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DC-SIGN and CD150 have distinct roles in transmission of measles virus from dendritic cells to T-lymphocytes.

de Witte L, de Vries RD, van der Vlist M, Yüksel S, Litjens M, de Swart RL, Geijtenbeek TB - PLoS Pathog. (2008)

Bottom Line: Using immunofluorescence microscopy, we demonstrate that DC-SIGN+ DCs are abundantly present just below the epithelia of the respiratory tract.DC-SIGN+ DCs efficiently present MV-derived antigens to CD4+ T-lymphocytes after antigen uptake via either CD150 or DC-SIGN in vitro.However, DC-SIGN+ DCs also mediate transmission of MV to CD4+ and CD8+ T-lymphocytes.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Cell Biology and Immunology, VU University Medical Center, Amsterdam, The Netherlands.

ABSTRACT
Measles virus (MV) is among the most infectious viruses that affect humans and is transmitted via the respiratory route. In macaques, MV primarily infects lymphocytes and dendritic cells (DCs). Little is known about the initial target cell for MV infection. Since DCs bridge the peripheral mucosal tissues with lymphoid tissues, we hypothesize that DCs are the initial target cells that capture MV in the respiratory tract and transport the virus to the lymphoid tissues where MV is transmitted to lymphocytes. Recently, we have demonstrated that the C-type lectin DC-SIGN interacts with MV and enhances infection of DCs in cis. Using immunofluorescence microscopy, we demonstrate that DC-SIGN+ DCs are abundantly present just below the epithelia of the respiratory tract. DC-SIGN+ DCs efficiently present MV-derived antigens to CD4+ T-lymphocytes after antigen uptake via either CD150 or DC-SIGN in vitro. However, DC-SIGN+ DCs also mediate transmission of MV to CD4+ and CD8+ T-lymphocytes. We distinguished two different transmission routes that were either dependent or independent on direct DC infection. DC-SIGN and CD150 are both involved in direct DC infection and subsequent transmission of de novo synthesized virus. However, DC-SIGN, but not CD150, mediates trans-infection of MV to T-lymphocytes independent of DC infection. Together these data suggest a prominent role for DCs during the initiation, dissemination, and clearance of MV infection.

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Dendritic cells mediate antigen presentation of measles virus to CD4+ T-lymphocytes.(A) DCs (1×105 cells) were incubated with different concentrations of UV-MV. After 24 hours, the cells were stained for CD150, HLA-DR, CD86 and DC-SIGN using specific antibodies and analyzed by flow cytometry. (B,C) DCs or autologous B cells (BLCL) (5×104 cells) were pre-incubated with mannan and incubated with UV-MV or a positive control peptide overnight. The MV-specific CD4+ T-cell clones, GRIM99, GRIM61 and GRIM76 and the non-specific T-cell clone LB5 were added and T cell activation was determined by ELISPOT. GRIM99, GRIM61 and LB5 were dependent on HLA-DQw1 as expressed by the DCs, whereas GRIM76 was HLA-mismatched to the DCs used. Images of the ELISPOT (left) and the mean of the spot counts per well are depicted (right). Error bars represent the standard deviation of duplicates. (D) DCs (1×104 cells) were pre-incubated with mannan, anti-DC-SIGN or anti-CD150 and subsequently incubated with different concentrations of UV-MV. After 6 hours the MV specific CD4+ T-cell clone (5×104 cells) was added to the wells. After 24 hours the supernatant was harvested and the amount of IFN-γ analyzed by ELISA. Standard deviations represent the standard deviation of duplicates. A representative experiment out of two is shown.
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ppat-1000049-g002: Dendritic cells mediate antigen presentation of measles virus to CD4+ T-lymphocytes.(A) DCs (1×105 cells) were incubated with different concentrations of UV-MV. After 24 hours, the cells were stained for CD150, HLA-DR, CD86 and DC-SIGN using specific antibodies and analyzed by flow cytometry. (B,C) DCs or autologous B cells (BLCL) (5×104 cells) were pre-incubated with mannan and incubated with UV-MV or a positive control peptide overnight. The MV-specific CD4+ T-cell clones, GRIM99, GRIM61 and GRIM76 and the non-specific T-cell clone LB5 were added and T cell activation was determined by ELISPOT. GRIM99, GRIM61 and LB5 were dependent on HLA-DQw1 as expressed by the DCs, whereas GRIM76 was HLA-mismatched to the DCs used. Images of the ELISPOT (left) and the mean of the spot counts per well are depicted (right). Error bars represent the standard deviation of duplicates. (D) DCs (1×104 cells) were pre-incubated with mannan, anti-DC-SIGN or anti-CD150 and subsequently incubated with different concentrations of UV-MV. After 6 hours the MV specific CD4+ T-cell clone (5×104 cells) was added to the wells. After 24 hours the supernatant was harvested and the amount of IFN-γ analyzed by ELISA. Standard deviations represent the standard deviation of duplicates. A representative experiment out of two is shown.

Mentions: DC-SIGN is an attachment receptor for MV and mediates infection of DCs through CD150 in cis [11]. We investigated whether MV capture by DC-SIGN+ DCs also leads to antigen processing and presentation to MV-specific CD4+ T-lymphocytes. We performed an antigen presentation assay using MV-specific CD4+ T cell clones. [18],[19]. As APCs we used moDCs, expressing high levels of DC-SIGN [20] or an autologous Epstein-Barr virus-transformed B-lymphoblastic cell line (BLCL). The APCs were incubated with different dilutions of MV that was UV-inactivated to exclude DC and T-lymphocyte infection. Subsequently the APCs were co-cultured with the MV-specific CD4+ T-cell clones. At the highest concentrations, UV-MV induced DC maturation, since CD86 was upregulated, whereas DC-SIGN was down-regulated (Figure 2A). Notably, in contrast to LPS stimulation, HLA-DR was not upregulated by MV. T-cell activation was measured by the detection of IFN-γ production by ELISPOT and ELISA. We used two different MV-specific T cell clones (GRIM99 and GRIM61) that matched and one that mismatched (GRIM 76) the HLA type of the donor DCs. DCs incubated with UV-inactivated MV specifically activated the HLA-matched MV-specific T cell clones, whereas the mismatched MV-specific T cell clone was not activated (Figure 2B and C). Moreover, an irrelevant T cell clone (LB5) was not activated by the DCs (Figure 2B). Thus, MV capture by DCs leads to specific antigen processing and presentation of MV peptides in the context of MHC-class II molecules. MV-derived antigen presentation by DCs was more efficient at low antigen concentrations than presentation by autologous BLCL (Figure 2B); while the peptide control response was not significantly different (239+/−16 versus 264+/−18).


DC-SIGN and CD150 have distinct roles in transmission of measles virus from dendritic cells to T-lymphocytes.

de Witte L, de Vries RD, van der Vlist M, Yüksel S, Litjens M, de Swart RL, Geijtenbeek TB - PLoS Pathog. (2008)

Dendritic cells mediate antigen presentation of measles virus to CD4+ T-lymphocytes.(A) DCs (1×105 cells) were incubated with different concentrations of UV-MV. After 24 hours, the cells were stained for CD150, HLA-DR, CD86 and DC-SIGN using specific antibodies and analyzed by flow cytometry. (B,C) DCs or autologous B cells (BLCL) (5×104 cells) were pre-incubated with mannan and incubated with UV-MV or a positive control peptide overnight. The MV-specific CD4+ T-cell clones, GRIM99, GRIM61 and GRIM76 and the non-specific T-cell clone LB5 were added and T cell activation was determined by ELISPOT. GRIM99, GRIM61 and LB5 were dependent on HLA-DQw1 as expressed by the DCs, whereas GRIM76 was HLA-mismatched to the DCs used. Images of the ELISPOT (left) and the mean of the spot counts per well are depicted (right). Error bars represent the standard deviation of duplicates. (D) DCs (1×104 cells) were pre-incubated with mannan, anti-DC-SIGN or anti-CD150 and subsequently incubated with different concentrations of UV-MV. After 6 hours the MV specific CD4+ T-cell clone (5×104 cells) was added to the wells. After 24 hours the supernatant was harvested and the amount of IFN-γ analyzed by ELISA. Standard deviations represent the standard deviation of duplicates. A representative experiment out of two is shown.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2277461&req=5

ppat-1000049-g002: Dendritic cells mediate antigen presentation of measles virus to CD4+ T-lymphocytes.(A) DCs (1×105 cells) were incubated with different concentrations of UV-MV. After 24 hours, the cells were stained for CD150, HLA-DR, CD86 and DC-SIGN using specific antibodies and analyzed by flow cytometry. (B,C) DCs or autologous B cells (BLCL) (5×104 cells) were pre-incubated with mannan and incubated with UV-MV or a positive control peptide overnight. The MV-specific CD4+ T-cell clones, GRIM99, GRIM61 and GRIM76 and the non-specific T-cell clone LB5 were added and T cell activation was determined by ELISPOT. GRIM99, GRIM61 and LB5 were dependent on HLA-DQw1 as expressed by the DCs, whereas GRIM76 was HLA-mismatched to the DCs used. Images of the ELISPOT (left) and the mean of the spot counts per well are depicted (right). Error bars represent the standard deviation of duplicates. (D) DCs (1×104 cells) were pre-incubated with mannan, anti-DC-SIGN or anti-CD150 and subsequently incubated with different concentrations of UV-MV. After 6 hours the MV specific CD4+ T-cell clone (5×104 cells) was added to the wells. After 24 hours the supernatant was harvested and the amount of IFN-γ analyzed by ELISA. Standard deviations represent the standard deviation of duplicates. A representative experiment out of two is shown.
Mentions: DC-SIGN is an attachment receptor for MV and mediates infection of DCs through CD150 in cis [11]. We investigated whether MV capture by DC-SIGN+ DCs also leads to antigen processing and presentation to MV-specific CD4+ T-lymphocytes. We performed an antigen presentation assay using MV-specific CD4+ T cell clones. [18],[19]. As APCs we used moDCs, expressing high levels of DC-SIGN [20] or an autologous Epstein-Barr virus-transformed B-lymphoblastic cell line (BLCL). The APCs were incubated with different dilutions of MV that was UV-inactivated to exclude DC and T-lymphocyte infection. Subsequently the APCs were co-cultured with the MV-specific CD4+ T-cell clones. At the highest concentrations, UV-MV induced DC maturation, since CD86 was upregulated, whereas DC-SIGN was down-regulated (Figure 2A). Notably, in contrast to LPS stimulation, HLA-DR was not upregulated by MV. T-cell activation was measured by the detection of IFN-γ production by ELISPOT and ELISA. We used two different MV-specific T cell clones (GRIM99 and GRIM61) that matched and one that mismatched (GRIM 76) the HLA type of the donor DCs. DCs incubated with UV-inactivated MV specifically activated the HLA-matched MV-specific T cell clones, whereas the mismatched MV-specific T cell clone was not activated (Figure 2B and C). Moreover, an irrelevant T cell clone (LB5) was not activated by the DCs (Figure 2B). Thus, MV capture by DCs leads to specific antigen processing and presentation of MV peptides in the context of MHC-class II molecules. MV-derived antigen presentation by DCs was more efficient at low antigen concentrations than presentation by autologous BLCL (Figure 2B); while the peptide control response was not significantly different (239+/−16 versus 264+/−18).

Bottom Line: Using immunofluorescence microscopy, we demonstrate that DC-SIGN+ DCs are abundantly present just below the epithelia of the respiratory tract.DC-SIGN+ DCs efficiently present MV-derived antigens to CD4+ T-lymphocytes after antigen uptake via either CD150 or DC-SIGN in vitro.However, DC-SIGN+ DCs also mediate transmission of MV to CD4+ and CD8+ T-lymphocytes.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Cell Biology and Immunology, VU University Medical Center, Amsterdam, The Netherlands.

ABSTRACT
Measles virus (MV) is among the most infectious viruses that affect humans and is transmitted via the respiratory route. In macaques, MV primarily infects lymphocytes and dendritic cells (DCs). Little is known about the initial target cell for MV infection. Since DCs bridge the peripheral mucosal tissues with lymphoid tissues, we hypothesize that DCs are the initial target cells that capture MV in the respiratory tract and transport the virus to the lymphoid tissues where MV is transmitted to lymphocytes. Recently, we have demonstrated that the C-type lectin DC-SIGN interacts with MV and enhances infection of DCs in cis. Using immunofluorescence microscopy, we demonstrate that DC-SIGN+ DCs are abundantly present just below the epithelia of the respiratory tract. DC-SIGN+ DCs efficiently present MV-derived antigens to CD4+ T-lymphocytes after antigen uptake via either CD150 or DC-SIGN in vitro. However, DC-SIGN+ DCs also mediate transmission of MV to CD4+ and CD8+ T-lymphocytes. We distinguished two different transmission routes that were either dependent or independent on direct DC infection. DC-SIGN and CD150 are both involved in direct DC infection and subsequent transmission of de novo synthesized virus. However, DC-SIGN, but not CD150, mediates trans-infection of MV to T-lymphocytes independent of DC infection. Together these data suggest a prominent role for DCs during the initiation, dissemination, and clearance of MV infection.

Show MeSH
Related in: MedlinePlus