Objective To construct a recombinant human interleukin-1 receptor antagonist(rhIL-1Ra)strain of kanamycin resistance,express,purify and identify rhIL-1Ra protein in order to reduce the risks of β-lactam antibiotics.Methods The ampicillin-resistant rhIL-1Ra(A-rhIL-1Ra)plasmid and PET-28a plasmid were used as templates to prepare the linearized vector and kanamycin gene fragment by PCR.The kanamycin resistant rhIL-1Ra plasmid(K-rhIL-1Ra)was constructed by homologous recombination,which was transformed into E.coli BL21(DE3)to construct the recombinant engineered bacteria after correct sequencing.The engineered bacteria were induced by IPTG and then purified by CM Bestarose Fast Flow and DEAE Bestarose Fast Flow column chromatography.The purified products were collected,and then detected for the purity by 15% SDS-PAGE,size-exclusion high-performance liquid chromatography(SEC-HPLC),and reversed-phase high-performance liquid chromatography(RP-HPLC),determined for the relative molecular mass by tandem mass spectrometry,identified for the specificity by Western blot,measured for the biological activity by reporter gene assay,and compared for the related impurities by liquid chromatography-mass spectrometry(LC-MS)analysis.Results Colony PCR and sequencing results showed that K-rhIL-1Ra plasmid was constructed correctly.The expressed K-rhIL-1Ra protein had a relative molecular mass of about 17 000,mainly existing in soluble form,and the expression amount accounted for more than 30% of the total bacterial proteins.The purity of K-rhIL-1Ra protein purified by two steps was 97% and 99%,the monomer content was 99.33%and 100%,and the chromatographic purity was 91.86% and 96.96%,respectively.The mass spectral molecular mass was consistent with that of the standard(A-rhIL-1Ra protein),and the protein reacted specifically relative with mouse antihuman IL-1Ra monoclonal antibody.The biological activity of the purified K-rhIL-1Ra protein was 1.29 × 105U/mg,and the chemical modification types of related proteins were the same as those of the standard(A-rhIL-1Ra protein).Conclusion The K-rhIL-1Ra strain was successfully constructed,and the expressed and purified protein was in line with the characteristics and quality standards of rhIL-1Ra,which lays a foundation for the study of rhIL-1Ra strain change and comparability

AIM: To understand the expression of ecto-5'-nucleotidase (CD73) and explore its effect on the motility and invasion ability of breast cancer cell lines. METHODS: ① CD73 mRNA expression was analyzed by semiquantitative RT-PCR. Surface CD73 activity in breast carcinoma cell lines was measured by HPLC according to the conversion of E-AMP to E-ADO. ② Plasmid pcDNA-NT5E was transfected into human low metastatic breast cancer cell line T47D with lipofectinamine2000. CD73 mRNA expression was detected by semiquantitative RT-PCR and surface CD73 activity was determined by HPLC before and after transfection. The effect of pcDNA-NT5E transfection on motility and invasion ability was observed by in vitro invasion assay. RESULTS: ① The RNA expression and enzyme activity of CD73 were variable in different breast cancer cell lines. ② The transfect-T47D overexpressed CD73 at mRNA level and with high enzyme activity. Constitutively active CD73 drove cells transmembrane and invasion into matrigel. Promotion effects were statistically significant (P

Objective: To discuss the application value with two-dimensional ultrasound speckle tracking technique in the evaluation of left ventricular function in patients with OSAS. Methods: Selected 110 cases of hospital patients with OSAS in recent years, used STI technical analysis of left ventricular myocardium using different directions overall resilience, Left ventricular torsion and systolic mitral annular midpoint of the maximum displacement (MAD) and left ventricular ejection fraction (LVEF) relationship, performed using SPSS 17.0 statistical correlation analysis. Results: MAD had a significant correlation with LVEF(r=0.81, P<0.01); GLS(r=-0.74, P<0.01),GRS GRS(r=0.57, P<0.01),GCS (r=-0.55, P<0.01) and LVtw(r=0.65, P<0.01)were significantly associated with LVEF;MAD and GLs(r=-0.69, P<0.01),GRs (r=0.49, P<0.01), LVtw (r=0.45, P<0.01)were significantly associated;MAD and GCs (r=-0.39, P<0.05)with a negative correlation. Conclusion:STI resilience through the myocardium, ventricular torsion and systolic mitral annular maximum displacement at the midpoint of the analysis, can accurately evaluate left ventricular longitudinal, radial and torsional motion synchronization, highly correlated with left ventricular ejection fraction, good consistency, with a very broad application prospects.

Objective To investigate the biological features of epidermal growth factor receptor inhibitor AG1478 on cervical carcinoma cell. Methods The proliferation of HeLa cells under AG1478 stimulation was determined by CCK8 assay. The expression of EGFR downstream signaling protein and apoptotic relative protein were examined by Western blot and transcription of apotosis?related genes were measured by RT?qPCR in AG 1478 treated HeLa cells. Nuclear transport of phosphorylated ERK were measured through ICC assay. TUNEL assay was used to determine early stage of apoptosis. Results CCK8 assay showed that AG1478 inhibit the proliferation of HeLa cells and also block phosphorylated level of EGFR ,ERK and AKT. Furthermore ,nuclear transport of phosphorylated ERK upon EGF stimulation were blocked and pro?apoptotic proteins were up?regluated with activat ed cleaved Caspases. Conclusion AG1478 inhibited the proliferation and induced apoptosis in HeLa cells and it could be a potential therapeutic drug for cervical carcinoma.

Objective:To understand the current situation of physicians′ competency in China and statistically analyze its influencing factors, hence providing referential evidences for promoting their competency.Methods:The evaluation scale for Chinese physicians′ competency was developed by referring to sophisticated overseas evaluation frameworks. Based on the electronic platform of the medical alliance, stratified sampling was conducted at 39 hospitals in 13 provinces, from which 2 156 physicians were surveyed, and statistical analysis was made on the current situation and influential factor of their competency in 6 dimensions(medical knowledge, diagnosis and treatment, scientific research and teaching, interpersonal communication, spirit of professionalism, population health)Results:The score range in multiple dimensions of physicians′ competency was between 2.85 and 3.89(5 in total), among which spirit of professionalism dimension scores the highest, and teaching and research dimension scores the lowest. Logistic regression analysis shows that their competency level in 6 dimensions is correlated to a range of factors, including gender, degree, title, supervisor qualifications, standardized training program of specialist physicians, and affiliation or not to teaching hospitals( P<0.05). Conclusions:Their overall competency is expected to be elevated. To name a few, they are lack of teaching and research abilities, and need to conduct clinical-oriented scientific research; they are expected to enrich knowledge structure, and need to embrace a wider concept of health; they need to improve their non-technical service abilities, and should further emphasize their humanistic quality; physicians′ practice competency depends upon their extent of medical education, and ensuring the quality of standardized training becomes the key measure.

ObjectiveTo summarize the experience of percutaneous nephrolithotomy under flexible ureteroscope in the treatment of staghorn calculi.Method The clinical data of 30 patieits with staghorn calculi undergoing percutaneous nephrolithotomy under flexible ureteroscope were reviewed for summarizing the surgical cooperation experience.Results All operations were successful,with the clearance rates of stage one and stage two calculi 86.0% and 95.0%,respectively.There occurred one case of massive hemorrhage,but no cases of kidney perforation,peritoneal perforation,liquid pneumothorax or organ damage.Conclusion Well-prepared surgical cooperation,full mastery of surgical procedures and skilled cooperation can increase surgical efficiency and safety by shortening surgical duration.

In order to promote our country's research on transitional care for patients with chronic kidney disease (CKD),the article summarized research progress on transitional care for patients with CKD,analysed the problems existed including lack of evidence-based basis,insugiciency of Intervention staff,one-sided intervention content and effect evaluation,etc,and put forward the corresponding countermeasures.

Objective To summarize the experience of intraoperative rapid parathyroid hormone (PTH) detection by immunochromatography in thyroid surgery.Methods Patients undergoing thyroid surgery in Shanghai Tenth People's Hospital from Nov.2016 to Jul.2017 were collected and the clinical data including level of blood calcium,serum phosphorus and PTH,and pathological findings were analyzed.Results In this study,im munochromatographic assay was used in 45 cases (experimental group) and nano carbon method was used in 47 cases (control group).In benign cases,levels of blood calcium and PTH in experimental group were higher than those in the control group ((2.07±0.09) vs (2.04±0.06) and (3.41±1.82) vs (3.34±1.84),P＞0.05).In malignant cases,blood calcium level in experimental group was higher than that in the control group ((2.02±0.13) vs (1.99±0.38),P＞0.05) and PTH level was significantly higher than that in the control group ((2.22±1.27) vs (1.44±1.44),P＜0.05).Serum phosphorus level in experimental group group was much lower than that in the control group ((1.02±0.17) vs (1.14±0.23),P＜0.05).Conclusions Using intraoperative rapid PTH detection by immunochromatography can identify parathyroid and avoid accidental injury of parathyroid gland,which can reduce the postoperative incidence of hypocalcemia.

Bronchial asthma is the most common chronic respiratory disease in children.With the increasing use of inhaled corticosteroids, asthma deaths in the world have decreased by nearly two thirds.Mortality of childhood asthma is very low, ranging from 0 to 0.7/100 000, however, most of risk factors of asthma death could be avoidable.Risk factors of asthma death include poor asthma control, poor treatment compliance, insufficient medicine prescribed by doctors, insufficient monitoring of lung function, mental and psychological problems, food allergy, insufficient recognition and treatment of asthma exacerbation, improper use of β 2-receptor agonist, insufficient application of asthma action plan, et al.The mortality and risk factors of asthma in children were reviewed, which could support further improving asthma management strategies.

Objective To evaluate the effect of Ji Tang Zhi on glucose metabolism in insulin resistance (IR) HepG 2 cell line,and to explore the related mechanism.Methods The HepG2 cells were incubated in culture medium addition of 10-7 mol/L insulin for 24 h to establish the IR cell model.Effect of Ji Tang Zhi on rate of glucose absorption in HepG2 cell was detected by the method of glucose oxidase-peroxidase (GOD-POD).We performed an MTT assay to determine cytotoxicity effects of Ji Tang Zhi on HepG2 cell line.The expression of p-IRS-1 Ser307,PI3K and GLUT-4 were detected by Western blotting.Results Incubated with 10-7 mol/L insulin for 24 h,the insulin resistance cell model had been built.Compared with model group,the rate of glucose absorption of cell treated with JTZ (30 ～ 120 μg/mL) was significantly improved.According to model cells,the expression of GLUT-4 and PI3K decreased significantly compared to control cells.While the expression of p-IRS-1 Ser 307 was inhibited and GLUT-4 and PI3K expression were increased in IR cells after treated with JTZ (30 ～ 120 μtg/mL).Conclusion JTZ exert beneficial effects on hyperglycosemia in IR cell line possibly through regulating the levels of GLUT-4,p-IRS-1 Ser307 and PI3K in HepG2 cell.