Animals ; Dementia, Vascular ; metabolism ; physiopathology ; Extracellular Signal-Regulated MAP Kinases ; metabolism ; Hippocampus ; cytology ; metabolism ; physiopathology ; Male ; Mice ; Neurons ; metabolism ; physiology ; Random Allocation

This study is to investigate the effect of baicalin (BL) against oxidative injury stress of SH-SY5Y cells induced by H2O2 and the possible mechanism. SH-SY5Y cells were pre-incubated with baicalin (25, 50, and 100 micromol x L(-1)) for 12 h prior to exposure to H2O2 (150 micromol x L(-1)) for 24 h. The viability of SH-SY5Y cells was measured by MTT assay. The contents of LDH and NO were determined. The percentage of apoptotic cells was assessed by flow cytometry (FCM). The content of Caspase-3 was tested by immunofluorescence histochemical method. BL at 50 and 100 micromol x L(-1) separately increased the cell viability and up-regulated SIRT1, reduced the contents of LDH, NO, Caspase-3 and the apoptotic percentage of SH-SY5Y cells. This study results suggest that baicalin could inhibit the H2O2-induced neuronal apoptosis. The further mechanism studies show that baicalin inhibit apoptosis via reducing Caspase-3 expression and up-regulating SIRT1.
Antioxidants ; isolation & purification ; pharmacology ; Apoptosis ; drug effects ; Caspase 3 ; metabolism ; Cell Line, Tumor ; Cell Survival ; drug effects ; Flavonoids ; isolation & purification ; pharmacology ; Humans ; Hydrogen Peroxide ; toxicity ; L-Lactate Dehydrogenase ; metabolism ; Neuroblastoma ; metabolism ; pathology ; Nitric Oxide ; metabolism ; Oxidative Stress ; drug effects ; Plants, Medicinal ; chemistry ; Scutellaria ; chemistry ; Sirtuin 1 ; metabolism ; Up-Regulation

OBJECTIVETo explore whether the constructed vector of short haprin in vivo can induce human glioma cell line BT325 to produce RNAi duplexes and reverse the expression of MDR1 gene.

METHODSThree 62nt oligonucleotide fragments (shRNA) were constructed according to GenBank MDR1 sequence and were cloned to the retrovirus-delivered vectors. After transfected these vectors directly into the human malignant glioma BT325 cells by lipofectamine 2000 with enhanced green fluorescence protein (EGFP) co-transfecting, the MDR1 gene silence effects were detected by the changing level of mRNA and P-glycoprotein including real time PCR (RT-PCR), Northern blot and Western blot analysis. To assess the multidrug resistance against adriamycin (ADR) and VCR, cell proliferation assays were performed by cell counting kit-8.

RESULTSThe RNAi plasmid vectors were constructed successfully. RT-PCR showed MDR1 mRNA was significantly reduced (P < 0.05). Northern blot analysis showed that the gene silence became most intense at 48 hours after transfection. Western blot analysis demonstrated that P-gp expression was reduced at different time to 12.9%, 30.3% and 4.8%, respectively. The chemosensitivity assays indicated that the transfected cells showed an enhanced sensitivity to ADR and VCR. Based on the value of IC(50), BT325 cells had significantly increased sensitivity to the drugs.

CONCLUSIONThe sequence specific RNAi can inhibit MDR1 mRNA and P-gp expression in the glioma cell line. It may reverse multidrug resistance phenotype, therefore, may provide promising therapeutic modalities in the treatment of human glioma.

ATP-Binding Cassette, Sub-Family B, Member 1 ; biosynthesis ; genetics ; Antibiotics, Antineoplastic ; pharmacology ; Antineoplastic Agents, Phytogenic ; pharmacology ; Cell Line, Tumor ; Doxorubicin ; pharmacology ; Drug Resistance, Multiple ; drug effects ; Drug Resistance, Neoplasm ; drug effects ; Glioma ; metabolism ; pathology ; Humans ; Plasmids ; RNA Interference ; RNA, Messenger ; biosynthesis ; genetics ; RNA, Small Interfering ; genetics ; pharmacology ; Transfection ; Vincristine ; pharmacology

OBJECTIVETo get a better understanding of the mechanisms underlying type 1 diabetes mellitus, the differentially expressed pancreatic proteins from multiple low-dose streptozotocin (MLD-SIZ) mouse and normal mouse were analyzed and compared.

METHODS20 male rats were separated into 2 groups (n=10): model mice treated with MLD-STZ and normal mice,differences of pancreatic proteome among in the two groups of mice, were analyzed by two dimensional polyacryamide gel electrophoresis (2DE). Protein quantification was analyzed and the differentially expressed spots were identified using mass spectrometry and MASCOT database searching.

RESULTSCompared with control group, 23 proteins had changed significantly in the model group, 8 proteins expression were up-regulated, 15 proteins expressions down-regulated significantly. Using MALDI-TOF-MS, 15 proteins with significant change were identified by peptide fingerprinting map and the results were searched in MASCOT database. The function analyzed showed that proteins with change were associated with metabolic, anti-oxidant, structural, catalytic enzymes and chaperone, et al.

CONCLUSIONType 1 diabetes is probably exerted via multi-target and multi-path mechanism. The proteins with significant change are newly target for type 1 diabetes early diagnosis and treatment.

Animals ; Diabetes Mellitus, Experimental ; physiopathology ; Diabetes Mellitus, Type 1 ; chemically induced ; metabolism ; physiopathology ; Male ; Mice ; Pancreas ; metabolism ; Proteins ; metabolism ; Proteomics ; methods ; Streptozocin

OBJECTIVETo prepare a time-resolved fluoroimmunoassay (TRFIA) kit for clinical detection of IgM antibodies to hepatitis B core antigen (HBc).

METHODSImmunocapture method was used to develop the TRFIA kit for detection of the anti-HBc IgM antibodies, and the precision, cross-reactivity and sensitivity of the kit were tested with the clinical serum samples.

RESULTSThe intra- and inter-assay coefficients of variation of the TRFIA kit were 4.8%-7.2% and 7.5%-8.6%, respectively, and no cross-reactivity with anti-HBs, anti-HBc-IgG or anti-HBe was found. Comparison of the results of the TRFIA kit and enzyme-linked immunosorbent assay (ELISA) demonstrated greater sensitivity of the kit than ELISA in detecting the anti-HBc IgM antibodies in 584 serum samples. According to the detection results in 300 serum samples from healthy donors, the cutoff value of the TRFIA kit was 4.5 times of the fluorescence value of the negative control.

CONCLUSIONThis TRFIA kit for detecting anti-HBc IgM antibodies meets the demand for clinical application and can replace the ELISA kits.

Fluoroimmunoassay ; methods ; Hepatitis B ; immunology ; virology ; Hepatitis B Antibodies ; blood ; Hepatitis B Core Antigens ; immunology ; Humans ; Immunoglobulin M ; blood ; Reagent Kits, Diagnostic ; Sensitivity and Specificity

OBJECTIVETo investigate morphological changes of capillary in aging brain and explore the role of vascular factor in brain aging.

METHODSTwenty-eight brains of individuals (mean age 65 years) who died without clinical or pathological involvement of nervous system and 6 brains of Alzheimer's disease (AD) patients (mean age 83 years) were obtained at autopsy. Sections from frontal lobe, occipital lobe, striatum and hippocampus of normal subjects and sections from hippocampus of AD patients were used for hematoxylin eosin (HE), lox fast blue (LFB), toluidine blue stains and ulex europaeus agglutinin (UEA) immunostaining. After observations of morphological changes of neuron and capillary, computer-aid image analysis was performed to quantify numerical density and area density of neuron and capillary in frontal lobe, occipital lobe, putamen, CA3 sector of normal subjects and CA3 sector of AD patients. Numerical ratio and area ratio of neuron and capillary were then calculated. Correlations between neuron/capillary ratio and age were estimated using Pearson's correlation test. Difference of neuron/capillary ratio in CA3 sectors between AD patients and advanced aged normal subjects (> 75 years) was analyzed with Student's t-test.

RESULTSSeveral pathological microvascular changes, including increased tortuosity, looping, bundling, stringing, and effacement of endothelia were seen in aged subjects and more prevalent in AD patients. Numerical ratio and area ratio of neuron and capillary of frontal lobe, occipital lobe and putamen significantly increased with age in normal aging subjects.

CONCLUSIONSMorphological changes and relative decrease in number and capacity of capillary in aging brain may reduce cerebral blood flow and metabolism, and consequently result in functional impairment of aging brain. Vascular factors may play an important role in the development of brain aging.

Adult ; Aged ; Aged, 80 and over ; Aging ; Alzheimer Disease ; etiology ; pathology ; Capillaries ; anatomy & histology ; pathology ; Cell Count ; Cerebral Cortex ; blood supply ; pathology ; Female ; Frontal Lobe ; blood supply ; pathology ; Hippocampus ; blood supply ; pathology ; Humans ; Image Processing, Computer-Assisted ; Male ; Middle Aged ; Neurons ; pathology ; Occipital Lobe ; blood supply ; pathology

OBJECTIVETo investigate the effect of Shuyusan decoction on neuropeptide Y (NPY) and serotonin (5-HT) expression in the hippocampus and plasma of rats with chronic mild unpredictable stressors depression.

METHODSFifty Wistar rats were randomly divided into 5 groups, namely the normal control group, model group, fluoxetine group, and high- and low-dose Shuyusan groups. Except for those in the normal control group, all the rats were subjected to chronic mild unpredicted stress for 21 consecutive days with corresponding treatments. Open-field test was used to assess the behavioral changes of the rats. The content of NPY in the hippocampus and plasma was detected by competitive enzyme-linked immunosorbent assay, and immunocytochemistry was used to determine the expression of 5-HT in the hippocampus.

RESULTSNPY levels in the hippocampus and plasma was significantly decreased in the model group as compared with that in the normal control group (P<0.05). Treatments with fluoxetine and high-dose Shuyusan both significantly increased NPY levels in the hippocampus and plasma in the depressive rats (P<0.05), resulting also in significantly increased 5-HT-immunoreactive neurons in the cerebral cortex and hippocampus and the average optical density (P<0.05).

CONCLUSIONShuyusan decoction can effectively increase plasma and hippocampus NPY levels and the number of 5-HT-positive neurons in the cerebral cortex and happocampus of rats with chronic mild unpredictable stress-induced depression.

Animals ; Depression ; etiology ; metabolism ; Drugs, Chinese Herbal ; pharmacology ; Female ; Hippocampus ; metabolism ; Male ; Neurons ; metabolism ; Neuropeptide Y ; metabolism ; Random Allocation ; Rats ; Rats, Wistar ; Serotonin ; metabolism ; Stress, Physiological

OBJECTIVETo characterize histopathologic features of non-Alzheimer type dementia.

METHODSBodian, Gallyas-Braak silver staining, tau and ubiquitin immunohistochemistry were applied in an analysis of 22 cases of autopsy-proven neurodegenerative dementia. Appearance, distribution and immunoreactivity of neuronal and glial inclusions in the brain were observed. The final histological diagnoses were made according to the pathological criteria for several types of common non-Alzheimer type dementia.

RESULTSAmong the 22 cases of neurodegenerative dementia, 12 cases were identified as non-Alzheimer type dementia, including Pick's disease (2 cases), progressive supranuclear palsy (3 cases) and corticobasal degeneration (3 cases), dementia with Lewy bodies (1 case), and Parkinson's disease (3 cases). Another 10 cases consisted of pure Alzheimer's disease (AD, 9 cases) and AD combined with argyrophilic grain disease (1 case). Characteristic neuronal and glial inclusions, such as classical and cortical Lewy body, Pick body, Globous NFTs, astrocytic plaque and tufted astrocyte, argyrophilic grain were found in the brains of non-Alzheimer type dementia. Classical and cortical Lewy bodies were not argyrophilic but were immunoreactive to ubiquitin. Pick bodies, Globous NFTs, astrocytic plaques, tufted astrocytes and argyrophilic grains were all argyrophilic. Pick bodies showed tau and ubiquitin immunoreactivity. However, Globous NFTs, astrocytic plaques, tufted astrocytes, and argyrophilic grains were reactive only to tau immunohistochemistry.

CONCLUSIONSFindings of characteristic neuronal and glial inclusions may help to differentiate non-Alzheimer type dementia from AD, and in conjunction with Gallyas-Braak staining and immunohistochemistry for tau and ubiquitin, to further define histopathologic subcategories of non-Alzheimer type dementia.

Aged ; Aged, 80 and over ; Brain ; pathology ; Dementia ; pathology ; Diagnosis, Differential ; Female ; Humans ; Lewy Body Disease ; pathology ; Male ; Neurodegenerative Diseases ; pathology ; Neurons ; pathology ; Parkinson Disease ; pathology ; Pick Disease of the Brain ; pathology ; Supranuclear Palsy, Progressive ; pathology

OBJECTIVETo study the expression changes of neuroglobin in rats with the model of diffuse traumatic brain injury and explore the relationship between the neuroglobin and neuron apoptosis in traumatic brain injury.

METHODSThe diffuse traumatic brain injury of rats was induced by the Marmarou's 'weight-drop' device. And the immunohistochemical technique was used to detect the expression changes of neuroglobin and neuron apoptosis in rat brain at different time points post-injury.

RESULTSThe expression of neuroglobin increased twice and reached peaks at 2 hours and 72 hours post-injury respectively. And the increased expression of neuroglobin from 30 minutes to 1 hour post-injury and from 48 hours to 72 hours post-injury accompanied with the decreased expression ratio of Bax to Bcl-2.

CONCLUSIONThe increased expression of neuroglobin in traumatic brain injury informed us that neuroglobin had anti-apoptosis action in post-injury neuron. It could protect the neuron from traumatic stress and secondary ischemia and hypoxia insults during ultra-early and acute stages.

Animals ; Apoptosis ; physiology ; Brain ; metabolism ; pathology ; Brain Injuries ; metabolism ; pathology ; Globins ; metabolism ; Male ; Nerve Tissue Proteins ; metabolism ; Neurons ; pathology ; Random Allocation ; Rats ; Rats, Sprague-Dawley

The aim of the present study is to investigate the effect of progesterone-induced expression of cyclin G1 on the proliferation of endometrial epithelial cells. To obtain mouse endometrial epithelial cells, the uteri were isolated from ovariectomized mice which were injected subcutaneously with 100 ng estradiol per day for two days. Then the uteri were digested by dispase and pancreatin respectively. Endometrial epithelial cells were cultured in DMEM/F12 containing 6% fetal bovine serum, and divided into four groups when they grew to confluence. Each of the groups was treated as follows: Group E was treated with 0.01 micromol/L estradiol only, group P was treated with 1 micromol/L progesterone, group EP was treated with both 0.01 micromol/L estradiol and 1 micromol/L progesterone, and group C was treated with 0.01% DMSO for control. Immunocytochemistry was used to examine the expression of cyclin G1 protein. MTT assay was used to evaluate metabolic activity of cells. Flow cytometry was used to check the number of cells distributing in each phase of the cell cycle. The result of immunocytochemistry showed that there was no expression of cyclin G1 protein in group C and group E, while cyclin G1 was obviously expressed in group P and group EP and localized in nucleus. In the MTT assay, compared with group C, the viability of group E significantly increased, while that of both group P and group EP decreased significantly. The results of flow cytometry were in accordance with those of MTT, which showed that compared with group C, group E had a higher proportion of cells in S phase, while group P, as well as group EP had a lower proportion of cells in S phase but a higher proportion in G1 phase and G2/M phase. These results indicate that progesterone could induce cyclin G1 expression in the primary culture of mouse endometrial epithelial cells, meanwhile inhibit the proliferation of cells and block the cell cycle progression. Thus, progesterone-induced expression of cyclin G1 is probably a negative factor in regulating cell cycle, which is involved in the inhibitory effect of progesterone on the proliferation of endometrial epithelial cells.
Animals ; Cell Cycle ; Cell Division ; Cell Proliferation ; drug effects ; Cyclin G1 ; metabolism ; Epithelial Cells ; cytology ; drug effects ; metabolism ; Estradiol ; pharmacology ; Female ; Flow Cytometry ; Mice ; Ovariectomy ; Progesterone ; pharmacology ; Uterus ; cytology