis of variance of repeated data measured by water maze with SPSS].

OBJECTIVE: To introduce the method of analyzing repeated data measured by water maze with SPSS 11.0, and offer a reference statistical method to clinical and basic medicine researchers who take the design of repeated measures. METHODS: Using repeated measures and multivariate analysis of variance (ANOVA) process of the general linear model in SPSS and giving comparison among different groups and different measure time pairwise. RESULTS: Firstly, Mauchly's test of sphericity should be used to judge whether there were relations among the repeatedly measured data. If any (P

[Effects of recipes for replenishing qi and activating blood on senescence related gene expressions in the liver of aging rats].

OBJECTIVE: To observe the effects of recipes for replenishing qi and activating blood on p16, p21, proliferating cell nuclear antigen (PCNA), cyclin D1 and cyclin E gene expressions in the liver of aging rats. METHODS: A recipe for replenishing qi and a recipe for activating blood were administered to aging rats respectively, and the effects of the above recipes on the expressions of senescence related genes (p16, p21, PCNA, cyclin D1 and cyclin E) were examined by RT-PCR and Western blotting methods. RESULTS: The expressions of p16, p21 and cyclin D1 mRNAs and proteins in the liver of the untreated aging rats were up-regulated, while the expressions of PCNA and cyclin E mRNAs and proteins decreased. As compared with the untreated aging rats, both recipes could down-regulate the expressions of cyclin D1 mRNA and protein and up-regulate the expressions of cyclin E mRNA and protein, but had no obvious effects on the expressions of mRNAs and proteins of p16, p21 and PCNA. CONCLUSION: Recipes for replenishing qi and activating blood can improve the liver cell proliferation of aging rats via down-regulating the expressions of cyclin D1 mRNA and protein and up-regulating the expressions of cyclin E mRNA and protein.

[Effect of heart benefiting recipe in controlling IL-1beta, IL-6 and APPmRNA expression in brain of beta-amyloid protein induced rat model of dementia].

OBJECTIVE: To investigate the neuro-immune regulatory mechanism of Heart Benefiting recipe (HBR), an effective recipe for treatment of Alzheimer's disease (AD). METHODS: Using immunohistochemical and RT-PCR methods, the neuro-immunological pathological changes in the AD rat model induced by beta-amyloid protein (A beta1-40) via lateral cerebral ventricle injection, including mainly the glial fibrillary acidic protein expression and inflammatory cytokines IL-1beta, IL-6mRNA and beta-amyloid protein precursor (APPmRNA) gene expression were studied. And the effects of HBR on these parameters were observed. RESULTS: Deposition of A beta in cerebral tissue could induce activation of stellate glial cells and abnormal increased levels of inflammatory cytokines (IL-1beta and IL-6mRNA), also the elevation of APPmRNA level. HBR could effectively control the above-mentioned pathological changes. CONCLUSION: HBR could effectively control the inflammation and the A beta immune cascade reaction in brain of AD patients, it is one of the important therapeutic mechanisms of the recipe.

[Establishment of analogous oxidative damaged Alzheimer's disease rat model and effect of tiaoxin recipe on it].

OBJECTIVE: To establish a convenient, economical and practical analogous oxidative damaged Alzheimer's disease rat model (AD model) for exploring the effect of Tiaoxin Recipe (TXR) on the spatial memory capacity and beta-amyloid protein (A beta) deposition in the model. METHODS: The AD model was established by left ventricular injection of DHF-FeCl3-ADP. Spatial memory and learning capacity of the model rat was observed by Morris water maze method, A beta deposition in its cerebral cortex was observed by immunohistochemistry, and the effect of TXR was analysed. RESULTS: Compared with the normal group, the spatial memory capacity in the model group was obviously decreased, with A beta widely deposited in cortex, immunohistochemical examination showed that the number of A beta positive cells and their mean optic density significantly increased. TXR displayed significantly improving effect on the above-mentioned changes. CONCLUSION: The oxidative damaged model could not only express the clinical characteristics (short-term memory impairment), but also partially reflex the pathological changes (A beta deposition) of AD, is an economical and practical analogous AD model. TXR has the effects of improving spatial memory impairment and lowering A beta deposition in the AD model rats.