hVEGF siRNA inhibits initial growth of A549 tumor cells in nude mice and angiogenesis on CAM assay / 浙江大学学报·医学版

<p><b>OBJECTIVE</b>To investigate the effect of small interfering RNA (siRNA) targeting human vascular endothelial growth factor (hVEGF) on A549 cell growth in nude mice and angiogenesis on chorioallantoic membrane (CAM) assay.</p><p><b>METHODS</b>Three pairs of hVEGF siRNA-plasmid and non-silencing-plasmid were constructed, and transfected into A549 cells through lipofectamine 2000, respectively. The most effective pair of hVEGF siRNA-plasmid was selected by ELISA and real-time RT-PCR. A549 cells transfected with selected hVEGF siRNA- plasmid, A549 cells transfected with non-silencing-plasmid and A549 cells without transfection were inoculated into nude mice, respectively. Chick embryos were randomly divided into four groups and CAM was treated by different solutions for 48 h: culture media DMEM as negative control group,un-transfected A549 cell culture supernatants as positive control group, hVEGF siRNA A549 cell culture supernatants as hVEGF siRNA group and nonsilencing siRNA A549 cell culture supernatants as non-silencing siRNA group. The CAMs were harvested on d12 for microscopic assays.</p><p><b>RESULT</b>Compared with control group, hVEGF siRNA-plasmid induced 48% reduction in hVEGF secretion by A549 cells accompanied by 70% reduction in hVEGF mRNA. Compared with non-silencing siRNA group, the mean tumor volume of murine xenograft was reduced by 58% in hVEGF siRNA group; time for xenografts growing to 50 mm(3)was delayed by 5.4 d. hVEGF contents in xenograft were reduced by 54%; but mean doubling time of tumors and the growth rate of tumors were not significantly reduced. In CAM assays, hVEGF content was zero in negative group, and in hVEGF siRNA group that was 40%-44% of non-silencing siRNA group or positive group; vessels branch points of CAM in hVEGF siRNA group or non-silencing siRNA group or positive group were increased by 45%-55% compared with negative group; total vessel length of CAM in hVEGF siRNA group was increased by 53% compared with negative group, while in non-silencing siRNA group or positive group that was increased by 97% or 99%. Compared with negative control group, the proliferation of microvessels was increased when cell culture supernatant with hVEGF was added in hVEGF siRNA group, significant proliferated vessels were observed in non-silencing siRNA group or positive group.</p><p><b>CONCLUSION</b>A plasmid-mediated hVEGF siRNA has been constructed and verified, which can effectively downregulate the expression of hVEGF in human A549 cells, resulting in the inhibition of angiogenesis. hVEGF siRNA can delay initial growth of A549 tumor xenograft but not reduce the growth rate.</p>

The effect of silicon dioxide on the activation of nuclear factor-kappaB in THP-1 cells / 中华劳动卫生职业病杂志

<p><b>OBJECTIVE</b>To study the effect of silicon dioxide (SiO(2)) on the activation of nuclear factor-kappaB (NF-kappaB) in THP-1 cell line.</p><p><b>METHODS</b>THP-1 cells were incubated with a series of doses of SiO(2) (0, 100, 200 micro g/ml). The location of NF-kappaB p65 subunit (NF-kappaB/p65) in THP-1 cells was detected by immunofluorescence and laser scanning confocal microscope (LSCM). The expression of NF-kappaB/p65 in nuclei was measured by Western blot analysis.</p><p><b>RESULTS</b>The majority of fluorescein isothiocyanate (FITC)-labelled NF-kappaB/p65 located in the nuclei 30 min after stimulation by 100 micro g/ml SiO(2), whereas the FITC-labelled NF-kappaB/p65 were mainly seen in the plasma of normal control cells. The expression of NF-kappaB/p65 in THP-1 nuclear protein was low in control group (0 micro g/ml SiO(2)) while it increased after stimulation by 100 micro g/ml SiO(2) and 200 micro g/ml SiO(2) for 15 min and 30 min. The level of NF-kappaB/p65 was comparatively increased with the increasing of doses and time. Lipopolysaccharides (LPS), an activator of NF-kappaB, had similar effect as SiO(2) on the activation of NF-kappaB/p65 in THP-1 cells.</p><p><b>CONCLUSION</b>SiO(2) could activate and internalize NF-kappaB in the THP-1 cell line.</p>

Effect of eucalyptus globulus oil on activation of nuclear factor-kappaB in THP-1 cells / 浙江大学学报·医学版

<p><b>OBJECTIVE</b>To study the effect of eucalyptus globulus oil on the activity of nuclear factor-kappaB(NF-kappaB) in THP-1 cell line.</p><p><b>METHODS</b>THP-1 cells were cultured with or without eucalyptus globulus oil at different concentrations (1, 10, 100 mg x L(-1), 30 min) before being stimulated with lipopolysaccharide (LPS, 1 mg x L(-1), 30 min). The location of NF-kappaB p65 subunit (NF-kappaB/p65) in THP-1 cells was detected by indirect immunofluorescence and laser scanning confocal microscope. The expression of NF-kappaB/p65 in nuclei was measured by Western-blot analysis.</p><p><b>RESULT</b>The FITC-label NF-kappaB/p65 was mainly located in the nuclei after THP-1 cells were stimulated with LPS. Whereas, no fluorescence were seen in the nuclei of cells pretreated with eucalyptus globulus oil. This effect on NF-kappaB/p65 nuclear translocation was in a concentration dependent manner.</p><p><b>CONCLUSION</b>Eucalyptus globulus oil inhibits the nuclear translocation of NF-kappaB induced by LPS in THP-1 cells.</p>