Objective To optimize the method of transcription activator‐like effector transcription factors (TALE‐TFs) con‐struction ,some improvement and adaption were made based on the traditional methods .Methods We first constructed the basic tandem fragments with different length ,including trimer ,tetramer ,pentamer and hexamer by Golden Gate cloning technique and PCR ,then the procedure with the highest efficacy was chosen to construct our TALE‐TFs .To determine the function of the TALE‐TFs ,the plasmid pminCMV with the specific binding sequence of TALE‐TFs was constructed by fragment substitution reaction (FSR) .The transcription activating function of TALE‐TFs was confirmed by the intensity of red fluorescence ,after TALE‐TFs , pEGFP‐N1 and pminCMV plasmid were co‐transfected into 293HEK cells .Results An optimized method for TALE‐TFs construc‐tion and functional assay was established .Conclusion This method can potentially be wildly used in fields that the expression of some constitutively expressed genes needs to be modified .

Objective To investigate the nutrition status and quality of life(QOL)in the perioperative patients with laryngeal cancer and to analyze the effect of nutrition status on quality of life.Methods Sixty-six patients with laryngeal cancer were surveyed respectively by self-designed general situation questionnaire,nutritional risk srcreening 2002(NRS2002)and European organization for research and treatment of cancer quality of life core questionnaire(EORTC QLQ-C30).The impact of nutritional status on QOL was analyzed by the multiple linear regression analysis.Results The total score of nutritional risk of perioperative patients with laryngeal cancer was 3.53±1.51 and the incidence of nutritional risk was 28.79%.The nutrition status influenced on QOL, physiological function,cognitive function and pain area(P<0.05).Conclusion Nurses should assess the nutritional status of the patients and correct malnutrition without delay in order to improve the physical function and QOL of the patients.

Hepatitis B virus (HBV) DNA replication takes place in the viral capsid that consists of 180 or 240 copies of HBV capsid (HBc or core) protein. The monomeric core protein contains an apical loop region that forms the spikes on the surface of viral capsid upon core dimerization and capsid assembly. To investigate the impact on HBV DNA replication through gene engineering at the spike of HBV capsid. plasmids expressing engineered HBc with linker-fused enhanced green fluorescent protein (EGFP) or shortened EGFP insertion at the spike region were constructed by Restriction Digestion and Ligation-independent Cloning (RLIC). The wildtype or mutant HBc construct was cotransfected with HBV1.1c(-), a plasmid containing 1.1 unit-length HBV genome with deficiency in HBc expression, into HEK293 cells, respectively. GFP signal was observed through a fluorescence microscope and HBV DNA replicative intermediates were assayed by Southern blotting to determine the expression and functions of different recombinants. Our results demonstrated that the RLIC method was effective to generate deletion or insertion in the apical loop region of HBc. Both HBc-EGFP recombinants with different linkers produced green fluorescence but with different subcellular distribution pattern. However, HBV DNA replication was not detected with the trans-complementation of these two HBc recombinants. In addition, other recombinants including the one only with the deletion of aa79-80 failed to support HBV replication. Taken together, our results suggest that RLIC is a robust method which can be broadly applied in gene engineering; different peptide linkers may have different influences on the functions of an engineered fusion protein; and HBc aa79-80 play a critical role for HBc to support HBV DNA replication.
Capsid Proteins ; genetics ; Cloning, Molecular ; Genetic Engineering ; methods ; Green Fluorescent Proteins ; biosynthesis ; genetics ; HEK293 Cells ; Hepatitis B Core Antigens ; biosynthesis ; genetics ; Hepatitis B virus ; genetics ; physiology ; Humans ; Mutation ; Recombinant Fusion Proteins ; biosynthesis ; genetics ; Transfection ; Virus Replication

OBJECTIVE:To explore the establishment of DUE criteria for the evaluation of glucocorticoids(GCs)use in oncol-ogy department of our hospital,and to evaluate the rationality of glucocorticoid use. METHODS:DUE criteria for GCs in our hos-pital was established preliminary,and used to evaluate medication indication,medication course,medication results,etc. Medical records of 300 patients receiving GCs were collected from oncology department of our hospital during Apr. to Dec. in 2013(before intervention)and Jun. 2014 to Mar. 2015(after intervention)via hospital information system. The application of GCs was evaluat-ed. RESULTS:Pexamethasone sodium phosphate injection was used most frequently in our hospital(82.34%),followed by Dexa-methasone acetate tablet(15.33%)and prednisone asetate tablets(2.33%). The mest common purpose of GCs use in oncology de-partment of our hospital is radiotherapy and chemotherapy induced vomiting prevention and treatment(55.55% of total case),fol-lowed by anti-inflammatory therapy(14.19%)and chemotheraphy drug pretreatment(12.76%). After intervention,rational use of GCs for tumor had been improved greatly,and the rate of unqualified drug dosage and medication course decreased from 6.87% to 3.95%,and that of decreased from 3.58% to 2.74%. CONCLUSIONS:The establishment of DUE criteria and continuous criteria improvement during clinical rational drug use guidance can promote rational use of GCs in oncology department.

Objective:To investigate the association between metabolically healthy obesity(MHO) and atherosclerosis risk among Chinese community population aged 40 or older.Methods:A total of 9 525 participants without cardiovascular diseases (3 621 men and 5 904 women) from Jiading community in Shanghai were enrolled to complete questionnaires, undergo extensive physical examination including brachial-ankle pulse wave velocity (baPWV) and blood pressure (BP) assessment, and laboratory screening. According to body mass index (BMI) and metabolic status, these participants were categorized into 4 groups including metabolically healthy non-obese (MHNO), metabolically unhealthy non-obese (MUNO), MHO, and metabolically unhealthy obese (MUO). High baPWV was defined as baPWV>1 400 mm/s, and high pulse pressure (PP) was defined as PP above fourth quartile of the population. Multivariate logistic regression model was conducted to explore the relationship between MHO and high baPWV as well as high PP after adjusting for confounders. Results:After multivariable adjustment, such as sex, age, current smoking, current drinking, and education, logistic regression analysis showed that MHO was significantly correlated with high baPWV ( OR=1.18, 95% CI 1.02-1.37) and high PP ( OR=1.72, 95% CI 1.43-2.08) in comparison with MHNO. Otherwise, both MUNO and MUO subjects were at higher risk for suffering from high baPWV (MUNO: OR=3.02, 95% CI 2.60-3.50; MUO: OR=3.26, 95% CI 2.87-3.70) and high PP (MUNO: OR=2.56, 95% CI 2.17-3.02; MUO: OR=3.49, 95% CI 3.01-4.06). Conclusion:On the basis of Chinese community population, there was a pronounced correlation between the MHO phenotype and the increased risk of developing atherosclerosis.

The experiment was designed to investigate the function of SREBP cleavage-activating protein (SCAP) mutant (D443N) by constructing an eukaryotic expressive vector using a smooth muscle specific promoter SM22 (pGL3-SM22-SCAP(D443N)). SM22 promoter (pSM22) was amplified from genome DNA of mice by nested PCR, and then cloned into pMD-T vector. The SM22 promoter fragment released from the vector by Kpn I and Hind III digestion was sub-cloned into pGL3-control-Luc vector, to form pGL3-SM22-Luc. The activity of pSM22 in human vascular smooth muscle cells (VSMCs) was tested using Dual-Luciferase Reporter System. SCAP(D443) mutant amplified from plasmid pTK-HSV-SCAP(D443N) and pSM22 from mice liver were cloned into pGL3-control vector to construct pGL3-SM22-SCAP(D443N) which was transfected into Chinese hamster ovary cells (CHO) to test SCAP(D443) expression by real-time PCR and Western blot. The sequence and construction of pGL3-SM22-SCAP(D443N) were correct. SM22 promoter activity initiated the expression of luciferase in VSMCs and also drove SCAP(D443) expression in transfected CHO cells. The pGL3-SM22-SCAP(D443N) eukaryotic expression vector was successfully constructed and the recombinant vector provides a powerful approach in investigating the function and regulation of SCAP and also in producing vascular smooth muscle specific SCAP transgenic mice.
Animals ; CHO Cells ; Cricetinae ; Cricetulus ; Genetic Vectors ; genetics ; Humans ; Intracellular Signaling Peptides and Proteins ; genetics ; physiology ; Membrane Proteins ; biosynthesis ; genetics ; physiology ; Mice ; Mice, Transgenic ; Microfilament Proteins ; genetics ; Muscle Proteins ; genetics ; Mutant Proteins ; biosynthesis ; genetics ; Promoter Regions, Genetic ; genetics ; RNA, Messenger ; biosynthesis ; genetics ; Recombinant Proteins ; biosynthesis ; genetics ; Transfection

Objective:To evaluate the accuracy of measuring the siae of atrial septal defect(ASD) by transthoracic echocardiography(TTE) on " compression of right superior pulmonary vein sheath during operation" (compression-size) using multi-layer perceptron.Methods:From January 2016 to January 2019, 460 cases of ASD with soft margin, thin growth or central type with atrial septal tumor in the First Hospital Affiliated of Air Force Medical University were analyzed retrospectively. The age was (34.32±18.84)years, 129 males and 331 females. Preoperative TTE and intraoperative TTE were performed to measure the size of ASD and compression-size. With the final occluder size selected as the dependent variable, the preoperative and intraoperative ultrasound measurements were used as covariates, and all patients were randomly divided into training set, test set and verification set according to the proportion of 5∶2∶3, which were imported into multi-layer perceptron. Gradient descent method was used to optimize and adjust the weight, and 10 operations were carried out respectively, and the average value was calculated to evaluate the two ultrasound methods for blocking.Results:The mean values of ASD were (15.26±5.33)mm for preoperative, (23.83±6.39)mm for compression-size intraoperative TTE, and (25.14±6.55)mm for the occluder, respectively. The difference was statistically significant (χ 2=850.450, P<0.001). There were significant differences between the pairwise ( P<0.001). Ten times of multi-layer perceptron were used to obtain a training model for predicting the size of the occluder based on preoperative transthoracic ultrasound and TTE measurement after right upper pulmonary vein sheath compression. The average relative error of the model obtained by preoperative ultrasound measurement in the validation set was (16.55±0.02)%, and that of the model obtained by intraoperative ultrasound measurement in the validation set was (4.81±0.01)%. The difference between the two methods was statistically significant ( t=16.185, P<0.001). Conclusions:It is more accurate to measure the ASD size for selection of occluder by the use of the right superior pulmonary vein sheath compression and TTE, especially for ASD patients with soft margin, thin growth or atrial septal tumor.

The persistent infection of hepatitis B virus (HBV) is the result of lacking specific immunity against the virus. This state is also called immune tolerance to HBV. In most cases, acute HBV infection in adults can induce specific immune response which can clear the virus. Perinatal HBV infection, however, usually progresses to chronic infection, indicating a defect in HBV-specific immune response. A typical specific immune response includes four processes, which were antigen presentation, specific CD4 + T cell activation, specific CD8 + T cell activation and B cell activation. There must be some dysfunctions in some or all of the four processes during chronic HBV infection. This article discussed the relationship between chronic HBV infection and cellular immunity, hoping to provide a reference for further study on the reconstitution of specific immunity against HBV.

Objective To investigate the association of age at menarche with the risks of diabetes and metabolic syndrome ( MS) in adulthood in Shanghai community postmenopausal women. Methods A total of 10375 residents over 40 years in Jiading, Shanghai were identified by a cluster sampling method. After taking standardized questionnaire surveys, physical examinations, and biochemical parameters testing, we gathered their lifestyle and medicine information. A total of 4723 postmenopausal women were enrolled in the study. The population was divided into three groups according to age at menarche, and their risks of MS and diabetes in various groups were compared. Results After multivariate adjustment, postmenopausal women with a menarche age<15 years had a 29％increased risk of diabetes(95％CI 1.03-1.61) and a 45％ increased risk of MS(95％ CI 1.20-1.76) in adulthood, compared with those with menarcheal age at 15-18 years. Among the five components of MS, earlier age of menarche was associated with higher risks of hypertriglyceridemia, low high density lipoprotein-cholesterol, and central obesity. Conclusions In Shanghai Jiading community postmenopausal women, age at menarche is associated with the risk of diabetes and MS, independent of the body mass index in adulthood.

Objective:To investigate the correlation between exposure to famine in early life and later risk of type 2 diabetes in adulthood.Methods:A cluster sampling method was used to include 8 868 residents who were lived in the Jiading community of Shanghai during the Great Famine from 1959 to 1962 in China. Subjects were divided into non-exposed group, fetal exposure group, childhood exposure group, and adolescent exposure group. Logistic regression model was used to analyze the relationship between famine exposure in early life and the risk of type 2 diabetes in adulthood. Results:Famine exposure during childhood and adolescent both increased the risk of developing type 2 diabetes in adulthood in women. No significant correlation was observed in men. In subjects with less physical activity and lower education level, the risk of developing type 2 diabetes mellitus in adulthood was significantly higher in the famine-exposed group than that of non-exposed groupand the interactions were statistically significant.Conclusion:Early life famine exposure increases the risk of developing type 2 diabetes in adults, especially in women.