Effects of Dendrobium officinale Polysaccharides on Gene Expression Profile of HUVEC / 中国药房

OBJECTIVE： To investigate the effects of Dendrobium officinale polysaccharides on gene expression profile of HUVEC. METHODS： HUVEC was selected as objects. MTS method was used to detect the effects of different doses of D. officinale polysaccharides （50， 100， 200， 400， 800 μg/mL） on the proliferation activity of HUVEC. The growth inhibitory concentration of 30％ cells （IC30） was calculated to screen the dose of follow-up tests. cDNA microarray assay was used to detect the changes of gene expression profile for HUVEC after treated with D. officinale polysaccharides for 24 h， so as to screen differentially expressed genes. GO enrichment analysis and KEGG pathway enrichment analysis were performed for top 5 differentially expressed genes by using DAVID bioinformatics resource database. Real-time fluorescence quantitative polymerase chain reaction （qRT-PCR） was used to validate the results of microarray detection with immunity-related differentially expressed genes as objects. RESULTS： After treated with 100， 200， 400， 800 μg/mL D. officinale polysaccharides， survival rate of HUVEC  were decreased significantly （P＜0.05 or P＜0.01）. IC30 value was 408 μg/mL. After treated with 400 μg/mL （by IC30） D. officinale polysaccharides， there were 91 differentially expressed genes in HUVEC cells， of which 84 were up-regulated and 7 were down-regulated. Top 5 genes of up-regulated and down- regulated expression were SELE， CCL2， CXCL6， IL8， ICAM1 as well as VWCE， CPT1A， CLU， CCL14， CINS4， which may be mainly associated with immune conditions and inflammatory responses. The differentially expressed genes mainly distributed in extracellular domain， and were enriched in biological processes such as production and response of cytokines and stimulus response， and played molecular functions such as chemokine and its receptor activity. The up-regulated genes as SELE， ICAM1 and CXCL2 were mainly enriched in TNF signaling pathway， influenza A （H1N1）， herpes simplex virus infection and other pathways. The down-regulated gene CCL14 was mainly enriched in chemokine signaling pathway. Results of qRT-PCR validation tests showed that relative expression of ICAM1 was increased significantly， while that of CCL14 was decreased significantly （P＜0.05）， which was in agreement with microarray detection results. CONCLUSIONS： After treated with D. officinale polysaccharides， the expression of 91 genes in HUVEC cells are different significantly， mainly being up-regulated. The differentially expressed genes may participate in immune regulation through TNF signaling pathway， influenza A （H1N1） and herpes simplex virus infection.

Reporter cell line-based screening for anti-varicella-zoster virus compounds / 中华微生物学和免疫学杂志

Objective To establish a novel method to screen for anti-varicella-zoster virus (VZV) compounds with our previously generated reporter cell line for VZV, MV9G. MethodsMV9G cells were directly infected with cell-free virus of Oka vaccine strain (vOka) for 2 hours( CFV direct-infection) or cocultured with vOka-infected MeWo cells containing cell-associated virus for 48 hours (CAV co-culture) to promote expression of the reporter gene firefly luciferase. Antiviral compounds including heparin, mannose-6-phosphate( M-6-P), acyclovir( ACV ), resveratrol and roscovitine were added in the medium before or after the virus infection. Inhibitory effects( IC50 ) of the antiviral compounds were analyzed by comparing firefly luciferase activities of MV9G cells in the presence of antiviral compounds with those in the absence. Results Antiviral compounds inhibited luciferase activities of MV9G cells activated by CFV direct-infection and/or CAV co-culture in different levels. The reductions of luciferase activities statistically correlated with those of viral foci shown by immunostaining with a monoclonal antibody against VZV immediate early 62 antigens (IE62) in controls. Among these compounds, heparin, M-6-P, and 2.5 μmol/L of roscovitine inhibited CFV-activated more strongly than CAV-activated luciferase activities, whereas ACV and resveratrol inhibited CAV-activated more strongly than CFV-activated luciferase activities. Cell-associated ACV-resistant strains,Kanno and rOka YSR, activated luciferase activities of MV9G cells, too. However, the inhibitory concentrations (IC50) of ACV to the ACV-resistant strains were much higher than those to the ACV-sensitive strains,pOka and CaGu. ConclusionThe CFV direct-infection and CAV co-culture assays were useful to screen for antiviral compounds targeting the early and late phases of VZV infection, respectively. The VZV reporter cell-based assays may provide a simple, rapid, sensitive, and high-throughput method to screen for anti-VZV compounds.

Expression and distribution of NF-κB in experimental hepatic fibrosis / 中国病理生理杂志

AIM: To investigate the expression, distribution and significance of nuclear factor κB (NF-κB) in experimental hepatic fibrosis.METHODS: Wistar rats were randomly divided into normal control group, hepatic fibrisis model group and the pyrrolidine-1-dithiocarboxylic acid ammonium sail (PDTC) group. The PDTC group was treated with subcutaneous injection of carboan tetrachloride, and treated with PDTC by oral administration. The content of hydroxyproline was measured. Endotoxin was determined with a Limulus amebocyte lysate test kit. The alanine aminotransferase (ALT) in plasma was measured by laishi method. The content of malondialdehyde (MDA) in liver tissue was detected by means of TBA method. The expression of NF-κB was determined by immunohistochemistry. The expression of connective tissue growth factor (CTGF) was measured by Western blotting.RESULTS: In control group, just a small amount of NF-κB p65 was expressed in the cytoplasm of a few hepatocytes around central veins. In model group, the positive staining of NF-κB p65 was expressed in cytoplasm and nucleus, mainly in fibrous stepta, hepatic sinusoid and partial vascular endothelial cells, part of proliferating ductular epithelial cells and impaired hepatocytes. The positive staining began to increase from the first week. The expression of NF-κB in the liver tissues in PDTC group was lower than that in model control group (P<0.05). The ET levels and expression of NF-κB and CTGF began to increase significantly in liver fibrosis group. The levels of plasma ET and expression of NF-κB and CTGF were correlated positively with each other. In PDTC group, ET content in plasma increased significantly at first, then began to decline at the end of the test. The expression of NF-κB and CTGF in liver tissues in PDTC groups was lower than that in model group. Furthermore, the expression of NF-κB in liver tissues in PDTC group was correlated positively with CTGF. The levels of plasma ET were not correlated with the expression of NF-κB and CTGF.CONCLUSION: ET may up-regulate the expression of CTGF by activating NF-κB.

Radiosensitizing effect of gemcitabine on human lung cancer cells : A preliminary in vitro study / 中华放射肿瘤学杂志

Objective To investigate whether gemcitabine (GEM) could enhance radiosensitivity of human non-small cell lung cancer cells and its related mechanism.Methods Clonogenic assay was used to analyze radiosensitivity enhancement by GEM on p53 mutant human lung adenocarcinoma cell line 973.Alterations of cell cycle distribution and apoptosis were measured by flow cytometry.Results Mild radiosesitizing effect was observed when 10 nmol/L GEM was administrated before or after irradiation.Marked radiosesitizing effect was demonstrated when 100 nmol/L GEM was administrated before or after irradiation, with much stronger effect of pre-irradiation GEM treatment.Mutation of p53 gene affected cell cycle redistribution and cell apoptosis, but had no relationship with radiosensitivity enhancement of GEM.Conclusions 100 nmol/L GEM could significantly enhance radiosensitivity of human lung cancer cells.However, this effect may not be associated with p53 gene mutation, cell cycle redistribution or cell apoptosis.

Expression and distribution of NF-?B in experimental hepatic fibrosis / 中国病理生理杂志

AIM:To investigate the expression,distribution and significance of nuclear factor ?B (NF-?B) in experimental hepatic fibrosis.METHODS:Wistar rats were randomly divided into normal control group,hepatic fibrisis model group and the pyrrolidine-1-dithiocarboxylic acid ammonium sail (PDTC) group. The PDTC group was treated with subcutaneous injection of carboan tetrachloride,and treated with PDTC by oral administration. The content of hydroxyproline was measured. Endotoxin was determined with a Limulus amebocyte lysate test kit. The alanine aminotransferase (ALT) in plasma was measured by laishi method. The content of malondialdehyde (MDA) in liver tissue was detected by means of TBA method. The expression of NF-?B was determined by immunohistochemistry. The expression of connective tissue growth factor (CTGF) was measured by Western blotting.RESULTS:In control group,just a small amount of NF-?B p65 was expressed in the cytoplasm of a few hepatocytes around central veins. In model group,the positive staining of NF-?B p65 was expressed in cytoplasm and nucleus,mainly in fibrous stepta,hepatic sinusoid and partial vascular endothelial cells,part of proliferating ductular epithelial cells and impaired hepatocytes. The positive staining began to increase from the first week. The expression of NF-?B in the liver tissues in PDTC group was lower than that in model control group (P