Objective To construct a recombinant eukaryotic expression vector of human tumor suppressor p53-binding protein 2(TP53BP2)and transfect human embryonic kidney Expi293F cells.High-purity recombinant human full-length TP53BP2 protein was obtained and its biological activity was identified.Methods The TP53BP2 gene sequence was queried on the UniProt website,and the Expi293F expression system was optimized.The TP53BP2 gene was connected to pcDNA3.1(+)-P2A-eGFP vector by homologous recombination,and identified by double enzyme digestion and sequencing.Transect pcDNA3.1(+)-P2A-eGFP-TP53BP2 plasmid into Expi293F cells of Polyethylenimine(PEI),observe the transfection efficiency with a fluorescence microscope,collected cells from the experiment group and control group.The expression level of TP53BP2 recombinant protein was detected by Western blot(WB).Protein was purified by His label purification kit and Superdex 20010/300 GL chromatographic column.Sodium dodecyl sulfate-polyacrylamide gel electrophoresis.The purified recombinant protein was identified by SDS-PAGE.Combining recombinant human full-length TP53BP2 protein with p65 protein was investigated for Co-immunoprecipitation(Co-IP)precipitation.Recombinant human full-length TP53BP2 protein was co-localized with p65 protein by Immunofluorescence(IF).The surface plasmon resonance(SPR)technique was used to detect the interaction between purified recombinant human full-length TP53BP2 protein and TP53BP2 antibody.Results The recombinant plasmid pcDNA3.1(+)-P2A-eGFP-TP53BP2 was successfully constructed by sequencing and double digestion.The fluorescence microscopy results showed that the transfection efficiency was about 60%.WB showed that the TP53BP2 protein was overexpressed in Expi293F cells,which proved that transfection was successful.SDS-PAGE results showed that the purity of the purified recombinant protein was above 90%,which proved that the purification was successful.Co-IP results showed that the TP53BP2 could interact with p65 protein.The results of IF showed that His tag protein,TP53BP2 protein,and p65 protein were co-located,indicating the interaction between the three proteins.SPR results showed that the purified TP53BP2 recombinant protein had good binding activity with the TP53BP2 antibody.These results all prove that the recombinant human full-length TP53BP2 protein has biological activity.Conclusion The eukaryotic expression vector of TP53BP2 gene was successfully constructed and the recombinant full-length human TP53BP2 protein with biological activity was successfully expressed in human embryonic kidney Expi293F cells.It lays a foundation for further study on the structure and function of TP53BP2.

Mycoplasma capricolum subsp. capripneumoniae (Mccp) is the cause of contagious caprine pleuropneumonia (CCPP) in goats. Inactivated vaccines and capsular polysaccharide (CPS) indirect hemagglutination reagents are available for prevention and serological detection, but high culture costs and complex antigen quantification have been plagued by production staff. In order to solve these problems in production practice, a sugar fermentation medium with an initial pH value of 7.8, which could improve the production of two antigens simultaneously, was screened out by changing the initial pH value based on previous Mccp metabolomics analysis. Since phenol red can be identified by UV absorption spectrum and cetyltrimethylammonium bromide (CTAB) can bind to anionic capsular polysaccharide, a UV spectrum measurement method for analyzing the culture stage reached by Mccp and a CTAB precipitation test for relative quantification of capsular polysaccharide antigen content in the fermentation broth were established. The UV spectrum observation method can guide the production of Mccp according to the growth curve of Mccp, which greatly reduces the monitoring time of the traditional CCU method and improves the accuracy of the original eye-observation method. The established CTAB precipitation test can complete the monitoring of CPS content within 5 hours, which greatly reduces the time required compared with the traditional differential technique, and its accuracy was verified by the phenol-sulfuric acid method. The optimized culture medium and the two correlation comparison methods established in this study can effectively reduce the production cost of Mccp and improve the production efficiency. The two assays have been used in the research at our laboratory, which provides experimental data for further improvement of the production process of CCPP inactivated vaccine and capsular polysaccharide as well as rapid quantification.
Humans ; Animals ; Goats ; Cetrimonium ; Mycoplasma ; Polysaccharides

Objective To study the regulatory effects of microRNA-200b (miR-200b) on hypoxia-inducible factors-1α (HIF-1α） in neonatal immature rats with hypoxic-ischemic brain damage (HIBD).Method A total of 240 three-day-old neonatal Sprague-Dawley (SD) rats were randomly assigned into six groups with 40 rats in each group:the hypoxic-ischemic group (HI group),intraventricular injection of miR-200b agomir,intraventricular injection of miR-200b antagomir,intraventricular injection of agomir negative control group,intraventricular injection of antagomir negative control group and the normal control group.The HIBD models of immature neonatal rats were established except for the normal control group.The relative expressions of HIF-1 α in brain tissues of each group were detected using quantitative real-time-PCR at 12 h,1 d,3 d and 7 d after ventricular injection,and the changes of HIF-1α expression in each group were compared.Result (1) Compared with the control group,the expression of HIF-1oα of the HI group began to increase 12 h after the injection of normal saline into the lateral ventricle (P＜0.05),and reached the peak at 1d,with statistically significant difference (P＜0.05),and then gradually decreased to the normal control group level at 7 d.(2) No significant differences of HIF-1α existed among the HI group and the HI+ agomir negative control group and the HI + antagomir negative control group (P＞0.05),and the miR-200b carrier had no significant effects on the expression of HIF-1α.(3)HIF-1α continued to be highly expressed after the injection of antagomir into the lateral ventricle of HI,and was significantly higher than the HI group at 12 h (P＜0.05).No significant differences existed between the HI+antagomir group and the H1 group at 1 d,3 d and 7 d after antagomir injection (P＞0.05).The expression of HIF-1α was constantly lower than the HI group after the injection of agomir,and significantly lower than the HI group at 1d after injection (P＜0.05).Conclusion MiR-200b overexpression inhibits the expression of HIF-1α,and the low expression of miR-200b can increase the level of HIF-1oα in a limited time window.Therefore,miR-200b may participate in the regulation of brain injury in neonatal rats after HIBD by regulating the expression of HIF-1α.

Objective To explore the features of resting state functional magnetic resonance imaging (rs-fMRI) in the patients with mild cognitive impairment (MCI) after during the process of converting to Alzheimer's disease (AD) dementia. To study the correlation of the brain function imaging and neuropsychological scores in converted MCI subjects with the purpose of finding the potential functional biomarkers which may predict the conversion of MCI to AD dementia in short term using resting state functional MRI. Methods Twenty-seven patients with MCI and 31 age, gender and years of education matched normal controls (NC) were enrolled in this study and neuropsychological assessment and rs-fMRI data were acquired respectively at the baseline and follow-up. Functional connectivity strength (FCS) analyses were used to assess the functional differences among the groups of MCI-converters (MCI-c) , MCI-non converters (MCI-nc) and NC. Variance analyses were used to compare the group differences in age, years of education and each neuropsychological assessment. In addition, Chi-squared test was used to compare group differences in gender. Then, Kruskal-Wallis test was used to compare group differences in the interval time of twice data collection. Group differences in FCS values were analyzed by general linear model. Finally, correlation analyses were used to evaluate the relationships between the FCS values of the brain regions with group differences and neuropsychological assessment scores in MCI patients. Results (1) Significant group differences of FCS among the MCI-c, MCI-nc and NC groups were observed in bilateral angular gyri, bilateral orbitofrontal cortex, right precentral gyrus, left paracentral lobule, left precuneus, right fusiform, right middle occipital gyrus, right superior occipital gyrus, left cerebellum. (2) The MCI-c showed decreased FCS in bilateral angular gyri, right middle occipital gyrus, right superior occipital gyrus, right inferior occipital gyrus, right fusiform, left precuneus, and left paracentral lobule, as compared to MCI-nc, the cluster was 149, 114, 186, 56, 48, 33, 38 and 102 voxels, respectively;P<0.05, respectively. The correlation analysis showed that the FCS values of the right angular gyrus were negative correlated with the MMSE scores (r=-0.561, P=0.017) . Conclusions This study revealed that the comparison of rs-fMRI imaging data between MCI-c and MCI-nc at baseline is of great benefits in analysing the imaging characteristics of patients during converting from MCI to AD. The feature of FCS in rs-fMRI in right angular gyrus might serve as early indicators for the dysfunction and progression from MCI to AD.

Objective To explore the features of resting state functional magnetic resonance imaging (rs-fMRI) in the patients with mild cognitive impairment (MCI) after during the process of converting to Alzheimer's disease (AD) dementia. To study the correlation of the brain function imaging and neuropsychological scores in converted MCI subjects with the purpose of finding the potential functional biomarkers which may predict the conversion of MCI to AD dementia in short term using resting state functional MRI. Methods Twenty-seven patients with MCI and 31 age, gender and years of education matched normal controls (NC) were enrolled in this study and neuropsychological assessment and rs-fMRI data were acquired respectively at the baseline and follow-up. Functional connectivity strength (FCS) analyses were used to assess the functional differences among the groups of MCI-converters (MCI-c) , MCI-non converters (MCI-nc) and NC. Variance analyses were used to compare the group differences in age, years of education and each neuropsychological assessment. In addition, Chi-squared test was used to compare group differences in gender. Then, Kruskal-Wallis test was used to compare group differences in the interval time of twice data collection. Group differences in FCS values were analyzed by general linear model. Finally, correlation analyses were used to evaluate the relationships between the FCS values of the brain regions with group differences and neuropsychological assessment scores in MCI patients. Results (1) Significant group differences of FCS among the MCI-c, MCI-nc and NC groups were observed in bilateral angular gyri, bilateral orbitofrontal cortex, right precentral gyrus, left paracentral lobule, left precuneus, right fusiform, right middle occipital gyrus, right superior occipital gyrus, left cerebellum. (2) The MCI-c showed decreased FCS in bilateral angular gyri, right middle occipital gyrus, right superior occipital gyrus, right inferior occipital gyrus, right fusiform, left precuneus, and left paracentral lobule, as compared to MCI-nc, the cluster was 149, 114, 186, 56, 48, 33, 38 and 102 voxels, respectively;P<0.05, respectively. The correlation analysis showed that the FCS values of the right angular gyrus were negative correlated with the MMSE scores (r=-0.561, P=0.017) . Conclusions This study revealed that the comparison of rs-fMRI imaging data between MCI-c and MCI-nc at baseline is of great benefits in analysing the imaging characteristics of patients during converting from MCI to AD. The feature of FCS in rs-fMRI in right angular gyrus might serve as early indicators for the dysfunction and progression from MCI to AD.

Objective To analysis the basic condition of the health resources in Chongqing ,and to explore the effect of the al‐location of health resources on doctor‐patient relationship .Methods Health statistics yearbook of Chongqing city was collected ,the fairness of the allocation of health resources in chongqing carried on the analysis and evaluation ,using the DEA model to analyze and evaluation the efficiency of the allocation of health resources in chongqing ,self‐designed questionnaire was used;hospitals ,doc‐tors and patients were investigated ,the descriptive and statistical analysis carried on .Results The distribution of health resources in Chongqing was inequitable on a regional .This inequitable distribution had been evaluated to the effect that ,as per the DEA mod‐el ,7 prefectures were effective with another 5 slightly effective and 26 as DEA ineffective .In the 8 hospitals that had been investiga‐ted ,the occurrence of medical disputes was undergoing a surge ,which indicates the tense relation between doctors and patients .Con‐clusion The total amount of health resources in Chongqing is fairly insufficient ,the high quality health resources are scarce .Unbal‐anced allocation of health resources and allocation inefficiency is the main reason for nervous doctor‐patient relationship and contra‐diction focus .

Studying effects of 50 Hz sinusoidal electromagnetic fields (SEMFs) with different intensities on peak bone mass (PBM) of rats may provide a theoretical basis for application of electromagnetic clinical field. 30 female SD rats, 6 weeks of age, were randomly divided into three groups: the control group, 0.1 mT electromagnetic field group (EMFs) and 0.6 mT EMFs. The EMFs groups were treated for 3 h/day. After 8 weeks, we examined their bone mineral densities (BMD) , measured their bone biomechanical properties, and made serum levels of osteocalcin (OC), tartrate-resistant acid phosphatase 5b (TRACP 5b), and histomorphometry. It was found that the BMD (P < 0.01), maximum mechanical load (P < 0.01) in the 0.1 mT group were significantly higher than those in the control group, and Yield strength (P < 0.05), the analyses of serum bone turnover markers and histomorphometric parameters were better than those in the control group (P < 0.05). However, the 0.6 mT group did not have significantly difference comparing with that in the control group. This study proved that 50 Hz 0.1 mT SEMFs can increased BMD, bone strength, and bone tissue microstructure. Therefore, 50 Hz 0.1 mT SEMFs can improve peak bone mass of rats.
Acid Phosphatase ; blood ; Animals ; Bone Density ; Bone and Bones ; physiology ; Electromagnetic Fields ; Female ; Isoenzymes ; blood ; Osteocalcin ; blood ; Rats ; Rats, Sprague-Dawley ; Tartrate-Resistant Acid Phosphatase

To meet the needs of the society and improve the competitiveness of nursing students , many nursing school carried out nursing English courses .Among the related practice and researches , the nursing English teaching methods studies were more active .To investigate the current application and research situation of nursing English teaching methods in China , we reviewed and analyzed the literature based on the modern foreign language teaching methods .We hope this paper can provide the evidence for the future direction of nursing English teaching practice and research .

Objective To assess the efficacy of proton pump inhibitors for preventing post-endoscopio retrograde cholangiopancreatography (post-ERCP) pancreatitis and hyperamylasemia. Methods A total of 250 patients, who underwent successful ERCP, were randomized into control group (n = 126) or treatment group (n=124) after ERCP. All patients received octreotide, the patients in treatment group received omeprazole 40 mg i. v. every 12 h; while the patients in control group received same amount of normal saline. The serum level of amylase at 4, 24 h and TNF-a were determined and the incidence of post-ERCP pancreatitis was documented. Results There were no differences in age, sex and proportion of therapeutic procedures performed between the two groups. The serum amylase and TNF-a levels at 4, 24h were (221 ± 31) U/L, (181 ±39) U/L, (0.264 ±0.052) ng/ml and (0.257 ±0.071) ng/ml in treatment group, respectively; which were significantly lower than (272 ±32) U/L, (227 ±30) U/L, (0. 372 ±0.047) ng/ml and (0.422 ±0.026) ng/ml in the control group (P<0.05). The incidence of pancreatitis in proton pump inhibitors group was 1.6%, which was significantly lower than that in control group (5.6% , P = 0. 04) . Conclusions Combination of proton pump inhibitors and octreotide could effectively decrease the incidence of post-ERCP hyperamylasemia and pancreatitis.

Objective To study the mechanism of Hezhong Jiangni Formnla(HJF) in protecting esophageal mucosa.Methods Rat models of reflux esophagitis were established by cardioplasty,pyloric ligation and roux-Y type gastrojejunostomy to evaluate the degree of esophageal mucosa injury.After treatment of HJF,the gastric juice was collected with the method of pyloric ligation to determine its volume and acidity and the activity of pepsin.The gastrointestinal motility was observed after oral use of charcoal powder.The level of malondialydehyde(MDA)in esophageal mucosa was measured to evaluate the esophageal antioxidative ability.Results HJF relieved the injury of esophageal mucosa dose-dependently;it had no obvious influence on the volume and acidity of gastric juice and the activity of pepsin,but it could obviously improve the gastrointestinal motility.MDA level of esophageal mucosa in the model group was increased significantly as compared with the control group.Pretreatment with HJF could greatly inhibited the formation of MDA.Conclusion HJF has protective effect on esophageal mucosa,which may be related to the improvement of gastrointestinal motility and its antioxidant action.