Objective To determine the infectiousness of the respiratory syncytial virus(RSV)18537 strain of subtype B in different host cell lines and evaluate its pathogenicity and pathological damage in various animal models.Methods The cytopathic features,viral plaque morphology,viral protein expression,and in vitro proliferation efficiency were assessed to determine the basic biological characteristics of such infections.Nasal drops were used to infect 10-month-old BALB/c mice and 6-week-old cotton mice.The viral load in lung tissue after infection was detected,and the pathological injury was analyzed to assess the pathogenicity.Results The RSV 18537 strain of subtype B strain induced polynuclear fusion in Hep-2 cells,and typical viral plaques were formed in BHK-21 cells.In addition,viral proteins could be detected in Hep-2 and A549 cells.In BALB/c and cotton mice infected with nasal drops,viral nucleic acids were detectable in lung tissue on day 5 post-infection.This dose caused mild thickening of alveolar walls with scattered lymphocytes and neutrophil infiltration.Conclusion The RSV 18537 strain of subtype B can be effectively proliferated in Hep-2 and A549 cells while infecting BALB/c mice and cotton mice,resulting in pathological injury to lung tissue.The 18537 strain of RSV subtype B is less contagious than the A2 strain of subtype A both in cells and animals.

Objective: Balb/c mice infected with respiratory syncytial virus(RSV) were used to investigate the metabolic changes in cecal luminal content. Methods: A total of 13 female Balb/c mice were randomly divided into Case group(n=7) and control(Ctrl) group(n=6). Animals in Case group were infected with RSV by using intranasal method, while mice in Ctrl group were treated with DMEM medium. Mice were anesthetized with intraperitoneal administration of 10% chloral hydrate and the cecal luminal contents were harvested under sterile conditions. Metabolite concentrations were measured by UPLC-QTOF/MS system. Univariate and multivariate statistical analysis were used to identify differential metabolites between Case and Ctrl groups. Results: The overall base peak chromatogram between Case and Ctrl groups had a clear disparity, and PCA and OPLS-DA analysis showed obviously discrepancy based overall metabolomic profile. L-serine, 2-ketobutyric acid, Oleic acid and Chenodeoxycholic acid glycine conjugate were enriched in Case group, whereas L-methionine, L-tyrosine and Nicotinic acid were depleted. Pathway analysis showed lysine degradation, Cysteine and methionine metabolism were enriched. Conclusion Lung injury induced by RSV infection may cause the endogenous metabolism disorder of cecal contents.

Objective:To evaluate the effect of intestinal carbapenem-resistant Enterobacteriaceae (CRE) active screening combined with enhanced intervention in the prevention and control of nosocomial infection in patients admitted to the hematological ward.Methods:Patients who were admitted to the Department of Hematology in a tertiary-care general hospital from March 1, 2017 to December 31, 2019 and underwent chemotherapy or immunosuppressive therapy comprised the intervention group. They were screened for intestinal CRE at least thrice. From December 1, 2016 to February 28, 2017, patients who underwent chemotherapy or immunosuppressive therapy without active intestinal CRE screening in the Department of Hematology formed the control group. Both the patient groups were monitored for CRE infection in real time. The χ2 test was used to compare the changes in the CRE infection rate and mortality in high-risk patients before and after the active screening. Results:During the intervention period, the CRE colonization rate of patients was 16.46% (66/401) ; in terms of disease distribution, the colonization rate of acute leukemia was the highest 23.03% (26/113) . Of the 66 colonized patients, 27 (40.9%) patients were identified as positive for CRE at the first screening, 15 (22.7%) were identified at the time of the second screening, and the remaining 24 (36.4%) were identified at the third or subsequent screening; Carbapenem-resistant Klebsiella pneumoniae (CRPK) strains were dominant among the pathogens, accounting for 54.55% (36/66) . During the active screening period, the CRE infection rate (2.49%) and mortality rate (50.00%) of high-risk patients were significantly lower than those of the controls (11.30% and 69.23%, respectively) . The pathogens of 10 CRE infection patients during the intervention period were exactly the same as the previous active screening pathogens, and the coincidence rate was 100%.Conclusion:The CRE colonization rate was the highest in patients with acute leukemia who were admitted in the hematology wards. CRPK is the main pathogen of CRE colonization, infection, and death. Increasing the frequency of screening can significantly raise the positive rate of screening, Active screening can effectively reduce the incidence and subsequent mortality of CRE in high-risk patients admitted in the hematological wards. High coincidence rate between CRE screening positive pathogens and subsequent CRE infection pathogens. Intestinal CRE screening can serve as an indicator of CRE bloodstream infection in patients with hematological diseases as well as provide information for antibiotics therapy.

Objective To explore the clinical significance of T lymphocyte subsets and cytokines in the identifica-tion of Acinetobacter baumannii colonization and infection. Methods A total of 109 patients in hospital who had A. baumannii in sputum were collected. All patients were divided into colonization group (53 cases) and infection group (56 cases) according to the diagnostic criteria of hospital-acquired pneumonia by respiratory society, Chinese medical association. Another 50 healthy cases as control group. T lymphocyte subgroup and cytokines in peripheral blood were measured, the differences of T lymphocyte subgroup or cytokines between the groups were also com-pared. The factors of T lymphocyte subgroup or cytokines were analyzed by Spearman correlation analysis. Logistic regression analysis was used for investigating the predictors for the infection of A. baumannii. Results Between colonization group and infection group, the significant differences were been found in T lymphocytes subgroup, TNF-α,IFN-γ and IL-17. The A. baumannii infection was positively associated with TNF-α ( rs=0.241, P =0.012), IFN-γ (rs=0.235, P=0.014), IL-2(rs=0.249, P=0.009), IL-4(rs=0.268, P=0.005) and IL-17 (rs=0.538, P=0.000), whereas which was negatively associated with CD3 +(rs= -0.193, P=0.045) and CD4 +/CD8 +(rs= -0.302, P=0.001). IFN-γ and IL-17 was the independent factors for discrimination of A. baumannii infection by multivariate Logistic regression analysis. Conclusion Serum IFN-γ, IL-17 can be used as an indicator of Acinetobacter baumannii infection and colonization, and can provide a basis for the rational use of antimicrobial agents and nosocomial infection prevention and control.

Objective To investigate choroid thickness at horizontal meridian with optical coherence tomography (OCT) and compare the choroid thickness difference between first visit myopia children with those children who wear orthokeratology lens for more than 1 year.Methods This retrospective study enrolled 68 myopia children with low to moderate myopia (-1.00--6.00 D) who visited our hospital and took choroid images by OCT.The total subjects were divided into 2 groups.The subjects of 34 children in group 1 visited for myopia initially and wear spectacles.The other one group wear orthokeratology lens more than 1 year.Only the data of right eye were analyzed.Scans through the fovea at horizontal meridian were acquired with OCT.Choroid images were detected by custom software with 500 μm intervals up to 3.5 mm around fovea.Choroid thickness (CT) was calc~ated based on the average of the 7 zones.Statistical analysis was performed to evaluate choroid thickness at each zone.ANOVA was used to compare choroid thickness differences between various zones in each group.Paired t test was used to compare choroid thickness difference at the same zone between two groups.Results The mean age of OK lens group was (12.3 ± 1.8) years old,and the spectacles group was (11.8 ± 1.4) years old,there was no statistical difference.From temple to nasal choroid,the mean CT of the orthokeratology lens group were (296.7 ± 61.8) μm (T3),(290.7 ± 58.9) μm (T2),(285.7 ± 57.4) μm (T1),(278.5 ±57.7) μm (M),(262.2 ±57.9) μm (N1),(239.8 ±59.7) μm (N2),(214.7 ±59.0) μm (N3);And the mean CT of the spectacles group were (294.2 ± 45.4) μm (T3),(292.0±44.0) μm (T2),(283.6 ±45.5) μm (T1),(272.0 ±51.6) μm (M),(255.2 ± 56.3) μm (N1),(236.5 ±58.1)μm (N2),(212.8 ±57.7) μm (N3),respectively.The thicknesses were significantly thicker in temple zones than that in nasal zones in each group (all P ＜ 0.05),but the CT was not significantly different between the two groups in each zone (all P ＞ 0.05).Conclusion The choroid thickness has regional deference in myopia children,the thickest is in the temple and the thinnest in the nasal region.There is no significant difference between the children who initially corrected by spectacles and those who wear OK lens more than 1 year.

Objective To study the antiviral effects of Pudilan xiaoyan oral liquid on Hep-2 cell models infected with respiratory syncytial virus (RSV), adenoviruses serotype 3 strains (ADV3) in vitro. Methods The cell cytotoxic and inhibition effect of Pudilan xiaoyan oral liquid on RSV or ADV3 were investigated by MTT assay and the inhibitory assay for cytopathic effect (CPE) in Hep-2 cell cultures to detect its antiviral effects. Results The median toxic concentration (TC50) of Pudilan xiaoyan oral liquid on Hep-2 cells was 776.97 mg/L. The median effective concentration (EC50) of inhibiting RSV and ADV3 were 28.08, 28.10 mg/L,whose therapeutic index (TI) were 27.67 and 27.65 respectively. The safety coefficient of Pudilan xiaoyan oral liquid is higher than that of control, ribavirin. Compared with the virus control group, Pudilan xiaoyan oral liquid can obviously produce actions of a dose-dependent relationship to inhibit CPE in Hep-2 cells infected with RSV or ADV3 virus. Conclusions Pudilan xiaoyan oral liquid significantly improves the protection against RSV and ADV3 virus infection in Hep-2 cells. And the inhibition of CPE induced by viruses infection increased with the elevation of higher drug concentration for its antiviral effect augmented in vitro.

OBJECTIVEDesign a convenient and stable eye refractometer based on the theory of blur circle.

METHODSAnalyze the retinal blur circle in both Emsly reduced eye model and Liou & Brennan 1997 eye model by ZEMAX. Design the coefficients including PD (pupil diameter) and NO' (length between node point and fovea) with the purpose of improving the accuracy. At last, compare the clinical optometry data from this refractor with the data obtained from optometry hospital in Wenzhou.

RESULTSThe blur circle diameters are nearly the same in both reduced eye model and the Liou & Brennan 1997 eye model. With the PD = 4 mm and NO' = 20 mm, the refractor shows a fine accuracy in optometry. The paired t test shows that the myopia group and the astigmatism axial direction group have no statistical difference between the data from the blur circle refractor and the hospital (P > 0.05), while the astigmatism degree group has the result of P = 0.41 which may be caused by the poor cooperation of pediatric patients. 80% of the astigmatism degree data differ from the data from the hospital in less than 0.75D.

CONCLUSIONThe blur circle refractor, with the features of convenience and fine accuracy, is promised to be a new style of refractometer in the future.

Astigmatism ; Equipment Design ; Fovea Centralis ; Humans ; Models, Statistical ; Myopia ; Ophthalmoscopes ; Ophthalmoscopy ; Refraction, Ocular ; Visual Acuity

Objective: To investigate the effect of cobalt chloride (CoCl2) on transgene expression and viral particle titers in tumor cells infected by conditionally replicating adenovirus expression vector with hypoxia response elements(HRE)-regulated E1AE1B expression (Ad-5HRE-E1AE1B-RFP) and non-HRE regulated replication-deficient adenovirus expression vector (Ad-EGFP, Ad-Luc) in vitro and in vivo. Methods: Ad-5HRE-E1AE1B-RFP had five duplicated HRE and mini CMV acted as a promoter to drive E1AE1B expression and constitutive expression of RFP as reporter. The hypoxia model was optimized by exposing tumor cells to different concentrations of CoCl2. The hypoxia-inducible factor 1 alpha (HIF-1α) protein expression was determined by Western blotting. Under the optimized hypoxia model, the positive expression of exogenous gene and virus replication of Ad-5HRE-E1AE1B-RFP or Ad-EGFP-infected tumor cells were examined by conversed microscopic observation, FACS analysis and plaques formation test. Furthermore, transgene expression induced by combined application of hypoxia-inducible replicative adenovirus and replication deficient adenovirus (Ad-Luc) was also evaluated by examining the lucifererse activity in xenografted tumor models in nude mice by micro PET. Results: Western blotting results showed that CoCl2 at 0.4 and 0.08 μg/mL could stabilize and acumulate HIF-1α protein in gastric cancer SGC7901 cells, which could better mimic hypoxia condition. The microscopic observation and FACS analysis showed that CoCl2 at 0.4 μg/mL could remarkably increase the transduction efficacy of Ad-5HRE-E1AE1B-RFP, which was verified by significant increase in the percentage of positive expression of exogenous gene RFP and fluorescence intensity. But plaques formation test showed that Ad-5HRE-E1AE1B-RFP had no replication. CoCl2 0.4 μg/mL augmented the tranduction efficacy and expression levels of non-HRE regulated replication deficient adenovirus Ad-EGFP and Ad-Luc. Combined intratumoral injection of Ad-5HRE-E1AE1B-RFP and Ad-Luc significantly increased the expression of Ad-Luc in nude mice.Conclusion: CoCl2 markedly enhances transgene expression of recombinant adenovirus. However, the underlying mechanism is not only related to the CoCl2-induced hypoxia, but also probably related to regulation of gene transcription.

BACKGROUND: Genetically engineered cells have been used in the replacement therapy and gene therapy. However, how to select proper donor cells, target cells, and corresponding viral vectors is the most difficult in the therapy.OBJECTIVE: To compare the transduction efficiency of recombinant adenovirus Ad5 and AdSF35, adeno-associated virus rAAV1/2 and rAAV2, and lentivirus LV in bone marrow mesenchymal stem cells (BMSCs) and exogenous gene expression level so as to select the vectors, which can efficiently transduce BMSCs.DESIGN, TIME AND SETTING: Gene engineering controlled observation, performed in the Central Laboratory, Shanghai First People's Hospital between October 2006 and March 2007.MATERIALS: Ten Sprague Dawley rats of clean grade were used to prepare BMSCs. All recombinant viral vectors carded enhanced green fluorescent protein (EGFP) report gene. Ad5 was prepared by the Central Laboratory, Shanghai First People's Hospital. Ad5F35 was gifted by professor Qian Qi-jun from the Second Military Medical University of Chinese PLA. rAAV2 and rAAVI/2 were the products of Benyuan Zhengyang Gene Technique Co.,Ltd. LV was gifted by professor Cuo Li-be from Shanghai Institute of Biochemistry and Cytobiology, Chinese Academy of Sciences.METHODS: Rat BMSCs were in vitro isolated and cultured by density gradient centrifugation. BMSCs of passage 4 were inoculated into 24-well plate at lxlO5/well. One day later, ceils adhered to the wall and allocated to 5 groups. Ad5-EGFP [10, 100,1 000 multiplicity of infection (MOI)], Ad5F35-EGFP (10,10, 1 000 MOI), rAAVI/2-EGFP (1×104,1x10× vg), rAAV2-EGFP(1×104, 1x105vg), and LV-EGFP (30 TU) were respectively added into the 5 groups. BMSCs were transduced for 2 days with Ad virus and separately for 6 days with rAAV and LV virus.MAIN OUTCOME MEASURES: EGFP-positive expression and fluorescence intensity.RESULTS: After twenty-four hours of Ad5-EGFP transduction, EGFP-positive cells were visible under the microscope. With virus dose increasing, EGFP-positive cells increased and fluorescence intensity strengthened. Twelve days later, EGFP-positive cells gradually reduced and fluorescence intensity weakened. For Ad5F35-EGFP, its transduction was basically similar to Ad5-EGFP, but EGFP-positive cell number and fluorescence intensity were increased. After 6 days of rAAV1/2-EGFP or rAAV2-EGFP transduction, EGFP-posidve cell number and fluorescence intensity were decreased. For LV-EGFP transduction, a small amount of EGFP-positive cells could be visible on the second day, and then EGFP-positive ceils and fluorescence intensity were gradually increased until the sixth day.CONCLUSION: Ad5, Ad5F35 and LV could effectively transduce BMSCs cultured in vitro and express exogenous gene.Furthermore, transduction efficiency was correlated with virus dose in dose-dependent manner, rAAVI/2 and rAAV2 had poor/in vitro transduction efficiency.

AIM: After human lung epithelial cells (A549) infected with respiratory syncytial virus (RSV), inducer cells over-express active transmitter. The study investigated the expression changes in nuclear factor ?B (NF-?B), inducible nitric oxide synthase (iNOS) mRNA and protein, nitric oxide (NO) and malonaldehyde (MDA) on human lung epithelial cells (A549) infected with RSV. METHODS: Experiments were performed at the Laboratory of Etiology and Immunology, Anhui Medical University between December 2006 and May 2007. ① Gyopreserved virus broliferated in Hep-2 cells after recovery. A549 cells infected with RSV in vitro were used to collect cells and cellular culture supernatant at hours 4, 8, 16 and 24. Cells non-infected with RSV were served as controls. ② Reverse transcription-polymerase chain reaction (RT-PCR) was used to evaluate the expression of iNOS mRNA. The expression of iNOS protein and NF-?B was detected by Western-Blot. The concentrations of NO and MDA were measured by nitrate reductase method and thiobarbituric acid method. RESULTS: ①The basal expression of NF-?B was increased with the prolongation of RSV infection to A549 cells. ②RSV infection to A549 increased the amounts of mRNA and protein of iNOS in a time-dependent manner. The expression of iNOS mRNA was about 12 times as many as basal expression. RSV infection promoted the expression of iNOS protein, which was higher than the control group. The activities of NF-?B significantly were positively correlated with the mRNA and protein expression of iNOS. ③RSV could improve A549 cells secreting NO and MDA, but the NO levels rose slowly during the 24 hours of infection. CONCLUSION: The inflammatory response of RSV can increase the activity of NF-?B and it is positive correlated with the increase in iNOS mRNA and protein levels.