0.05). Compared with control cells, the relative luciferase activity of NF-?B in virus-infected cells was apparently up-regulated (P

Objective:To observe the influence of ginsenoside Rg1 on transcriptional activation of NF-κB induced by hydrogen peroxide (H_2O_2) in 293T cell,and probe into the antioxidant mechanism of ginsenoside Rg1.Methods:In the experiment,cells was exposed to H_2O_2 after pretreatment with Rg1,cell proliferation and cytotoxicity studies were detected by MTT and Trypan blue.The quantities of generation of intracellular reactive oxygen species (iROS) was analyzed by flow cytometric analysis measured with fluorescent probe 2,7-dichlorofluorescin diacetate (DCFH-DA).NF-κB-responsive element-luciferase reporter gene was transfected and dual-luciferase cis-reporting systems were used to assay the transcriptional activity of NF-κB under the stimulated circumstance of oxidative stress induced by H_2O_2.Results:The results of MTT showed that ginsenoside Rg1 apparently protected the proliferation of 293T cell,which were repressed by H_2O_2 (P＜0.05).The results by trypan blue showed that H_2O_2 stimulated substantial cytotoxicity.This effect was markedly attenuated by treatment with ginsenoside Rg1.Oxidant production,measured as the fluorescence of dichlorofluorescein,was significant increased by 40%-50% through H_2O_2 stimulation.The decrease in iROS generation was significant blocked by 35%-40% through Rg1 and antioxidant.The relative luciferase reporter assay of NF-κB was apparently improved by H_2O_2-induced(P＜0.05),but Ginsenoside Rg1 significantly repressed the relative value of luciferase (P＜0.05).Conclusion:Ginsenoside Rg1 has the obvious protective function from the damage of oxidative stress damage,whose possible mechanism is to eliminate excessive free radicals of the cells effectively,to reduce transcriptional activation of nuclear factor kappa B(NF-κB),and subsequently to suppress the NF-κB circuit activation.

Objective To identify the species of cultured cells and to detect inter-species cross contamination by polymerase chain reaction (PCR).Methods From references and NCBI database,we outsourced 32 pairs of species-specific primers targeted to genomic DNA of ten common species of cultured cells.Then we screened for optional primers with high specificity and high sensitivity.PCR amplification with these primers,the genomic DNAs isolated from the tested cell line and agarose gel electrophoresis of the PCR products were done.Mixed DNA of 10 species was used as positive-control template,and water as negative-control template.Results Ten pairs of species-specific and highly sensitive primers were selected.By PCR amplification with these primers and agarose gel electrophoresis,we may easily identify the origin of cell lines and find whether the tested cell lines are contaminated by cells of other species.Conclusion This PCR assay provides a simple,rapid,sensitive,and cost-effective method to identify cell species and detect interspecies cross-contamination.

Objective Determining the characteristics of human fetal hepatocytes in vitro. Methods Isolating and culturing human fetal hepatocytes by stepwise trypsinization of liver fragment in vitro; collecting the culture medium to determine the secretion of AFP, ALB and the functional enzymes (including ALT, AST, GGT, ALP and LDH) of different generations in culture; determining the expression of cytochrome C by immunohistochemistry; testing the effects of sodium byturate on human fetal hepatocytes. Results Human fetal hepatocytes were polygonal epithelial cells in DMEM medium. They could be maintained for 5.5 months (about 30 passages) in vitro. They secreted ALB and functional enzymes all over their cultivation. Conclusion Human fetal hepatocytes can be maintained keeping function in vitro for several months.

Objective:To artificially design and express a recombinant protein named as ScFv-pLLO by fusing ScFv gene of Rituximab(C2B8)and dominant antigen epitopes from listeriolysin O(LLO),and studying its anti-tumor activity.Methods:VH and VL gene sequences of C2B8 against CD20 were acquired by searching United States Patent database,and ScFv sequence was constructed by linking VL and VH with a short peptide linker.Two CD4+T cell epitopes from LLO were selected and designed to splice ScFv sequence.The recombinant gene of ScFv-pLLO was cloned into prokaryotic expression vector and purified after induction.The capacity of ScFv-pLLO target-binding to B-cell lymphomas was evaluated by flow cytometry ( FCM ) and co-immunoprecipitation ( Co-IP ) .The effects of ScFv-pLLO on B-cell lymphomas proliferation and apoptosis were detected respectively.The immunogenicity of ScFv-pLLO was assessed by lymphocyte proliferation assay.Results: ScFv-pLLO was successfully expressed.It could bind to different B-cell lymphomas cell lines and obviously inhibit the growth of Raji cells as well as inducing apoptosis.Moreover,ScFv-pLLO was able to stimulate proliferation of spleen lymphocytes of immunized mice.Conclusion: The recombinant protein ScFv-pLLO can target-bind to B-cell lymphomas,and perform inhibitory effect and induce apoptosis on Raji cells that indicate ScFv-pLLO retain the capacity of ScFv derived from monoclonal antibody against CD20.Besides, ScFv-pLLO can induce immune response.This study provides a basis for further research about the role of ScFv-pLLO on simulating tumor cell antigens as well as being tumor vaccine adjuvant.

Aim To explore the influencing factors of major adverse cardiovascular events(MACE)after drug-e-luting stent(DES)implantation in young and middle-aged patients with coronary artery disease(CAD).Methods Retrospective study data from the First Affiliated Hospital of Zhengzhou University were extracted from the Dryad database,and young and middle-aged CAD patients who received DES were divided into MACE group(n=57)and non MACE group(n=321)according to whether MACE occurred during the follow-up period.Clinical data including age,gender etc.were compared between the two groups,and the variables with significant differences between the two groups were substitu-ted into Logistic regression model to screen the influencing factors of MACE after DES implantation in young and middle-aged CAD patients.Value of influencing factors predicting occurrence of MACE after DES implantation in young and mid-dle-aged CAD patients was evaluated by plotting ROC curve and calculating the area under the curve(AUC).Results Stent diameter in MACE group was significantly smaller than that in non MACE group(P＜0.001),the number of left main coronary artery lesion in MACE group was significantly higher than that in non MACE group(P＜0.05).Multivariate Lo-gistic regression analysis indicated that stent diameter(OR=0.184,95％CI:0.084～0.405,P＜0.001)and left main coronary artery lesion(OR=9.319,95％CI:2.291～37.904,P=0.002)were the independent risk factors of MACE after DES implantation in young and middle-aged CAD patients.The risk of MACE after DES implantation in young and middle-aged CAD patients significantly decreased when the stent diameter was＞3 mm.The AUC of stent diameter com-bined with left main coronary artery lesion predicting occurrence of MACE after DES implantation in young and middle-aged CAD patients was 0.700(95％CI:0.623～0.776).Conclusion The smaller stent diameter and left main coronary artery lesion are the influencing factors of the occurrence of MACE after DES implantation in young and middle-aged pa-tients with CAD,and should be emphasized by clinical practitioners.

3' Untranslated Regions ; Animals ; Biomarkers ; metabolism ; Cell Proliferation ; Cell Survival ; Embryonic Stem Cells ; cytology ; metabolism ; Induced Pluripotent Stem Cells ; cytology ; metabolism ; Mice ; Swine ; Transcription Factors ; genetics ; metabolism