Release of chemokines into the supernatants of DC and MP infected with LASV and MOPV.

<p>The levels of CC and CXC chemokines were quantified by ELISA in the supernatants of MP (A) and DC (B) 24 and 72 h after mock (white bars), MOPV (gray bars), and LASV (black bars) infection. <a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0002637#s3" target="_blank">Results</a> are expressed in pg/ml and ng/ml for CC and CXC chemokines, respectively. Significant differences are indicated as follows: * (<i>p</i><0.05), ** (<i>p</i><0.01), and *** (<i>p</i><0.001).</p

Production of chemokines by APC infected with recombinant LASV.

<p>The production of CC and CXC chemokines by DC (A, B) and MP (C, D) was assessed after mock infection (white bars), or infection with recombinant wild-type LASV (gray bars) or rLASV NP-D389A/G392A (rNP LASV) (black bars). (A, C) The synthesis of mRNAs was analyzed by RT-qPCR 24 h after infection. (B, D) The protein levels released in the supernatants were quantified by ELISA 24 h (only for CXCL9, 10 and 11) and 48 h after infection. <a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0002637#s3" target="_blank">Results</a> are expressed as the mean ± SE of 4 (A, C) and 3–7 (B, D) independent experiments. Significant differences are indicated as follows: * (<i>p</i><0.05), ** (<i>p</i><0.01) and *** (p<0.001).</p

Production of type I IFN and chemokine mRNA in DC and MP infected with LASV and MOPV.

<p>We obtained mRNA from DC and MP 24(black bars), MOPV (gray bars), or after mock infection (white bars). (A) The amounts of type I IFN mRNA produced by DC and MP are shown as a gene mRNA/β-actin mRNA ratio. The synthesis of chemokine mRNA was evaluated with the GAPDH gene used as a housekeeping gene, in both DC (B) and MP (C). <a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0002637#s3" target="_blank">Results</a> are expressed as the mean ± standard error (SE) of 4 and 5 independent experiments, for LASV- and MOPV-infected cells, respectively. Significant differences between mock- and virus-infected cells are indicated as follows: * (<i>p</i><0.05), ** (<i>p</i><0.01), and *** (<i>p</i><0.001).</p

Role of type I IFN in the production of chemokines in MOPV- and LASV-infected iDC/T-cell cocultures.

<p>(A) The synthesis of mRNAs for CCL4, CCL7, CXCL10 and CXCL11 was evaluated in mock- (gray bars) or MOPV- (black bars) infected iDC cocultured with naïve T cells for 2 days in the presence of neutralizing Ab against CD118. (B) The levels of these mRNAs then were quantified 2 days after restimulation with mock- (gray bars) or inactivated MOPV- (black bars) pulsed iDC of T cells previously cocultured with mock- or MOPV-infected iDC in the presence of irrelevant IgG2a or CD118-neutralizing Ab. <a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0002637#s3" target="_blank">Results</a> are expressed as the mean ± SE of 3 independent experiments.</p

Production of chemokines by MOPV- and LASV-infected DC cocultured with T cells and correlation with type I IFN synthesis.

<p>(A) Levels of CC chemokine and CXCL11 mRNA were quantified relative to GAPDH mRNA levels, after 2 days of coculture of mock- (white bars), MOPV- (gray bars), or LASV- (black bars) infected iDC and autologous T cells. Levels of CXCL10 mRNA were evaluated in iDC/T cell coculture 2 days after each stimulation of the T cells with infected or inactivated virus-stimulated iDC. <a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0002637#s3" target="_blank">Results</a> are expressed as the mean ± standard error (SE) of 4 independent experiments. Significant differences are indicated as follows: * (<i>p</i><0.05) and ** (<i>p</i><0.01). (B) The levels of CC and CXC chemokines were quantified by ELISA in the supernatants after 2 days of coculture of mock- (white bars), MOPV- (gray bars), or LASV- (black bars) infected iDC and autologous T cells. <a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0002637#s3" target="_blank">Results</a> are expressed in pg/ml, except for CXCL10 expressed in ng/ml. Significant differences are indicated as follows: * (<i>p</i><0.05). (C) Correlation between CCL4, CCL7, CXCL10, and CXCL11 mRNA levels and type I IFN synthesis by MOPV- or LASV-infected iDC cultured with naïve T cells 2 days after infection, represented by a linear regression with a correlation coefficient <i>r</i> and a probability of correlation <i>p</i>.</p