HIV-1 Inhibits Phagocytosis and Inflammatory Cytokine Responses of Human Monocyte-Derived Macrophages to P. falciparum Infected Erythrocytes

HIV-1 infection increases the risk and severity of malaria by poorly defined mechanisms. We investigated the effect of HIV-1Ba-L infection of monocyte-derived macrophages (MDM) on phagocytosis of opsonised P. falciparum infected erythrocytes (IE) and subsequent proinflammatory cytokine secretion. Compared to mock-infected MDM, HIV-1 infection significantly inhibited phagocytosis of IE (median (IQR) (10 (0–28) versus (34 (27–108); IE internalised/100 MDM; p = 0.001) and decreased secretion of IL-6 (1,116 (352–3,387) versus 1,552 (889–6,331); pg/mL; p = 0.0078) and IL-1β (16 (7–21) versus 33 (27–65); pg/mL; p = 0.0078). Thus inadequate phagocytosis and cytokine production may contribute to impaired control of malaria in HIV-1 infected individuals

Effects of Anacetrapib in Patients with Atherosclerotic Vascular Disease

BACKGROUND: Patients with atherosclerotic vascular disease remain at high risk for cardiovascular events despite effective statin-based treatment of low-density lipoprotein (LDL) cholesterol levels. The inhibition of cholesteryl ester transfer protein (CETP) by anacetrapib reduces LDL cholesterol levels and increases high-density lipoprotein (HDL) cholesterol levels. However, trials of other CETP inhibitors have shown neutral or adverse effects on cardiovascular outcomes. METHODS: We conducted a randomized, double-blind, placebo-controlled trial involving 30,449 adults with atherosclerotic vascular disease who were receiving intensive atorvastatin therapy and who had a mean LDL cholesterol level of 61 mg per deciliter (1.58 mmol per liter), a mean non-HDL cholesterol level of 92 mg per deciliter (2.38 mmol per liter), and a mean HDL cholesterol level of 40 mg per deciliter (1.03 mmol per liter). The patients were assigned to receive either 100 mg of anacetrapib once daily (15,225 patients) or matching placebo (15,224 patients). The primary outcome was the first major coronary event, a composite of coronary death, myocardial infarction, or coronary revascularization. RESULTS: During the median follow-up period of 4.1 years, the primary outcome occurred in significantly fewer patients in the anacetrapib group than in the placebo group (1640 of 15,225 patients [10.8%] vs. 1803 of 15,224 patients [11.8%]; rate ratio, 0.91; 95% confidence interval, 0.85 to 0.97; P=0.004). The relative difference in risk was similar across multiple prespecified subgroups. At the trial midpoint, the mean level of HDL cholesterol was higher by 43 mg per deciliter (1.12 mmol per liter) in the anacetrapib group than in the placebo group (a relative difference of 104%), and the mean level of non-HDL cholesterol was lower by 17 mg per deciliter (0.44 mmol per liter), a relative difference of -18%. There were no significant between-group differences in the risk of death, cancer, or other serious adverse events. CONCLUSIONS: Among patients with atherosclerotic vascular disease who were receiving intensive statin therapy, the use of anacetrapib resulted in a lower incidence of major coronary events than the use of placebo. (Funded by Merck and others; Current Controlled Trials number, ISRCTN48678192 ; ClinicalTrials.gov number, NCT01252953 ; and EudraCT number, 2010-023467-18 .)

Henipavirus V Protein Association with Polo-Like Kinase Reveals Functional Overlap with STAT1 Binding and Interferon Evasion▿

Emerging viruses in the paramyxovirus genus Henipavirus evade host antiviral responses via protein interactions between the viral V and W proteins and cellular STAT1 and STAT2 and the cytosolic RNA sensor MDA5. Polo-like kinase (PLK1) is identified as being an additional cellular partner that can bind to Nipah virus P, V, and W proteins. For both Nipah virus and Hendra virus, contact between the V protein and the PLK1 polo box domain is required for V protein phosphorylation. Results indicate that PLK1 is engaged by Nipah virus V protein amino acids 100 to 160, previously identified as being the STAT1 binding domain responsible for host interferon (IFN) signaling evasion, via a Thr-Ser-Ser-Pro motif surrounding residue 130. A distinct Ser-Thr-Pro motif surrounding residue 199 mediates the PLK1 interaction with Hendra virus V protein. Select mutations in the motif surrounding residue 130 also influenced STAT1 binding and innate immune interference, and data indicate that the V:PLK1 and V:STAT complexes are V mediated yet independent of one another. The effects of STAT1/PLK1 binding motif mutations on the function the Nipah virus P protein in directing RNA synthesis were tested. Remarkably, mutations that selectively disrupt the STAT or PLK1 interaction site have no effects on Nipah virus P protein-mediated viral RNA synthesis. Therefore, mutations targeting V protein-mediated IFN evasion will not alter the RNA synthetic capacity of the virus, supporting an attenuation strategy based on disrupting host protein interactions

Time course of the effect of HIV-1 on cytokine secretion of MDM exposed to antibody opsonised IE.

<p>MDM from 3 donors cultured as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0032102#pone-0032102-g001" target="_blank">Figure 1</a> were infected with HIV-1_Ba-L (•) or mock-infected (○) and primed for 48 hrs with IFNγ then exposed to IE opsonised with 9% pooled immune serum (IE-PPS) at a target to cell ratio of 20∶1. At the indicated time points culture supernatants were collected for (<b>A</b>) IL-1β, (<b>B</b>) TNF and (<b>C</b>) IL-6 cytokine measurement. Data are means of MDM derived from 3 donors plus-minus SEM. Statistical analysis was carried out using the generalised estimating equations method which indicated that mock-infected MDM secreted significantly higher levels of cytokines compared to HIV-1 infected MDM (p = 0.026, IL-1β; p = 0.047, TNF; p = 0.019, IL-6).</p

HIV-1 infection of MDM inhibits phagocytosis of antibody opsonised CS2 infected erythrocytes.

<p>(<b>A</b>). MDM from 11 donors were cultured for 5 days and infected with HIV-1_Ba-L (•) at a MOI of between 0.1 and 1.0 or mock-infected (○) then cultured for an additional 7 days. IE opsonised with 9% PPS were added at a 20∶1 ratio and phagocytosis was measured at 1 hour. Phagocytic index represents the number of ingested IE per 100 MDM. Statistical comparisons were performed in a pair wise manner using the Wilcoxon matched pairs test. (<b>B</b>). In 6 selected cultures analysed for phagocytosis, XTT assay was used to measure viability of MDM infected with HIV-1_Ba-L (•) or mock-infected (○) MDM (3 mg/mL XTT, 4 hr, absorbance was read at 450 nm using a reference wavelength of 650 nm). (<b>C</b>). Effect of IFNγ on HIV-1_Ba-L production by MDM. MDM were cultured in triplicate in 96-well plates for 5 days, infected with HIV-1_Ba-L for a further 5 days then primed with 100 ng/mL IFNγ where indicated. After 72 hrs media from triplicate wells was collected and analysed by micro RT assay. Comparisons between unprimed and primed MDM were made using the Wilcoxon matched pairs test and a significant difference (p&lt;.05) is indicated.</p

The effect of HIV-1 on cytokine mRNA levels of MDM exposed to antibody opsonised IE.

<p>MDM from 6 donors cultured in 24-well plates were infected with HIV-1_Ba-L (•) or mock-infected (○) and primed for 48 hrs with IFNγ then exposed to IE opsonised with 9% pooled immune serum (IE-PPS) at a target to cell ratio of 20∶1 for 2 hrs followed by preparation of RNA and cDNA for analysis of (<b>A</b>) IL-1β, (<b>B</b>) TNF and (<b>C</b>) IL-6 cytokine mRNA levels by qPCR. Comparisons of mRNA levels between cultures infected with HIV-1_Ba-L or mock-infected exposed to IE-PPS were made using the Mann-Whitney non-parametric U test.</p

The effect of HIV-1 on cytokine secretion of MDM exposed to antibody opsonised IE.

<p>MDM from 8 donors cultured as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0032102#pone-0032102-g001" target="_blank">Figure 1</a> were infected with HIV-1_Ba-L (•) or mock-infected (○) and primed for 48 hrs with IFNγ then exposed to unopsonised (IE) or IE opsonised with 9% pooled immune serum (IE-PPS) at a target to cell ratio of 20∶1 for 24 hrs followed by collection of culture medium for (<b>A</b>) IL-1β, (<b>B</b>) TNF, (<b>C</b>) IL-6 and (<b>D</b>) IL-8 cytokine secretion analysis. Comparisons of cytokine secretion between cultures infected with HIV-1_Ba-L exposed to IE or IE-PPS, and between HIV-1_Ba-L infected and uninfected cultures exposed to IE-PPS were made using the Wilcoxon matched pairs test.</p

PBMC derived from G5–7 secrete higher levels of CXCL8, CCL8, IFNγ and CXCL10.

<p>PBMC (2×10⁵/well) were exposed to 6×10⁵ unopsonised IE or IE opsonised with 9% HI autologous plasma (IE-IgG). After 48 h, culture supernatants were quantified for (<b>A</b>) CXCL8, (<b>B</b>) CCL8, (<b>C</b>) IFNγ and (<b>D</b>) CXCL10. Both CXCL8 and CCL8 were quantified using ELISA while IFNγ and CXCL10 were quantified using a commercial multi-plex assay. Cytokine production from resting PBMC was subtracted from IE induced cytokine to eliminate the contribution of constitutive cytokine production and data is presented on a log₁₀ scale. Comparison between three groups (G1, G2–4, G5–7) was achieved using a non-parametric One Way ANOVA: Kruskal-Wallis test and the Dunn's Multiple Comparison post-test. * p&lt;0.05, *** p&lt;0.001.</p