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Background & objectives: Recently, there has been a surge to develop new devices and techniques for the diagnosis of peripheral pulmonary lesions such as the combination of LungPoint navigation and endobronchial ultrasound with a guide sheath (EBUS-GS). The present study aimed to explore the diagnostic value of LungPoint navigation in combination with EBUS-GS and rapid on-site evaluation (ROSE) particularly for peripheral pulmonary nodules. Methods: Patients (n=108) with pulmonary nodules (10 mm ? nodal diameter ?30 mm) presenting to Henan Provincial People’s Hospital were detected using chest computed tomographic (CT) scanning and bronchoscopy. All patients were evaluated using LungPoint navigation, EBUS-GS and ROSE techniques to evaluate the positive rate of combined diagnosis using the three methods. Results: A total of 108 patients participated in this study and successfully underwent all the three procedures. Of these, 82 patients were accurately diagnosed, making the overall diagnostic rate of 75.9 per cent for combined LungPoint navigation, EBUS-GS, and ROSE analyses. Further subgroup analysis of the diagnostic rate of the three combined techniques were conducted based on the size of the nodules which showed a diagnostic rate of 65.3 per cent for 10 mm ? nodule diameter ?20 mm and 85.7 per cent for 20 mm ? nodal diameter ?30 mm. Of the 108 patients, 85 had solid nodules and 23 had ground-glass nodules; the positive rate of diagnosis of solid nodules was the highest. The patients ultimately were diagnosed with lung cancer with a positive rate of 83.5 per cent. The sensitivity, specificity and positive and negative predicted values for ROSE were 90.3, 78.3, 84.8 and 83.6 per cent, respectively. Interpretation & conclusions: The combined use of the three techniques can effectively shorten the duration of the total diagnosis period and improve the safety of diagnosis without affecting the detection rate.
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Objective:To observe the difference in drying dampness between Scutellariae Radix and Atractylodis Rhizoma in model rats with spleen-stomach dampness-heat syndrome and clarify their property-efficacy relationship. Method:Sixty-four healthy male SD rats were randomized into the blank group， model group， high-， medium-， and low-dose Scutellariae Radix groups， as well as high-， medium-， and low-dose Atractylodis Rhizoma groups. The rats were exposed to high-fat and high-sugar diet and external dampness-heat environment for 20 days for inducing the spleen-stomach dampness-heat syndrome. The macroscopic manifestations of rats were observed and the morphological changes in stomach and colon were detected under a light microscope after hematoxylin-eosin （HE） staining， followed by the calculation of pathological scores. The serum tumor necrosis factor-<italic>α</italic> （TNF-<italic>α</italic>）， interleukin-4 （IL-4）， and interferon-<italic>γ</italic> （IFN-<italic>γ</italic>） levels were determined by enzyme-linked immunosorbent assay （ELISA）. The protein and mRNA expression levels of aquaporin-4 （AQP4） in the gastric tissue were measured by Western blot and Real-time polymerase chain reaction （Real-time PCR）， respectively. Result:Rats in the model group presented with the manifestations of dampness-heat syndrome. The inflammatory reaction in stomach and colon was obvious， and the pathological score was significantly increased （<italic>P</italic><0.01）. The serum IFN-<italic>γ</italic>， IL-4， and TNF-<italic>α</italic> levels were elevated （<italic>P</italic><0.05）， and so were the AQP4 protein and mRNA expression levels in the gastric tissue except that there was no statistical difference. The clinical symptoms of rats in the medication groups were alleviated. Scutellariae Radix significantly relieved the gastric and colonic inflammation in model rats. Atractylodis Rhizoma inhibited the colonic inflammation in model rats to a certain extent， but it had no obvious effect on gastric inflammation. The pathological score of each Scutellariae Radix group was decreased. In terms of the pathological score of gastric tissue， only the high-dose Scutellariae Radix produced a significant difference （<italic>P</italic><0.01）， and the pathological scores of the three Atractylodis Rhizoma groups were not significantly different from that in the model group. As for the pathological score of colonic tissue， all the medication groups except for the low-dose Atractylodis Rhizoma group exhibited a significant difference in comparison with that of the model group （<italic>P</italic><0.01）. Scutellariae Radix and Atractylodis Rhizoma at each dose reduced not only the serum IFN-<italic>γ</italic>， IL-4， and TNF-<italic>α</italic> levels （<italic>P</italic><0.05， <italic>P</italic><0.01）， but also the AQP4 protein expression in gastric tissue of model rats （<italic>P</italic><0.01）. The AQP4 mRNA expression in the gastric tissue of model rats declined in the high- and low-dose Scutellariae Radix groups， while that in the medium-dose Scutellariae Radix group and each Atractylodis Rhizoma group rose without statistical difference. Conclusion:Scutellariae Radix and Atractylodis Rhizoma display a certain property-efficacy relationship in drying dampness of rats with spleen-stomach dampness-heat syndrome. Specifically， the efficacy of drying dampness is related to their cold/heat property， and the resulting outcome of bitter-cold Scutellariae Radix is better than that of bitter-warm Atractylodis Rhizoma.
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A 90-year-old man was diagnosed with primary gastric diffuse large B-cell lymphoma (PGDLBL) by PET/CT examination, gastroscopy, biopsy and histopathological analysis at a regular physical check in April, 2016. The patient received R-CO chemotherapy (rituximab, cyclophosphamide, and vincristine) and radiotherapy subsequently, with enteral nutritional treatment through 3-cavity nasogastric tube due to development of pyloric obstruction. To satisfy patient's strong desire of eating by himself, we performed surgery of exploratory laparotomy and Roux-en-Y gastric bypass (RGB) to relieve pylorus obstruction. Postoperatively, the patient resumed oral feeding, supplemented by nasogastric tube feeding at 1350 - 1550 Kcal daily. He is now 94 years old with fairly well nutrition and normal communication. The outcome of 4 year follow-up suggests that nutritional treatment and palliative medicine are important for improving prognosis and life-quality of very elderly patients with end-stage tumors apart from the effective chemotherapy, radiotherapy, and surgery.
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Objective:To explore the effect of methylphenidate (MPH) sustained-release tablets on the sleep characteristics of children with attention deficit hyperactivity disorder (ADHD).Methods:A total of 23 children with ADHD were included in the MPH group, and treated with sustained variable doses of methylphenidate sustained-release tablets, and overnight polysomnograms (PSG) were performed at baseline, 2 weeks of treatment, and 6 weeks of treatment.At the same time, 21 matched normal children were recruited as control group for PSG.SPSS 22.0 was used for statistical analysis, and the mean ± standard deviation, M( P25, P75) and percentage were used for statistical description according to the type of data.Paired t test or analysis of variance was used for comparison of sleep parameters. Results:At baseline, children in MPH group had lower sleep efficiency (SE (%)) ((86.7±12.4)%, (90.9±7.5)%), longer sleep latency (SOL) ((21.1±12.4) min, (18.1 ±13.0) min), shorter rapid eye movement onset latency (ROL) ((145.2±35.2)min, (155.6±57.7)min), and the differences were statistically significant (all P<0.05). Compared with baseline, children in MPH group after 2 weeks of treatment had prolonged SOL ((27.2±6.4)min, (21.1±12.4) min), shortened ROL((127.2±32.1) min, (145.2±35.2) min) and REM phase time((70.5±13.8) min, (79.8±15.3)min) and significantly lower SE (%) ((83.1±15.4)%, (86.7±12.4)%), and the differences were statistically significant (all P<0.05). There was no significant difference in the SOL, ROL, REM phase time, and SE (%) sleep parameters of children in the MPH group after 6 weeks of treatment and baseline (all P>0.05). The sleep parameters such as %TST, number of awakenings and time of awakening of WASO in N1, N2, N3 and R phase of MPH group were not statistically significant among baseline, 2 weeks of treatment and 6 weeks of treatment (all P> 0.05). Conclusion:Children with ADHD present lower sleep efficiency, more difficulty to fall asleep, and shortened REM latency.MPH has a certain negative effect on the sleep of ADHD children in the first 2 weeks, which worsens the sleep problems of ADHD children.After 6 weeks of continuous treatment, the effect of MPH on sleep gradually improved.
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Objective:To investigate the effects of tranexamic acid on the drainage duration after primary unilateral total knee arthroplasty (TKA).Methods:From June 2017 to December 2018, a total of 182 patients (42 males and 140 females) who underwent primary TKA were included. The age of patients was 68.1±7.1 years (60-76 years). According to the random number table, the subjects were divided into four groups: 45 cases in tranexamic acid group 1 (TXA1), 46 cases in tranexamic acid group 2 (TXA2), 46 cases in tranexamic acid group 3 (TXA3) and others in none tranexamic acid group (NTXA). The patients in TXA1-3 groups received intravenous infusion combined with local application of tranexamic acid. The patients in NTXA group were locally perfused in the joint cavity with 100 ml normal saline after surgery. The drainage volume, total blood loss, invisible blood loss, degree of postoperative joint swelling, Hospital for Special Surgery (HSS) and visual analogue scale (VAS) of pain were compared among the four groups.Results:The postoperative drainage volume of TXA1-3 groups was 85.5±34.3 ml, 189.4±72.3 ml and 215.3±93.4 ml, respectively, which were less than that of the NTXA group (351.3±113.5 ml) with significant difference ( F=11.5, P=0.005). The postoperative drainage volume of TXA1 group was less than that of TXA2-3 groups. The total postoperative blood loss was 699.0±255.7 ml, 710.4±296.1 ml, and 715.8±248.2 ml in the TXA1-3 groups, respectively, which were less than that of the NTXA group (1 130.5±354.2 ml) with significant difference ( F=13.1, P=0.001). At the 4th day after TKA, the knee swelling degree of TXA1-3 groups was 1.25±0.07, 1.13±0.12, and 1.12±0.13, respectively, which were less severe than that of the NTXA group (1.43±0.22) with significant difference ( F=8.23, P=0.015). There were 2 cases with positive bacterial culture in NTXA group and TXA3, 3 cases with hematoma and 1 case with delayed wound healing in the NTXA group. Conclusion:The application of tranexamic acid after TKA can reduce postoperative drainage volume, latent blood loss and total blood loss. The removal of the drainage at 18 h after the operation can not only drain completely, relieve pain and promote knee joint function, but also effectively reduce the risk of infection associated drainage.
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Objective To investigate the effects and regulating mechanisms of matrix metalloproteinase 9 (MMP-9) gene silencing on aquaporin 4(AQP4) expression in astrocytes induced by oxygen-glucose deprivation.Methods Cerebral cortical astrocytes from 2 days newborn SD rats were undergone the primary culture.The ischemic cell model was established by oxygen-glucose deprivation.This experiment were divided into control group,negative control group and MMP-9 gene silencing group.The leakage rate of lactated dehydrogenase (LDH)was detected by chromatoptometry.The MMP-9 gene silencing was carried out by Lentivirus transfection.Real-time fluorescence quantitative PCR and Western blot were used to detect the expression levels of AQP4 and MMP-9.The expressions of PKA,PKC,PKG and CaMK Ⅱ were determined by Western blot.Results Compared with control group,LDH leakage rate was significantly higher in astrocytes induced by oxygen-glucose deprivation(t=13.35,P＜0.01).The expression of MMP-9 mRNA in astrocytes in MMP-9 gene silencing group(0.412±0.297) decreased significantly compared with that in negative control group(1.118 ± 0.240) (t =-4.964,P＜ 0.05).The expression of AQP4 mRNA in astrocytes in MMP-9 gene silencing group(1.002±0.082) decreased significantly compared with that in negative control group(1.442±0.066) (t=-9.886,P＜0.01).The expression of AQP4 protein in astrocytes in MMP-9 gene silencing group(0.643±0.036)decreased significantly compared with that in negative control group(1.000± 0.069)(t=-11.073,P＜0.01).The expression of PKC protein in astrocytes in MMP-9 gene silencing group (0.198±0.110)decreased significantly compared with that in negative control group (0.980± 0.232) (t =-7.218,P＜0.01).The expressions of PKA(t=0.875),PKG(t=0.818) and CaMK Ⅱ (t=0.933) protein had no statistically significant difference between negative control group and MMP-9 gene silencing group(all P＞0.05).Conclusion The permeability of astrocytes is increased by oxygen-glucose deprivation.Gene silencing MMP-9 could induce expression of AQP4 mRNA and protein decreased,and MMP-9 may regulate AQP4 expression by regulating PKC activity.
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Glioma is the most common primary intracranial tumor, among which glioblastoma (GBM) is the most malignant subtype. Because of its high heterogeneity and invasiveness, GBM can't be completely removed by surgical resection and is also resistance to chemotherapy and radiotherapy. Even after a standard therapy, the median survival time is only 14.6 months, the five-year survival rate is less than 10%, and the relapse of GBM is common. Immunotherapy, a new treatment paradigm, treats cancer through regulating the autologous immune system and the tumor microenvironment. As a promising method to improve the prognosis of GBM, immunotherapy has attracted more and more attention. This paper gives a review to the difficulty, the mainly existing strategies and the bottlenecks in GBM immunotherapy, aiming at providing new direction to improve the prognosis of GBM patients.
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Objective To evaluate senile plaque formation and compare the sensitivity of three different β-amyloid (Aβ) labeling methods (antibody staining, Gallyas silver staining, and thioflavin-S staining) to detect Aβ deposition.Methods APPswe/PSEN1dE9 transgenic mice (APP/PS1) of different ages were used to examine spatiotemporal changes in Aβ plaque deposition. Antibody staining, Gallyas silver staining, and thioflavin-S staining were used to detect Aβ plaque deposition in the same brain region of adjacent slices from model mice, and the results were compared.Results With aging, Aβ plaques first appeared in the cortex and then the deposition increased throughout the whole brain. Significantly greater plaque deposition was detected by 6E10 antibody than that analyzed with Gallyas silver staining or thioflavin-S staining (P<0.05). Plaque deposition did not show significant difference between the APP/PS1 mice brains assayed with Gallyas silver staining and ones with thioflavin-S staining (P=0.0033).Conclusions The APP/PS1 mouse model of Alzheimer's disease could mimick the progress of Aβ plaques occurred in patients with Alzheimer's disease. Antibody detection of Aβ deposition may be more sensitive than chemical staining methods.
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Objective To investigate the effects of different abstinent duration on brain functional network of heroin addicts at resting state.Methods Sixteen heroin addicts during protracted abstinence for 11-13 months (PA12 group) and twenty heroin addicts during protracted abstinence for 5-7 months (PA6 group) were recruited in the resting-state functional MRI study.Graph theoretical methods were applied to construct topological organization of whole brain network and nodes betweenness of the networks in all subjects,and the between-group differences were analyzed.The correlation of the node betweenness with the abstinence duration was conducted.Results There was no significant difference (P＞0.05) in the small world characteristic (γ≈1,λ》1) between two groups.Compared with the PA6 group,the PA12 group demonstrated significantly decreased nodal betweenness in regions of left parahippocampal gyrus,left precentral gyrus,and significantly enhanced nodal betweenness in regions of the left cuneus,left temporal and right middle occipital gyrus (all P＜0.05).Moreover,the betweenness of the left precentral gyrus (r =-0.52,P =0.001) and parahippocampal gyrus (r=-0.49,P=0.002) were negatively related with the abstinence duration,the betweenness of the right middle occipital was positively correlated with the abstinence duration (r=0.49,P=0.003).Conclusion The brain network small world topology of heroin addicts tend to be stable after 5-7 months of abstinence.Long-term abstinence may minimize the addicfs memory of drugs and potential drug seeking behavior,and recover addicts' visual spatial attention function.
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Objective To discuss the treatment of periprosthetic femoral fractures (PFF) with unstable prosthesis by replacement of long-stem femoral prosthesis and internal fixation.Methods From December 2005 to December 2014,15 PFF patients with unstable prosthesis (15 hips) following were treated at our department.They were 10 men and 5 women,aged from 64 to 89 years (mean,76.2 years).Their primary surgeries included total hip arthroplasty in 13 cases and biological bi-polar replacement of femoral head in 2.Two prostheses were cement and 13 biological.By Vancouver classification,9 cases were type B2,and 6 type B3.The unstable prostheses in the 15 cases were replaced by long-stem femoral ones,followed by internal fixation.At the last follow-ups,clinical outcome were evaluated by Harris scoring and images of PFF by Beals & Tower criteria.Complications were documented.Results One died 4 months after operation.The other 14 patients were followed up for an average of 4.5 years (from 6 months to 9 years).Fracture union was achieved in 12 cases after an average of 3.9 months (from 3 to 9 months).Nonunion occurred in 2 cases.Imaging evaluation revealed 9 excellent cases,3 good ones and 2 poor ones.The Harris scores at the last follow-up averaged was 82.3 points (from 50 to 100 points).Deep vein thrombosis occurred preoperatively in one case and posterior tibial vein thrombosis occurred in 2 cases respectively on day 3 and day 10 postoperatively.No such complications occurred as malunion,fixation failure,dislocation or prosthesis loosening.Conclusion Satisfactory outcomes can be achieved by replacement of long-stem femoral prosthesis combined with appropriate fixation for treatment of PFF with unstabrosthesis.
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Objective To establish a joint wavelength switching HPLC gradient elution method for simultaneous determination of darutoside, kirenol, darutigenol, fangchinoline and d-tetrandrine in Xixian Fengshi Wan. Methods A Kromasil C18 column (4. 6 mm × 250 mm, 5 μm) was used with a mobile phase A; acetonitrile-methano (2: 1) and mobile phase B: 0. 05% acetic acid solution with gradient elution. The flow rate was 1. 0 mL/min; the injection volume was 20 μL; darutoside, kirenol and darutigenol were detected at 215 nm, and fangchinoline and d-tetrandrine were detected at 280 nm. Results In the given concentration range, the linearity ranges of darutoside, kirenol, darutigenol, fangchinoline and d-tetrandrine were 6. 45-129. 00 μg/mL (r=0. 999 5), 5. 61-112. 20 μg/mL (r=0. 999 9), 4. 25-85. 00 μg/mL (r=0. 999 3), 9. 19-183. 80 μg/mL (r =0.999 8), and 11. 05-221. 00 μg/mL (r=0. 999 7), respectively; and their average recoveries and RSD were 98. 73% (1. 43%), 97. 63% (1. 28%), 99. 44% (1. 29%), 98. 33% (1. 38%), and 97. 36% (1. 37%), respectively. Conclusion The established joint wavelength switching HPLC gradient elution method is simple, accurate and reproducible; it can simultaneously determine the contents of darutoside, kirenol, darutigenol, fangchinoline and d-tetrandrine in Xixian Fengshi Wnn and can be used for quality control of Xixinn Fengshi Wan.
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Objective:To study the cause of the medical disturbances and its influential factors, to formulate the corresponding prevention strategy and measures. Methods:Total 32 medical disturbances in a hospital were an-alyzed and evaluated. Results:The number of medical disturbances in this hospital was declining. The department of maternity, orthopedic, pediatrics had the most medical disturbances. The compensation for the patients with med-ical disturbances was 2. 18 times as the patients without medical disturbances. Conclusion:It should be clear the causes of the medical disturbances and countermeasures should be proposed according to the reasons, rationally re-solve doctor-patient conflicts.
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Objective To evaluate the preoperative value of the sleep respiration monitoring and upper airway obstruction positioning system ApneaGraph 200 (AG 200,JC Medical,America) in disease diagnosis (severity) and the location of upper airway obstruction plane in patients with obstructive sleep apnea hypopnea syndrome (OSAHS). Methods A total of 121 suspected patients with OSAHS were randomly selected. They were examined by Müller's maneuver in the upper airway obstruction site. Then they underwent one-night (7 h) sleeping monitor by polysomnography (PSG, D-22525, Weinmann GmbH, Germany) and AG 200 simultaneously. The data obtained by PSG and AG 200 were compared, including apnea-hypopnea index (AHI), mean arterial blood oxygen saturation (mean SaO2, MSaO2), lowest arterial oxygen saturation (lowest SaO2, LSaO2), percentage of the cumulative time of SaO2≤90% to total monitoring time (CT90%) and diagnosis of the properties and the severity of upper airway obstruction. The constituent ratios of the different obstructive plane measured by AG 200 and the upper airway obstructive sites determined by Müller's maneuver were also compared. Results The AHI, MSaO2, LSaO2, CT90% and the properties and the severity of upper airway obstruction by the AG 200 and PSG were similar. The positive rates of the upper plane obstructive sites determined by the endoscopy with Müller's maneuver and AG 200 were the same. The positive rate of the down plane obstructive sites determined by AG 200 was significantly higher than that determined by the endoscopy with Müller's maneuver (P <0.05). Conclusion One-night PSG and AG 200 monitoring can simultaneously achieve accurate qualitative and primary positioning diagnosis of OSAHS. AG 200 can identify the level of upper airway obstruction more accurately when combined with the endoscopy with Müller's maneuver.
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Objective To evaluate the clinical effect and safety of transcatheter thrombus-breaking combined with local infusion of reteplase (rPA) in treating severe pulmonary thromboembolism (PTE). Methods During the period from June 2010 to June 2013 at authors’ hospital, transcatheter thrombus-breaking combined with local infusion of reteplase was carried out in 26 patients (17 males and 9 females) with severe pulmonary embolism. The changes of pulmonary circulation and the cardio - pulmonary hemodynamics after the treatment were analyzed. Results The preoperative mean pulmonary artery pressure, arterial blood oxygen partial pressure and the blood pressure were (63.78 ± 6.89) mmHg, (73.23 ± 11.51) mmHg and (87.35 ± 10.92) mmHg respectively, while the postoperative ones were (26.23 ± 10.27) mmHg, (93.48 ± 6.17) mmHg and(127.14 ± 13.15) mmHg respectively, and the differences between preoperative and postoperative ones were statistically significant (P<0.01). All the patients were followed up for 6 - 36 months. Significant stable clinical improvement was obtained in 25 patients, and recurrence was seen in one patient. Conclusion Transcatheter thrombus-breaking combined with local infusion of reteplase can quickly improve the pulmonary circulation and the clinical conditions for patients with severe pulmonary thromboembolism, and it has no obvious complications. This technique carries excellent value in maintaining the stability of hemodynamics.
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<p><b>OBJECTIVE</b>To evaluate the performance of visual inspection with acetic acid (VIA) in preliminary screening of cervical cancer and its precancerous lesions among Chinese women by meta-analysis of diagnosis.</p><p><b>METHODS</b>Pubmed, Cochrane, Wanfang, CNKI and Weipu databases were employed to search for citations using the MeSH terms as "acetic acid", "cervical intraepithelial neoplasia", and "cervical cancer" both in Chinese and English. Additional relevant references cited in retrieval articles were also searched.40 pieces of research paper related with screening of cervical cancer and precancerous lesions of cervical cancer in Chinese women by VIA were collected. Bivariate random effects model was adopted using SAS 8.02.</p><p><b>RESULTS</b>Twenty-two studies including 23 330 cases were finally selected in the analysis, among which 19 studies were reported in Chinese and the other 3 in English. These studies were reported from 2004 to 2010 and the age-range of subjects was between 15 and 81 years old. Stratified analysis of diagnosis threshold showed that the pooled diagnostic odds ratio (DOR) of VIA for CIN1+ (4.11, 95%CI: 3.20 - 5.04) was similar to that for CIN2+ (4.45, 95%CI: 3.73 - 5.15). Either CIN1+ or CIN2+, the DOR in younger women (≤ 40 year) (4.22, 95%CI: 3.29 - 5.16; 4.53, 95%CI: 3.46 - 5.47) was also similar to it in older women (> 40 year) (3.66, 95%CI: 2.27 - 5.37; 4.26, 95%CI: 3.32 - 5.26). There was no difference in the screening performance between county-level doctors (DOR = 4.62, 95%CI: 3.13 - 5.93) and municipal-level doctors (DOR = 4.48, 95%CI: 3.71 - 5.16).</p><p><b>CONCLUSION</b>The screening performances of VIA were relatively consistent among different lesion grades and aging groups of Chinese women. After professional training, there was no difference in performance between county-level hospitals and municipal-level hospitals.</p>
Subject(s)
Adolescent , Adult , Aged , Aged, 80 and over , Female , Humans , Middle Aged , Young Adult , Acetic Acid , Asian People , Uterine Cervical Dysplasia , Diagnosis , Mass Screening , Methods , Uterine Cervical Neoplasms , DiagnosisABSTRACT
<p><b>OBJECTIVE</b>To investigate the effects of a recombinant adenovirus-mediated triple mutant hypoxia-inducible factor-1α (HIF-1α) on the proliferation and vascular endothelial growth factor (VEGF) expression in human microvascular endothelial cells (hMVECs).</p><p><b>METHODS</b>The adenovirus vector of the triple mutant HIF-1α (Ad-HIF-1α(564/402/803)), adenovirus vector of wild-type HIF-1α (Ad-HIF-1α(nature)), Ad-lacZ and Ad-Null were amplified in HEK293A cells, and the adenoviruses were purified and titrated. Dual luciferase reporter assay system was employed to detect the transcriptional activities of wild-type and triple mutant HIF-1α. After infection of the hMVECs with the adenoviruses, the cellular protein expressions of HIF-1α and VEGF were detected using Western blotting, and the cell proliferation was assessed by MTS assay.</p><p><b>RESULTS</b>The transcriptional activity of the triple mutant HIF-1α was significantly higher than that of wildtype HIF-1α in the infected hMVECs (P<0.001). The protein levels of HIF-1α and VEGF in cells infected with Ad-HIF-1α(564/402/803) were significantly higher than those in cells infected with other adenoviruses, and HIF-1α dose-dependently up-regulated VEGF protein expression. The absorbance was significantly higher in Ad-HIF-1α(564/402/803) group than in the other groups (P<0.01) on the third and fifth days after infection.</p><p><b>CONCLUSION</b>The recombinant adenovirus-mediated triple mutant HIF-1α expression is stable under normoxic condition. The triple mutant HIF-1α can up-regulate the expression of VEGF protein in hMVECs to promote the cell proliferation.</p>
Subject(s)
Humans , Adenoviridae , Genetics , Cell Proliferation , Endothelial Cells , Cell Biology , Metabolism , Endothelium, Vascular , Cell Biology , Genetic Vectors , HEK293 Cells , Hypoxia-Inducible Factor 1, alpha Subunit , Genetics , Pharmacology , Microvessels , Cell Biology , Recombinant Proteins , Genetics , Transfection , Vascular Endothelial Growth Factor A , MetabolismABSTRACT
BackgroundAn ideal animal model is very important for the investigation of the immune mechanism of high-risk rejection corneal transplantation.ObjectiveThis study was to compare three methods of creating a high-risk corneal transplantation model in rabbits to study high-risk rejection corneal transplantation.MethodsForty-five New Zealand white rabbits were utilized and assigned randomly to three groups of different modeling methods,with 15 rabbits for each group.The high-risk corneal transplantation models were created by suturing with 5-0 silk thread in 4 quadrants,inducing alkali burn with 1 mol/L NaOH or corneal xenotransplantation.In the suturing group and alkali burning group,the rabbits received a unilateral 7.25 mm diameter corneal allograft after corneal neovascularization was induced,and in the xenotransplantation group,corneas from cats were used as donors.Rabbits were followed-up for 4 weeks in all groups.Corneal neovascular area was calculated and compared among the three groups.The amount of rejection,inflammatory index ( IF),neovascularization and histology of grafts were clinically scored to calculate the reject index (RI).ResultsThere were 14,15 and 15 rabbits that survived the high-risk penetrating corneal transplantation,respectively,in the suturing group,alkali burning group and xenotransplantation group.Two weeks after operation,the IF scores were 0.543 ± 0.103,0.811 ± 0.054 and 0.191 ±0.087,and the RI were 2.111±0.928,7.0±0.816 and 3.182±0.751 in the suturing group,alkali burning group and xenotransplantationgroup,respectively,showingstatisticallysignificantdifferencesamongthethreegroups (x2 =25.736,22.432,P =0.000).The IF value was lower in the xenotransplantation group compared with the suturing group and alkali burning group (Z =3.841,3.993,P =0.000),and that of the suturing group was lower than the alkali burning group (Z =3.568,P =0.000).The RI value of the xenotransplantation group was significantly raised in comparison with the suturing group and declined in comparison with the alkali burning group (Z =2.373,P =0.018;Z =3.936,P =0.000),and that of the suturing group was lower than the alkali burning group (Z =3.729,P =0.000 ).The survival times of the grafts were ( 17.9±2.0 ) days,( 13.4 ±2.4) days and ( 15.5 ±2.0 ) days in these three groups with a significant difference among them ( F =9.474,P =0.001 ).The neovascularization area in the xenotransplantation group was smaller than the suturing group and alkali burning group (P＜ 0.05 ).Histological examination revealed a large number of inflammatory cells infiltration in the grafts 2 and 4 weeks after transplantation in the suturing group and alkali burning group,but less inflammatory cells were seen in the xenotransplantaion group.Immunofluorescence staining showed abundant CD4+ T positive cells in the grafts in the three groups.Conclusions The cat-rabbit corneal xenotransplantation can induce stable and moderate immune rejection.This animal model has milder inflammatory response and less corneal neovascularization than the suture and alkali burn models.This method therefore is an ideal model for high-risk corneal transplantation.
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<p><b>OBJECTIVE</b>To investigate the influence of lyophilization on the biological activity of recombinant adenovirus-mediated triple mutant of hypoxia inducible factor-1α (Ad-HIF-1α-564/402/803).</p><p><b>METHODS</b>Ad-HIF-1α-564/402/803 was amplified from HEK293A cells and purified by ultracentrifugation in CsCl gradient solutions. The infection efficiency was observed by X-gal staining. The lyophilized adenovirus was prepared under appropriate conditions. Before and after lyophilization, the effect of Ad-HIF-1α-564/402/803 on hMVEC proliferation was evaluated by MTS assay. The recombinant adenovirus was confirmed by PCR and DNA sequence analysis before and 1 day, 6 months and 12 months after lyophilization, and hMVECs infected with Ad-HIF-1α-564/402/803 at these time points were examined for HIF-1α protein expression using Western blotting.</p><p><b>RESULTS</b>No significant changes were observed in the effect of lyophilized Ad-HIF-1α-564/402/803 on hMVECs proliferation at the optimal multiplicity of infection of 100 pfu/cell (P>0.05). At the 4 time points, the recombinant adenovirus HIF-1α showed no structural alterations or significant changes in the expression level of HIF-1α protein in the transfected hMVECs (P>0.05).</p><p><b>CONCLUSION</b>Lyophilized Ad-HIF-1α-564/402/803 can maintain its biological activities for a long time.</p>
Subject(s)
Humans , Adenoviridae , Genetics , Metabolism , Antibodies, Monoclonal , Genetics , Freeze Drying , Genetic Vectors , HEK293 Cells , Hypoxia-Inducible Factor 1, alpha Subunit , Genetics , Mutant Proteins , Genetics , MetabolismABSTRACT
<p><b>OBJECTIVE</b>To investigate the effect of recombinant adenovirus-mediated triple mutant of hypoxia inducible factor-1alpha (Ad-HIF-1alpha-564/402/803) in modulating angiogenesis in vitro.</p><p><b>METHODS</b>The recombinant adenoviruses Ad-lacZ, Ad-Null, Ad-HIF-1alpha-nature, and Ad-HIF-1alpha-564/402/803 were amplified in HEK293A cells and purified by ultracentrifugation in CsCl step gradient solutions, and the adenoviral titer was determined by end-point dilution assay. The recombinant adenovirus was confirmed by PCR and DNA sequence analysis, and the infection efficiency was observed by X-gal staining. Human microvascular endothelial cells (HMVECs) were infected with Ad- HIF-1alpha-564/402/803, Ad- HIF-1alpha-nature, or Ad-Null to compare the number of capillary-like tube structures in vitro. The effect of Ad- HIF-1alpha-564/402/803, Ad-HIF-1alpha-nature, and Ad-Null on angiogenesis was evaluated using a chick embryo chorioallantoic membrane (CAM) model.</p><p><b>RESULTS</b>PCR and gene sequencing suggested the correct construction of the recombinant adenovirus HIF-1alpha, and the adenoviral titer reached 1011-1012 PFU/ml. Infection of the hMVECs with Ad-HIF-1alpha-564/402/803 at the optimal multiplicity of infection of 100 pfu/cell resulted in a significantly greater number of capillary-like tube structures than infection by Ad-HIF-1alpha-nature and Ad-Null (P=0.000). Ad-HIF-1alpha-564/402/803 group showed significantly higher microvessel density than Ad-HIF-1alpha-nature, Ad-Null, and PBS groups, with also higher angiogenesis area to CAM area ratio (P=0.01, 0.000, and 0.000, respectively).</p><p><b>CONCLUSION</b>The triple mutant Ad-HIF-1alpha-564/402/803 can obviously promote the formation of capillary-like tube structures in vitro and modulate angiogenesis in the CAM model, suggesting the capacity of Ad-HIF-1alpha-564/402/803 in promoting angiogenesis under normoxic condition.</p>
Subject(s)
Animals , Chick Embryo , Humans , Adenoviridae , Genetics , Metabolism , Capillaries , Chorioallantoic Membrane , Endothelial Cells , Cell Biology , Metabolism , HEK293 Cells , Hypoxia-Inducible Factor 1, alpha Subunit , Genetics , Mutation , Neovascularization, Physiologic , Genetics , Recombinant Proteins , GeneticsABSTRACT
Objective To investigate in evaluation of various capacity requirement of probationer nurs-ing students by nurses in operation room in order to improve clinical teaching work. Methods A self-de-signed questionnaire was applied, 191 qualified teaching nurses from nine grade three A general hospitals in Beijing were invited to fdl the questionaires. The results underwent subsequent analysis. Results The teach-ers had high expectations for various capacity of the probationer nursing students. Conclusions Clinical teachers should establish feasible teaching scheme according to the existing teaching syllabus in order to in-crease teaching quality, make students gain new insights through reviewing old materials, so that they can mas-ter various technique in operation room in short time.