China is among the middle-high endemic regions of HBV infection.The pathological outcomes of chronic HBV infection have been shown to be greatly influenced by several important factors,including HBV genotype,sub-genotype and gene viability mutation.HBV genome mutation,on the one hand,could alter its replication and secretion and thus change viral pathogenicity.In addition,host immune microenvironment and host-virus interaction,disease progression and the effect of antiviral therapy could be adapted at the same time.The detection of HBV genotypes,genetic subtypes and the key hotspot mutation is helpful to clinical risk assessment and prognosis prediction of HBV-related end-stage liver diseases (cirrhosis and hepatocellular carcinoma),it is also helpful to auxiliary predict the liver diseases recurrence and metastasis after treatment.Thus persistent care should be taken on the HBV mutation and its clinical translation so as to provide solid evidences for the personalized,standardized and fine management of HBV-related liver diseases.

The natural history of chronic HBV infection is diverse and variable, ranging from inactive carriers to progressive chronic hepatitis B ( CHB), cirrhosis and hepatocelluar carcinoma.It is estimated that 93 million people are chronically infected with HBV and 20 million cases suffering from chronic hepatitis B in China.Hepatocelluar carcinoma has been the second leading cause of death for male in China.Liver cirrhosis and HCC which have high mortality and morbidity have become the heavy burden for the limited medication resource of China.Here the current clinical applications and consensus progression based on antigen and nuclear acid detection were acknowledged.The reasonable application as well as appropriate clinical interpretation are emphasized indicating that laboratory medicine practitioners should be more actively involved in clinical diagnosis and treatment.More efforts and contributions should be made by the laboratory medicine practitioner for optimizing clinical management of HBV-related diseases in future.

Objective To investigate the oxidative mechanism of homocysteine ( Hey) -induced apoptosis in endothelial progenitor cells( EPCs). Methods Total mononuclear cells were isolated from mouse bone marrow by Ficoll density gradient centrifugation and were cultured in vitro for 7 d. Adherent cells were harvested and identified by fluorescence microscopy. EPCs were cultured with Hey (0, 50, 100 and 500 μmol/L) for 12, 24 and 48 h, or pretreated with NAC (1 mmol/L), DPI( 10 μmol/L) or SB203580 (10 μmol/L) for 30 min, then cultured with 500 μmol/L Hey for 24 h. Apoptosis was detected by Annexin-V/PI flow cytometry, levels of reactive oxygen species (ROS) in cells were measured using H2DCF-DA as a fluorescence probe, NADPH oxidases were evaluated with lucigenin-enhanced chemilumine9cence, and NO in the supernatant was determined by nitrate reductase assay. Results Hey induced EPCs apoptosis, ROS accumulation, NADPH oxidase activation and decrease of NO in a time-dose dependent manner( P <0.05 or P < 0.01). Pretreatment with NAC, DPI and SB203580 could inhibit these effects (P < 0.05 or P < 0.01). Conclusion Hey could activate NADPH oxidase, induce ROS increase and NO decrease, and activate p38MAPK to enhance EPCs apoptosis.

Objective To observe the expression and distribution of IL 10 in local vessel after balloon injury in rat and study its significance in the response of artery injury Methods RT PCR and Western Blot as well as immunohistochemical method were used to assay the changes of IL 10′s mRNA as well as expression and distribution of protein Results There is no expression of IL 10 in vascular wall of normal rat After Balloon injury, the level of IL 10 mRNA and product of IL 10 was up regulated, and immunohistochemiscal staining showed immunoreactive IL 10 mainly in smooth muscle cells Conclusion IL 10 is expressed in balloon injured aorta and may contribute to the modulation of the local inflammatory response

To evaluate the feasibility of using polyethyleneimine (PEI) coated magnetic iron.oxide nanoparticles (polyMAG-1000) as gene vectors. The surface characteristics of the nanoparticles were observed with scanning electron microscopy. The ability of the nanoparticles to combine with and protect DNA was investigated at different PH values after polyMAG-1000 and DNA were combined in different ratios. The nanoparticles were tested as gene vectors with in vitro transfection models. Under the scanning electron microscope the nanoparticles were about 100 nm in diameter. The nanoparticles could bind and condense DNA under acid, neutral and alkaline conditions, and they could transfer genes into cells and express green fluorescent proteins (GFP). The transfection efficiency was highest (51%) when the ratio of nanoparticles to DNA was 1:1 (v:w). In that ratio, the difference in transfection efficiency was marked depending on whether a magnetic field was present or not: about 10% when it was absent but 51% when it was present. The magnetic iron oxide nanoparticles coated with PEI may potentially be used as gene vectors.
Breast Neoplasms/metabolism ; Breast Neoplasms/*pathology ; Cell Line, Tumor ; Ferric Compounds/*chemistry ; Ferric Compounds/metabolism ; Gene Targeting ; Genetic Vectors ; Green Fluorescent Proteins ; Magnetics ; Nanotechnology ; Particle Size ; Polyethyleneimine/chemistry ; Transfection/methods

The aim of this study was to evaluate the feasibility and efficacy of using TRAIL gene to treat breast cancer mediated with a novel carrier - magnetic iron oxide nanoparticles (poly-MAG-1000) coated with PEI. The magnetic iron oxide nanoparticles were used as gene carrier to transfect TRAIL gene into MCF-7 cells. The polyMAG-1000 without TRAIL gene was transfected into the tumor cells as negative control. TRAIL gene transfection with liposome as carrier served as positive control. The apoptosis of cells was detected with TUNEL method. The apoptosis ratio of tumor cells was measured with flow cytometry (FCM). It was found that the apoptosis occurred in the tumor cells after transfection of TRAIL gene mediated by both polyMAG-1000 and liposome. The apoptosis ratio in the group with polyMAG-1000 as gene carrier was (25.11+/-2.85) %, whereas it was (5.06+/- 1.05) % in the control group with polyMAG-1000 (P<0.01). The apoptosis ratio was as low as (18.31+/-2.44) % in the group with liposome as gene carrier (P<0.05, as compared with the group with polyMAG-1000 as gene carrier). It is suggested that TRAIL gene may induce apoptosis in MCF-7 breast cancer cells. The magnetic iron oxide nanoparticles coated with PEI may be a potential gene carrier with high transfection efficacy for cancer gene therapy..

Objective To explore the relationship between type 2 diabetes mellitus(T2DM)and thyroid diseases by analyzing the abnormal status of thyroid hormone and antibodies in T2DM patients and the healthy control group.Methods The serum thyroid stimulating hormone (TSH),free triiodothyronine (FT3),free thyroxine (FT4),anti-thyroglobulin antibody (TGAb)and anti -thyroid peroxidase antibody (TPOAb)were measured in 396 patients with T2DM and 411 healthy persons.Results In the healthy group,the incidence rate of abnormal thyroid hormone was 7.5%,in which the male was 5.5% and female was 9.4%,the incidence rate of subclinical hypothyroidism in females was 4.7%,which was higher than 1.5% in males,there was statistically significant difference between them(P <0.05);the positive rate of TPOAb in the healthy control group was 10.2% and the positive rate of TGAb was 6.6%;in T2DM patients,the incidence rate of thyroid hormone abnormalities was 16.2%,which in males and females were 12.3% and 20.5% respectively,among them the incidence rate of subclinical hypothyroidism in females was 9.2%,which was significantly higher than 4.3% in males,the difference had statistical significance (P <0.01),T2DM patients with subclinical hypo-thyroidism accounted for 40.6% of thyroid dysfunction incidence rate;the positive rate of TPOAb was 15.2% and the positive rate of TGAb was 7.1 %.Conclusion The incidence rate of thyroid diseases in T2DM patients is significantly increased compared with the healthy control group,subclinical hypothyroidism in females is predominant;screening thyroid autoantibodies and monitoring FT3,FT4 and TSH at regular intervals in T2DM have the important clinical significance for evaluating the disease condition,jud-ging prognosis and guiding treatment.

Objective To observe the difference of lung injury between toll-like receptor 4 (TLR4) mutant mice and wild type mice in a model of lipopolysaccharide (LPS) induced acute lung injury (ALI),discuss the role of TLR4 in ALI.Methods Different doses of LPS solution (1,5mg?kg~(-1)) were injected in vein tail to reproduce ALI model in both TLR4 mutant (TLR4~(-/-)) and wild type (WT,TLR4~+(/+)) mice.Lung tissues were collected for gross and micrographic histological injury analysis and for assessment of lung edema.Meanwhile,the levels of myeloperoxidase (MPO) in lung tissues in both strains were assessed to evaluate the extent of polymorphological neutrophils (PMN) infiltration.Results The gross and micrographic injury of lung was milder in TLR4 mutant mice than that in wild type mice.The extent of lung edema (W/D) was also reduced compared with wild type mice,especially in 5 mg?kg~(-1) group [(4.08?0.1)vs.(4.55+0.2),n=10,t=12.71,P＜0.01].With high dosage of LPS,the value of W/D in both mice strains was higher than that in sham operation group (P＜0.01).The extent of PMN infiltration in lung tissues in TLR4 mutant mice was reduced compared with wild type mice.But they were higher than sham operated mice (P＜0.01).Conclusion TLR4 May involve in the development of ALI,by sequestration of PMN into lung tissues.

BACKGROUND: Inflammatory and proliferating effect after mechanical injury of vascular wall is the major cause of restenosis. Nuclear factor-kappa B (NF-κB) in the NF-κB/Rel family is expressed in a variety of cell types and activates a series of target genes, which are related to the pathophysioiogy of vascular wall.OBJECTIVE: To investigate the effect of antisense and decoy NF-κB oligonucleotides on vascular smooth muscle cell (VSMS) proliferation in vitro and neointimal proliferation and monocyte chemotactic protein-1 (MCP-1) in the balloon-injured carotid artery of rats.DESTGN: Randomized controlled animal trial.SETTTNG: Department of Cardiology, Ruijin Hospital, Medical College, Shanghai Jiaotong University.MATERTALS: Totally 126 male Sprague-Dawley (SD) rats, aged 3 months, weighing 350 to 380 g, were involved in this study. Synthesis of primer and oligonucleotide: they were synthesized and designed by Shanghai Bioengineering Co. Ltd according to literatures and international internet cDNA library.METHODS: This study was carried out in the Laboratory of Cell Biology, Medical College, Shanghai Jiao Tong University and Cardiovascular Laboratory, Ruijin Hospital Affiliated to Shanghai Jiaotong University from May 2001 to March 2003.Rat aortic smooth muscle cells were isolated from May 2001 to March 2003. Rat thoracic aorta vascular smooth muscle cells were cultured by primary-explant method. And the third to fifth generations of VSMCs were involved in the experiment. Proliferating cell nuclear antigen (PCNA) NF-κB p65 protein synthesis in proliferating smooth muscle cells were detected. SD rat carotid artery underwent balloon injury. The involved 126 rats were randomly divided into 7 groups with 18in each group: normal group: normal group (the procedure was the same as other group except for balloon injury), sense group, antisense group, decoy group, scramble group, antisense plus decoy group, model group. Each group includes 6time points (6 hours, and 1,3,5,7,14 days, n =3). Then, the effect of antisense and decoy NF-κB oligonucleotides on intimai proliferation and MCP-1 and NF-κB p65 and extracellular signal regulated kinase(ERK2) expression in the balloon-injured carotid artery of rats were detected.MAIN OUTCOME MEASURES: ①Effect of oligonucleotide of NF-κB p65 on VSMCs proliferation; ② NF-κB p65 gene expression and protein synthesis; ③ Patho-morphological change after carotid balloon-injury. ④ Vascular MCP-1 mRNA Expression in balloon-injured rat carotid artery; ⑤ MCP-1 immunoreactivity in the injured arterial wall detected by immunohistochemistry; ⑥ NF-κBp65 and ERK2 protein synthesis after balloon-injury detected by Western blot in injured rat carotid arteries.RESULTS: ①PCNA protein synthesis increased in proliferating smooth muscle cells. ②NF-κB p65 gene expression was found in the cytoplasm and nucleus of proliferating smooth musclecells by in situ hybridization and NF-κB p 65 protein level increased in proliferating smooth muscle cells by flow cytometry. NF-κB p65 gene expression in antisense group decreased 53.66% compared with in sense group; it decreased 57.35% in decoy group compared with in scramble group. There were all statistical differences(P＜0.05).③ PCNA expression were inhibited in proliferating smooth musclecells by antisense and decoy oligonucleotides. Compared with positive control group, PCNA protein expression in antisense group and decoy group decreased 45.12% and 45.05%,respectively. ④ In model group, sense group and scramble group, vessel intimal area, medial area and intimal area/medial area increased at the 5th day after balloon-injury and reached the maximum at the 7th day after injury. The intimal area/medial area was significantly decreased in the antisense group and decoy group. The effect of antisense plus decoy oligonucleotides was more obvious than that of antisense group and decoy group alone but there were not significant differences among three groups. ⑤ Reverse transcription-polymerase chain reaction showed that MCP-1 mRNA expression was significantly increased 6 hours after balloon-injury, but not evident after 1 day. It was increased at the 3th, 5th and 7th days continuously, but decreased at the 14th day. MCP-1 mRNA expression was decreased at each time point in antisense group, decoy group, antisense plus decoy group (P＜0.05). ⑥Western blot analysis showed that NF-κB p65 was weakly expressed at 6 hours after vascular balloon-injury, increased significantly at 1 day, reached the peak at 7 days and weakened at 14 days, while ERK2 protein was weakly expressed, a little increased at 1 day, reached the peak at 7 days and weakened at 14 days. Treatment of antisense group, decoy group and antisense plus decoy group inhibited protein synthesis more significantly than those of model group, sense group and scramble group (P＜0.05).CONCLUSTON: NF-κB expression increases in proliferating smooth muscle cells. NF-κB modulates genes expression and protein synthesis of MCP-1 and ERK2. Cellular proliferation in vessel wall dynamically changes after balloon angioplasty injury. Antisense and decoy oligonucleotide of NF-κB by local lipofectamine transfer inhibit the expression of regulated target gene.

Objective To investigate the therapeutic effect of frozen-thawed murine islets which were transplanted into diabetic rats after cultured with hyperbaric oxygenated rotary cell culture system (HORCCS). Methods The purified rat islets were divided into two groups: A. In vitro experiment groups (IvEG) : The rat islets in each subgroup were cultured in HORCCS or common medium for 30 days, then evaluated for the intracellular DNA and insulin contents of islets, and the viability and insulin secreting level of islets. B. Islet transplantation experimental groups (TxEG) : The frozen-thawed islets were cultured in HORCCS or common medium for 7 days, and then transplanted into the recipients. We observed the blood glucose level (BGL) and insulin secreting level in the recipients as well as the uhrastructure change of islets in TxEG. Results The viability and insulin secreting level of islets cultured with HORCCS at 14th day were much higher than those cultured with common medium (P <0.05). The blood glucose level in recipients transplanted with islets cultured with HORCCS recovered to normal value at the 2nd week and lasted for 8 weeks. All these recipients maintained the normal glucose tolerance curve. Electronic microscopy found microchannel outlets on the surface of the frozen-thawed islets cultured with HORCCS. Conclusions Frozen-thawed islets cultured with HORCCS could establish nutrient transmission microchannels, which were not only capable of oxygen and nutrients transmission, but also improving cryopreservation solution to diffuse inside the islet cells evenly and uniformly. So this method not only lessens islet damage from cryopreservation, but also improves the effect of transplantation.