To investigate a baculovirus insect cell system for expressing an interferon alpha 2b (IFNa2b)/immunoglobulin G-4 (IgG4) Fc fusion protein, which has long-acting antiviral effects. Human IFNa2b and IgG4 Fc cDNAs were generated by molecular cloning and inserted into a baculovirus shuttle vector, which was then transposed into the DH10 Bac strain to form recombinant Bacmid-IFN/Fc. The Bacmid-IFN/Fc was transfected into High five insect cells, and expression of the IFN/Fc fusion protein was detected by Western blotting and its biological activity was assessed by the cytopathic effect inhibition method. The IFNa2b and IgG4 Fc cDNA fragments were successfully amplified by RT-PCR using human peripheral lymphocytes. After cloning into the baculovirus shuttle vector, pFastBac1, and transforming into DH10 Bac competent cells, screening identified positive clones carrying the recombinant Bacmid-IFN/Fc. A Bacmid-IFN/Fc clone was successfully transfected into the High five insect cells and packaged into the baculovirus for expression of the IFN/Fc fusion protein. Western blotting revealed that the fusion protein expression was specific, and yielded a protein of 45 kD in size. The in vitro antiviral activity of the IFN/Fc fusion protein was 580 IU/mL. A novel IFN/Fc fusion protein was successfully generated using a baculovirus insect cell system, which may prove useful for providing future experimental data for development of a new long-acting interferon to treat chronic viral hepatitis.
Animals ; Antiviral Agents ; metabolism ; Baculoviridae ; genetics ; Cell Line ; Cloning, Molecular ; Gene Expression ; Gene Fusion ; Genetic Vectors ; Humans ; Immunoglobulin Fc Fragments ; biosynthesis ; genetics ; Immunoglobulin G ; biosynthesis ; genetics ; Insecta ; Interferon-alpha ; biosynthesis ; genetics ; Recombinant Fusion Proteins ; biosynthesis ; genetics ; Recombinant Proteins ; biosynthesis ; genetics ; Reverse Transcriptase Polymerase Chain Reaction ; Transfection

Adaptor Proteins, Signal Transducing ; metabolism ; Carcinoma, Hepatocellular ; metabolism ; pathology ; Case-Control Studies ; Humans ; Liver Neoplasms ; metabolism ; pathology ; Membrane Proteins ; metabolism ; RNA, Messenger ; genetics

OBJECTIVETo investigate the distribution of hepatitis delta virus (HDV) marker among hepatitis B virus (HBV) infected patients and to reveal its clinical significance.

METHODTo collect the clinical data and sera samples of HBV infected patients and to detect HDAg, Anti-HDV as well as HBV infection markers by means of enzyme-linked immunosorbnent assay. These data combined with clinical diagnostic results and biochemical index were then analyzed.

RESULT462 samples of HBV infected patients were collected including 210 HBV carriers without symptom, 175 chronic HBV infections, 35 acute HBV infections and 42 liver fibrosis. The HDV infection rate was 4.8% overall. The highest infection rate of 9.5% was found in the group of liver fibrosis whereas the lower rate of 6.9% was found in HBV chronic carriers. HDV infection rate was 7.8% among the population of 40-60 years old, obviously higher than any other age groups.

CONCLUSIONHDV infection was significantly higher in the chronic HBV patients and liver fibrosis patients. Because HDV infection was highly associated with the progress of liver disease, we suggest the screen of HDV markers among hepatitis patients and discriminate whether the patient was co-infected with HDV.

Adolescent ; Adult ; Aged ; Biomarkers ; blood ; Child ; Coinfection ; blood ; diagnosis ; immunology ; virology ; Female ; Hepatitis Antibodies ; immunology ; Hepatitis B ; blood ; diagnosis ; immunology ; virology ; Hepatitis B Antigens ; blood ; Hepatitis B virus ; immunology ; isolation & purification ; physiology ; Hepatitis D ; blood ; diagnosis ; immunology ; virology ; Hepatitis Delta Virus ; immunology ; isolation & purification ; Humans ; Male ; Middle Aged ; Young Adult

OBJECTIVE: To investigate the incidence of severe neonatal hyperbilirubinemia and the management on the treatment and follow-up of this disease in Jiangsu Province, China. METHODS: The neonates with severe hyperbilirubinemia who were admitted to 13 hospitals in Jiangsu Province from January to December, 2018, were enrolled as subjects. A retrospective analysis was performed on their mediacal data and follow-up data. RESULTS: In 2018, 740 neonates with severe hyperbilirubinemia were reported from the 13 hospitals in Jiangsu Province, accounting for 2.70% (740/27 386) of the total number of neonates admitted to the department of neonatology. Among these neonates, 620 (83.8%) had severe hyperbilirubinemia, 106 (14.3%) had extremely severe hyperbilirubinemia, and 14 (1.9%) had hazardous hyperbilirubinemia. Four neonates (0.5%) were diagnosed with acute bilirubin encephalopathy. A total of 484 neonates (65.4%) were readmitted due to severe hyperbilirubinemia after discharge from the delivery institution, with a median age of 7 days, among whom 214 (44.2%) were followed up for jaundice at the outpatient service before readmission, with a median age of 6 days at the first time of outpatient examination. During hospitalization, 211 neonates (28.5%) underwent cranial MRI examinations, among whom 85 (40.3%) had high T1WI signal in the bilateral basal ganglia and the globus pallidus; 238 neonates (32.2%) underwent brainstem auditory evoked potential examinations, among whom 14 (5.9%) passed only at one side and 7 (2.9%) failed at both sides. The 17 neonates with acute bilirubin encephalopathy or hazardous hyperbilirubinemia were followed up. Except one neonate was lost to follow-up, and there were no abnormal neurological symptoms in the other neonates. CONCLUSIONS Neonates with severe hyperbilirubinemia account for a relatively high proportion of the total number of neonates in the department of neonatology. Jaundice monitoring and management after discharge from delivery institutions need to be strengthened. For neonates with severe hyperbilirubinemia, relevant examinations should be carried out more comprehensively during hospitalization and these neonates should be followed up comprehensively and systematically after discharge.
Bilirubin ; China ; Evoked Potentials, Auditory, Brain Stem ; Humans ; Hyperbilirubinemia, Neonatal ; Infant, Newborn ; Retrospective Studies

This study was purposed to evaluate the effects of recombinant human granulocyte colony-stimulating factor (rhG-CSF) on hematopoietic reconstruction and survival in beagles exposed to mixed fission neutron and γ-ray. 13 beagles were unilaterally exposed to single dose of 2.3 Gy 90% neutrons. The experiments were divided into 3 groups: irradiation control group (no any treatment, n = 4), supportive care group (n = 5) and rhG-CSF plus supportive care group (n = 4, abbreviated as rhG-CSF group) in which the beagles were subcutaneously injected with 200 µg/kg of rhG-CSF early at half an hour and 24 hours post-irradiation respectively. The results showed that 2.3 Gy 90% neutron irradiation induced a severe acute radiation sickness of bone marrow type. The administration of rhG-CSF increased the survival rate from 60% in supportive care group to 100%. Twice injection of rhG-CSF in the first 24 hours reduced duration of neutropenia, enhanced neutrophil nadir and promoted neutrophil recovery when compared with control cohort administered clinical support. The number of colony-forming cells (CFU-GM, CFU-E, and BFU-E) in peripheral blood of rhG-CSF treated canines increased 2-to 5-fold relative to those of the supportive care group on day 3. All canines treated with rhG-CSF achieved hematopoietic reconstruction as evidenced by the pathological section of sternum while severe shortage of hemopoietic cells remained in the cohorts given supportive care alone. It is concluded that the combination of supportive care and high-dose rhG-CSF can accelerate hematopoietic recovery and enhance survival of dogs exposed to 2.3 Gy mixed neutron and gamma ray.
Animals ; Dogs ; Gamma Rays ; adverse effects ; Granulocyte Colony-Stimulating Factor ; administration & dosage ; pharmacology ; Hematopoietic System ; drug effects ; radiation effects ; Neutron Diffraction ; Recombinant Proteins ; administration & dosage ; pharmacology ; Survival Rate

The aim of this study was to investigate the effect of recombinant human granulocyte stimulating factor (rhG-CSF) on blood coagulation of beagles irradiated by 2.3 Gy neutron so as to provide new therapy for blood coagulation disorder after neutron irradiation. 10 beagles were exposed to 2.3 Gy neutron, and then randomly assigned into supportive care group and rhG-CSF-treated group. The rhG-CSF-treated cohorts were injected subcutaneously with rhG-CSF (10 µg/kg·d) beginning at the day of exposure for 21 consecutive days. Peripheral blood platelet counts were examined once every two days. In vitro platelet aggregation test, thromboelastography and blood clotting tetrachoric tests were also performed. The results indicated that the blood clotting system of irradiated dogs was in hypercoagulable state in the early days after 2.3 Gy neutron irradiation, and became hypocoagulable at crisis later and were mainly on intrinsic coagulation pathway. Blood fibrinogen increased markedly during the course of disease, while platelet counts and aggregation function were decreased remarkably. rhG-CSF administered daily could correct hypercoagulable state induced by 2.3 Gy neutron irradiation at the early time post exposure, shortened the thromboplastin generation time and clotting formation, down-regulated the abnormal high fibrinogen in blood, and improved platelet aggregation function. It is concluded that rhG-CSF can improve coagulation disorders of irradiated dogs.
Animals ; Blood Coagulation ; drug effects ; Bone Marrow ; radiation effects ; Dogs ; Granulocyte Colony-Stimulating Factor ; pharmacology ; therapeutic use ; Humans ; Leukocyte Count ; Neutron Diffraction ; Platelet Count ; Radiation Dosage ; Radiation Injuries, Experimental ; physiopathology ; Recombinant Proteins

Objective To examine the relationship between serum vitamin D level and immune imbalance in advanced schistosomiasis patients with liver fibrosis. Methods A total of 120 advanced schistosomiasis patients with liver fibrosis that were admitted to the Department of Schistosomiasis of The First Hospital of Jiaxing City from May 2016 to September 2018 were recruited as the observation group, and 50 healthy volunteers randomly sampled from the hospital during the same period served as the control group. The serum IgG antibody, IgA antibody, C3 complement, C4 complement, CD4⁺ cell proportion, CD8⁺ cell proportion, 25-hydroxyvitamin D [25(OH)D] levels were compared between the two groups. Liver fibrosis was classified into grade I, II and III according to the classification criteria of liver fibrosis by ultrasonography, and the serum IgG antibody, IgA antibody, C3 complement, C4 complement, CD4⁺ proportion, CD8⁺ proportion, 25(OH)D levels were compared among patients with grade I, II and III liver fibrosis. In addition, all patients were classified into the sufficient group, the insufficient group and the deficient group according to the serum vitamin D level, and the serum IgG antibody, IgA antibody, C3 complement, C4 complement, CD4⁺ proportion, CD8⁺ proportion, 25(OH)D levels were compared among these three groups. Moreover, the associations of the serum vitamin D level with these immune indicators were examined. Results The 120 advanced schistosomiasis patients with liver fibrosis included 58 men and 62 women, and had a mean age of (72.00 ± 3.00) years. There were 32 cases with grade I liver fibrosis, 46 cases with grade II liver fibrosis, and 42 cases with grade III liver fibrosis. There were no significant differences between the observation group and the control group in terms of serum D-dimer, total cholesterol (TC), triglyceride (TG), C3 complement or C4 complement levels (t = 2.467, 0.322, 0.790, -2.432 and -2.630, all P values > 0.05); however, there were significant differences seen in alanine aminotransferase (ALT), aspartate aminotransferase (AST), blood calcium, blood phosphorus, IgG antibody, IgA antibody, CD4⁺ proportion, CD8⁺ proportion, and 25(OH)D levels (t = 5.130, 6.382, -1.341, 2.361, 8.708, 11.783, -2.995, -6.543 and -3.022, all P values < 0.05). In addition, there were significant differences in AST, ALT, blood phosphorus, IgA antibody, C3 complement, CD8⁺ cell proportion and 25(OH)D levels among patients with grades I, II and III liver fibrosis (F = 19.704, 16.254, 62.669, 49.347, 5.430, 5.434 and 5.783, all P values < 0.05). There were significant differences in ALT, blood phosphorus, IgA antibody, CD8⁺ cell proportion and 25(OH)D levels between patients with grades I and III liver fibrosis (all P values < 0.05), and significant differences were seen between patients with grades II and III liver fibrosis in terms of blood phosphorus, IgA antibody and CD8⁺ cell proportion (all P values < 0.05), while there was a significant difference in the CD8⁺ cell proportion between patients with grades I and II liver fibrosis (P < 0.05). Moreover, there were significant differences among the sufficient, insufficient and deficient groups in terms of IgG antibody, IgA antibody, C3 complement, CD4⁺ cell proportion and CD8⁺ cell proportion (F = 13.303, 59.623, 8.698, 9.969 and 12.805, all P values < 0.05), and there was a significant difference in the CD8⁺ cell proportion between the insufficient and deficient groups (P < 0.05). Pearson correlation analysis revealed that serum 25(OH)D level were negatively associated with IgG and IgA antibody levels (r = -0.754 and -0.773, both P values < 0.05), and positively associated with C3 complement, CD4⁺ cell proportion and CD8⁺ cell proportion in advanced schistosomiasis patients with liver fibrosis (r = 0.827, 0.850 and 0.830, all P values < 0.05). Conclusion Immune imbalance occurs in advanced schistosomiasis patients with liver fibrosis, and serum vitamin D level may correlate with immune imbalance in advanced schistosomiasis patients with liver fibrosis.

OBJECTIVE: To analyze and characterize the separation effectiveness of self-constructed asymmetrical flow field-flow fractionation system on proteins and lipoproteins, to achieve the optimization of the experimental conditions when separating lipoproteins by orthogonal design test and to investigate the carrier viscosity's influence on separation effectiveness. METHODS: The evaluation of asymmetrical flow field-flow fractionation separation capacity was conducted by using two standard proteins (carbonic anhydrase and thyroglobulin). Under the optimized separation conditions of carbonic anhydrase and thyroglobulin, the channel actual thickness (after assembling, the actual thickness of separation channel was less than initial thickness) was calculated by the analytes' elution time based on the hydrokinetic theory. With orthogonal design test the optimized experimental conditions were studied and statistical analysis was carried on to find out the factors with statistical significance which needed further exploration. RESULTS: According to the hydrodynamics principle and Stoke's function, the channel actual thickness was measured to be 164 μm by separating the two standard proteins, carbonic anhydrase and thyroglobulin, under proper experimental conditions. By the optimization based on orthogonal design test, base-line separation (the resolution had to be higher than 1.50) was achieved. The resolutions of the two experiments were 1.61 and 1.58. According to previous study/ pre-study and supporting theory, in the orthogonal design test, the total 5 factors were integrated for comprehensive investigation: the total flow rate (3.00, 3.50, 4.00, 4.50 mL/min), focus time (3.00, 3.50, 4.00, 4.50 min), transition time (0.5, 1.0, 1.5, 2.0 min), pH of the carrier fluid(6.8, 7.00, 7.20, 7.40) and viscosity of the carrier fluid hydroxypropylmethylcellulose concentration: 0.00%, 0.03%, 0.06%, 1.00%). Among the 5 factors, viscosity was found to have the statistical significance on separation effectiveness which was further investigated. The resolution of high density lipoprotein and low density lipoprotein was increased by the increasing viscosity which also caused more obvious negative spikes. CONCLUSION The separating capacities of self-constructed asymmetrical flow field-flow fractionation system on lipoproteins were verified to be effective and an optimized experimental condition was found to achieve the base-line separation of high density lipoprotein and low density lipoprotein. Viscosity of the carrier fluid was proved to have the statistical significance on lipoprotein separation.
Fractionation, Field Flow ; Lipoproteins ; Lipoproteins, LDL

OBJECTIVE: To investigate the clinical manifestations pathologic features, treatment options and prognosis of patients with bone lymphoma. METHODS: The clinical characteristics, pathologic features, treatment and prognosis of 34 BL patients diagnosed by histopathologic method or/and PET-CT and treated in first hospital of peking university from January 2004 to April 2018 were analyzed retrospectively. RESULTS: The median age of 34 BL patients was 56 years old, the male and female ratio was 1.43∶1 (24 /10). Among 34 patients, the patients with primary bone lymphoma(PBL) were 8 cases, the patients with secondary bone lymphoma(SBL) was 26 cases, the PBL and SBL ratio was 0.31∶1. Bone lymphoma lacks typical systemic symptoms, and its onset began mostly from bone pain and pathologic bone fracture. The most frequent pathological type of bone lymphoma in our study was diffuse large B-cell lymphoma (DLBCL), accounting for 55.88%. At present, the conventional treatment for bone lymphoma includes chemotherapy, or chemotherapy combined with radiotherapy and surgery, as well as hematopoietic stem cell transplantation. The average and median OS time of BL patients were 349 years and 3 years respectively, meanwhile the OS rate for three years and two years were 56.25% and 78.16%, respectively. Factors that affect survival of BL patients were PBL and SBL classification, pathological type, blood LDH level, and treatment methods. CONCLUSION Bone lymphoma is usually concealed onset，an adequate and adequate combination therapy can improve the survival rate and transplantation therapy plays an important role. Primary bone lymphoma is rare, the prognosis of patients with primary bone lymphoma is good, whereas the prognosis of patients with secondary bone lymphoma is poor.
Bone Neoplasms ; Female ; Hematopoietic Stem Cell Transplantation ; Humans ; Lymphoma, Large B-Cell, Diffuse ; Male ; Middle Aged ; Positron Emission Tomography Computed Tomography ; Prognosis ; Retrospective Studies

BACKGROUND: Prospective analyses have yet to identify a consistent relationship between sleep duration and the incidence of gastrointestinal (GI) cancers. The effect of changes in sleep duration on GI cancer incidence has scarcely been studied. Therefore, we aimed to examine the association between baseline sleep duration and annual changes in sleep duration and GI cancer risk in a large population-based cohort study. METHODS: A total of 123,495 participants with baseline information and 83,511 participants with annual changes in sleep duration information were prospectively observed from 2006 to 2015 for cancer incidence. Cox proportional-hazards models were used to calculate hazard ratios (HRs) and their confidence intervals (CIs) for GI cancers according to sleep duration and annual changes in sleep duration. RESULTS: In baseline sleep duration analyses, short sleep duration (≤5 h) was significantly associated with a lower risk of GI cancer in females (HR: 0.31, 95% CI: 0.10-0.90), and a linear relationship between baseline sleep duration and GI cancer was observed (P = 0.010), especially in males and in the >50-year-old group. In the annual changes in sleep duration analyses, with stable category (0 to -15 min/year) as the control group, decreased sleep duration (≤-15 min/year) was significantly associated with the development of GI cancer (HR: 1.29; 95% CI: 1.04-1.61), especially in the >50-year-old group (HR: 1.32; 95% CI: 1.01-1.71), and increased sleep duration (>0 min/year) was significantly associated with GI cancer in females (HR: 2.89; 95% CI: 1.14-7.30). CONCLUSIONS Both sleep duration and annual changes in sleep duration were associated with the incidence of GI cancer.
Cohort Studies ; Female ; Gastrointestinal Neoplasms/etiology* ; Humans ; Incidence ; Male ; Middle Aged ; Proportional Hazards Models ; Prospective Studies ; Risk Factors ; Sleep