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Objective To evaluate the expression of epidermal growth factor-like domain 7 (Egfl7) in atherosclerotic plaques and effects of its small interference RNA (siRNA) on angiogenesis gene expression in human endothelial cell line (HUVEC). MethodsEgfl7 expression in atheroscleroticplaquesweredetectedinhumaniliacarteryandmousearteriaeusing immunohistochemistry and immunofluorescence stainings.The siRNA targeting Egfl7 was transfected into HUVEC by lipofectamine with non-transfected cells and unconcerned siRNA as controls.At 0 h,12 h,24 h and 48 h after intervention,the levels of mRNA and protein of Egf17,vascular endothelial growth factor(VEGF),platelet derived growth factor-A (PDGF-A),platelet derived growth factor-B (PDGF-B),vascular cell adhesion molecule(VCAM) and intercellular adhesion molecule (ICAM)were measured by RT-PCR and Western blotting,respectively. ResultsThe expressions of Egfl7 in human iliac artery and mouse arteriae were increased.The expressions of Egfl7 in HUVEC at the levels of mRNA were[(0.14±0.02),(0.09±0.01),(0.02±0.01)]and the levels of protein[(0.71±0.11),(0.39±0.09),(0.07±0.01)]at 12 h,24 h and 48 h after siRNA intervention,respectively,which were decreased as compared with 0 h intervention [(0.31 ±0.05) and (0.93±0.08) ].Other genes such as VEGF,PDGF-A and PDGF-B were reduced or silenced at the levels of protein and mRNA in HUVEC with siRNA longer interventions(all P＜0.05).ConclusionsThe expression of Egfl7 in atherosclerotic plaques is increased.The siRNA inhibiting Egfl7 gene expression results in silence of other factors involved in angiogenesis.
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AIM:To investigate the effect of tumor-suppressor RUNX3 on the transcription of apoptosis-related genes bcl-2,bax,caspase-3,caspase-8,caspase-9 in human gastric carcinoma cells BGC823,and to reveal the apoptosis molecular mechanism promoted by RUNX3. METHODS:The eukaryotic expression vector of human Runx3 gene pcDNA3.1-Runx3 was constructed. pcDNA3.1-Runx3 and blank vector pcDNA3.1 were transfected into BGC823 cells,respectively. After 48 h,the total mRNA and protein were acquired and the expression level of Runx3 was determined by RT-PCR and Western blotting. Then,the mRNA and protein expression of bcl-2,bax,caspase-3,caspase-8 and caspase-9 was determined by RT-PCR and Western blotting. ?-actin was used as a control. RESULTS:The eukaryotic expression vector pcDNA3.1-Runx3 was constructed successfully and transfected into BGC823 cells. RT-PCR and Western blotting confirmed that RUNX3 level was higher in pcDNA3.1-Runx3 transfected BGC823 cells than that in blank vector-transfected cells (P
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AIM:To establish the gastric cancerous multidrug resistance cell stain BGC823/5-FU and investigate the relationship between the resistance and the expression of apoptosis related protein Survivin, Bcl-2, Bax and caspase-3. METHODS: Human gastric cancer cell line BGC823 was induced into MDR cell line by intermittent administration of high dose of 5-FU. MTT assay was used to detect the sensitivity of these MDR cells to some chemotherapeutic agents. Flow cytometry was used to detect the expression of P-glucoprotein and the accumulative value of intracellular daunorubicin (DNR) in these MDR cells. Western blotting was used to detect the expression of Survivin, Bcl-2, Bax and caspase-3. RESULTS: The resistance cell stain BGC823/5-FU was established, which possessed the ability of 10.82 fold resistance to 5-FU and cross-resistance to adriamycin, mitomycin C and cisplatin. The expression of P-glucoprotein was higher in BGC823/5-FU cells than that in BGC823 cells, while the accumulative value of intracellular DNR was decreased in BGC823/5-FU cells. Compared with its parent cells, expressions of Bax and caspase-3 in BGC823/5-FU cells were significantly down-regulated, surviving and Bcl-2 were upregulated in BGC823/5-FU cells. CONCLUSION: Gastric cancer cell line BGC823 has been induced into MDR cell line BGC823/5-FU. P-glucoprotein, Survivin, Bcl-2/Bax ratio and caspase-3 may play an important role in MDR of BGC823/5-FU cells.