This study is to report the preparation of complexes of Ad5 and anionic liposomes (AL-Ad5), the amplification of adenoviruses with enhanced green fluorescent protein (eGFP) reporter gene performed by HEK 293 cells, the adenoviral vectors purified by cesium chloride gradient centrifugation, and the titer of adenovirus determined by cytopathic effect (CPE) method, hexon capsid immunoassay and quantitative-PCR (Q-PCR), separately. The prescription and experiment conditions were optimized by central composite design (CCD). The complexes of Ad5 and AL-Ad5 were formulated by the calcium-induced phase change method. The morpholopy, particle size and zeta potential were detected by dynamic light scattering (DLS) and transmission electron microscopy (TEM), respectively. Additionally, the bicolourable fluoresce-labeled complexes (F(labeled)-AL-Ad5) were prepared and their intracellular location in MDCK cells was detected by confocal laser scanning microscopy (CLSM). The results indicate that the complexes of AL-Ad5 exhibited a uniform distribution with a particle size of 211 +/- 10 nm and a zeta potential of -41.2 +/- 2.2 mV. The result of CLSM demonstrates that the intracellular location of red fluoresce-labeled adenovirus was consistent with that of green fluoresce-labeled liposomes suggesting that the naked adenovirus was well encapsulated by the anionic liposomes in complexes of AL-Ad5.

OBJECTIVETo investigate the early clinical effects and radiological outcome of dynamic cervical implant (DCI) internal fixation in treating cervical spondylosis, and evaluate its safety and efficiency.

METHODSFrom June 2009 to December 2011, 19 patients with cervical spondylosis correspond to the indication of DCI internal fixation in the study, including 5 cases of cervical spondylotic myelopathy and 14 cases of cervical spondylotic radiculopathy. There were 8 males and 11 females, aged from 35 to 54 years with a mean of 43.2 years. Pathological segments included C3,4 in 1 case, C4,5 in 6, C5,6 in 6, C6,7 in 4, C3,4 and C5,6, C6,7 in 2. All patients were treated with anterior discectomy and decompression and DCI internal fixation, meanwhile, 2 cases of them with anterior cervical corpectomy and fusion plate fixation. Clinical evaluation included Modified Japanese orthopedics association (mJOA), neck disability index (NDI), visual analogue scale (VAS) score and patient satisfaction index (PSI) at pre-operation and final follow-up. Radiographic evaluation included flexion/extension lateral view at operative level and adjacent segment. The adjacent level degeneration was analyzed according to Miyazaki classification on MRI images.

RESULTSAll patients were followed up from 12 to 42 months with an average of 19.8 months. Preoperative mJOA score was 13.6±1.1 and at final follow-up was 16.3±1.2 with improvement rate of 85.0%. Preoperative VAS,NDI was 6.6± 1.4, 17.1±7.4 and at final follow-up was 1.4±0.8, 6.1±3.9, respectively; there was statistical significance in all above-mentioned results between preoperative and final follow-up (P<0.05). Preoperative ROM at operation level was (7.6±1.9)° and final follow-up was (7.8+2.1)°; preoperative ROM at C2-C7 was (38.6±7.2)° and final follow-up was (39.9±6.4)°; there was no statistical significance in all above-mentioned results between preoperative and final follow-up (P>0.05). Preoperative DHI at operation level was (6.3±1.1) mm and final follow-up was (7.1±0.8) mm, there was statistical significance in DHI between preoperative and final follow-up (P<0.05). No heterotopic ossification was found. All patients followed up MRI, degeneration of 3 segments aggravated 1 degree in 38 adjacent segments, without clinical symptom.

CONCLUSIONTreatment of cervical spondylosis with dynamic cervical implant can got satisfactory outcome in early follow-up. Activity of operative segment obtain reservation in some degree. The incidence of adjacent segment degeneration is lower and no adjacent segment disease occur. Nevertheless a longer follow-up time should be needed to assess the long term functionality of the DCI and the influence on adjacent levels.

Adult ; Cervical Vertebrae ; surgery ; Female ; Fracture Fixation, Internal ; methods ; Humans ; Male ; Middle Aged ; Prostheses and Implants ; Range of Motion, Articular ; Spondylosis ; physiopathology ; surgery

Objective To summarize the characteristics and clinical value of multi-slice spiral computed tomography (MSCT) examination in the biliary gallbladder-duodenal fistula.Methods The imaging data of 28 patients with gallbladder-duodenal fistula who were admitted to the Wuxi No.2 Hospital of Nanjing Medical University between June 2012 and March 2015 were retrospectively analyzed.All the 28 patients received MSCT examinations,and the imaging changes were observed and analyzed,including the location of lesions,figures of fistulous tract,shrinking or enlarging gallbladder,pneumotosis and stones of gallbladder or bile duct.Results Of the 28 patients,fistula located at the duodenal bulb were detected in 14 patients,junction of the bulb and the descending part of the duodenum in 2 patients,ascending duodenum in 7 patients,horizontal part in 5 patients.Indirect signs of biliary gallbladder-duodenal fistula included that gallbladder volume in 28 patients was significantly reduced,cross sectional area of gallbladder was 2 cm × 1 cm-6 cm × 2 cm,and gallbladder wall was thickened with an average thickness of 5 mm (range,4-9 mm).Adhesion of gallbladder and duodenum,unclear boundary,structure disorder and visible effusion surrounding gallbladder were detected.Among 21 patients with biliary gas,19 patients had pneumotosis of gallbladder and 17 had biliary pneumatosis.Biliary stones were detected in 23 patients including cholecystolithiasis in 19 patients,gallbladder neck stones in 6 patients,common bile duct stones in 13 patients and intra-and extra-hepatic cholangiolithiasis in 1 patient.The diverticulum signs appeared in the duodenum of 11 patients.The direct signs of MSCT in the biliary gallbladder-duodenal fistula included that fistulous tract of 13 patients clearly showed and some were dumbbell-shaped.Two and 2 patients were complicated with gallstone ileus and multiple liver abscesses,respectively.The diagnostic results of MSCT in 28 patients were compared with the results of operative exploration,with an diagnostic concordance rate of 78.6％ (22/28),and the diagnostic concordance rate of gallbladder stones was 82.1％ (23/28).Conclusions The indirect signs of MSCT in patients with biliary gallbladder-duodenal fistula include pneumotosis of gallbladder or/ and biliary gas,gallbladder neck stones or common bile duct stones,gallbladder shrank,adhesion of gallbladder and duodenum,unclear boundary,diverticulum signs in the adhesions of duodenum and gallbladder,and clear orificium fistulae of gallbladder-duodenum is a direct sign of MSCT.

Aim To investigate the effect and mecha-nism of valproic acid on neuroglobin expression, and the neuroprotective role of valproic acid against H2 O2-induced neurotoxicity. Methods Western blot, RT-PCR and luciferase assay were used to detect the pro-tein levels, mRNA levels and promoter activity of mouse and human neuroglobin induced by valproic acid. Luciferase assay was used to investigate the role of transcription factor CREB in the up-regulation of neuroglobin by valproic acid. MTT assay was used to evaluate the effect of valproic acid against H2 O2-in-duced neurotoxicity. Results VPA treatment marked-ly increased the protein levels, mRNA levels and pro-moter activity of Ngb in mouse N2 a cells and human SKNSH cells. CREB specific inhibitor KG501 or CREB dominant negative mutant KCREB attenuated VPA-induced Ngb promoter activity. VPA could pro-tect N2a cells from H2 O2-induced neurotoxicity. Con-clusion CREB mediates VPA-induced Ngb up-regula-tion, which may contribute to the neuroprotective effects of VPA in oxidative stress in neurons.

Objective To study the radiologic features and the diagnostic value of multi-slice spiral CT (Multi-slice CT, MSCT) in cholecysto-duodenal fistula.Methods A retrospective analysis was conducted on 33 patients with cholecysto-duodenum fistula.Plain and IV enhanced MSCT were carried out on these patients.Results Of the 33 patients, the fistula was located at the duodenal bulb in 15 patients (45.5％) , the junction of the bulb and the descending part of the duodenum in 3 patients (9.1％) , the horizontal part in 5 patients (15.1％) and the ascending of the duodenum in 10 patients (30.3％).The CT signs of cholecysto-duodenum fistula included in 16 cases.The fistulae were clearly displayed including some fistulae being dumbbell-shaped.The indirect signs of cholecysto-duodenum fistula included in 2 cases the gallbladders were unclearly shown.In 1 case the gallbladder volume increased because of cancer and in another case because of acute cholecystitis.In 29 cases, the gallbladder volume was significantly reduced,with an average volume which ranged from 6 cm × 2 cm to 2 cm × 1 cm, and an average gallbladder wall thickening of 5 cm.There were extensive adhesions between the gallbladder and duodenum with visible effusion.In 26 cases, gas was present in the biliary system with 22 cases showing gallbladder gas, and 19 cases showing biliary pneumatosis.Biliary stones were present in 26 patients (gallbladder stones in 22 cases, gallbladder neck stones in 6 cases, common bile duct stones in 13 cases).At the site between the duodenum and the gallbladder there were radiological changes simulating a diverticulum.In 11 cases the changes were like a duodenal diverticula.The complications of cholecysto-duodenal fistula included 5 cases of gallstone ileus and 2 cases of multiple liver abscesses.Conclusions MSCT is important in depicting presence and location of cholecysto-duodenal fistula.The morphology and shape of the gallbladder, the presence of gas in the biliary system, the presence of stones and the surrounding adhesions could be fully demonstrated by MSCT.MSCT are important for diagnosing cholecysto-duodenal fistula and in the planning of surgery.

A hybridoma cell line 1G4A7 secreting monoclonal antibody (McAb) specific to hemagglutinin of avian paramyxovirus type 2 (APMV-2) was developed by fusing the spleen cells of APMV-2 immunized BAlb/c mice with SP2/0 myeloma cells. The immunoglobulin subclass of 1G4A7 was IgG1 with light chain kappa and the affinity constant against APMV-2 was 1.02 X 10(10). Identified by HI and indirect ELISA, the McAb titers in ascities were 10 log 2 and 1 : 10(6) respectively. The McAb did not cross react with the common avian viruses, showing good specificity. There existed obvious differences in antigenitic relationship among APMV-2 viruses analyzed by HI and indirect ELISA using McAb 1G4A7.
Animals ; Antibodies, Monoclonal ; biosynthesis ; immunology ; Avulavirus ; immunology ; Enzyme-Linked Immunosorbent Assay ; Hemagglutination Inhibition Tests ; Hemagglutinins, Viral ; immunology ; Mice ; Mice, Inbred BALB C

OBJECTIVETo investigate the hemodynamic changes of primary hepatocellular carcinoma (HCC) evolved from hepatic cirrhosis using CT perfusion imaging.

METHODSThirty-two patients with primary hepatocellular carcinoma evolved from virus-induced fibrosis or cirrhosis underwent dynamic CT scanning of the target slices for 60 min. The perfusion parameters of the hepatic parenchyma and HCC including the blood flow (BF), blood volume (BV), mean transit time (MTT), permeability-surface area product (PS), hepatic arterial fraction (HAF), IRF time of arrival (IRF TO) were obtained. Paired-sample t test was used to determine the differences in the perfusion parameters between the hepatic parenchyma and the primary HCC mass.

RESULTSCompared with hepatic BF (117.13-/+31.05 ml/100 mg/min), BV (14.73-/+3.91 ml/100 mg), PS (31.93-/+5.91 ml/100 mg/min), HAF (25.02-/+8.19%), MTT (12.79-/+3.31 s), IRF TO (3.14-/+1.09 s), the primary HCC mass showed significant increments in the BF (239.69-/+96.07 ml/100 mg/min), BV (20.26-/+6.73 ml/100 mg), PS (37.50-/+9.50 ml/100 mg/min), HAF (68.97-/+15.22%) with decreased MTT (7.17-/+1.38 s) and IRF TO (2.42-/+0.94 s). Significant differences were found in all the perfusion parameters between the hepatic parenchyma and HCC (P<0.05).

CONCLUSIONLiver perfusion parameters can represent the hemodynamic changes in the HCC derived from hepatic cirrhosis.

Adult ; Aged ; Carcinoma, Hepatocellular ; diagnostic imaging ; etiology ; physiopathology ; Female ; Hemodynamics ; Hepatitis ; complications ; Humans ; Image Interpretation, Computer-Assisted ; methods ; Liver Cirrhosis ; complications ; Liver Neoplasms ; diagnostic imaging ; etiology ; physiopathology ; Male ; Middle Aged ; Perfusion ; methods ; Radiographic Image Enhancement ; instrumentation ; methods ; Tomography, X-Ray Computed

This study is to report the preparation of complexes of Ad5 and anionic liposomes (AL-Ad5), the amplification of adenoviruses with enhanced green fluorescent protein (eGFP) reporter gene performed by HEK 293 cells, the adenoviral vectors purified by cesium chloride gradient centrifugation, and the titer of adenovirus determined by cytopathic effect (CPE) method, hexon capsid immunoassay and quantitative-PCR (Q-PCR), separately. The prescription and experiment conditions were optimized by central composite design (CCD). The complexes of Ad5 and AL-Ad5 were formulated by the calcium-induced phase change method. The morpholopy, particle size and zeta potential were detected by dynamic light scattering (DLS) and transmission electron microscopy (TEM), respectively. Additionally, the bicolourable fluoresce-labeled complexes (F(labeled)-AL-Ad5) were prepared and their intracellular location in MDCK cells was detected by confocal laser scanning microscopy (CLSM). The results indicate that the complexes of AL-Ad5 exhibited a uniform distribution with a particle size of 211 +/- 10 nm and a zeta potential of -41.2 +/- 2.2 mV. The result of CLSM demonstrates that the intracellular location of red fluoresce-labeled adenovirus was consistent with that of green fluoresce-labeled liposomes suggesting that the naked adenovirus was well encapsulated by the anionic liposomes in complexes of AL-Ad5.
Adenoviridae ; genetics ; ultrastructure ; Animals ; Anions ; Cytopathogenic Effect, Viral ; Dogs ; Drug Compounding ; methods ; Genetic Vectors ; Green Fluorescent Proteins ; chemistry ; HEK293 Cells ; Humans ; Liposomes ; chemistry ; pharmacokinetics ; ultrastructure ; Madin Darby Canine Kidney Cells ; Microscopy, Confocal ; Microscopy, Electron, Transmission ; Particle Size ; Polymerase Chain Reaction ; methods ; Recombinant Fusion Proteins ; ultrastructure

OBJECTIVETo study the factors affecting the management and treatment of immigrant workers with tuberculosis, to determine the most effective measures and to provide an academic basis for tuberculosis control and prevention strategies targeting the immigrant population.

METHODSA self-designed questionnaire was administered to 1364 immigrant workers with tuberculosis (study group) and 436 local residents (control group) with tuberculosis.

RESULTSWhether a patient's tuberculosis status was discovered or not was related to the medical facilities initially visited, job mobility, and the individual worker's economic condition. The percentage of cases discovered was relatively low among those who made their first visit to a private clinic (39.0%), and 58.4% (796) of immigrant workers delayed their first consultation mainly due to neglecting symptoms (55.9%, 445), subjective perception of symptoms being not severe enough (19.3%, 154) to visit a physician and the inconvenience of visiting a hospital because of its being far away from home (15.8%, 47). As comparing immigrant workers with local residents, the differences of the delay reason between the two groups were statistically significant (chi(2) = 21.49, P < 0.01). And 39.9% (544) of immigrant workers and 49.1% (214) of local residents had had late confirmation. The differences of the delay reasons between the two groups were insignificant (chi(2) = 7.31, P = 0.293). And 17.6% (240) patients of immigrant workers and 13.1% (57) patients of local residents did not keep to their drug regimens in a timely fashion. The differences of the reasons between the two groups were insignificant (chi(2) = 6.66, P > 0.05). And 77.5% (1057) of immigrant workers and 31.8% (138) of local residents were considered that taking medicine in supervision spot might have impact on their lives and work. The differences of the reasons between the two groups were significant (chi(2) = 9.71, P < 0.05). All 79.2% (1080) of immigrant workers and 63.3% (276) of local residents did not obtain medicine according to prescriptions. The differences of reasons between the two groups were statistically significant (chi(2) = 24.84, P < 0.01). And 51.2% (699) of immigrant workers and 46.1% (201) of the registered population did not follow up with doctors' directions for lab tests. The differences of the reasons between the two groups were insignificant (chi(2) = 3.26, P > 0.05).

CONCLUSIONThe influential factors in tuberculosis management and treatment were complex. Prevention strategies should focus on health education and promotion activities to improve awareness in seeking medical services. Also, developing and standardizing reference mechanisms for patients, and alleviating the economic burden of the workers will be critical to reduce the tuberculosis incidence.

China ; epidemiology ; Factor Analysis, Statistical ; Humans ; Transients and Migrants ; Treatment Outcome ; Tuberculosis, Pulmonary ; epidemiology ; prevention & control ; therapy ; Urban Population

To compare the DAAO expression level in different Pichia pastoris host strains, the gene encoding DAAO from Trigonopsis variabilis was cloned into plasmid pPIC3.5k and then transformed into P. pastoris GS115 and KM71 respectively. The positive transformants PDK13 (MutS) and PD27 (Mut+) were obtained by PCR analysis. Their optimal and different expression conditions were investigated. To compare with PD27, PDK13 was determined to poss a slower consumption of methanol, a longer induction time, a lower oxygen request and apparently higher expression of DAAO. The highest expression levels were reached up to 2700, 2500 IU/L in shaking flask and 10140, 8463.5 IU/L in fermentor respectively. The over-expression of DAAO can meet its large demand for production of 7-ACA, alpha-keto acid and L-amino acid. In addition, the phenylpyruvate and L-phenylalanine were obtained by crude DAAO reacting with DL-phenylalanine at 37 degrees C for 3h.
D-Amino-Acid Oxidase ; genetics ; Fermentation ; Methanol ; metabolism ; Phenylalanine ; metabolism ; Pichia ; genetics ; Polymerase Chain Reaction