Mice lacking RPMs exhibit wild type infection kinetics.

<p>(A) Parasitemia courses in 129Sv <i>SpiC^+/−</i> (<i>n</i> = 4) and <i>SpiC^−/−</i> (<i>n</i> = 5) mice are represented as geometric means with standard deviations and Mann-Whitney <i>p</i>-value. (B) Ly6c^lo monocyte (CD11b⁺ F4/80⁺ Ly6g⁻ SSC^lo Ly6c^lo) frequencies in blood and spleen of 129Sv <i>SpiC^+/−</i> (white bars) and <i>SpiC^−/−</i> mice (black bars) during the course of infection. Days depicted in blue and orange represent a 1.5-fold decrease or increase, respectively, in Ly6c^lo monocyte frequencies in blood and spleen of mice infected with <i>P. chabaudi</i>. (C) Ly6c^lo monocyte frequencies on day 12 post-infection. Means are presented with standard errors; <i>p</i>-values represent a two-tailed <i>t</i>-test assuming unequal variances. Data represent three independent experiments (<i>n</i> = 6–7 mice per group total).</p

T1IFNs contribute to control of <i>P. chabaudi</i> infection.

<p>Infected mice were monitored for parasitemia by thin blood smear and survival. Wild type C57BL/6 and congenic knockout mice were infected and monitored for percent parasitemia (<i>n</i> = 5 per strain), which are represented as geometric means with standard deviations and Mann-Whitney <i>p</i>-value. A representative experiment of two is shown. Crosses indicate deaths due to parasitemia.</p

<i>P. chabaudi</i> infection induces IFNB production in pDCs and RPMs.

<p>(A) No splenocytes are YFP⁺ in mock-infected samples, but some splenocytes become YFP⁺24 h after <i>P. chabaudi</i> infection of C57BL/6 <i>Ifnb</i> reporter mice. 2.5×10⁶ events are depicted in each dot plot. (B) pDCs and RPMs constitute over 90% of YFP⁺ events in C57BL/6 mice. (C) Both pDCs and RPMs contribute to splenic IFNB production in 129Sv mice. pDCs were depleted with a single 500 µg dose of anti-mPDCA1 antibody 18 h before infection with <i>P. chabaudi</i>. Grey dots represent individual mice, with horizontal bars representing the mean (B) or geometric mean (C).</p

Cellular requirements for splenic T1IFN transcriptional induction.

<p>(A) RPMs, but not other macrophage or dendritic cell subsets, induce <i>Ifna</i> and <i>Ifnb</i> at 24 h post-infection with <i>P. chabaudi</i> as detected by qRT-PCR in C57BL/6 mice. Fold mRNA induction represents fold induction of transcript in infected versus mock-infected normalized to beta-actin. Grey dots represent independent experiments conducted on different days; black bars denote the geometric means of the fold inductions. (B) pDCs are not required for splenic <i>Ifna</i> or <i>Ifnb</i> transcriptional induction in response to <i>P. chabaudi</i> in C57BL/6 mice. (C) Genetic deletion of RPMs in 129Sv mice results in diminished T1IFN transcriptional induction. (D) Genetic deletion of <i>Myd88</i> from dendritic cells does not impact transcriptional induction of T1IFNs in spleens of C57BL/6 mice. (E) Genetic deletion of <i>Myd88</i> from macrophages/neutrophils decreases transcriptional induction of T1IFNs by an order of magnitude in C57Bl/6 mice. Grey bars represent geometric means with 95% confidence intervals. Asterisks represent <i>p</i><0.05 in a Student’s <i>t</i>-test against control samples.</p

T1IFNs are produced during <i>P. chabaudi</i> infection.

<p>(A) Kinetics of early <i>Ifna, Ifnb</i>, and <i>Ifng</i> transcription using whole spleen qRT-PCR. Fold mRNA induction represents the ratio of transcript in infected over mock-infected C57BL/6 mice. (B) Reproducibility of T1IFN transcript induction as detected by whole spleen qRT-PCR. Each point represents an independent experiment with 4–6 animals, with horizontal bars displaying the geometric mean. (C) Plasma IFNA and IFNB at 24 h post-infection. Data are combined from two independent experiments with each point representing one animal. N.D. = not detected (<i>n</i> = 8).</p

T1IFN and IFNG signaling redundantly regulate early gene expression responses to <i>P. chabaudi</i> infection.

<p>A representative set of ISG is shown for the gene expression response in whole blood from animals infected for 24 h with <i>P. chabaudi</i> in C57BL/6 knockout mice. Each column represents an individual mouse.</p

Molecular requirements for splenic T1IFN transcriptional induction.

<p>(A) <i>Tlr9</i> and <i>Myd88</i> are required for full transcriptional induction of T1IFNs in spleens of <i>P. chabaudi</i>-infected C57Bl/6 mice. Grey dots represent the means of independent experiments using 4–6 total mice, with T1IFN fold mRNA induction in knockout mice represented as a percentage of induction in wild type animals. Black bars represent means; asterisks represent <i>p</i><0.05 as compared to wild type induction in a two-tailed Student’s <i>t</i>-test assuming unequal variances. (B) <i>Irf7</i> is required for full T1IFN induction, and <i>Irf3</i> is required for full <i>Ifna</i> but not <i>Ifnb</i> induction. (C) <i>Ifnar1</i> is required for full <i>Ifna</i> induction but dispensable for <i>Ifnb</i> induction, whereas <i>Ifngr1</i> is dispensable for all T1IFN induction.</p

Synthesis and Evaluation of 7-Substituted 4-Aminoquinoline Analogues for Antimalarial Activity

We previously reported that substituted 4-aminoquinolines with a phenyl ether substituent at the 7-position of the quinoline ring and the capability of intramolecular hydrogen bonding between the protonated amine on the side chain and a hydrogen bond acceptor on the amine’s alkyl substituents exhibited potent antimalarial activity against the multidrug resistant strain <i>P</i>. <i>falciparum</i> W2. We employed a parallel synthetic method to generate diaryl ether, biaryl, and alkylaryl 4-aminoquinoline analogues in the background of a limited number of side chain variations that had previously afforded potent 4-aminoquinolines. All subsets were evaluated for their antimalarial activity against the chloroquine-sensitive strain 3D7 and the chloroquine-resistant K1 strain as well as for cytotoxicity against mammalian cell lines. While all three arrays showed good antimalarial activity, only the biaryl-containing subset showed consistently good potency against the drug-resistant K1 strain and good selectivity with regard to mammalian cytotoxicity. Overall, our data indicate that the biaryl-containing series contains promising candidates for further study