Objective To construct the recombinant plasmid of YTH domain family 2(YTHDF2)and express it in E.coli in order to obtain YTHDF2 fusion protein that was capable of binding m 6A-modified RNA.Methods The coding region of YTHDF2 gene was amplified by RT-PCR.The recombinant plasmid pET-28a-YTHDF2 was constructed and expressed in E.coli.The fusion protein was purified by Ni2+-NTA resin affinity chromatography, while the fusion protein activity was analyzed by Ni2+-NTA magnetic spheres.Results and Conclusion The recombinant YTHDF2 protein was expressed in E.coli BL21(DE3)and purified.YTHDF2 fusion protein was capable of binding RNA with m 6A-modification. The preparation of YTHDF2 fusion protein provides an essential tool to study the biological function of RNA with m6A-modification.

BACKGROUND: Three-dimensional magnetic field positioning and three-dimensional electric field positioning are two main technologies used in heart ablation, but their required equipments are very expensive. OBJECTIVE: To develop a new method of ablation catheter positioning by combining ultrasonic probe with ablation catheter. METHODS: Three-dimensional data of human heart were obtained from the anatomical data set of the body, and then a three-dimensional vector model of the endocardial surface was established. The basic parameters and scanning strategy of ultrasonic probe were designed and calculated, to compile a corresponding simulation software. Based on the three-dimensional endocardial vector model, the detection data of the ultrasonic probe were obtained by simulation calculation and processed. Then, the target surface characteristics were reconstructed, matching with the surface features of the proposed vector model. The position of the probe was finally determined. RESULTS AND CONCLUSION: The three-dimensional model of the endocardium was established based on the anatomical data set, and the ultrasonic surface detection data were obtained by the simulated ultrasonic irradiation. The reconstructed surface model with high accuracy exhibited a desired match with the target area in the three-dimensional model of endocardium. The error of the probe position and its theoretical position fulfilled the requirement of positioning accuracy. In conclusion, the use of ultrasonic imaging technology can accurately position the catheter probe in the heart during heart ablation, providing an accurate and reliable navigation for the implementation of heart ablation.

ngthen cellular immunity, especially Th1-type immune response to HPV11-E7 DNA vaccine in mice.

The preparation of tetracalcium phosphate (Ca4(PO4)2O, TTCP)was studied. Then calcium phosphate cement (CPC) was prepared. The setting time, pH value, compressive strength, X-ray diffraction (XRD) and scanning electron microscope (SEM) analysis of CPC were studied. The results show that TTCP containing small amount of CaO can be successfully attained heated at 1 500 degrees C for 6 h in vacuum condition. The initial setting time and final setting time of CPC is 4 min and 15 min, respectively. Its compressive strength is 20 MPa after 1-day immersion and 35 MPa after 7-day immersion. The pH value of the solution changes between 6.4 and 8.9. These properties can satisfy the clinical requirements of CPC. The final product of CPC is flake-like or needle-like hydroxyapatite (Ca5(P04)3OH, HA). The continuous network structure of HA appears in the microstructure, this leads to increase the strength of the material. This CPC can be used as bone substitute material.
Bone Cements ; chemical synthesis ; chemistry ; Calcium Phosphates ; chemical synthesis ; chemistry ; Compressive Strength ; Materials Testing ; Surface Properties ; Tensile Strength

Objective To reveal the relationships between epileptic seizure in patients with tuberous sclerosis complex (TSC) and their intelligence.Methods A retrospective analysis was performed on the clinical data of 110 patients with seizure secondary to TSC,admitted to our hospitals from May 2007 to May 2012; the prognostic factors for mental retardation were analyzed.Results There were 67 males and 43 females in the TSC patients with seizures.The age of seizure onset ranged from 10 days to 28 years; the onset age was less than 1 year in 30％ and less than 3 years in 65％ of TSC patients.Twelve patients presented epileptic spasms; 40 had normal intelligence and 51 had mental retardation.In patients having onset age younger than 1 year,80％ had mental retardation; 92％ patients with epileptic spasms had mental retardation; 65％ patients with two seizure patterns had mental retardation; 62％ TSC patients with seizure frequency exceeding 4 times per month and 63％ patients with seizure history exceeding 5 years both had mental retardation.Conclusion The patients with seizure onset age less than 1 year old and having a history of epileptic spasms are more prone to mental retardation; so early control of seizure should be done to improve the intelligence.

To express and secrete native HBscFv (anti-HBsAg single-chain Fv) in P. pastoris, HBscFv was amplified from plasmid pGEM-HBscFv, and then sub-cloned into expression vector pPICZalphaA. The resulting plasmid pPIC-HBscFv was linearized and transformed into P. pastoris GS115. The recombinant Pichia strains, identified by direct PCR and Zeocin-resistant screening of Pichia transformants, were cultured and induced with methanol. It was found that recombinant HBscFv, lead by alpha-factor, could be secreted into the culture supernatant to a level of 80mg/L. The bioactivity of Pichia produced HBscFv was confirmed by indirect ELISA, which also suggested that the bioactivity of HBscFv in the culture supernatant reached its peak in 72h and decreased in the late-stage of the induction. PAS staining suggests that HBscFv produced by yeast is poorly glycosylated or none-glycosylated protein.
Blotting, Western ; Electrophoresis, Polyacrylamide Gel ; Hepatitis B Surface Antigens ; immunology ; Humans ; Pichia ; genetics ; metabolism ; Polymerase Chain Reaction ; Single-Chain Antibodies ; genetics ; immunology ; metabolism

OBJECTIVETo investigate the effect of high mobility group box protein 1 (HMGB1) on the production of pro-inflammatory cytokines by Kupffer cell (KC) of rats with severe burn and the role of receptor for advanced glycation end products (RAGE) in the process.

METHODSModel of 30% TBSA full-thickness burn was reproduced in 32 SD rats through immersing the back in 98°C water for 12 s. KC (32 samples) was isolated from rat liver 24 h after injury and inoculated in 24-well plate in the concentration of 1×10(6) cell per well. (1) Cells were divided into control group (cultured with 1 mL PBS) and HMGB1 group (stimulated with 100 ng/mL HMGB1 in the volume of 1 mL) according to the random number table, with 8 samples in each group. At post culture hour (PCH) 48, the expression of RAGE (denoted as grey value ratio) was detected with Western blotting. (2) Another portion of cells were divided into control group (cultured with 1 mL PBS), HMGB1 group (treated with 100 ng/mL HMGB1 in the volume of 1 mL), HMGB1 + anti-RAGE antibody group (treated with 100 ng/mL HMGB1 in the volume of 1 mL after being pre-incubated with 20 µg/mL anti-RAGE monoclonal antibody in the volume of 1 mL for 2 hours), HMGB1 + recombinant rat RAGE/Fc chimera (rrRAGE/Fc) group (treated with the mixture of 100 ng/mL HMGB1 in the volume of 0.5 mL and 5 µg/mL rrRAGE/Fc in the volume of 0.5 mL which were pre-incubated for 2 hours) according to the random number table, with 8 samples in each group. At PCH 48, the protein levels of TNF-α and IL-1β in supernatant were determined with enzyme-linked immunosorbent assay, while the mRNA expression of TNF-α and IL-1β (denoted as grey value ratio) were determined with Northern blotting. Data were processed with one-way analysis of variance, t test, and LSD test.

RESULTS(1) The expression of RAGE in HMGB1 group (1.036 ± 0.101) was significantly higher than that of control group at PCH 48 (0.191 ± 0.024, t = -23.158, P = 0.000). (2) In HMGB1 group, HMGB1 + anti-RAGE antibody group, and HMGB1 + rrRAGE/Fc group, the contents of TNF-α in supernatant were respectively (10.59 ± 1.39), (9.91 ± 1.68), (11.51 ± 2.27) ng/mL; the contents of IL-1β in supernatant were respectively (2.49 ± 0.33), (2.08 ± 0.32), (2.42 ± 0.42) ng/mL; the mRNA levels of TNF-α in cells were respectively 0.311 ± 0.009, 0.301 ± 0.047, 0.326 ± 0.016; the mRNA levels of IL-1β in cells were respectively 0.237 ± 0.021, 0.244 ± 0.041, 0.245 ± 0.013. There were no statistically significant differences in the above indexes among these three groups (with P values all above 0.05). Their levels were all significantly higher than those of control group [with contents of TNF-α and IL-1β in supernatant respectively (2.69 ± 0.14), (0.43 ± 0.05) ng/mL, and mRNA levels of TNF-α and IL-1β in cells respectively 0.140 ± 0.022, 0.077 ± 0.005, P values all below 0.01].

CONCLUSIONSHMGB1 can induce the production of pro-inflammatory cytokines TNF-α and IL-1β from the KC in rats with severe burn. However, RAGE does not play a predominant role in this process.

Animals ; Burns ; metabolism ; Cytokines ; metabolism ; Disease Models, Animal ; HMGB1 Protein ; pharmacology ; Interleukin-1beta ; metabolism ; Kupffer Cells ; drug effects ; metabolism ; Male ; Rats ; Rats, Sprague-Dawley ; Receptor for Advanced Glycation End Products ; Receptors, Immunologic ; metabolism ; Tumor Necrosis Factor-alpha ; metabolism

Adult ; Aged ; Carcinoma, Hepatocellular ; metabolism ; Female ; Humans ; Liver Neoplasms ; metabolism ; Male ; Middle Aged ; RNA, Messenger ; biosynthesis ; genetics ; Receptor, ErbB-2 ; biosynthesis ; genetics ; Receptor, ErbB-3 ; biosynthesis ; genetics

Although many studies have investigated intensity-modulated radiation therapy (IMRT) for nasopharyngeal carcinoma (NPC), sample sizes in the reported studies are usually small and different in outcomes in different T and N subgroups are seldom analyzed. Herein, we evaluated the outcomes of NPC patients treated with IMRT and further explored treatment strategy to improve such outcome. We collected clinical data of 865 NPC patients treated with IMRT alone or in combination with chemotherapy, and classified all cases into the following prognostic categories according to different TNM stages: early stage group (T1-2N0-1M0), advanced local disease group (T3-4N0-1M0), advanced nodal disease group (T1-2N2-3M0), and advanced locoregional disease group (T3-4N2-3M0). The 5-year overall survival (OS), local relapse-free survival (LRFS), and distant metastases-free survival (DMFS) were 83.0%, 90.4%, and 84.0%, respectively. The early disease group had the lowest treatment failure rate, with a 5-year OS of 95.6%. The advanced local disease group and advanced nodal disease group had similar failure pattern and treatment outcomes as well as similar hazard ratios for death (4.230 and 4.625, respectively). The advanced locoregional disease group had the highest incidence of relapse and death, with a 5-year DMFS and OS of 62.3% and 62.2%, respectively, and a hazard ratio for death of 10.402. Comparing with IMRT alone, IMRT in combination with chemotherapy provided no significant benefit to locoregionally advanced NPC. Our results suggest that the decision of treatment strategy for NPC patients should consider combinations of T and N stages, and that IMRT alone for early stage NPC patients can produce satisfactory results. However, for advanced local, nodal, and locoregional disease groups, a combination of chemotherapy and radiotherapy is recommended.
Adolescent ; Adult ; Aged ; Carcinoma ; Chemoradiotherapy ; Chemotherapy, Adjuvant ; Disease-Free Survival ; Female ; Humans ; Lymphatic Metastasis ; Male ; Middle Aged ; Nasopharyngeal Neoplasms ; drug therapy ; pathology ; radiotherapy ; Neoplasm Recurrence, Local ; Neoplasm Staging ; Radiotherapy Dosage ; Radiotherapy, Intensity-Modulated ; Survival Rate ; Young Adult

OBJECTIVETo understand the current epidemic situation and high risk factors of human immunodeficiency virus (HIV) infection among paid blood donors living in Shandong provincial China Comprehensive Response (CARES) Project Areas.

METHODSAll residents between 20 to 60 years old were selected from one or several counties in August 2003.

RESULTSThere were 19 HIV(+) infections among 661 subjects interviewed. HIV prevalence rate among paid blood donors was higher (3.98%) than that of others (0.48%) while HIV prevalence rate was higher in plasma donors (7.24%) than that in both plasma and full blood donors (2.90%). There was no infections identified in full blood donors. Donors who donated blood during 1994 - 1995 had a higher prevalence rate (7.07%) than those who started donation after 1995 and those began donation before 1993 (0.99%). The rate of condom use was lower among paid blood donors (13.72%) than that of others. There was a lower rate of voluntary HIV testing among paid blood donors than that of others.

CONCLUSIONThe main high risk population of HIV infection were paid donors in CARES Project Areas, whose risk factor was plasma donation and were infected mainly before 1995. Both the rate of condom use and HIV testing were all very low, indicating that they knew little on HIV/AIDS and paid less attention to self-proctection. It is necessary to enhance HIV/AIDS health education and behavior intervention on paid blood donors.

Adult ; Blood Donors ; China ; epidemiology ; Female ; HIV Infections ; epidemiology ; HIV Seroprevalence ; HIV-1 ; Humans ; Incidence ; Male ; Middle Aged ; Plasma Exchange ; adverse effects ; Risk Factors ; Seroepidemiologic Studies