Two fragments G1 and G2 of the glycoprotein G gene of bovine respiratory syncytial virus(BRSV) were selected for expression in Escherichia coli based on the analysis of glycoprotein G by DNA Star software.Then the two fragments of glycoprotein G were amplified by PCR with synthesized G gene of BRSV as the template.The amplified fragments G1 and G2 are 570bp and 308bp in length,respectively.The PCR products were cloned into pET30a vector and expressed in soluble form in E.coli after induction of cultured E.coli with IPTG.Both of the recombinant proteins G1 and G2 were purified by immobilized Ni ion affinity chromatography under native conditions.Then the purified proteins were analysed by Western blotting.The results showed that the purified recombinant protein G1 retained good antigenicity and specificity.But the purified recombinant protein G2 didn't possess biological activity.Antibodies against BRSV were detected in suspected bovine serum samples in China by using indirect ELISA and Western blotting with the purified recombinant protein G1.The purified recombinant protein G1 might be used as antigen for establishing serological methods for diagnosis of BRSV infection.And the purified recombinant protein G1 might also be used for preparing polyclonal and monoclonal antibodies for research on biological functions of glycoprotein G of BRSV.

Objective The aim of this study was to investigate how different concentrations of dextran sulfate sodi-um ( DSS) influence the establishment of mouse model of inflammatory bowel disease ( IBD) and the effect of DSS on the expression of colitis-associated immune factors.Methods The DSS solution in different concentrations (3%, 5%, 7%) were given to male C57BL/6J mice to generate mouse inflammatory bowel disease model.The IBD mice were observed by defecation characteristics, body weight, and survival time.The animals were sacrificed at 6 days after the start of DSS drinking.The general appearance of colons was observed and scored.Moreover, the pathological changes of the colon were examined and analyzed by routine histology.The expression of immune factors in the spleen was detected by real-time PCR.Results The mice in the 3%, 5%, 7% DSS groups developed murine colitis.In addition, the incidence of IBD and mouse mortality rate was directly proportional to the increase of DSS concentration.Furthermore, the higher concentra-tion of DSS induced the expression of proinflammatory factors including TNF-α, IFN-γand IL-17A, but cause a decrease of anti-inflammatory factors such as IL-4, IL-10 and Treg-related transcription factor Foxp3.Conclusions Our data suggest that giving 5%DSS solution to C57BL/6J mouse is appropriate to efficiently establish a murine IBD model.This laid an important foundation for further studies of the pathogenesis of IBD, biological characteristics, and intervention factors.

Budd-Chiari Syndrome ; Humans

0.05).There was significant difference in the 3-year survival rate between A and C group. Conclusions The 3-year survival rate was dramatically increased with combined therapy mainly by cisplatin, the dose of 60～80mg is tolerant for the elderly aged above seventy years, and perioperation complications can be cured.

Cell Line ; Cytochrome P-450 CYP1A1 ; drug effects ; metabolism ; Hepatocytes ; cytology ; metabolism ; Humans ; Microsomes, Liver ; drug effects ; metabolism ; Quercetin ; pharmacology

OBJECTIVEExpression of vimentin in carcinoma cells is a marker for poor prognosis in patients. The aim of this investigation was to assess the influence of vimentin on the characteristics of carcinoma cells.

METHODSThe full-length vimentin gene open reading frame (1401 base pairs) was cloned into the plasmid vector pcDNA 3.1 (+), and these vectors were used to stably transfect the human hepatocellular carcinoma HepG2 cell line. Vimentin gene expression was evaluated by RT-PCR and Western blot. Proliferative activity and invasive potential of tumor cells were determined by the CellTiter 96 aqueous one solution cell proliferation assay and BioCoat GFR Matrigel invasion chamber, respectively.

RESULTSDNA sequencing and restriction endonuclease digestion analysis demonstrated that the recombinant vector was correctly cloned. The stable cell line demonstrated a higher vimentin RNA and protein expression. However, both proliferative and invasive abilities of the cells were reduced in vitro ( P < 0.05).

CONCLUSIONA recombinant plasmid pcDNA3. 1-VIM is successfully constructed and a carcinoma cell line HepG2-pV highly expressing vimentin is obtained. Recombinant vimentin protein suppresses the proliferative and invasive abilities of HepG2 cells, suggesting that it might decrease malignant phenotype of tumor cells in vitro. This work makes a foundation for further study on the relationship between vimentin and biological phenotype of carcinoma cells.

Cell Movement ; Cell Proliferation ; Gene Expression Regulation, Neoplastic ; Genetic Vectors ; Hep G2 Cells ; Humans ; Neoplasm Invasiveness ; Open Reading Frames ; genetics ; Plasmids ; RNA, Messenger ; metabolism ; Recombinant Proteins ; genetics ; metabolism ; Transfection ; Vimentin ; genetics ; metabolism

OBJECTIVETo determine the therapeutic effectiveness and safety of polyethylene glycol 4000 (forlax) in the treatment of constipation in children over 8 years old.

METHODSThis study was designed as a randomized, positive medicine (lactulose) controlled multicenter trial. A total of 216 children with constipation from 8-18 years old from 7 hospitals across China who were matched with a uniform entry criteria were enrolled in this study. The 216 patients were randomized to receive either oral forlax (20 g/d, n=105) or lactulose (15 mL/d, n=111) for 2 weeks. The therapeutic effects, including bowel movement frequency, stool consistency, clinical complete remission rate of constipation and abdominal symptoms, and the safety of forlax and lactulose were evaluated at 1 and 2 weeks of treatment.

RESULTSThe median weekly frequency of bowel movement in the forlax group increased by 4 and 5 times respectively after 1 and 2 weeks of treatment, and increased by 3 and 4 times in the lactulose group (P < 0.05). The stool consistency of the two groups was both improved significantly after treatment. The Bristol score of stool consistency of the forlax and lactulose groups were 3.41+/-1.11 and 3.64+/-1.33 respectively (P < 0.05) after 1 week of treatment, and were 4.26+/-0.89 and 3.63+/-1.33 respectively (P < 0.05) after 2 weeks of treatment. The clinical complete remission rate of constipation in the forlax and lactulose groups was 70% and 40% respectively (P < 0.05) by week 1 of treatment, and that was 72% and 41% respectively (P < 0.05) by week 2 of treatment. Abdominal pain disappeared in 75% of patients in the forlax group but in only 57% in the lactulose group by week 2 of treatment (P < 0.05). No serious adverse events happened and no abnormalities were found in laboratory tests and physical examinations in the two groups after medication.

CONCLUSIONSForlax is safe and effective in the treatment of constipation in children over 8 years old.

Adolescent ; Cathartics ; adverse effects ; therapeutic use ; Child ; Constipation ; therapy ; Female ; Humans ; Lactulose ; therapeutic use ; Male ; Polyethylene Glycols ; adverse effects ; therapeutic use

Acute myeloid leukemia (AML) is a heterogeneous disease characterized by the accu-mulation of immature myeloid progenitor cells in the bone marrow,compromising of normal hematopoi-esis and ultimately resulting in bone marrow failure. Chemotherapy is the mainstay treatment for all AML patients,however,drug resistance and clinical relapse limits its efficacy.The 5-year survival rate of AML patients is only 26.6%.Survival rates are even lower among patients ages 65 to 74 years (5.3%)and 75 years or older(1.6%).Therefore,exploring novel therapeutic agents is urgent for improving the outcome of patients with AML. Saponins are amphipathic glycosides found in traditional Chinese medicines. In the present study, we isolated a panel of saponins from Paris forrestii (Takht.) H. Li, a unique plant found in Tibet and Yunnan provinces, China. By examining their activities in suppressing acute myeloid leukemia cell proliferation, total saponins from Paris forrestii (TSPf) displayed more potent activity than individual ones.TSPf induced more than 40% AML cell apoptosis within 24 h and decreased the viability of all leukemia cell lines. TSPf-induced apoptosis was confirmed by both Annexin V staining and caspase-3 activation.TSPf downregulated pro-survival proteins Mcl-1,Bcl-xL and Bcl-2,but upreg-ulated the expression of tumor suppressor proteins p53,p27,Bax and Beclin 1.The AKT/mTOR signaling pathway is frequently over activated in various AML cells,and TSPf was found to suppress the activa-tion of both AKT and mTOR,but had no effects on their total protein expression.This was further con-firmed by the inactivation of 4EBP-1 and p70S6K,two typical downstream signal molecules in the AKT/mTOR pathway. More specifically, TSPf-inactivated AKT/mTOR signaling was found to be associated with downregulated RNF6, a recently identified oncogene in AML. RNF6 activated AKT/mTOR, and consistently, knockdown of RNF6 led to inactivation of the AKT/mTOR pathway. Furthermore, TSPf suppressed the growth of AML xenografts in nude mice models. Oral administration of 100 mg·kg-1 body weight almost fully suppressed tumor growth within 14 d, without gross toxicity. This study thus demonstrated that TSPf displays potent anti-AML activity by suppressing the RNF6/AKT/mTOR pathway. Given its low toxicity,TSPf could be developed for the treatment of AML.

OBJECTIVETo search for the genes which could interact with newly found homo sapiens cross-immune reaction antigen (PCIA1) gene and accordingly to provide insights into the study of the gene function.

METHODSThe Stratagene's BacterioMatch Two-Hybrid System and BacterioMatch Fetal Kidney Library were adopted and the recombinant bait plasmid pBT-PCIA1 was cotransformated with the target plasmid pTRG-cDNA library DNA into the reporter stain. After screening and isolation of positive pTRG clones, the target genes were identified by DNA sequencing and bioinformation analysis.

RESULTSAmong all the seven detected target genes, three genes' function were not known, the other four genes had important functions. Their mutations or abberant expression resulted in severe diseases and overexpression of ACTN4 (actinin, alpha 4), PSAP (prosaposin) or EIF3S10 (eukaryotic translation initiation factor 3, subunit 10 theta) could promote tumor development and progression.

CONCLUSIONThe bacterial two-hybrid system technique is an efficient method, which can provides insights into the study of novel genes' function by detecting protein-protein interactions. This study indicates that PCIA1 gene expression correlates with tumor formation, invasion and metastasis.

Bacteria ; genetics ; metabolism ; Computational Biology ; DNA Restriction Enzymes ; metabolism ; Gene Library ; Genetic Vectors ; Humans ; Neoplasms ; genetics ; pathology ; Plasmids ; genetics ; Sequence Analysis, DNA ; Two-Hybrid System Techniques

Objective To study the features of Yersinia pestis(Y.pestis)in areas along Qinghai-Tibet Railroad in Qinghai Province.Methods To identify the biologic types and the molecular biological feathers of Y.pestis isolated from areas along Qinghai-Tibet Railroad in Qinghai from 2001-2006.Results All the tested Y.pestis was biologically of classical type and ecologically of Qinghai-Tibet plateau type.The Y.pestis had high virulence.The Y.pestis of 65×106 plasmids was distributed in the Tanggula area,the Y.pestis of 52×106plasmids,in Tianjun and Delingha areas.The Y.pestis srains carried 52 × 106 plasmids.except the two containing 65 X 106 plasmids in Wulan County.The genetic type of Y.pestis in Tanggula was type 5 and that in Zongwulong of Delingha,Saishike,Keke,Tongpu of Wulan was type 8 except 2 strains of Y.pestis isolated from woodchuck and the patients in Dananwan of Tongpu,Wulan County were type 15.Conclusion The Y.pestis in the area along Qinghai-Tibet Railroad in Qinghai belongs to Qinghai-Tibet plateau type with high virulence.