HSV-2 Infection of Dendritic Cells Amplifies a Highly Susceptible HIV-1 Cell Target

Herpes simplex virus type 2 (HSV-2) increases the risk of HIV-1 infection and, although several reports describe the interaction between these two viruses, the exact mechanism for this increased susceptibility remains unclear. Dendritic cells (DCs) at the site of entry of HSV-2 and HIV-1 contribute to viral spread in the mucosa. Specialized DCs present in the gut-associated lymphoid tissues produce retinoic acid (RA), an important immunomodulator, able to influence HIV-1 replication and a key mediator of integrin α4β7 on lymphocytes. α4β7 can be engaged by HIV-1 on the cell-surface and CD4+ T cells expressing high levels of this integrin (α4β7high) are particularly susceptible to HIV-1 infection. Herein we provide in-vivo data in macaques showing an increased percentage of α4β7high CD4+ T cells in rectal mucosa, iliac lymph nodes and blood within 6 days of rectal exposure to live (n = 11), but not UV-treated (n = 8), HSV-2. We found that CD11c+ DCs are a major target of HSV-2 infection in in-vitro exposed PBMCs. We determined that immature monocyte-derived DCs (moDCs) express aldehyde dehydrogenase ALDH1A1, an enzyme essential for RA production, which increases upon HSV-2 infection. Moreover, HSV-2-infected moDCs significantly increase α4β7 expression on CD4+ T lymphocytes and HIV-1 infection in DC-T cell mixtures in a RA-dependent manner. Thus, we propose that HSV-2 modulates its microenviroment, influencing DC function, increasing RA production capability and amplifying a α4β7highCD4+ T cells. These factors may play a role in increasing the susceptibility to HIV-1

Pivotal role of Ccr1 in murine lupus nephritis

Systemic Lupus Erythematosus is a chronic, inflammatory autoimmune disease that affects multiple organs including the kidney. Mononuclear cell infiltration in both glomerular and tubulointerstitial compartments characterizes human and experimental lupus nephritis and is associated with a progressive loss of renal function. Molecular recruitment mechanisms into chronically inflamed kidneys have not been fully characterized especially in the lupus-prone New Zealand Black/New Zealand White (NZB/W) mouse model. By combining pharmacologic and functional approaches, we uncover a previously unappreciated role for the chemokine receptor Ccr1 in renal infiltration of myeloid and T cells in nephritis NZB/W mice. We revealed a functional expression of Ccr1 on peripheral T cells, macrophages and neutrophils from NZB/W mice. Acute treatment of nephritis NZB/W mice with the orally available Ccr1 antagonist BL5923 reduced kidney recruitment of myeloid and T cells, while sparing B cells. Late onset BL5923-based treatment delayed proteinuria and death in nephritis mice. This is likely related to the beneficial effect of Ccr1 blockade on interstitial infiltration of T cells and macrophages as well as on tubulointerstitial and glomerular injuries. In contrast, systemic and renal humoral autoimmunity was unaffected in BL5923-treated mice. Altogether, these findings highlight a pivotal role for Ccr1 in recruiting T and myeloid cells to inflamed kidneys of NZB/W mice and that such activity contributes to the progression of renal injury

PolyICLC Exerts Pro- and Anti-HIV Effects on the DC-T Cell Milieu In Vitro and In Vivo.

Myeloid dendritic cells (mDCs) contribute to both HIV pathogenesis and elicitation of antiviral immunity. Understanding how mDC responses to stimuli shape HIV infection outcomes will inform HIV prevention and treatment strategies. The long double-stranded RNA (dsRNA) viral mimic, polyinosinic polycytidylic acid (polyIC, PIC) potently stimulates DCs to focus Th1 responses, triggers direct antiviral activity in vitro, and boosts anti-HIV responses in vivo. Stabilized polyICLC (PICLC) is being developed for vaccine adjuvant applications in humans, making it critical to understand how mDC sensing of PICLC influences HIV infection. Using the monocyte-derived DC (moDC) model, we sought to describe how PICLC (vs. other dsRNAs) impacts HIV infection within DCs and DC-T cell mixtures. We extended this work to in vivo macaque rectal transmission studies by administering PICLC with or before rectal SIVmac239 (SIVwt) or SIVmac239ΔNef (SIVΔNef) challenge. Like PIC, PICLC activated DCs and T cells, increased expression of α4β7 and CD169, and induced type I IFN responses in vitro. The type of dsRNA and timing of dsRNA exposure differentially impacted in vitro DC-driven HIV infection. Rectal PICLC treatment similarly induced DC and T cell activation and pro- and anti-HIV factors locally and systemically. Importantly, this did not enhance SIV transmission in vivo. Instead, SIV acquisition was marginally reduced after a single high dose challenge. Interestingly, in the PICLC-treated, SIVΔNef-infected animals, SIVΔNef viremia was higher, in line with the importance of DC and T cell activation in SIVΔNef replication. In the right combination anti-HIV strategy, PICLC has the potential to limit HIV infection and boost HIV immunity

CXCR4 dysfunction in non-alcoholic steatohepatitis in mice and patients

Abstract Homing of inflammatory cells to the liver is key in the progression of non-alcoholic steatohepatitis (NASH). An abnormal response of CD4 + T-cells from obese mice to the chemotactic effect of CXCL12 has been reported but the mechanism involved in this process and relevance in patients are unknown. We aimed to explore the mechanism involved in the abnormal chemotaxis of CXC chemokine ligand 12 (CXCL12) in several mouse models of NASH and the relevance in the context of human non-alcoholic fatty liver disease (NAFLD). We assessed chemotactic responsiveness of CD4 + T-cells to CXCL12, the effect of AMD3100, a CXC chemokine receptor 4 (CXCR4) antagonist, in mice and lymphocytes from patients with NAFLD, and the affinity of CXCL12 for CXCR4. CXCL12-promoted migration of CD4 + T-cells from three different mouse models of NASH was increased and dependent of CXCR4. CD4 + T-cells from patients with NASH, but not from patients with pure steatosis, responded more strongly to the chemotactic effect of CXCL12, and this response was inhibited by AMD3100. Treatment with AMD3100 decreased the number of CD4 + T-cells to the liver in ob/ob mice. CXCL12 expression in the liver, CXCR4 and CXCR7 expression in CD4 + T-cells were not increased in three different mouse models of NASH. However, the affinity of CXCL12 for CXCR4 was increased in CD4 + T-cells of ob/ob mice. In conclusion, the CXCL12/CXCR4 pathway contributes in both mice and patients to the enhanced recruitment of CD4 + T-cells in NASH. An increased affinity of CXCL12 to CXCR4 rather than a higher expression of the chemokine or its receptors is involved in this process

Rhesus macaques used in challenge studies.

<p>Rhesus macaques used in challenge studies.</p

Rectal CD169 and β₇ expression correlate with systemic virus replication but do not predict infection.

<p>CD169 mRNA levels were measured in (A) rectal tissues and (B) inguinal LNs from macaques infected with SIVwt and SIVΔNef at different times post-infection and in uninfected macaques (baseline of the infected and other macaques that did not become infected within the study). Correlations between rectal CD169 level and viral replication in (C) SIVwt and (D) SIVΔNef-infected macaques are shown. (E) Relationship between baseline rectal CD169 expression and infection outcome for SIVwt and SIVΔNef. (F) Correlation between baseline rectal β₇ level and peak viral loads in SIVwt and SIVΔNef-infected macaques is shown. (G) Relationship between baseline rectal β₇ level and infection outcome. (H) Correlation between rectal CD169 and β₇ levels at baseline for all animals challenged with SIVwt and SIVΔNef. In (A-H), samples from all challenged macaques were not available at every time point. In (A-B), statistical analyses used the Kruskal Wallis test and Dunns post-test. In (A), Dunns comparisons not made were SIVwt W8 vs. SIVΔNef W20 and SIVwt W28 vs. SIVΔNef W6. In (C), (D), (F), and (H), Spearman correlation coefficient was determined. *<i>P</i><0.05, **<i>P</i><0.01, ***<i>P</i><0.001.</p

dsRNAs mediate changes in HIV location and T cell phenotype within co-cultures.

<p>HIV-pulsed DCs were co-cultured with autologous CD4⁺ T cells in the presence or absence of dsRNAs as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0161730#pone.0161730.g001" target="_blank">Fig 1</a>. After 24 hours, cells were collected, surface stained, and intracellularly stained for p24. (A) Conjugate frequency within DC-T cell co-cultures was defined as the proportion of live CD3⁺CD4⁺ large cells in the co-cultures (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0161730#sec002" target="_blank">Methods</a>). (B) Frequency of p24⁺ cells within the populations of free T cells (single) and conjugated T cells (conj). (C) CD69 GMFI and (D) the percentage of α₄β₇^highCD45RO⁺ CD4⁺ T cells were monitored within the single and conjugated T cell populations. For (A-D), 5 donors and the medians are shown, and the Friedman test with Dunns post-test was used to analyze the data. Dunns post-test excluded comparisons of PIC vs. piclcDC and PICLC vs. picDC. *<i>P</i><0.05, **<i>P</i><0.01.</p

Rectal PICLC modestly decreases SIV transmission but increases SIVΔNef replication in infected animals and promotes the vaccine effect.

<p>(A) Macaques were rectally challenged with 3000 TCID₅₀ SIVmac239 (SIVwt) coincident with (n = 7, “Coincident”) or 24 hours after (n = 7, “24h pre”) rectal PICLC or 24 hours after rectal PBS (n = 8). The fraction of PBS vs. PICLC-treated macaques that became infected is shown as a percent, and the number of animals infected is above each bar. (B) SIV RNA copies/ml were measured over time in each animal shown in (A). (C) Plasma viremia in infected animals in each group is shown at peak (highest observed viremia, 2–4 weeks post-challenge in all macaques, left) and 16 weeks post-challenge (middle), and as the area under the curve (AUC) of viremia over the whole observation period (right). The timing of PICLC administration is denoted by the symbols used in (B). (D) Macaques were rectally challenged with 3000 TCID₅₀ SIVmac239ΔNef (SIVΔNef) 24 hours after rectal PICLC (n = 7) or PBS (n = 7). 12 weeks after SIVΔNef challenge, all animals were rectally challenged with 3000 TCID₅₀ SIVwt. The fraction of PBS vs. PICLC-treated animals that became infected with SIVΔNef is shown as a percent, and the number of SIVΔNef-infected macaques is above each bar. (E) SIV RNA copies/ml of SIVΔNef (open symbols) and SIVwt (filled symbols) were measured over time in each animal. The two SIVΔNef-infected macaques not protected from SIVwt are shown in red. (F) SIVΔNef plasma viremia in each group is shown at peak (2–4 weeks post-challenge, left) and 16wks post-challenge (4 weeks post-SIVwt challenge, middle), and as AUC (right). The two macaques not protected from SIVwt are shown in red. (G) Fraction of SIVΔNef-infected animals that subsequently also became infected with SIVwt is shown by treatment group. In (C) and (F), P values were derived from Mann Whitney test comparisons of the control group with each of the treated groups.</p

Rectally applied PICLC induces rapid local and systemic immune changes.

<p>Macaques (n = 11) were bled 4 hours (4h) and 24h after rectal PBS vs. PICLC application. Either 4h (n = 6) or 24h (n = 5) after receiving treatment, rectal biopsies were also collected. (A) Blood mDCs were characterized at the indicated times post-treatment by their frequency (%Lin^-HLA-DR⁺CD11c^high) and expression of CD80 and CCR7. (B) Blood and (C) rectal CD4⁺ T cells were characterized by their expression of CD69 and CCR7 and the frequency of α₄β₇^highCD95⁺ cells. (D) mRNA levels of the markers shown were measured in rectal tissue. (E) In a separate group of macaques biopsied 5 weeks before (Pre) and 24 hours after (Post) a single rectal application of 2 mg (filled symbols) or 4 mg (open symbols) PICLC, mRNA levels of the markers from (D) were measured in cells isolated from rectal tissue. In (A-E), statistical analyses using the Wilcoxon Signed Rank test compared the post-PICLC time points with control post-PBS time points in each animal. *<i>P</i><0.05, **<i>P</i><0.01, ***<i>P</i><0.001.</p