Respiratory virus poses a serious threat to human life and health. Airway epithelial cells are the body's first line of defense from a wide variety of foreign pathogens, such as viruses and bacteria. Therefore, successful airway epithelial cell culture can provide a model for investigating the mechanisms underlying respiratory pathogenic diseases following airway virus infection. This respiratory disease model can also be used for the potential development of novel therapeutics. Here we provide a brief review of recent developments on the culture of cells derived from human trachea-bronchial airway epithelium, and the application of this model for studying respiratory virus and disease.
Animals ; Cell Culture Techniques ; Epithelial Cells ; virology ; Humans ; Respiratory Tract Diseases ; virology ; Virus Diseases ; virology ; Virus Physiological Phenomena ; Viruses ; genetics ; isolation & purification

Objective To assess the risk assessment of stroke associated pneumonia (SAP ) in patients with ischemic stroke using A2DS2 score.Methods A total of 1279 patients with ischemic stroke who were admitted and treated in our department from 2009 to 201 1 were retrospectively analyzed and applicated A2DS2 score.A2DS2 score was calculated as follows:Age ≥75 years=1,atrial fibrillation=1, dysphagia=2,male sex=1;stroke severity:NIHSS score 0-4=0,5-15 =3,≥16=5.The patients were divided into three groups according to A2DS2 score:620 in score 0 group,383 in score 1-9 group,276 in score ≥10 group.The three groups were comparatively analyzed.The diagnostic criteria for SAP were as follows:newly emerging lesions or progressively infiltrating lesions in post-stroke chest images combined with more than two of the following clinical symptoms of infection:(1 )fever ≥38 ℃;(2 )newly occurred cough,productive cough or exacerbation of preexisting respiratory tract symptoms with or without chest pain;(3)signs of pulmonary consolidation and/or moist rales;(4)peripheral white blood cell count≥10 ×109 L -1 or≤4 ×109 L-1 with or without nuclear shift to left,while excluding some diseases with similar clinical manifestations to pneumonia, such as tuberculosis, pulmonary tumors, non-infectious interstitial lung disease,pulmonary edema,pulmonary embolism and atelectasis.Analysis of the incidence and mortality of SAP as well as the correlation with ischemic stroke site were performed in the three groups,respectively. Mean ± standard deviation (x ±s)was used to represent measurement data with normal distribution and t test was used.Percentage was used to represent enumeration data and χ2 test was used.Results The incidence of SAP was significantly higher in A2DS2 score ≥10 group compared with those in score 1-9 and score 0 groups (7 1.7%vs..22.7%,7 1.7%vs..3.7%,respectively),whereas the mortality in score ≥10 group was significantly higher than those in score 1-9 and score 0 groups (16.7%vs.4.96%,16.7%vs.0.3%,respectively).The incidences of cerebral infarction in posterior circulation and cross-MCA, ACA distribution areas were significantly higher in SAP group compared with those in non-SAP group (35.1%vs.10.1%,11.4%vs.7.5%,respectively).The incidence of non-fermentative bacteria infection was significantly increased in score ≥10 group.Conclusions A2DS2 score provides a basis for risk stratification of SAP.The prevention of SAP needs to be strengthened in acute ischemic stroke patients having a A2DS2 score ≥10.

Objective To explore the effectiveness of transcranial magnetic stimulation (TMS) combined with hyperbaric oxygen (HBO) in treating cerebral infarction.Methods Totally 120 patients with cerebral infarction were randomly divided into a control group,a treatment group A and a treatment group B,40 cases in each group.The control group was given conventional treatment,the treatment group A was treated by HBO on the basis of conventional therapy,and the treatment group B was treated by HBO combined with TMS,10 d for a period of treatment.Before and after 2 courses of treatment,all patients' neurological defect was evaluated using National Institutes of Health Stroke Scale (NIHSS).Their central motor conduction time (CMCT) and the expression of brain derived neurotrophic factor (BDNF) and nerve growth factor (NGF) were also assessed.Results After treatment,the average NIHSS score and effective rate of the treatment group B was (7.35 ± 1.98) and 92.5％ respectively,significantly better than the other two groups.Moreover,the average BDNF content [(4.96 ± 1.20) ng/ml] and NGF content [(152.36 ± 18.01)pg/ml] of the treatment group B after treatment were significantly better than before treatment,and those of the other two groups after treatment.Conclusion TMS combined with HBO is effective in treating cerebral infarction.This may relate to its strengthening the expression of neurotropic factors.

Pigs are increasingly recognized as "natural" hosts of infection by human respiratory viruses because of their similarities to humans in terms of lung physiology, airway morphology, cell types, and distribution of cell receptors in the respiratory tract. We wished to explore the mechanisms of infection by respiratory viruses and screening of drug that could be used to treat respiratory-system diseases. Hence, we developed a model of well-differentiated porcine airway epithelial cells (PAECs) derived from pig-lung tissue and cultured them with serum-free medium under an air-liquid interface condition in vitro. We identified the PAEC model using scanning electron microscopy, electrophysiology, and immunohistology. To evaluate application of gene therapy of adeno-associated virus (AAV)6 on the PAEC model, we generated recombinant adeno-associated virus 6-green fluorescent protein (rAAV6-GFP) using the three-plasmid transfection method and infected PAECs from the apical surface with rAAV6-GFP. Results demonstrated that the PAEC model comprised a multilayer epithelial structure containing ciliated mucous secretory cells, with basal cells located directly beneath the multilayer. rAAV6-GFP could infect PAECs from the apical surface and efficiently transduce PAECs to mediate the long-term expression of the exogenous gene. Establishment of a model of well-differentiated PAECs in vitro could lay a solid foundation for the study of infection by respiratory pathogens, as well as the screening and gene therapy of agents used to treat diseases of the respiratory system.
Animals ; Cell Differentiation ; Dependovirus ; genetics ; Epithelial Cells ; cytology ; metabolism ; Green Fluorescent Proteins ; genetics ; HEK293 Cells ; Humans ; Lung ; cytology ; Membrane Potentials ; Mucins ; metabolism ; Swine ; Transduction, Genetic ; Tubulin ; metabolism

Objective To establish animal models in order to provide an experimental study basis for both the pathogenesis study and taking effective prevention scheme for doxorubicin extravasation injury. Methods A total of 20 Kunming mice for experiments on doxorubicin extravasation injury were divided in-to four groups, I.e., high dose group(2 g/L), medium dose group(1 g/L), low dose group(0.5 g/L) and the control group (injection with water). Dosages were administered with subcutaneous injection on both sides of mice abdomen. The adverse reaction of body, damage areas of extravasation injury, recovery period were observed and histopathologic slides of animal models on both 4 days and 11 days after experiment were performed and compared. Results No significant adverse body reaction was observed after injection for all groups. The damage areas due to extravasation injury were dosage and concentration dependent. In addi-tion, significant differences in recovery period were observed for mice in different groups, that is, the higher injection concentration and dose led to the longer recovery period. Results from the histopathologic study in-dicated that the putrescence of damage area was developed in high dose group mice, and the ulcer occurred after 4 d of dosage in medium dose group mice, respectively. In contrast, no ulcer was observed in low dose group mice. Conclusions It would be feasible to establish a prevention model for mice on doxorubicin extravasation injury by subcutaneous injection at a dosage of 0.05ml(1 g/L).

Objective To investigate the effect of diabetes on short-term prognosis of transient ischemic attack (TIA) in elderly patients.Methods From January 2006 to June 2010,126 patients with TIA aged over 60 years were selected.Patients were divided into diabetic group and non-diabetic group according to past history,blood glucose and glycosylated hemoglobin levels.The cumulative ischemic stroke incidences were analyzed by Kaplan-Meier survival analysis 30 days and 90 days after the first TIA.The risk factors for short-term stroke after TIA were analyzed by Cox regression analysis.Results Among 126 patients with TIA,31 cases (24.6％) had diabetes.The cumulative ischemic stroke incidences were significantly higher in diabetic group than in non-diabetic group 30 days and 90 days after the first TIA (54.8％ vs.22.1％,61.3％ vs.28.4％,both P＜0.01).Cox regression analysis revealed that diabetes and cerebral arterial stenosis were the risk factors for recurrent stroke within 90 days.Conclusions The short-term stroke incidence is significantly higher in elderly diabetic patients than in non-diabetic patients.Diabetes is the independent risk factor for recurrent stroke after TIA.

Objective To investigate the bactericidal mechanism of electrolyzed oxidizing water (EOW) against Pseudomona aeruginosa (P.aeruginosa).Methods Bactericidal mechanism of EOW against P.aeruginosa was studied through intracellular protein leakage,nucleic acid,and cell membrane calcium ion permeability,2 ％ glutaraldehyde was used as positive control group,and normal saline (NS) was used as negative control group.Results The killing rates of EOW and 2％ glutaraldehyde to P.aeruginosa were both＞99.99％ with 30-second contact time,and 100.00％ with 60-second contact time.After 60-second contact with EOW,NS,and 2％ glutaraldehyde,the protein leakage of P.aeruginosa detected by bicinchoninic acid (BCA) were (96.00 ± 7.42),(94.15 ± 7.49),and (216.97 ± 10.35)μg/mL,respectively,difference was significant(F =613.20,P＜0.01),2％ glutaraldehyde group was higher than EOW group and NS group;protein leakage did not change with the increase of contact time(all P＞0.05).Electrophoretogram of random amplified polymorphic DNA showed high intensity dense band between 500-1000 Kb in EOW group and NS group,while 2％ glutaraldehyde group was without amplified bands.The fluorescence intensity of calcium ion of EOW group and 2％ glutaraldehyde group were both lower than that of NS group.Conclusion Bactericidal mechanism of EOW may be due to the damage of membrane permeability of P.aeruginosa,which causes Ca2+ leakage,but fails to cause protein leakage,the damage to nucleic acid is not obvious,DNA may not be a bactericidal target of EOW.

Objective:To explore the application value of problem-based learning (PBL) controlled by closed-loop feedback in the teaching of respiratory medicine.Methods:In PBL teaching, after students' open inquiry, discussion and PBL self-study, closed-loop feedback was given by organizing PPT report, written summary and mechanism diagram display of medical students. The participation of teachers and students, teaching quality, the degree of students' identification of key knowledge points, the breadth and depth of mastering the characteristics of key symptoms and the satisfaction of PBL teaching work were investigated, and the differences were compared before and after the closed-loop feedback. GraphPad Prism 5.01 was used in the analysis.Results:It was found that closed-loop feedback could improve the self-evaluation of tutor's teaching participation [(7.97±0.91) vs. (8.77±0.64), P < 0.001] and students' evaluation on teaching participation of tutor [(8.09±0.79) vs. (8.74±0.45), P < 0.001]. At the same time, students' evaluation on the teaching quality of tutors was also improved [(88.61±6.53) vs. (92.59±5.44), P < 0.001]. After closed-loop feedback, the students' identification of the required knowledge points in the syllabus was significantly increased [(84.00±21.75) vs. (90.22±16.18), P = 0.017]. The overall satisfaction with PBL teaching was also improved obviously [(8.93±0.67) vs. (9.37±0.64), P < 0.001]. Conclusion:Practice has proved that the closed-loop controlled PBL teaching approach has a good effect on the teaching of respiratory medicine, and it's worth popularizing in clinical teaching.

Objective To build a bio-material artificial chest wall and discuss its feasibility and efficiency in reconstruction of huge bony defects of chest wall in mongrels in comparison with traditional "sandwich" procedure. Methods (1) The procine osteal and membranate tissues were treated with epoxy cross linking method and their surfaces were modified with amino acid solutions at various concen-trations and at different temperatures to obtain an artificial pleura and artificial ribs for construction of the artificial chest wall. (2) The huge bony defects (5 cm×5 cm) were created in chest wall of five Chinese mongrels. (3) Three mongrels in test group was repaired with artificial chest wall, while two mongrels in control group was repaired with traditional "sandwich" complex. A follow-up was carried out to observe reconstruction effect and rejection in both groups at 3,6 and 12 months after implantation. Results There was no death found during the perioperative period and at 12 month follow-up in test group, with abnormal contour of chest wall and good thoracic activity after reconstruction. In the meantime, there found no rejection, collapse in the repaired region or paradoxical respiration. The postoperative X-rays at 3,6,12 and 24 months showed a good integrity of the thorax, with no collapse, deformation or abnormal movement. Meanwhile, the follow-up of control group showed a normal contour but slight collapse, with no paradoxical respiration. The Chest X-ray examination revealed that the bone cement in" sandwich" complex was X ray opaque and showed mild abnormal movement with breathing. The common blood test and immune items showed no abnormal. Conclusions The bio-material artificial chest wall is a safe and effective reconstruction technique for bony defects of thoracic wall in mongrels, with no acute or chronic rejection.

Objective To explore the value of color Doppler ultrasonography(CDU)as preoperative diagnosing and postoperative monitoring in patients with stenosis of subclavian or innominate artery(SIA)before and after percutaneous transluminal angioplasty and stent(PTAS).Methods A total of 45 patients with stenosis of SIA were selected.Their extracranial vertebral artery and subclavian artery were observed with CDU.In 36 patients with one-sided stenosis of SIA.blood flow direction and spectrum of extracranial vertebraI artery were analyzed.Subclavian steal grade confirmed by spectrum change of vertebral artery was compared with the stenosis grade on quantitative angiography(QA).Among 45 patients with stenosis of SIA,PTAS was performed successfully in 36 patients(36 stents implanted).After stents implanted, hemodynamie parameters such as peak systolic velocity(PSV)in stent, follow-up velocity ratio in stem(VR)were measured.Restenosis in stent were observed.Results In 36 patients with one-sided stenosis of SIA,the ultrasonographic steal grade correlated with the QA stenosis grade significantly(r=0.752,P<0.01).Among 36 patients with stent implanted,follow up 8-12 months, 4 patients with restenosis in stent were found.Compared with the patients without restenosis in stent,PSV and VR in stent increased significantly(P<0.01).Conclusions As a fast and non-invasive imaging technique,CDU could allow diagnosis of stenosis of SIA before PTAS,and detection of restenosis after PTAS.