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Dermal-type macrophages expressing CD209/DC-SIGN show inherent resistance to dengue virus growth.

Kwan WH, Navarro-Sanchez E, Dumortier H, Decossas M, Vachon H, dos Santos FB, Fridman HW, Rey FA, Harris E, Despres P, Mueller CG - PLoS Negl Trop Dis (2008)

Bottom Line: Here, we showed that dermal macrophages bound recombinant envelope E glycoprotein fused to green fluorescent protein.The macrophages were able to internalize the virus, but progeny virus production was undetectable in the infected cells.In addition, no IFN-alpha was produced in response to the virus.

View Article: PubMed Central - PubMed

Affiliation: CNRS, Laboratory of Therapeutic Immunology and Chemistry, IBMC, Université Louis Pasteur, Strasbourg, France.

ABSTRACT

Background: An important question in dengue pathogenesis is the identity of immune cells involved in the control of dengue virus infection at the site of the mosquito bite. There is evidence that infection of immature myeloid dendritic cells plays a crucial role in dengue pathogenesis and that the interaction of the viral envelope E glycoprotein with CD209/DC-SIGN is a key element for their productive infection. Dermal macrophages express CD209, yet little is known about their role in dengue virus infection.

Methods and findings: Here, we showed that dermal macrophages bound recombinant envelope E glycoprotein fused to green fluorescent protein. Because dermal macrophages stain for IL-10 in situ, we generated dermal-type macrophages from monocytes in the presence of IL-10 to study their infection by dengue virus. The macrophages were able to internalize the virus, but progeny virus production was undetectable in the infected cells. In addition, no IFN-alpha was produced in response to the virus. The inability of dengue virus to grow in the macrophages was attributable to accumulation of internalized virus particles into poorly-acidified phagosomes.

Conclusions: Aborting infection by viral sequestration in early phagosomes would present a novel means to curb infection of enveloped virus and may constitute a prime defense system to prevent dengue virus spread shortly after the bite of the infected mosquito.

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Related in: MedlinePlus

MDdMφ are resistant to permissive DV infection.A. 5×105 MDDC and MDdMφ were exposed to DV1 at an MOI of 1 for 2 h, washed, and after 40 h processed for intracellular DV antigen detection by FACS. Graph shows the mean %±SD of intracellular DV antigens from 4 donors. * is significant (p<0.05) by the two-tailed student's t-test. B. Focus/plaque-forming assays (FFU/PFU) showing the mean titers±SD of DV1, 2 and 3 from supernatants of MDDC and MDdMφ (3 donors each). 5×105 cells were exposed to DV1 and DV3 at an MOI of 1 and to DV2 at an MOI of 2 for 2 h, washed, and after 40 h, the supernatant was collected. The infectious titer was determined on AP61 cells (DV1 and DV3) and on BHK cells (DV2). C. IFN-α secretion of non-infected (Mock) or infected MDDC and MDdMφ was determined with increasing MOI of DV1. The data is expressed as the mean±SD of triplicate values and is representative of 3 donors. D. Both cell-types were exposed to DV1 at an MOI of 100. Transmission electron micrograph showing virions bound to plasma membranes at 4°C, and internalization into vesicles 1 h after exposure to virus at 37°C.
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pntd-0000311-g003: MDdMφ are resistant to permissive DV infection.A. 5×105 MDDC and MDdMφ were exposed to DV1 at an MOI of 1 for 2 h, washed, and after 40 h processed for intracellular DV antigen detection by FACS. Graph shows the mean %±SD of intracellular DV antigens from 4 donors. * is significant (p<0.05) by the two-tailed student's t-test. B. Focus/plaque-forming assays (FFU/PFU) showing the mean titers±SD of DV1, 2 and 3 from supernatants of MDDC and MDdMφ (3 donors each). 5×105 cells were exposed to DV1 and DV3 at an MOI of 1 and to DV2 at an MOI of 2 for 2 h, washed, and after 40 h, the supernatant was collected. The infectious titer was determined on AP61 cells (DV1 and DV3) and on BHK cells (DV2). C. IFN-α secretion of non-infected (Mock) or infected MDDC and MDdMφ was determined with increasing MOI of DV1. The data is expressed as the mean±SD of triplicate values and is representative of 3 donors. D. Both cell-types were exposed to DV1 at an MOI of 100. Transmission electron micrograph showing virions bound to plasma membranes at 4°C, and internalization into vesicles 1 h after exposure to virus at 37°C.

Mentions: Monocyte-derived dMφ (MDdMφ) and monocyte-derived DC (MDDC) were analyzed for DV infection using low-passage DV1 and DV3 strains grown in mosquito cells [3] as well as the prototype DV2 strain 16681 [15]. The cells were exposed to DV1 at a multiplicity of infection (MOI) of 1 for 2 h, washed, and then cultured for 40 h. As shown in Figure 3A, intracellular viral antigen was clearly detected in MDDC by flow cytometry, whereas no specific immuno-labeling was observed in MDdMφ. An analysis of DV replication in these cells infected at an MOI of 1 (DV1 and DV3) or 2 (DV2) showed that MDDC were highly permissive to productive infection (∼105 FFU/ml or PFU/ml) (Fig. 3B); in contrast, progeny virus production was undetectable in DV-infected MDdMφ (<103 FFU/ml or PFU/ml). Consistent with this finding, no IFN-α was produced by DV-infected MDdMφ, even at an MOI of 10, whereas MDDC readily released IFN-α when infected with DV at an MOI of 1 or 10 [16] (Fig. 3C). To verify that MDdMφ acquired the virus, both myeloid cell-types were exposed to high DV input (MOI of 100) and electron microscopy analysis was performed after 30 min at 4°C and after 1 h at 37°C (Fig. 3D). Cell surface-bound (at 4°C) and endosomal vesicle-associated virus particles (at 37°C) were clearly detected in both cell-types. Thus, internalization of DV can occur in MDdMφ but does not result in productive infection.


Dermal-type macrophages expressing CD209/DC-SIGN show inherent resistance to dengue virus growth.

Kwan WH, Navarro-Sanchez E, Dumortier H, Decossas M, Vachon H, dos Santos FB, Fridman HW, Rey FA, Harris E, Despres P, Mueller CG - PLoS Negl Trop Dis (2008)

MDdMφ are resistant to permissive DV infection.A. 5×105 MDDC and MDdMφ were exposed to DV1 at an MOI of 1 for 2 h, washed, and after 40 h processed for intracellular DV antigen detection by FACS. Graph shows the mean %±SD of intracellular DV antigens from 4 donors. * is significant (p<0.05) by the two-tailed student's t-test. B. Focus/plaque-forming assays (FFU/PFU) showing the mean titers±SD of DV1, 2 and 3 from supernatants of MDDC and MDdMφ (3 donors each). 5×105 cells were exposed to DV1 and DV3 at an MOI of 1 and to DV2 at an MOI of 2 for 2 h, washed, and after 40 h, the supernatant was collected. The infectious titer was determined on AP61 cells (DV1 and DV3) and on BHK cells (DV2). C. IFN-α secretion of non-infected (Mock) or infected MDDC and MDdMφ was determined with increasing MOI of DV1. The data is expressed as the mean±SD of triplicate values and is representative of 3 donors. D. Both cell-types were exposed to DV1 at an MOI of 100. Transmission electron micrograph showing virions bound to plasma membranes at 4°C, and internalization into vesicles 1 h after exposure to virus at 37°C.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2553280&req=5

pntd-0000311-g003: MDdMφ are resistant to permissive DV infection.A. 5×105 MDDC and MDdMφ were exposed to DV1 at an MOI of 1 for 2 h, washed, and after 40 h processed for intracellular DV antigen detection by FACS. Graph shows the mean %±SD of intracellular DV antigens from 4 donors. * is significant (p<0.05) by the two-tailed student's t-test. B. Focus/plaque-forming assays (FFU/PFU) showing the mean titers±SD of DV1, 2 and 3 from supernatants of MDDC and MDdMφ (3 donors each). 5×105 cells were exposed to DV1 and DV3 at an MOI of 1 and to DV2 at an MOI of 2 for 2 h, washed, and after 40 h, the supernatant was collected. The infectious titer was determined on AP61 cells (DV1 and DV3) and on BHK cells (DV2). C. IFN-α secretion of non-infected (Mock) or infected MDDC and MDdMφ was determined with increasing MOI of DV1. The data is expressed as the mean±SD of triplicate values and is representative of 3 donors. D. Both cell-types were exposed to DV1 at an MOI of 100. Transmission electron micrograph showing virions bound to plasma membranes at 4°C, and internalization into vesicles 1 h after exposure to virus at 37°C.
Mentions: Monocyte-derived dMφ (MDdMφ) and monocyte-derived DC (MDDC) were analyzed for DV infection using low-passage DV1 and DV3 strains grown in mosquito cells [3] as well as the prototype DV2 strain 16681 [15]. The cells were exposed to DV1 at a multiplicity of infection (MOI) of 1 for 2 h, washed, and then cultured for 40 h. As shown in Figure 3A, intracellular viral antigen was clearly detected in MDDC by flow cytometry, whereas no specific immuno-labeling was observed in MDdMφ. An analysis of DV replication in these cells infected at an MOI of 1 (DV1 and DV3) or 2 (DV2) showed that MDDC were highly permissive to productive infection (∼105 FFU/ml or PFU/ml) (Fig. 3B); in contrast, progeny virus production was undetectable in DV-infected MDdMφ (<103 FFU/ml or PFU/ml). Consistent with this finding, no IFN-α was produced by DV-infected MDdMφ, even at an MOI of 10, whereas MDDC readily released IFN-α when infected with DV at an MOI of 1 or 10 [16] (Fig. 3C). To verify that MDdMφ acquired the virus, both myeloid cell-types were exposed to high DV input (MOI of 100) and electron microscopy analysis was performed after 30 min at 4°C and after 1 h at 37°C (Fig. 3D). Cell surface-bound (at 4°C) and endosomal vesicle-associated virus particles (at 37°C) were clearly detected in both cell-types. Thus, internalization of DV can occur in MDdMφ but does not result in productive infection.

Bottom Line: Here, we showed that dermal macrophages bound recombinant envelope E glycoprotein fused to green fluorescent protein.The macrophages were able to internalize the virus, but progeny virus production was undetectable in the infected cells.In addition, no IFN-alpha was produced in response to the virus.

View Article: PubMed Central - PubMed

Affiliation: CNRS, Laboratory of Therapeutic Immunology and Chemistry, IBMC, Université Louis Pasteur, Strasbourg, France.

ABSTRACT

Background: An important question in dengue pathogenesis is the identity of immune cells involved in the control of dengue virus infection at the site of the mosquito bite. There is evidence that infection of immature myeloid dendritic cells plays a crucial role in dengue pathogenesis and that the interaction of the viral envelope E glycoprotein with CD209/DC-SIGN is a key element for their productive infection. Dermal macrophages express CD209, yet little is known about their role in dengue virus infection.

Methods and findings: Here, we showed that dermal macrophages bound recombinant envelope E glycoprotein fused to green fluorescent protein. Because dermal macrophages stain for IL-10 in situ, we generated dermal-type macrophages from monocytes in the presence of IL-10 to study their infection by dengue virus. The macrophages were able to internalize the virus, but progeny virus production was undetectable in the infected cells. In addition, no IFN-alpha was produced in response to the virus. The inability of dengue virus to grow in the macrophages was attributable to accumulation of internalized virus particles into poorly-acidified phagosomes.

Conclusions: Aborting infection by viral sequestration in early phagosomes would present a novel means to curb infection of enveloped virus and may constitute a prime defense system to prevent dengue virus spread shortly after the bite of the infected mosquito.

Show MeSH
Related in: MedlinePlus