This study aimed to construct full-length cDNA clones of the Japanese encephalitis virus (JEV). SA14-14-2 strain and discuss the feasibility of constructing chimeric viruses for exogenous gene expression based on the JEV genetic skeleton. Long-fragment RT-PCR techniques were applied to amplify JEV cD-NAs, and two amplified fragments with corresponding restriction endonuclease sites at both ends were cloned into the pACYC184 vector sequentially. Using standard molecular techniques, the enhanced green fluorescent protein (EGFP) gene was inserted into the 3' non-coding region of JEV as a reporter gene. After in vitro transcription and transfection procedures, wild-type JEV and chimeric JEV that expressed the EGFP as the reporter gene were successfully rescued. The recovered viruses were characterized by RT-PCR, plaque assays, and direct fluorescence microscopy. After six serial passage generations, the stability of the recovered viruses were studied in terms of virus growth characteristics and structural gene expression. The results showed that cDNA clones of rJEV and rJEV-EGFP were successfully constructed and rescued in BHK-21 cells after in vitro transcription and transfection. Each generation of the recovered viruses was stable and the chimeric virus rJEV-EGFP could stably express EGFP. The findings of this study indicate that both rJEV and rJEV-EGFP could be constructed and rescued in BHK-21 cells, and the JEV SA14-14-2 strain could be obtained as a viral vector to express foreign genes.
Cloning, Molecular ; DNA, Complementary ; genetics ; metabolism ; Encephalitis Virus, Japanese ; genetics ; metabolism ; Encephalitis, Japanese ; virology ; Gene Expression ; Genetic Vectors ; genetics ; metabolism ; Green Fluorescent Proteins ; genetics ; metabolism ; Humans ; Vaccines, Attenuated ; genetics ; metabolism ; Viral Vaccines ; genetics ; metabolism

To observe the morphology of corneal endothelial cells after femtosecond laser in situ keratomileusis ( Femto-LASlK) .METHODS:From May to September in 2013, 88 eyes of 45 patients with myopia who accepted Femto-LASlK were enrolled in this study. The morphology of central corneal endothelial cells was measured by a non-contact corneal endothelial cell analyzer before, 1mo and 1a after surgery. The number, density, average size, size standard deviation, size coefficient of variation and hexagonality of the corneal endothelial cells were observed. All the measurements were analyzed by statistical analysis.RESULTS: All patients underwent operation smoothly, and no complication was observed during and after surgery;One month after surgery, the endothelial cell density (ECD) was 2815. 34±297. 07/mm2, which had a 2. 64% decrease compared with preparative ECD (t=4. 60, P= 0. 00), there was no statistical significance between ECD measurements of 1mo and 1a after surgery ( P>0. 05 ); One month after surgery, the size standard deviation ( SSD ) was 118. 47 ± 31. 58μm2 , which increased significantly compared with the preoperative SSD(t=-3. 87, P=0. 03), there was no statistical significance between SSD measurements of 1mo and 1a after surgery ( P>0. 05 );After surgery, the number, hexagonality, average size and size coefficient of variation of the corneal endothelial cells didn't change statistically(P>0. 05).CONCLUSlON: ln the early period after Femto-LASlK, the ECD decreased slightly, however this kind of surgery did not have significant harm to the function of corneal endothelial cells, and the surgery didn't cause the progressive loss of corneal endothelial cells.

AIM: To explore the effects of selenium-enriched Spirulina platensis (Se-SP) on proliferation of hepatocytes in rat hepatectomy. METHODS: Rat hepaectomy model was conducted using male Wistar rats. The rats were randomized into five groups: operation groups with 150 (H), 50 (M) and 15 (L) mg?kg-1?d-1 of Se-SP, placebo-control (P) and sham operation group (F). Activities of glutathione peroxidase (GPx) and thioredoxin reductases (TR) in hepatocytes were determined by chemical colorimetry. The expression index of proliferating cell nuclear antigen (PCNA) in hepatocytes was detected by immunohistochemistry, and the level of [3H]-TDR incorporation in regenerative hepatocytes was analyzed by radio-immunity. RESULTS: Activity of GPx and TR, PCNA expression index as well as [3H]-TDR insertion in hepatocytes (in vitro) were obviously higher (P

AIM: To investigate the effect of Se-containing spirulina phycocyanin (Se-SPC) on liver injury of mice induced by carbon tetrachloride (CCl 4). METHODS: The mouse model was conducted by intragastric feeding with 2% CCl 4 oil for three times, meanwhile Se-SPC, spirulina phycocyanin (SPC) and Na 2SeO 3 were injected (ip) to various groups for 7 days. Then selenium (Se), glutathione peroxidase (GPx), superoxide dismutase (SOD), alanine aminotransferase (ALT), malondiaoldehyde (MDA) and nitric oxide (NO) levels in blood and liver were measured. RESULTS: The level of Se, GPx and SOD activities were obviously higher ( P

Objective To explore the diagnosis value of X-ray breast imaging reporting and data system (BI-RADS) combined with wire-guided localization biopsy for breast microcalcifications in impalpable breast cancer.Methods 192 palpation negative patients with 205 microcalcification lesions were detected by mammography.All lesions were classified according to BI-RADS descriptors for calcification and were categorized by the BI-RADS.The patients with BI-RADS category 4a and above underwent X-ray positioning guide wire-guided biopsy and pathological diagnosis.Results In 205 microcalcification lesions,74 (36.1％) were malignant lesions,131 (63.9 ％) were less than benign lesions.The positive predictive value of malignant breast lesions in clustered,segmental,regional linear branching calcifications were higher [83.3 ％ (5/6),100.0 ％ (11/11),100.0 ％ (1/1)],followed by clustered,linear,segmental,regional pleomorphic calcifications [55.9 ％ (38/68),50.0 ％ (1/2),40.0 ％ (8/20) and 33.3 ％ (4/12),respectively].The positive predictive values of malignant in linear branching calcifications and pleomorphic calcifications were significantly higher than those of coarse heterogeneous calcifications,amorphous or indistinct calcifications (x2 values were 34.44,51.87,16.71,29.86,all P ＜ 0.05).The linear branching calcification had the highest possibility.The proportions of malignant lesions in four different types of glands were extremely dense 40.5 ％ (30/74),heterogeneously dense 39.2 ％ (29/74),scattered areas of fibroglandular density 10.8 ％ (8/74) and fat 9.5 ％ (7/74),respectively.Conclusions BI-RADS categorization for breast microcalcification lesions can improve the detection rate of impalpable breast cancer.Linear branching calcification has higher predictive value for malignant lesions.Dense breast is the risk factors of breast cancer,which should be attached great importance.

Objective To explore the effects of selenium enriched Spirulina platensis(Se-SP) on(antioxidation) and regeneration of rat hepatocytes after partial hepatectomy.Methods Sixty-seven percent(hepatectomy) was performed in rats after high dose(H) and low dose(L) Se-SP was given via gastric tube for 7 days.Then selenium(Se),glutathione peroxidase(GPx),superoxide dismutase(SOD) and(malondiaoldehyde)(MDA) in hepatocytes were determined.The results were compared with normal saline(giving) via gastric tube before hepatectomy(C) and sham operation(S) groups.Hepatocytes were obtained(before) operation and 24h after operation,and the expression index of proliferating cell nuclear antigen((PCNA)) in hepatocytes was detected by immunohistochemistry,and the DNA damage of hepatocytes induced by peroxide was analyzed by comet assay.Results After partial hepatectomy,Se content,GPx and SOD(activity),and PCNA expression index were obviously higher(P

Objective To investigate the effect of combination of ganglioside and hyperbaric oxygen on spastic cerebral palsy. Methods 128 infants with spastic cerebral palsy were divided into treatment group (n=70) and control group (n=58), who received ganglioside and hyperbaric oxygen or cytidine diphosphate choline. The infants in the treatment group were divided into young group (6～12 months, n=38) and old group (1～3 years, n=32) according to their age. They were assessed with Ashworth scale and Functional Independence Measure (FIM) before and 3 courses after treatment. Results The scores of Ashworth scale and FIM improved more in the treatment group than in the control group (P<0.01) after treatment, as well as in the young group than in the old group (P<0.01). Conclusion The combination of ganglioside and hyperbaric oxygen is effective on spasticity and activities of daily living in infants with spastic cerebral palsy. The younger the infants are, the more effective it is.

To evaluate the effect of Hexue mingmu tablets on traumatic hyphema caused by blunt ocular trauma.
●METHODS: Totally 150 patients of traumatic hyphema were divided into seven types by using ultrasound biomicroscopy combining with anterior segment abnormalities, each type was randomly classified as trial group and control group. The trial group was administered Hexue mingmu tablets, control group was treated by hemocoagulase.
●RESULTS: The absorbing time of trial group was shorter than that of the control group. And there was statistical significance between the two groups (P<0. 05).
● CONCLUSlON: Hexue mingmu tablets is an effective medicine to treat traumatic hyphema. Ultrasound biomicroscopy can be used as a routine examination method in traumatic hyphema.

Objective To explore the effect of nitric oxide (NO) on ?-aminobutyric acid B receptor (GABA_BR) subunits during recurrent febrile seizures (FS).Methods Twenty-four Sprague-Dawley rats aged 21 days were randomly divided into 4 groups: control group (37.0 ℃ water,n=8), FS group (45.2 ℃ water,n=8), FS + SNP group (45.2 ℃ water,n=8), FS+L-NMMA group (45.2 ℃ water,n=8). FS rats were induced 10 times in a warm-water bath, once every 2 days. The plasma level of NO was detected by the spectrophotometer. The expressions of GABA_BR subunit mRNA and c-fos gene were examined by in situ hybridization. The expressions of GABA_BR subunit and Fos protein were observed by immunohistochemistry. Results The plasma level of NO increased in FS + SNP group while decreased in FS+L-NMMA group compared with that in FS group. The expressions of GABA_BR_2 were down-regulated in FS+SNP group, while GABA_BR_1 hardly changed compared with those in FS group. In FS+L-NMMA group, both the expression of GABA_BR_2 and GABA_BR_1 up regulated compared with those in FS group. The expressions of c-fos gene and Fos protein were significantly enhanced after recurrent FS. SNP elevated the expressions of c-fos gene and Fos protein, while L-NMMA down regulated the expressions of them.Conclusion NO may play a regulatory role through modulating GABA_BR function in the pathogenesis of recurrent FS.

Objective To explore the influence of ?-aminobutyric acid B receptor(GABA_BR)on carbon monoxide (CO)/heme oxygenase(HO-1)system during recurrent febrile seizures (FS).Methods Sprague-Dawley rats aged 21 days were randomly divi- ded into 4 groups:control group and FS group,FS+baclofen group,FS+phaclofen group.FS in rats were induced 10 times in a bath of warm water, once every 2 days.The plasma level of CO was detected by the dual wave lengh spectrophotometer;the expressions of GABA_BR and HO-1 mRNA were examined by insitu hybridization;the expressions of GABA_BR and HO-1 protein were observed by immunohistochemistry.Results The plasma level of CO increased in FS+baclofen group,while decreased in FS+phaclofen group compared with FS group.The expressions of GABA_BR and HO-1 upregulated in FS+baclofen group,while decreased in FS+phaclofen group compared with FS group.There were significant difference (All P