Nerve growth factor is one of the most important factors playing an important role in regulating the growth, development and survival of the neuron. The purified NGF from human placenta has been reported, the tissue from which can be isolated the NGF is very limited. It is important for basic research and clinic application to expression hNGF by genetic engineering. By polymerase chain reaction,gene fragment encoding the mature part of ?-NGF was amplified using the DNA of human placenta as template. The fragment was sequenced and inserted into expression vector pET-15b, and the recombinant expression vector pET15b-NGF was transformed into E.coli BL21(DE3)pLysS. After inducing with IPTG the NGF was higher expressed up to 25% of the total cell proteins. The expression product was purified with metal chelate affinity chromatography on Ni-IDA agarose under denaturing condition. The purity of rNGF was higher than 90% and yield of rNGF was 4.56mg/L expressing bacteria. SDS-PAGE revealed the NGF expression product had a Mr 16kDa. Western-blot displayed the recombinant product had strong immunological activity with rabbit anti-human ?-NGF polyclonic antibodies. The expression products were dealed with solubilizing inclusion bodies and refolding protein. The test of nerve fiber growth of chicken embryo DRG neurons displayed rNGF had biological activity.

Objective: To establish a blood-brain barrier (BBB) model for screening the effective ingredients of total salvianolic acids components probably effecting central nervous system (CNS), so as to provide basis for the rapid high throughput screening of CNS drugs. Methods: The permeability parameters and transendothelial electrical resistance (TEER) values showed that the co-cultured human brain microvascular endothelial cells (HBMEC)/HA (human astrocytes) model was of integrity and vitality. The active components in total salvianolic acids were screened with this in vitro BBB model, and then analyzed by HPLC-MS. Results: HA induced HBMEC to produce higher TEER in the first 10 d up to 300 Ω/cm2, sodium fluorescein was used to detect the formation of tight junctions of endothelial cells, and the permeability coefficient of its cross-vitro BBB model was (2.659 ± 0.730) × 10-3 cm/min. We found that six compounds at least from total salvianolic acids could permeate across BBB model, in which danshensu, protocatechuic acid, caffeic acid, isoferulic acid, lithospermic acid, and rosmarinicacid were identified. Conclusion: The co-cultured HBMEC/HA model combined with HPLC-MS analysis is a potential tool for screening of BBB permeable drug candidates from plant-derived materials.

OBJECTIVEThis study aimed to explore the effects of hyperoxia on the development of fetal lung by investigating the changes of morphological and cell proliferation induced by hyperoxia in cultured fetal lungs as well as the effects of dexamethasone on hyperoxia-exposed lungs.

METHODSHuman fetal lung explants at the pseudoglandular stage of development were cultured randomly either in normoxia (21% O2/5% CO2) or hyperoxia (95% O2/5% CO2) for 72 hrs. Dexamethasone was added into the feeding medium at the concentration of 10(-6)M. Harvested tissues were stained for pancytokeratin to identify epithelial cells, with Ki-67 as a marker of proliferation. The effects of lung morphometry were analyzed using computer assisted image analysis. The mean airway thickness, the proportion of the surface area occupied by airways, the mean airway surface area and the index of the epithelium proliferation were measured.

RESULTSThe lung architectures remained unchanged after 72 hrs normoxia culture, whereas hyperoxia culture resulted in significant dilation of airways and thinning of epithelium, with the surface area of airways of 6662 microm(2) vs 2728 microm(2) and the thickness of airways of 7.8 microm vs 8.1 microm (P < 0.05). Hyperoxia culture also resulted in an increase in the proportion of the surface area occupied by airways than normoxia culture (35.2% vs 23.4%; P < 0.05). The surface area of airways (3174 microm(2)) and the proportion of the surface area occupied by airways (23.9%) decreased significantly in hyperoxia-cultured lungs after dexamethasone administration (P < 0.05). The epithelium proliferation index in hyperoxia-cultured lungs (21.8%) was higher than that in normoxia-cultured lungs (5.1%) and dexamethasone-treated hyperoxia-cultured lungs (7.4%) (P < 0.05).

CONCLUSIONSThe exposure of pseudoglandular lungs to hyperoxia modulates the lung architecture to resemble saccular lungs with higher epithelium proliferation index. Dexamethasone may inhibit the effects induced by hyperoxia.

Cell Differentiation ; drug effects ; Dexamethasone ; pharmacology ; Female ; Humans ; Hyperoxia ; pathology ; Lung ; drug effects ; embryology ; pathology ; Pregnancy

OBJECTIVETo study the clinical feature and more reasonable diagnostic typing criteria for patients with liver failure.

METHODS13/21 cases of ALF, SALF with no past liver disease, 49/72 cases of with chronic hepatitis, and 23/73 cases ALF, SALF with liver cirrhosis, were analyzed respectively.

RESULTS1 ALF patients (1). There exist significant statistic differences in ALB, ALT, CHE in three ALF groups.(2). It had statistic differences in those patients with hepatic encephalopathy.(3). The prognosis of the patients with chronic hepatitis group (42.85 percent) was best than that of chronic cirrhosis (26.09 percent) and no past liver disease (15.38 percent). (2) In SALF patients (1). There exist significant statistic differences in ALB, GLO, ALT, AST, BDIL, GLU and CHE in three SALF groups.(2). It had statistic differences in those patients with hepatic encephalopathy in three SALF groups.(3). The prognosis of the patients with chronic hepatitis group (51.39 percent) was best than that of chronic cirrhosis (36.85 percent) and no past liver disease (33.33 percent).

CONCLUSIONThere are different clinic feature and prognosis in three ALF or SALF groups, so we suggest that it were clinic practicability and science in classify of liver failure at present.

Humans ; Liver Failure ; classification ; Liver Failure, Acute ; classification ; Prognosis

OBJECTIVETo study the betulonic acid on the cell cycle and related protein expressions on mice of bearing H22 tumor cells.

METHODFlow cytometray was used to observe the changes of betulonic acid on the cell cycle and P53 of H22 tumor cells. Immunohistochemistry was determined the espressions of PI3K and AKT.

RESULTIncreasing the doses of betulonic acid, the number of H22 cells in S phase and G2 phase was increasing gradually, it can speculate that when the betulonic acid act on cells, the cells were blocked in S and G2 phase and inhibited the protein expressions of PI3K and AKT.

CONCLUSIONBetulonic acid may be up-regulate the activity of P53 and inhibite the expressions of PI3K and AKT, so that it inhibited the survival pathway of tumor cells.

Animals ; Betula ; chemistry ; Cell Cycle ; drug effects ; Cell Line, Tumor ; Cell Survival ; drug effects ; Drugs, Chinese Herbal ; chemistry ; pharmacology ; Female ; Flow Cytometry ; G2 Phase ; drug effects ; Gene Expression Regulation ; drug effects ; Immunohistochemistry ; Male ; Mice ; Neoplasm Transplantation ; Phosphatidylinositol 3-Kinases ; metabolism ; Proto-Oncogene Proteins c-akt ; metabolism ; Tumor Suppressor Protein p53 ; metabolism

OBJECTIVETo detect the expression of MAP3K5 and miR-BART22 encoded by Epstein-Barr virus and explore their relationship in nasopharyngeal carcinomas (NPCs).

METHODSFifty-three archived specimens of NPCs and 30 nasopharyngitis specimens were collected for detecting the expression of EBERs and miR-BART22 by in situ hybridization, and the expression of MAP3K5 was detected using immunohistochemistry. Ten fresh NPC and 10 fresh nasopharyngitis specimens were also obtained for determining the protein expression of MAP3K5 by Western blotting.

RESULTSEBERs were positive in all the 53 NPC specimens, and miR-BART22 was positive in 49 specimens; all the 30 nasopharyngitis specimens were negative for EBER or miR-BART22. In the 53 NPC tissues, 50 were negative for MAP3K5 expression in the cancer areas but positive in the adjacent mucosal areas, with the other 3 specimens showing a weak positivity (+). In the 30 nasopharyngitis specimens, 25 showed strong MAP3K5 positivity, 3 showed weak positivity and 2 were negative for MAP3K5 (P<0.001). Western blotting showed that the expression of MAP3K5 protein was significantly higher in nasopharyngitis than in NPC tissues (P=0.029). The expression of MAP3K5 and miR-BART22 was inversely correlated (P<0.001).

CONCLUSIONCompared with the adjacent mucosal tissues, NPC tissues have a lower expression of MAP3K5 but a higher expression of miR-BART22. The expression of MAP3K5 and miR-BART22 is inversely correlated, suggesting the possibility of MAP3K5 to serve as target gene of EBV miR-BART22. miR-BART22 may inhibit the expression of MAP3K5, thus reducing the protein phosphorylation of MAPK pathway downstream genes, inhibiting NPC cell apoptosis, preventing their differentiation and promoting their escape from immune surveillance.

Adult ; Aged ; Female ; Gene Expression Regulation, Viral ; Herpesvirus 4, Human ; genetics ; Humans ; MAP Kinase Kinase Kinase 5 ; genetics ; metabolism ; Male ; MicroRNAs ; genetics ; Middle Aged ; Nasopharyngeal Neoplasms ; metabolism ; virology ; Tumor Escape ; Viral Matrix Proteins ; metabolism ; Young Adult

BACKGROUNDAutism spectrum disorders (ASD), which include autism, asperger syndrome (AS) and pervasive developmental disorder-not otherwise specified (PDD-NOS), are devastating neurodevelopmental disorders of childhood resulting in deficits in social interaction, repetitive patterns of behaviors, and restricted interests and activities. Single photon emission computed tomography (SPECT) is a common technique used to measure regional cerebral blood flow (rCBF). Several studies have measured rCBF in children with ASD using SPECT, however, findings are discordant. In addition, the majority of subjects used in these studies were autistic. In this study, we aimed to investigate changes in rCBF in children with ASD using SPECT.

METHODSA Technetium-99m-ethyl cysteinate dimmer (⁹⁹m)Tc-ECD) brain SPECT study was performed on an ASD group consisting of 23 children (3 girls and 20 boys; mean age (7.2 ± 3.0) years) who were diagnosed according to Diagnostic and Statistical Manual of Mental Disorders, 4th edition (DSM-IV) criteria and an age-matched control group with 8 children (1 girl and 7 boys, mean age (5.5 ± 2.4) years). Image data were evaluated with Statistical Parametric Mapping, 5th version (SPM5). A Student's t test for unpaired data was used to compare rCBF and asymmetry in the autism and corresponding control group. The covariance analysis, taking age as covariance, was performed between the ASD and control group.

RESULTSThere was a significant reduction in rCBF in the bilateral frontal lobe (frontal poles, arcula frontal gyrus) and the bilateral basal ganglia in the autism group, and a reduction in the bilateral frontal, temporal, parietal, legumina nucleus and cerebellum in the AS group compared to the control. In addition, asymmetry of hemispheric hypoperfusion in the ASD group was observed. Inner-group comparison analysis revealed that rCBF decreased significantly in the bilateral frontal lobe (42.7%), basal nucleus (24.9%) and temporal lobe (22.8%) in the autism group, and in the bilateral cerebellum (22.8%), basal nucleus (19.3%) and right thalamencephalon (16.6%) in the AS group (P < 0.05).

CONCLUSIONSThe decrease in rCBF in ASD is a global event, which involves the bilateral frontal, temporal, limbic system and basal ganglias. Asymmetry of hemispheric hypoperfusion was more obvious in the AS group than the autism group, which indicates a different neurobiological mechanism from that of autism.

Cerebrovascular Circulation ; physiology ; Child ; Child Development Disorders, Pervasive ; pathology ; physiopathology ; Child, Preschool ; Cysteine ; analogs & derivatives ; Female ; Humans ; Male ; Organotechnetium Compounds ; Regional Blood Flow ; physiology ; Tomography, Emission-Computed, Single-Photon ; methods

OBJECTIVETo evaluate the effect of bronchoalveolar lavage (BAL) through bronchoscopy in diagnosis and treatment of accidental inhalation of liquid and powdery foreign bodies in children.

METHODTwenty-one cases of accidental inhalation were classified into the following 4 groups according to the materials inhaled: 3 cases inhaled the lipidic material; 2 cases inhaled chemical material; 15 cases inhaled hydrosoluble materials; 1 case inhaled powdery material. Eighteen cases were hospitalized within 3 days after the accidents, which included 16 cases treated within 1 day in hospital. And 3 cases were hospitalized 3 days after the accident. After hospitalization, all patients were immediately treated with direct aspiration and lavage clearance method through bronchoscopy. The inhaled foreign materials were cleared as soon as possible, which reduced the absorption, stimulation and infection of respiratory mucosa. The therapeutic effect was evaluated by endoscopic findings, clinical manifestation, laboratory examination and chest imaging. At the same time, decision was made on the second BAL and the prognosis was estimated.

RESULTTwenty-one cases had dyspnea of certain degree. The bronchial mucous membrane of all the 21 cases showed hyperemia, edema and exudate attached. Lavage fluid looked like broth and contained much of the inhaled material. After two to five times treatment, 18 cases were cured (86%), but 3 cases died (14%). One of these 3 cases fell into an oil well, 1 into mosquito repellent solution, another into manure pit. The patients who could not survive had severe airway obstruction, which was difficult to clean and made the problem deteriorated progressively. One of the patients was sent to a hospital 3 days after the accident and another case developed fungal infection, pneumothorax and finally respiratory failure.

CONCLUSIONBAL through bronchoscopy is necessary for children who accidentally inhaled liquid and powdery materials. This treatment is safe and effective in diagnosis and treatment of such problems in children. The effect of treatment is influenced by types of inhaled material, the time of lavage clearance through bronchoscopy, and certain complications, such as infection etc.

Bronchoalveolar Lavage ; Bronchoscopy ; Child ; Child, Preschool ; Female ; Foreign Bodies ; therapy ; Humans ; Infant ; Inhalation Exposure ; Male ; Powders

OBJECTIVETo identify independent risk factors for myocardial infarction (MI) in Chinese men and to develop a model to predict risk profile of an individual suffering MI.

METHODSStudy sample included 5 137 men aged 45.2 +/- 7.8 years who came from a cohort in Beijing Capital Steel and Iron Company, based on the three surveys on coronary heart disease conducted in 1974, 1979 and 1980, respectively. Demographic data and other risk factors, such as life style, medical history, blood pressure, total serum cholesterol level (TC), etc. were collected according to the same protocol in 1980. All the participants were followed up for MI in an average period of 20.84 years until 2001.

RESULTS(1) There were 122 cases with MI identified during the period of follow-up, with an incidence of MI 117.4 per 100 000 person-years. Age of more than 50, smoking, higher systolic and diastolic blood pressure (SBP and DBP) levels, higher TC all were identified as important risk factors of MI. (2) Incidence of MI increased with TC. An increment of 0.52 mmol/L of TC significantly increased relative risk of MI by approximately 40% after adjusted for age, blood pressure and smoking. (3) An increment of 20 mm Hg in SBP or 10 mm Hg in DBP associated with a 40% increase in incidence of MI, adjusting for age, TC and smoking. (4) Smoking was the most risky factors for MI. Smokers had 2.3 times risk of MI, after as compared to non-smokers (or its incidence increased by 137%), after adjusting for blood pressure, TC and age, etc. (5) Incidence of MI increased by 20% with increment of five-year of age in those aged over 50 (P < 0.05), after adjusting for blood pressure, TC and smoking. And, (6) finally, based on multivariate logistic and Cox regression analyses, a model containing several risk factors, such as age, blood pressure, TC and smoking, was developed to predict individual's risk for afflicting MI.

CONCLUSIONSResults of this prospective study showed several established risk factors for MI, including age, blood pressure, TC and smoking all as independent predictors of MI in Chinese men. It is clear and rational that intervention and modification of those traditional risk factors can lead to a decrease in coronary events in Chinese population.

Adult ; Age Factors ; Blood Pressure ; physiology ; China ; epidemiology ; Cohort Studies ; Follow-Up Studies ; Humans ; Incidence ; Iron ; Male ; Metallurgy ; Middle Aged ; Multivariate Analysis ; Myocardial Infarction ; epidemiology ; etiology ; Proportional Hazards Models ; Prospective Studies ; Risk Factors ; Smoking ; adverse effects ; Steel ; Triglycerides ; blood

To identify Salvia shandongensis and its relatives at molecular level, the psbA-trnH intergenic region of three species including Salvia shandongensis, Salvia miltiorrhiza and S. miltiorrhiza f. alba were amplified and sequenced. Sequences were assembled with CodonCode Aligner. The K2P genetic distances between Salvia shandongensis and its relatives were calculated and UPGMA tree was performed by MEGA5.0. The results indicated that the lengths of psbA-trnH regions of Salvia shandongensis were about 391 bp, while the lengths of psbA-trnH regions of Salvia miltiorrhiza and S. miltiorrhiza f. alba were about 386 bp. The psbA-trnH sequences showed considerable variations between species and thus were revealed as a promising candidate for barcoding of Salvia shandongensis and its relatives. The intra-specific genetic distances of Salvia shandongensis were 0, while the intra-specific genetic distances of Salvia miltiorrhiza and S. miltiorrhiza f. alba were 0.002 and 0.001 respectively. Additionally, the genetic distance of Salvia shandongensis and Salvia miltiorrhiza ranged from 0.034 to 0.04, and the genetic distance of Salvia shandongensis and S. miltiorrhiza f. alba ranged from 0.005 to 0.008, the intra-specific genetic distances of Salvia shandongensis were much smaller than that of Salvia miltiorrhiza and S. miltiorrhiza f. alba; clustering results showed that there were obvious differences between Salvia shandongensis, Salvia miltiorrhiza and S. miltiorrhiza f. alba, which was consistent with morphological characteristics. This study not only firstly provides the scientific basis for establishing the taxonomy position in molecular level and revealing their genetic relationships of S. shandongensis, S. miltiorrhiza and S. miltiorrhiza f. alba; but also provides DNA molecular identification scientific basis for the development of new medicinal plant resources of Salvia shandongensis. Our results suggest that the psbA-trnH intergenic spacer region can be used as a barcoding to identify Salvia shandongensis, Salvia miltiorrhiza and S. miltiorrhiza f. alba.
Base Sequence ; DNA Barcoding, Taxonomic ; DNA, Intergenic ; genetics ; DNA, Plant ; genetics ; Genetic Variation ; Phylogeny ; Plants, Medicinal ; classification ; genetics ; Plastids ; genetics ; Salvia ; classification ; genetics ; Sequence Analysis, DNA ; Species Specificity