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

Workplace

Objective To study the epidemiological and clinical characteristics of patients with severe acute respiratory syndrome(SARA). Methods Clinical data from 682 patients with SARS were retrospectively analyzed. Results The patients ranged from 13 to 76 years old, 387 male and 295 female. A total of 356 patients (52.2) had a history of close contact with diagnosed SARS patients, and 113 (25.1%) had been to hospital ever. The most common symptom was fever (99.3%), followed by cough (44.4%), shortness of breath (12.2% ), diarrhea (8.9%). 596 patients (87.4%) had normal or decreased white blood cell counts. Serum ALT and CPK levels were elevated in 112 patients (16.4%)and 17 patients (2.5%) respectively. Infiltrates on chest radiography were seen in all patients, with 69.8% involved both lungs. Six patients (0.9%) died of SARS. Conclusion SARS is infectious. Hospital is an important place where SARS transmits. Fever, cough and infiltrative changes on chest radiography are mainly symptoms and signs.

Objective To study the influence and dose effect of ultrasound on the contraction of uterine smooth muscle in rats.Methods Estradiol benzoate was injected into rats three days before conducting an in-vitro experiment.Their uteri were resected and irradiated with ultrasound(0.8 MHz,3 W/cm~2,0-40 rain).The contrac- tion frequency and amplitude were recorded using an MS-302 biological experiment system.Results It could be seen that the contraction frequency and amplitude,and general contractile activity were significantly increased during ultrasonic irradiation(P＜0.01).The increased contraction frequency and amplitude lasted for ten minutes,and then the normal contraction pattern resumed.The contraction frequency as well as the percentage change in eontraction fre- quency were highest during the first 15 minutes of ultrasonic irradiation;the contraction amplitude as well as the per- centage change in amplitude were highest during 40 minutes of ultrasonic irradiation.Contraction activity was at its highest for 30 minutes,but the percentage change in activity was highest for 20 minutes.Conclusions Ultrasound can induce uterine smooth muscle contraction in rats.This biological effect is related to the irradiation time.

OBJECTIVETo observe the changes in expression of microRNA-203 and P63 in human epidermal stem cells and KCs, and to investigate their effects and significance in the epidermal proliferation and differentiation.

METHODS(1) Five normal foreskin tissue specimens were collected from 5 patients by circumcision in Department of Urinary Surgery of the First Affiliated Hospital of Nanchang University from March to June in 2013. Then single cell suspension was obtained by separating epidermis with trypsin digestion method. The cells were divided into quick adherent cells and non-quick adherent cells by type IV collagen differential adherent method. The biological characteristics of cells were observed by inverted phase contrast microscope immediately after isolation and on post culture day (PCD) 3. The expression of CD29, keratin 19, keratin 1, and keratin 10 was identified by immunocytochemical staining. The expression of microRNA-203 and mRNA of P63 was determined by real-time fluorescent quantitative RT-PCR. The protein expression of P63 was determined by Western blotting. Data were processed with t test and Pearson correlation analysis.

RESULTS(1) Immediately after isolation, quick adherent cells were small, round, and dispersed uniformly. On PCD 3, the cells adhered firmly, and they grew in clones. Immediately after isolation, non-quick adherent cells appeared in different shapes and sizes, and dispersed unevenly. On PCD 3, the cells adhered precariously and did not show clonal growth. Quick adherent cells showed positive expression of CD29 and keratin 19, while non-quick adherent cells showed positive expression of keratin 1 and keratin 10. Quick adherent cells were identified as epidermal stem cells, and non-quick adherent cells were identified as KCs. (2)The expression level of microRNA-203 in epidermal stem cells (0.74 ± 0.20) was lower than that in KCs (3.66 ± 0.34, t =16.582, P <0.001). The mRNA expression level of P63 in epidermal stem cells (4. 16 ± 0.28) was higher than that in KCs (2.90 ± 0.39, t =5. 850, P =0.001). The protein expression level of P63 in epidermal stem cells (1.42 ± 0.05) was higher than that in KCs (0.73 ± 0.03, t =26.460, P <0. 001). (3) The expression level of microRNA-203 was in significantly negative correlation with the expression levels of mRNA and protein of P63 (with r values respectively - 0. 94 and -0.98 , P values below 0.05).

CONCLUSIONSThe expression levels of microRNA-203 and P63 in human epidermal stem cells and KCs were significantly different, which might be related to the different characteristics of proliferation and differentiation of the cells.

Cell Differentiation ; Cells, Cultured ; Epidermis ; cytology ; growth & development ; Epithelial Cells ; cytology ; metabolism ; Gene Expression Profiling ; Gene Expression Regulation, Developmental ; Humans ; Integrin beta1 ; Keratin-10 ; genetics ; metabolism ; Keratin-19 ; genetics ; metabolism ; Keratinocytes ; Male ; Membrane Proteins ; genetics ; metabolism ; MicroRNAs ; genetics ; metabolism ; Stem Cells ; cytology ; metabolism

Objeetive To investigate the effect of alteration of blood flow in the hepatic artery on the therapeutic effect of cryoablation in VX2 hepatic tumor rabbit model.Methods Thirty rabbits with VX2 hepatic tumor were divided into three groups according to hepatic artery blood flow:complete occlusion of the hepatic artery(group A),paaial occlusion of the hepatic artery(group B),and no occlusion of the hepatic artery(group C).With conventional CT scau and perfusion scan,the values of blood flow(BF)and blood volume(BV)of VX2 tumor were computed and the differences among the three groups were analyzed.After cryoablation,the animals were euthanized and the livers were removed.The hepatic tissue from the cryoablation area and surrounding area underwent both methyl thiazolyl tetrazolium(MTT)diaphorase staining and triphenyl tetrazolium chloride(TTC)staining.The gross pathology and histopathological 3.14)ml/100 g in group C,respectively.There were significant differences among the three groups in the BF and BV(F value was 452.16 and 421.33 in the BF and BV,respectively,P＜0.01);(2)The maximum diameter of cryoablation-induced necrosis was(2.3±0.3)cm in group A,(1.5±0.2)cm in group B,and(0.8±0.1)cm in group C,respectively.The difference was significant among the groups (F value was 315.32,P＜0.01).(3)There were well-defined frozen areas.bordering areas and normal surrounding areas in MTT staining.In group C,positive staining around some blood vessels could be seen.Conclusion Alteration of the blood flow in the hepetatic artery can affect the cryoablation efficacy.With the decrease of hepatic artery blood flow,the efficacy of cryoablation on liver tumor increased.

OBJECTIVETo compare the changes of digestive and absorptive functions in three different models of Pi-deficiency syndrome (PDS).

METHODSExperimental mice were divided into four groups, the control group(CG), the rhubarb group (RG), the exhaustion group (EG) and the over-exertion group(OG). Criteria including general physical signs, D-xylose excretion rate, serum amylase activity, velocity of stomach emptying and enterokinesia, serum gastrin content and indexes of organs were determined before and after treatment.

RESULTSDecrease of D-xylose excretion rate and increase of stomach emptying and enterokinesia velocity appeared in all the three PDS models. As compared with CG, changes of all indices in OG were significant, while the decreasing of spleen index and serum amylase activity in EG, and the changes of serum gastrin content and thymus index in RG were insignificantly different.

CONCLUSIONAll the changes in various criteria showed that PDS mice model established by over-exertion was superior to that established by frequently used methods as purging with rhubarb and exhausting by swimming.

Animals ; Digestion ; physiology ; Disease Models, Animal ; Intestinal Absorption ; physiology ; Medicine, Chinese Traditional ; Mice ; Mice, Inbred ICR ; Splenic Diseases ; Syndrome ; Xylose ; urine ; Yang Deficiency

OBJECTIVETo evaluate and generalize the application of intermaxillary fixation screw in the jaw fracture.

METHODS41 cases of jaw fracture have been treated with intermaxillary fixation screw.

RESULTSBoth function and appearance have recuperated in 40 cases except 1 case has light malocclusion.

CONCLUSIONSApplication of intermaxillary fixation screw advanced the traditional therapy.

Adolescent ; Adult ; Bone Screws ; standards ; Female ; Humans ; Jaw Fractures ; surgery ; Male ; Maxillary Fractures ; surgery ; Middle Aged ; Orthopedic Fixation Devices ; standards ; Reconstructive Surgical Procedures ; instrumentation ; methods ; Treatment Outcome

Objective To investigate the antiproliferation and induction differentiation of human gas- tric carcinoma which human gastric lower-differentiation mucinous carcinoma MGC-803 cells transplanted in- to nude mice by using rosiglitazone(ROS),and to preliminarily explore the mechanism of differentiation. Methods The mice were randomly divided into five groups:model,ATRA,ROS 25 mg?kg~(-1),ROS 50 mg/kg, ROS 100 mg/kg.After that the volumes were measured and inhibition rates were calculated.The cell cycle was detected by FCM.The protein expression level of Mucin SAC was detected by immunohistochemistry. Results The volume of tumor decreased significantly in ROS treatment groups,the differences had statistical significance compared with model group(P0.05).The xenograft tumors of ROS groups demonstrated the characteristics of differentiation.Xenograft tumor cells were arrested in G_0/G_1 phase,and the cells in S phase decreased significantly,and up-regulated Mucin SAC gene expression.Conclusion ROS could inhibit the growth of tumor,and the effect were dose-dependent with ROS.ROS could induce the differentiation of Xenograft tumor cells of gastric cancer.Its mechanism might be related to the inhibit of transition from G_1 to S phase,degrade the activity of proliferation,regulate the expres- sion of Mucin 5AC.

Objective To investigate the expression of hypoxia inducible factor 1α (HIF-1α) and glucose transporter 1 (GLUT1) in lung adenocarcinoma and its correlation with tumor metastasis. Methods SP immunohistochemistry was used to detect GLUT1 and HIF-1α protein expression in 125 lung adenocarcinoma, including 41 cases without metastasis, 38 cases with lymphatic metastasis and 46 cases with brain metastasis. The correlation of GLUT1 and HIF-1α in lung adenocarcinoma metastasis was analyzed by using x 2 test and Pearson correlation analysis. Results Most lung adenocarcinoma were histologically heterogeneous, which contained more than one adenocarcinoma type. 73.2 % (30/41) cases were acinar predominant adenocarcinoma in lung adenocarcinoma without metastasis; 53.6 % (15/38) cases were acinar predominant adenocarcinoma and 26.3 % (10/38) cases were solid predominant adenocarcinoma in lung adenocarcinoma with lymphatic metastasis; 47.8 % (22/46) cases were papillary predominant adenocarcinoma and 34.8 % (16/46) cases were solid predominant adenocarcinoma in lung adenocarcinoma with brain metastases. The expression level of GLUT1 and HIF-1α in lung adenocarcinoma with lymphatic metastasis group was higher than that of the group without tumor metastasis (P< 0.05); the expression of GLUT1 and HIF-1α were positively correlated (r=0.407, P=0.000). Conclusions Papillary adenocarcinoma is the most histological type in lung adenocarcinoma with brain metastasis, suggesting that papillary adenocarcinoma is more prone to brain metastasis. The expression of GLUT1 and HIF-1α play an important role in lymph node metastasis and brain metastasis of lung adenocarcinoma.