Objective To observe the effects of PTD-hepatitis B core antigen (HBcAg) induced murine bone marrow-derived dendritic cells (DCs) maturation on T lymphocyte proliferation in vitro.Methods Bone marrow derived DCs isolated from BALB/c mice were cultured with recombinant granu|ocyte-macrophage colony-stimulating factor (rGM-CSF) and recombinant interleutin-4 (rIL-4)for 5 days.Tumor necrosis factor (TNF)-a,HBcAg and PTD-HBcAg were added to induce DCs maturation.The distribution and localization of intracellular immunofluorescence were observed by confocal microscopy,and DCs phenotypes were analyzed by flow cytometry.The level of IL-12 p70 in the supernatant was detected by enzyme linked immunosorhent assay (ELISA).The proliferation of T lymphocytes was performed by using cell counting kit-8 (CCK-8).All data were analyzed using t test.Results DCs were cultured and identified successfully.Recombinant PTD-HBcAg could penetrate into DCs cytoplasm while recombinant HBcAg was detected on the surface of cells.DCs surface molecules,such as CD80,CD86 and major histocompability complex (MHC) II were upregulated by PTDHBcAg;IL-12 p70 levels induced by 50 mg/L and 100 mg/L recombinant PTD-HBcAg were (142.50±18.31) ng/L and (124.30±15.12) ng/L,respectively,which were significantly higher than those induced by recombinant HBcAg [(42.31±4.21 ) ng/L,t = 9.234 and 9.045,respectively,P＜0.05].The proliferation of T lymphocytes induced by PTD-HBcAg was much higher than that in HBcAg group or positive control TNF-a group.Conclusions PTD-HBcAg could penetrate membrane of DCs and promote the differentiation and maturation of DCs.PTD-HBcAg could up-regulate the expressions of costimulatory molecules on cell surface of DCs,and enhance the ability of DCs on stimulating T lymphocytes proliferation and IL-12 p70 production.

Objective To investigate the effect of hepatitis B virus core protein (HBc) dimer interfaces amino acids mutation on nucleocapsid assembly and HBV DNA replication. Methods Based on HBc three dimension structure, four HBc dimer interfaces domain mutation plasmids, pHBc14-18M,pHBc120-135M,pHBc23-39M and pHBc122-139M were constructed in pcDNA3.1 vector by PCR site-directed mutagenesis, there was a flag-tag at the C-terminal of all mutants for easy detection. Wild type core protein plasmid 1-183flag was also constructed as a positive control. The 4 mutants were cotransfected HepG2 cells with pHBV1.2 core negative plasmid (pHBV1.2-core-) ,which contained 1.2 copies of HBV whole genome but the core protein would not express due to a stop codon. The capsid formation, HBV pregenome(pgRNA) and HBV DNA replication mediate were analyzed by native agarose gel electrophoresis and Western blot, Northern blot and Southern blot , respectively. The 4 mutants were also cotransfected HepG2 cells with HBV wild type plasmid pHBV1.2 and examined by Southern blot. Virions in the medium were determined by native agarose gel electrophoresis and Western blot. Results Cotransfecting HepG2 cells with pHBV1.2-core- plasmid, pHBc14-18M,pHBc120-135M and pHBc122-139M mutant groups formed nucleocapsid-like structure but pHBc23-39M could not, Northern and Southern blot displayed no signal in all mutants except 1-183flag conrol group. In pHBV1.2 cotransfection experiment, HBV DNA replication was blocked in pHBc14-18M, pHBc120-135M and pHBc122-139M mutant groups, sharply decreased in pHBc120-135M and pHBc122-139M groups, correspondingly virons production in medium were also inhibited. pHBc23-39M mutant exerted no influence on HBV replication. Conclusion pHBc23-39M mutant can neither form nucleocapsid-like structure nor interact with wild type HBc dimmer to interfere HBV replication.On the contrast, pHBc14-18M, pHBc120-135M and pHBc122-139M mutants can form nucleocapsid-like structure by themselves, but this structure does not support HBV DNA synthesis. Besides, they can effectively inhibit wild type HBV DNA replication by contacting with wild HBc dimmers resulting in nucleocapsid dysfunction.

Objective To investigate the mortality risk factors of nosocomial infection patients in intensive care unit (ICU), and to guide clinicians to take effective control measures. Methods A retrospectively cohort study was conducted. The relevant information of patients with nosocomial infection treated in ICU of Hengshui Harrison International Peace Hospital Affiliated to Hebei Medical University from June 2009 to December 2015 was analyzed. The patients who admitted to ICU again, with length of ICU stay less than 48 hours, without first etiology of screening within 48 hours of ICU admission, or without complete pathogenic information were excluded. The gender, age, diagnosis, length of ICU stay, invasive operation, nutritional status, acute physiology and chronic health evaluation Ⅱ (APACHEⅡ) score, sequential organ failure assessment (SOFA) score, distribution and drug resistance of the pathogens, and procalcitonin (PCT) levels at 7 days after nosocomial infection were recorded. The risk factors leading to death in patients with nosocomial infection were analyzed by logistic regression, and the receiver operating characteristic curve (ROC) was drawn to evaluate the predictive value of all risk factors on the outcome of patients with nosocomial infection. Results In 864 enrolled patients with male of 54.75% and mean age of (63.50±15.80) years, 732 (84.72%) patients survived and 132 (15.28%) died. Compared with survivors, the non-survivors had higher age (years: 65.47±15.32 vs. 58.15±13.27), incidence of urgent trachea intubation (32.58% vs. 22.81%), deep venous catheterization (83.33% vs. 63.25%), and multiple drug-resistant infection (65.91% vs. 33.20%), longer length of ICU stay (days: 13.56±4.29 vs. 10.29±4.32) and duration of coma (days: 7.36±2.46 vs. 5.48±2.14), lower albumin (g/L: 23.64±8.47 vs. 26.36±12.84), higher APACHEⅡ score (19.28±5.16 vs. 17.56±5.62), SOFA score (8.55±1.34 vs. 6.43±2.65), and PCT (μg/L: 3.06±1.36 vs. 2.53±0.87, all P < 0.05). There was no significant difference in gender and urinary tract catheterization between survivors and non-survivors (both P > 0.05). The low respiratory tract was the most common site of infection followed by urinary tract and bloodstream in both groups. It was shown by logistic regression analysis that prolonged ICU stay [odds ratio (OR) = 2.039, 95% confidence interval (95%CI) = 1.231-3.473, P = 0.002], APACHEⅡ score (OR = 1.683, 95%CI= 1.002-9.376, P = 0.000), SOFA score (OR = 2.060, 95%CI = 1.208 -14.309, P = 0.041), PCT (OR = 2.090, 95%CI = 1.706-13.098, P = 0.004), and multi-drug resistant pathogens infection (OR = 5.245, 95%CI = 2.213-35.098, P = 0.027) were independent risk factors for ICU mortality in patients with nosocomial infection. The area under ROC curve (AUC) of length of ICU stay, APACHEⅡ score, SOFA score, and PCT level for predicting death of nosocomial infection patients was 0.854, 0.738, 0.786, and 0.849, respectively, the best cut-off value was 16.50 days, 22.45, 6.37 and 3.38 μg/L, respectively, the sensitivity was 83.6%, 90.0%, 81.1%, and 89.6%, and the specificity was 70.3%, 75.6%, 71.3%, and 85.4%, respectively. Conclusions Prol onged ICU stay, nosocomial infection with secondary sepsis and multiple organ dysfunction syndrome were the leading causes of death for nosocomial infection patients in ICU. Prolonged ICU stay, APACHE Ⅱ score, SOFA score, and PCT level could effectively predict death risks for nosocomial infection patients.

Objective To study the nucleotide and amino acid sequences of norovirus G Ⅱ.4/Sydney 2012 variants,in China.Methods Twenty-two stool specimens,confirmed as G Ⅱ.4/Sydney 2012-positive were collected from Beijing in the winter of 2012-2013.RT-PCR was performed to target the complete capsid gene.G Ⅱ.4/Sydney 2012 strains from other regions in China were searched and obtained from the GenBank.Nucleotide and amino acid sequences of G Ⅱ.4/Sydney 2012 strains were analyzed,using the CLUSTAL X (Version 1.83)and followed by phylogenetic analysis using Mega version 5.1.Results The complete major capsid nucleotide sequences of thirty-eight G Ⅱ.4/Sydney 2012 strains from seven regions in China were obtained.The VP1 nucleotide and amino acid sequences diversity were 0.1％-3.3％ and 0-3.1％,respectively.Result from phylogenetic analysis demonstrated that the G Ⅱ.4/Sydney 2012 variant shared a common ancestor with both the dominant norovirus G Ⅱ.4 variants Apeldoom 2008 and the New Orleans 2009.G Ⅱ.4/Sydney 2012 variants appeared to have had two A/D/E site combinations at the amino acid level,TSRN-GTT-SNT and TSRN-STT-SNT.Conclusion G Ⅱ.4/Sydney 2012 variant had been circulating in many regions in China.There seemed a high nucleotide and amino acid identity among G Ⅱ.4/Sydney 2012 strains collected from China.G Ⅱ.4/Sydney 2012 variants showed different A/D/E site combination from other pandemic G Ⅱ.4 variants.

Objective To specifically cleave the intracellular HBV DNA using the clustered regularly interspaced short palindromic repeat ( CRISPR )/Cas9 genome editing technology. Methods Using of Lenti CRISPRv2 system, we designed 3 highly conserved guide RNAs ( gRNAs ) to specifically cleave HBV genome.The cleavage efficacy of CRISPR/Cas9 targeting at HBV DNA in plasmids, the episomal HBV DNA or integrated HBV DNA in chromosome was evaluated in the cell cultures of HepG2.2.15 and HepDES19, and in a model of C57BL/6 mouse infected with AAV-HBV1.3.Results Viral proteins were inhibited by 30% -50% by a single site CRISPR/Cas9 cleavagein the HepG2 cells transiently co-transfected with HBV DNA, multiple sites of cleavage significantly elevated the efficiency to 70%-90%.Compared with that of HepG2.2.15 cells, HBeAg in HepDES19 cells was inhibitedby at least three-fold higher.Conclusion Multiple sites cleavage significantly improve the efficacy of CRISPR/Cas9 systems,and this system is more efficient to cleave the extra-chromosome HBV DNA than the integrated HBV DNA in chromosomes.

Objective : To explore the clinical effect of personalized titanium mesh combined with free flap in the repair of maxillary defect. Methods: 36 cases of maxillary defect patients as the research object were selected in our hospital during May of 2010 to May of 2016. 36 defect cases were repaired with personalized titanium mesh combined with free flap, and summarize the treatment programs to explore the value of clinical application. Results : By the end of follow-up, all of the flap survived, tumor recurrence rate was 5.56% with paitient satisfaction was 100%; Diplopia and dysphagia occurred in no cases; Masticatory function declined accompanied with a longer chewing time but language communication was not affected. Conclusion The application of personalized titanium mesh and free flap repair methods in maxillary defect is significant, which effectively improve the quality of life of patients whereas still have difficulties in late denture at the same time.

Psoraleae Fructus (PF) is an important traditional herbal medicine with a long history of clinical application. It is widely used to treat various diseases, such as osteoporosis, leucoderma and diarrhea. As a traditional nontoxic herb, it has aroused worldwide concern about the potential risks due to increasing adverse reaction events. This article reviews the botany, ancient records of medical uses, adverse reactions, toxicological research advance and detoxification methods of PF. According to clinical studies, liver injury is the most predominant in PF-related adverse reactions. The underlying mechanisms include bile acid metabolism and transport disorders, oxidative stress, mitochondrial damage, inhibition of liver cell regeneration and inflammatory reactions. Furthermore, the potential toxins of PF are summarized. Traditional methods of processing and compatibility will provide reference for reducing the toxicity of PF, which requires further research. In sum, this work systematically summarizes the reserach progress on the safety of PF, which will provide comprehensive insights into the toxicity of PF and facilitate its safe use and future development.
Drugs, Chinese Herbal/toxicity* ; Fruit/toxicity* ; Liver

OBJECTIVES: To explore the effects of hypoxic and hypobaric conditions on blood gas and erythrocyte-related indicators in rats. METHODS: SD male rats were exposed to low-pressure hypoxic conditions simulating an altitude of 6500 m in a small or a large experimental cabin. Abdominal aortic blood samples were collected and blood gas indicators, red blood cells (RBCs) count, and hemoglobin (Hb) content were measured. The effects of exposure to different hypoxia times, different hypoxia modes, normal oxygen recovery after hypoxia, and re-hypoxia after hypoxia preconditioning on blood gas indicators, RBCs count and Hb content were investigated. RESULTS: The effect of blood gas indicators was correlated with the length of exposure time of hypoxia and the reoxygenation after leaving the cabin. Hypoxia caused acid-base imbalance and its severity was associated with the duration of hypoxia; hypoxia also led to an increase in RBCs count and Hb content, and the increase was also related to the time exposed to hypoxia. The effects of reoxygenation on acid-base imbalance in rats caged in a small animal cabin were more severe that those in a large experimental cabin. Acetazolamide alleviated the effects of reoxygenation after leaving the cabin. Different hypoxia modes and administration of acetazolamide had little effect on RBCs count and Hb content. Normal oxygen recovery can alleviate the reoxygenation and acid-base imbalance of hypoxic rats after leaving the cabin and improve the increase in red blood cell and hemoglobin content caused by hypoxia. The improvement of hypoxia preconditioning on post hypoxia reoxygenation is not significant, but it can alleviate the acid-base imbalance caused by hypoxia in rats and to some extent improve the increase in red blood cell and hemoglobin content caused by hypoxia. CONCLUSIONS Due to excessive ventilation and elevated RBCs count and Hb content after hypoxia reoxygenation, oxygen partial pressure and other oxygenation indicators in hypoxic rats are prone to become abnormal, while blood gas acid-base balance indicators are relatively stable, which are more suitable for evaluating the degree of hypoxia injury and related pharmacological effects in rats.
Rats ; Animals ; Male ; Acetazolamide ; Hypoxia ; Oxygen ; Erythrocytes ; Hemoglobins ; Acid-Base Imbalance