RESUMEN
<p><b>OBJECTIVE</b>To explore the change of endogenic nerve growth factor (NGF) expression in human glioma cells infected with human cytomegalovirus (HCMV).</p><p><b>METHODS</b>U251 cells were cultured in RPMI 1640 culture medium and infected with HCMV AD169 strain in vitro to establish a cell model of viral infection. Morphologic changes of U251 cells were observed under inverted microscope before and after infection with HCMV. Expression of NGF gene and protein of cells was detected by RT-PCR and Western blotting before and after infection with HCMV.</p><p><b>RESULTS</b>The cytopathic effects of HCMV-infected cells appeared on day 5 after infection. However, differential NGF expression was evident on day 7. NGF expression was decreased significantly in U251 cells on day 7 after infection in comparison with control group (P < 0.05).</p><p><b>CONCLUSION</b>HCMV can down-regulate endogenous NGF levels in human glioma cell line U251.</p>
Asunto(s)
Humanos , Actinas , Metabolismo , Línea Celular Tumoral , Citomegalovirus , Fisiología , Infecciones por Citomegalovirus , Patología , Técnica del Anticuerpo Fluorescente , Regulación Neoplásica de la Expresión Génica , Glioma , Genética , Patología , Virología , Modelos Biológicos , Factor de Crecimiento Nervioso , Genética , Metabolismo , ARN Mensajero , Genética , Metabolismo , Reproducibilidad de los Resultados , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
The objective of present study is to investigate the effect of human cytomegalovirus (HCMV) infection on human hippocampus neural stem cells NSCs differentiation in vitro, Fetal hippocampus tissue was dissociated mechanically and then cultured in proliferation medium with EGF and bFGF. Immunofluorescence method was used to detect the expression of NSCs marker-Nestin within these cells. Cultured in 10% FBS, NSCs began to differentiate. On the onset of the differentiation, HCMV AD169 (MOI=5) was added into the differentiation medium. After 7 days differentiation, the effect of HCMV infection on NSCs differentiation was observed by detecting the rate of nestin, GFAP and HCMV immediate-early (IE) positive cells with confocal microscopy and immunofluorescence method. The resucts showed most of the cells (passage 4-6 ) were Nestin positive and could differentiate into NSE-positive neurons and GFAP-positive astrocytes. On day 7 postinfection, 86% +/- 12% of infected cells were IE positive. The percentage of Nestin-positive cells was 50% +/- 19% and 93% +/- 10% (t= 6.03, P<0.01)and those of GFAP-positive cells was 81% +/- 11% and 55 +/- 17% (t=3.77, P<0.01) in uninfected and infected cells respectively. These findings indicated that NSCs were HCMV permissive cell and HCMV AD 169 infection suppressed the differentiation of Hippocampus-genetic human neural stem cells into astrocytes.
Asunto(s)
Humanos , Astrocitos , Biología Celular , Diferenciación Celular , Células Cultivadas , Citomegalovirus , Fisiología , Factor de Crecimiento Epidérmico , Farmacología , Factor 2 de Crecimiento de Fibroblastos , Farmacología , Hipocampo , Biología Celular , Proteínas de Filamentos Intermediarios , Metabolismo , Microscopía Fluorescente , Células Madre Multipotentes , Biología Celular , Metabolismo , Virología , Proteínas del Tejido Nervioso , Metabolismo , Nestina , Neuronas , Biología CelularRESUMEN
During the infection of host cells, IE2 protein is one of the first and most abundantly expressed products of HCMV genome, which plays an important role in the controlling of cell cycle and apoptosis. But the correlation between expression level and anti-apoptotic activity of IE2 protein is still not clear. In this study, we successfully established a HCMV IE2 protein expression cell line that was controlled by Tet-On system. The effect of IE2 protein on cell apoptosis and the expression of p53 was detected under different condition of induction. Our results showed that the IE2 protein could inhibit cell apoptosis induced by TNF-alpha. Additionally, the anti-apoptotic activity of IE2 protein seemed to be relevant to its expression level. However, we failed to detect any difference of p53 expression between the IE2 protein expression and non-expression cells. These data indicated that the IE2 protein might inhibit cell apoptosis through regulating different signal pathways.
Asunto(s)
Humanos , Antibacterianos , Farmacología , Apoptosis , Genética , Doxiciclina , Farmacología , Regulación de la Expresión Génica , Genética , Células HeLa , Proteínas Inmediatas-Precoces , Genética , Metabolismo , Plásmidos , Genética , Transactivadores , Genética , Metabolismo , Proteína p53 Supresora de Tumor , MetabolismoRESUMEN
Human herpes simplex virus 1 (HSV-1) causes facial,ocular,and encephalitic disease and is associated with latent infection and cancer.Here,we developed a means of studying the pathogenesis of HSV-1 infection at the protein level by using the SELDI Protein Chip to detect changes of protein expression in Vero cells cultured in vitro.After infection with HSV-1 and culture for 12,24 or 48 h,cells were harvested and lysed.IMAC3 arrays were applied to SELDI-TOF-MS to detect proteomic differences before and after infection.The chip detected a series of differentially expressed protein peaks.Interestingly,both peaks at 16 912 Da and 17 581 Da corresponded precisely with the molecular mass of ISG 15,which may participate in antiviral activity during the process of infection.Thus,the results we obtained can serve as a basis to study the pathogenesis of HSV-1 and the interaction between the virus and its host.In addition,they can help in the discovery of new therapeutic targets for treatment of HSV-1 infection.