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IFI6 Inhibits Apoptosis via Mitochondrial-Dependent Pathway in Dengue Virus 2 Infected Vascular Endothelial Cells.

Qi Y, Li Y, Zhang Y, Zhang L, Wang Z, Zhang X, Gui L, Huang J - PLoS ONE (2015)

Bottom Line: Interferon (IFN)-stimulated genes (ISGs) induced by dengue virus (DENV) exert antiviral effects.We observed that Bcl-2 expression was increased in IFI6+/+ and decreased in IFI6-/- cells.By contrast, Bax expression was decreased in IFI6+/+ and increased in IFI6-/- cells.

View Article: PubMed Central - PubMed

Affiliation: Institute of Immunology, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou, PR China; Key Laboratory of Tropical Diseases Control, Ministry of Education, Guangzhou, PR China.

ABSTRACT
Dengue hemorrhagic fever (DHF)/Dengue shock syndrome (DSS) is a fatal infectious disease that demands an effective treatment. Interferon (IFN)-stimulated genes (ISGs) induced by dengue virus (DENV) exert antiviral effects. Among ISGs, IFN-α inducible gene 6 (IFI6) was increased in DENV infected human umbilical vascular endothelial cells (HUVECs) by microarray analysis in our previous study. However, its function is incompletely understood. In this study, we confirmed that IFI6 was markedly induced in DENV infection of both primary HUVECs and EA.hy926 cell lines. Recombinant EA.hy926 cell lines in which IFI6 was either over-expressed (IFI6+/+) or knocked-down (IFI6-/-) were generated. The activation of caspase-3 and intrinsic apoptosis-related protein caspase-9 were down-regulated in IFI6+/+ but up-regulated in IFI6-/- cells at 24-48 hrs post-infection. After incubation with DENV for 48 hrs, the mitochondrial membrane potential (Δψ(m)) was more stable in IFI6+/+ cells but reduced in IFI6-/- cells, as assayed by fluorescence staining with JC-1. We observed that Bcl-2 expression was increased in IFI6+/+ and decreased in IFI6-/- cells. By contrast, Bax expression was decreased in IFI6+/+ and increased in IFI6-/- cells. It is presumed that the anti-apoptotic function of IFI6 is expressed by regulating the rheostatic balance between bcl-2/bax expression and inhibition of Δψ(m) depolarization during DENV infection of vascular endothelial cells(VECs). In addition, the pro-apoptotic protein X-linked Inhibitor of Apoptosis (XIAP)-Associated Factor 1(XAF1) expression had been reported to be up-regulated and led to the induction of apoptosis in DENV2-infected VECs,but the relationship between XAF1 and IFI6 dengue virus-induced apoptosis in VECs warrants further study.

No MeSH data available.


Related in: MedlinePlus

Identification of recombinant vascular endothelial EA.hy926 cell lines with stable over-expression/knock-down of IFI6.(a and b) mRNA expression of IFI6 was detected using qRT-PCR in EA.hy926 cells with vector-hi(a), vector-low(b) and the recombinant cell lines IFI6+/+ (a) and IFI6-/- (b). (c and d) Protein expression of IFI6 was detected using immunoblotting in the indicated lysate (c) and the gray scale scanning data were shown below and normalized to β-actin (d). ***P<0.01.
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pone.0132743.g002: Identification of recombinant vascular endothelial EA.hy926 cell lines with stable over-expression/knock-down of IFI6.(a and b) mRNA expression of IFI6 was detected using qRT-PCR in EA.hy926 cells with vector-hi(a), vector-low(b) and the recombinant cell lines IFI6+/+ (a) and IFI6-/- (b). (c and d) Protein expression of IFI6 was detected using immunoblotting in the indicated lysate (c) and the gray scale scanning data were shown below and normalized to β-actin (d). ***P<0.01.

Mentions: To further understand the role of IFI6 in apoptosis that was induced by DENV2-mediated infection of VECs, we constructed two stable cell lines of EA.hy926 IFI6+/+ and IFI6-/- cells, which were transfected with IFI6 over-expression (pMSCV-neo-IFI6+) and IFI6 knock-down (pMSCV-neo-IFI6-) plasmids. Empty vectors pMSCV-neo and pSUPERretro transfected cell lines were also generated as controls, which were referred to as “vector-high” and “vector-low”, respectively. Next, cells were subjected to qRT-PCR analysis of total RNA extracts and immunobloting of protein lysates in order to confirm alteration of IFI6 expression. The results were shown in Fig 2. Both mRNA and protein levels of IFI6 were significantly increased in IFI6+/+ cells and decreased in IFI6-/- cells.


IFI6 Inhibits Apoptosis via Mitochondrial-Dependent Pathway in Dengue Virus 2 Infected Vascular Endothelial Cells.

Qi Y, Li Y, Zhang Y, Zhang L, Wang Z, Zhang X, Gui L, Huang J - PLoS ONE (2015)

Identification of recombinant vascular endothelial EA.hy926 cell lines with stable over-expression/knock-down of IFI6.(a and b) mRNA expression of IFI6 was detected using qRT-PCR in EA.hy926 cells with vector-hi(a), vector-low(b) and the recombinant cell lines IFI6+/+ (a) and IFI6-/- (b). (c and d) Protein expression of IFI6 was detected using immunoblotting in the indicated lysate (c) and the gray scale scanning data were shown below and normalized to β-actin (d). ***P<0.01.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4526556&req=5

pone.0132743.g002: Identification of recombinant vascular endothelial EA.hy926 cell lines with stable over-expression/knock-down of IFI6.(a and b) mRNA expression of IFI6 was detected using qRT-PCR in EA.hy926 cells with vector-hi(a), vector-low(b) and the recombinant cell lines IFI6+/+ (a) and IFI6-/- (b). (c and d) Protein expression of IFI6 was detected using immunoblotting in the indicated lysate (c) and the gray scale scanning data were shown below and normalized to β-actin (d). ***P<0.01.
Mentions: To further understand the role of IFI6 in apoptosis that was induced by DENV2-mediated infection of VECs, we constructed two stable cell lines of EA.hy926 IFI6+/+ and IFI6-/- cells, which were transfected with IFI6 over-expression (pMSCV-neo-IFI6+) and IFI6 knock-down (pMSCV-neo-IFI6-) plasmids. Empty vectors pMSCV-neo and pSUPERretro transfected cell lines were also generated as controls, which were referred to as “vector-high” and “vector-low”, respectively. Next, cells were subjected to qRT-PCR analysis of total RNA extracts and immunobloting of protein lysates in order to confirm alteration of IFI6 expression. The results were shown in Fig 2. Both mRNA and protein levels of IFI6 were significantly increased in IFI6+/+ cells and decreased in IFI6-/- cells.

Bottom Line: Interferon (IFN)-stimulated genes (ISGs) induced by dengue virus (DENV) exert antiviral effects.We observed that Bcl-2 expression was increased in IFI6+/+ and decreased in IFI6-/- cells.By contrast, Bax expression was decreased in IFI6+/+ and increased in IFI6-/- cells.

View Article: PubMed Central - PubMed

Affiliation: Institute of Immunology, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou, PR China; Key Laboratory of Tropical Diseases Control, Ministry of Education, Guangzhou, PR China.

ABSTRACT
Dengue hemorrhagic fever (DHF)/Dengue shock syndrome (DSS) is a fatal infectious disease that demands an effective treatment. Interferon (IFN)-stimulated genes (ISGs) induced by dengue virus (DENV) exert antiviral effects. Among ISGs, IFN-α inducible gene 6 (IFI6) was increased in DENV infected human umbilical vascular endothelial cells (HUVECs) by microarray analysis in our previous study. However, its function is incompletely understood. In this study, we confirmed that IFI6 was markedly induced in DENV infection of both primary HUVECs and EA.hy926 cell lines. Recombinant EA.hy926 cell lines in which IFI6 was either over-expressed (IFI6+/+) or knocked-down (IFI6-/-) were generated. The activation of caspase-3 and intrinsic apoptosis-related protein caspase-9 were down-regulated in IFI6+/+ but up-regulated in IFI6-/- cells at 24-48 hrs post-infection. After incubation with DENV for 48 hrs, the mitochondrial membrane potential (Δψ(m)) was more stable in IFI6+/+ cells but reduced in IFI6-/- cells, as assayed by fluorescence staining with JC-1. We observed that Bcl-2 expression was increased in IFI6+/+ and decreased in IFI6-/- cells. By contrast, Bax expression was decreased in IFI6+/+ and increased in IFI6-/- cells. It is presumed that the anti-apoptotic function of IFI6 is expressed by regulating the rheostatic balance between bcl-2/bax expression and inhibition of Δψ(m) depolarization during DENV infection of vascular endothelial cells(VECs). In addition, the pro-apoptotic protein X-linked Inhibitor of Apoptosis (XIAP)-Associated Factor 1(XAF1) expression had been reported to be up-regulated and led to the induction of apoptosis in DENV2-infected VECs,but the relationship between XAF1 and IFI6 dengue virus-induced apoptosis in VECs warrants further study.

No MeSH data available.


Related in: MedlinePlus