Objective: To reconstruct adenovirus vector of breast eukaryotic initiation factor 4E and to observe its effect on the metastasis ability of breast cancer cell line MCF-7. Methods: eIF-4E gene was constructed into adenovirus vector pAD-X by gene recombination technique, which was transformed into 293 packaged cell for high titer adenovirus. Real-time PCR was applied to detect eIF-4E gene expression. eIF-4E siRNA was applied and then transwell cabin assay was used to observe changes of invasion and motor ability of MCF-7 cells transfected with reconstruction adenovirus. Result:The finding of digestion was coincided with expected. eIF-4E gene over-expression was detected in transfected MCF-7 cells with real-time PCR. And the invasion and motor abilities of transfected MCF-7 cells were more significantly inhibited in transwell cabin assay (respectively p

Objective To study the value of~1H-magnetic resonance spectroscopy(~1H-MRS) and diffusion weighted imaging(DWI) in evaluating the changes of pathological structures of hippocampal sclerosis.Methods Twelve normal controls and twelve cases with temporal lobe epilepsy were examined by conventional MRI,~1H-MRS and DWI.The NAA/(Cr+Cho) ratios and ADC values in the hippocampus were measured and compared between patients and normal controls.Results The NAA/(Cr+Cho) ratios of hippocampus in patients with hippocampal sclerosis were significant decrease in comparison with controls and the contralateral side,and the ADC values were also significant increase.There was correlation between the NAA/(Cr+Cho) ratios and ADC values in the disordered hippocampus(r=-0.79,P=0.002).Conclusion There is a significant correlation between the NAA/(Cr+Cho) ratios and ADC values in the hippocampal sclerosis.~1H-MRS and DWI capture partially complementary aspects of hippocampal pathology noninvasively in vivo.

Objective To explore the neural mechanism underlying the craving of heroin addicts induced by picture-cue and the correlation between the brain activation degree in nucleus accumbens (NAc)/the ventral striatum and the scores of patients' self-report craving. Methods Twelve active heroin addicts and 12 matched healthy controls underwent fMRI scan while viewing drug-related pictures and neutral pictures presented in a block design paradigm after anatomical scanning in GE 3.0 T scanner. The fMRI data were analyzed with SPM 5. The change of craving scores was tested by Wilcoxon signed rank test. The Pearson correlation between the activation of NAc/the ventral striatum and the heroin craving score was tested by SPSS 13.0. Results The craving scores of heroin addicts ranged from 0 to 3.70(median 0.15) before exposed to drug cue and 0 to 5.10(median 3.25) after viewing drug-related pictures and showed statistical significance(Z = -2.666, P < 0.05). There were 16 activated brain areas when heroin dependent patients exposed to visual drug-related cue vs. neutral visual stimuli. The activation brain regions belonged to two parts, one was limbic system (amygdale, hippocampus, putamen, anterior cingulate cortex and caudate), another was brain cortex (middle frontal cortex, inferior frontal cortex, precentral gyrus, middle temporal cortex, inferior temporal cortex, fusiform gyrus, precuneus and middle occipital gyrus). The MR signal activation magnitude of heroin addicts ranged from 0.19 to 3.50. The result displayed a significant positive correlation between the cue-induced fMRI activation in NAc/the ventral striatum and heroin craving severity (r=0.829, P < 0.05). Conclusion Heroin shared the same neural circuitry in part with other drugs of abuse for cue-induced craving, including brain reward circuitry, visualspatial attention circuit and working memory region. In addition, the dysfunction of NAc/the ventral striatum may attribute to heroin-related cue induced craving.

OBJECTIVETo investigate the safety and efficacy of percutaneous incisional needle biopsy (PINB) in the parapharyngeal region under CT guide for highly suspicious nasopharyngeal carcinoma (NPC) or recurrence of NPC after radiotherapy.

METHODSPINB under CT guide was performed in 32 highly suspicious NPC or recurrence of NPC after radiotherapy through three puncture routes: posterolateral maxillary sinus fatty area, mandibular fossa area, and anterior-mastoid area. Specimens were fixed by 95% alcohol and then underwent pathologic examination.

RESULTSCT guided PINB was successfully performed in every patients with a technical successful rate of 100%. Definitive histopathologic diagnosis was obtained in 30 patients: squamous-cell carcinoma 21, undifferentiated carcinoma 5 and adenocarcinoma 4. The remaining two negative cases were confirmed as fibrosis after radiotherapy. Complications included persistent bleeding of puncture point in one patient and bloody sputum in 3 patients which subsided after symptomatic management. None of these patients was found to have symptoms of nerve injury caused by PINB procedure.

CONCLUSIONThe CT guided percutaneous incisional needle biopsy in parapharyngeal region through the above three puncture routes for highly suspicious nasopharyngeal carcinoma is safe, rapid and effective.

Adult ; Aged ; Biopsy, Needle ; methods ; Female ; Humans ; Male ; Middle Aged ; Nasopharyngeal Neoplasms ; pathology ; Pharynx ; diagnostic imaging ; pathology ; Radiography, Interventional ; Tomography, X-Ray Computed

OBJECTIVETo investigate the antiviral effects of the aqueous extract of Spatholobus suberectus Dunn. (A.E.), a Chinese medicinal herb, against coxsackievirus B3 (CVB3).

METHODSThe antiviral effects of A.E. against CVB3 in vitro (primarily cultured myocardial cells) and in vivo (BALB/c mice) were determined. Serum pharmacological method was also adopted by in vitro experiments. The effects of A.E. inhibiting the CVB3 mRNA expression were compared by RT-PCR in mice in vivo.

RESULTSA.E. exhibited obvious antiviral: effects in vivo, and serum samples obtained from the rats with oral administration of A.E. (10 μg/mL, 5 μg/mL), reduced the virus titers in the infected myocardial cells (3.00±0.70, 3.55±0.52, P<0.01). Meanwhile, the viral myocarditis induced by CVB3 was inhibited significantly by A.E., and the 15-day mortality was reduced to 40% and 45% (P<0.01) in mice treated with A.E. at doses of 50 mg/kg and 100 mg/kg, respectively, while the 30-day mortality was decreased to 45% and 50%, respectively (P<0.01). Moreover, the mRNA expression of Coxsackie virus B3 was significantly inhibited by A.E.

CONCLUSIONAqueous extract of Spatholobus suberectus Dunn. (A.E.) has inhibitory effect on CVB3 both in vitro and in vivo.

Animals ; Antiviral Agents ; pharmacology ; therapeutic use ; Body Weight ; drug effects ; Cercopithecus aethiops ; Coxsackievirus Infections ; blood ; drug therapy ; pathology ; virology ; Enterovirus ; drug effects ; Fabaceae ; chemistry ; Gene Expression Regulation ; drug effects ; Male ; Mice ; Mice, Inbred BALB C ; Myocardium ; pathology ; Organ Size ; drug effects ; Phytotherapy ; Plant Extracts ; pharmacology ; therapeutic use ; RNA, Messenger ; genetics ; metabolism ; Rats ; Rats, Sprague-Dawley ; Reverse Transcriptase Polymerase Chain Reaction ; Survival Analysis ; Vero Cells ; Viral Load

A novel mucoadhesive microcapsule with drug-resin complex core loaded with berberine hydrochloride (BH) was developed and optimized. Drug-ion exchange resin (IER) complex was prepared by static method which stirring IER in drug solution at certain conditions. The influences of different IERs, different temperature, pH values and concentrations of drug solution on the drug loading were investigated. IER complex was coated by emulsion-solvent evaporation method. The coating fluid formulation was optimized using central composite design-response surface methodology, where the ratio between Carbopol 934 and IER (X1), the ratio between Eudragit and IER (X2) and the ratio between Eudragit RL and RS (X3) were taken as independent variables. Time of cumulative release 85% (Y1) and percentage of gastric retention (Y2) were taken as response variables. Drug loading achieved a high level and more drug available in the condition of IER (IRP 88), 37 degrees C, pH 5 and 1.0 mg x mL(-1) drug solution. When X1 = 0.75, X2 = 0.9, X3 = 0.6, the time of cumulative release reached 85% at 300 min, the highest percentage of gastric retention in the range of this experiment were procured.
Acrylates ; chemistry ; Animals ; Berberine ; administration & dosage ; pharmacokinetics ; Capsules ; Delayed-Action Preparations ; Drug Compounding ; methods ; Drug Delivery Systems ; Emulsions ; Gastric Mucosa ; metabolism ; Hydrogen-Ion Concentration ; Ion Exchange Resins ; chemistry ; Male ; Polymers ; chemistry ; Rats ; Rats, Sprague-Dawley ; Temperature

Production of Hepatitis E Virus capsid protein by high cell density culture in recombinant E. coli has been studied in 10L and 30L fermentors. The effects of different factors on growth and producing recombinant protein of E. coli have been studied by batch culture, such as different media, the ratio of phosphate and Magnesium sulfate. Comparison of fermentation performance for recombinant E. coli in different fed-methods culture has been investigated by fed-batch culture. The effects of inducing at different stages of growth and time of inducing on growth and producing recombinant protein, also obtained by fed-batch culture. At last, the solubility of inclusion body in different urea concentrations also has been obtained by fed-batch culture. The results show that the concentration of phosphate and Magnesium sulfate in the optimal media is 80mmol/L and 20mmol/L in batch culture respectively, that induction with 1.0mmol/L IPTG at mid log phase (about 45 OD at 600nm) is suitable for growth and recombinant protein expression, the cells were approaching stationary growth phase and the maximum cell OD at 600nm of 80 was achieved in 5h of fed-batch culture, and the expression level is 29.74%. The results also indicate that the solubility of inclusion body in 4mol/L urea solution induced at 37 degrees C reaches 14mg/mL, over 80% inclusion body was resolved. The culture process achieved in 10L fermentor could be successfully scaled up to 30L fenmentor with good reproducibility.
Bioreactors ; microbiology ; Colony Count, Microbial ; Escherichia coli ; genetics ; metabolism ; Hepatitis E virus ; genetics ; Nucleocapsid Proteins ; biosynthesis ; genetics ; Protein Engineering ; methods ; Recombinant Fusion Proteins ; biosynthesis ; genetics

This study was aimed to investigate the expression of Na(+)/H(+) exchanger 1 (NHE1) in K562 and HL-60 cells undergoing DNA damage induced by etoposide and to elucidate the regulating mechanism. Real-time quantitative PCR (RQ-PCR) and Western blot methods were used to determine the expression of NHE1 in K562 cells after the treating with etoposide. Meanwhile, the flow cytometry was used to detect the apoptosis of leukemic cells. The luciferase reporter vector containing NHE1 promoter was constructed to measure relative luciferase activity after treating with different etoposide concentrations. The results showed that the mRNA and protein of NHE1 increased in accordance with apoptosis ratio in HL-60 cells after treated with etoposide (p < 0.05), but no such obvious increase in K562 cells. Treatment with NHE1 specific inhibitor could block etoposide induced alkalization and reduce the apoptosis ratio of HL-60 cells. The expression pattern and apoptosis alteration was not similar in K562 cells. Relative luciferase activity of reporter vector containing NHE1 promoter however increased in K562 cells after treated with etoposide. It is concluded that the expression of NHE1 is up-regulated in the process of apoptosis of HL-60 cells induced by etoposide and depends on the pHi increasing caused by NHE1 up-regulation which is not found in K562 cells although the transcriptional activity increased.
Apoptosis ; Cation Transport Proteins ; metabolism ; DNA Damage ; Etoposide ; HL-60 Cells ; Humans ; K562 Cells ; Promoter Regions, Genetic ; Sodium-Hydrogen Exchanger 1 ; Sodium-Hydrogen Exchangers ; metabolism

This study was purposed to explore the changes of possible angiogenetic factors other than VEGF after inhibition of NHE1 and their related mechanisms. The K562 cells were treated by NHE1 specific inhibitor cariporide, the angiogenesis factors after inhibition of NHE1 were screened by using protein chip, the IL-8 expression level after cariporide treatment was detected by real-time quantitative PCR; the K562 cells with stable interference of NHE1 were constructed, the IL-8 expression level after interference of NHE1 was detected by real-time quantitative PCR; the p38 phosphorylation level in K562 cells treated with cariporide was detected by Western blot. After treatment of K562 cells with p38 inhibitor SB203580, the IL-8 expression level was decreased by real-time quantitative PCR. The results of protein chip showed that IL-8 expression decreased after cariporide treatment. Real-time quantitative PCR confirmed this inhibitory effect. The p38 phosphorylation level increased after cariporide treatment. The down-regulation of IL-8 expression induced by cariporide treatment was partially restored after K562 cells were treated with p38 inhibitor SB203580. It is concluded that the inhibition of NHE1 can inhibit IL-8 expression through up-regulation of p38 phosphorylation.
Cation Transport Proteins ; antagonists & inhibitors ; Down-Regulation ; Guanidines ; pharmacology ; Humans ; Imidazoles ; pharmacology ; Interleukin-8 ; metabolism ; K562 Cells ; Phosphorylation ; drug effects ; Pyridines ; pharmacology ; Sodium-Hydrogen Exchanger 1 ; Sodium-Hydrogen Exchangers ; antagonists & inhibitors ; Sulfones ; pharmacology ; p38 Mitogen-Activated Protein Kinases ; metabolism

The human UDP-glucuronosyltransferase 1A1 (UGT1A1), one of the most essential conjugative enzymes, is responsible for the metabolism and detoxification of bilirubin and other endogenous substances, as well as many different xenobiotic compounds. Deciphering UGT1A1 relevance to human diseases and characterizing the effects of small molecules on the activities of UGT1A1 requires reliable tools for probing the function of this key enzyme in complex biological matrices. Herein, an easy-to-use assay for highly-selective and sensitive monitoring of UGT1A1 activities in various biological matrices, using liquid chromatography with fluorescence detection (LC-FD), has been developed and validated. The newly developed LC-FD based assay has been confirmed in terms of sensitivity, specificity, precision, quanti-tative linear range and stability. One of its main advantages is lowering the limits of detection and quantification by about 100-fold in comparison to the previous assay that used the same probe substrate, enabling reliable quantification of lower amounts of active enzyme than any other method. The precision test demonstrated that both intra- and inter-day variations for this assay were less than 5.5％. Further-more, the newly developed assay has also been successfully used to screen and characterize the regu-latory effects of small molecules on the expression level of UGT1A1 in living cells. Overall, an easy-to-use LC-FD based assay has been developed for ultra-sensitive UGT1A1 activities measurements in various biological systems, providing an inexpensive and practical approach for exploring the role of UGT1A1 in human diseases, interactions with xenobiotics, and characterization modulatory effects of small mole-cules on this conjugative enzyme.