Clinical characteristics of patients with persistent HBV infection (n = 201).

<p>Characteristics are shown for all patients as well as inactive HBsAg carriers (n = 48) and active CHB patients (n = 46) in whom longitudinal data were available.</p><p>IC, inactive HBsAg carriers; CHB, active chronic hepatitis B;</p>*<p>HBV genotype was available in 67 patients (25 in IC and 18 in CHB).</p

Association of <i>HLA-DPA1</i> variant rs3077 and <i>HLA-DPB1</i> variant rs9277535 with HBV infection.

*<p>controls include either healthy subjects with negative serological HBV markers (vs. HBV patients) or inactive HBsAg carriers in the subgroup analysis of patients with HBV infection (vs. active CHB patients);</p>**<p>to sustain the conventional gene description from the 5′ end to the 3′ end we changed the SNP denomination in comparison to previous publications. That is, the rs3077T/C alleles in our study correspond to the A/G alleles, respectively, in previously published studies <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0032605#pone.0032605-Kamatani1" target="_blank">[9]</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0032605#pone.0032605-Guo1" target="_blank">[11]</a>.</p

Potent inhibition of HIV-1 replication by the maC46 fusion inhibitor and antisense-Env VRX494 vectors.

<p>Non-transduced CEMx174 cells (NT) and CEMx174 cells transduced with vectors expressing the indicated inhibitor gene or a control vector (HJ57, GFP) were infected with HIV-1_NL4-3 at MOIs of (A) 10⁻⁴, (B) 10⁻³, and (C) 10⁻² TCID₅₀ per cell. Viral replication was assessed by measuring HIV-1 Gag p24 antigen in cell-free culture supernatants. The minimum level of detection for HIV-1 Gag p24 was 15 pg/ml. The data are representative of at least two experiments.</p

Schematic diagrams of the viral transfer plasmids.

<p>The lentiviral vectors HIV-shI-GFP, VRX494, M589, HJ57, and M420, are based on HIV-1_NL4-3 or HIV-1_HXB2. The viral inhibitors encoded by each of the experimental vectors are noted in parentheses. Where indicated, vectors use a heterologous CMV promoter to initiate transcription of the genomic RNA with a self-inactivating 3′ LTR. The VRX494 vector uses a functional HIV-1 LTR, which is upregulated after viral infection <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0012357#pone.0012357-Braun2" target="_blank">[36]</a>. All vectors contain cis-acting regulatory domains (the ψ packaging signal, the central polypurine tract [cppt], the Rev response element [RRE], and, in some cases, the Woodchuck post-transcriptional regulatory element [wPRE]), and eGFP. The vector HIV-shI-GFP contains the U6 promoter regulating a shRNA targeting exon 1 of HIV-1 <i>tat</i> and <i>rev</i> (shI) <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0012357#pone.0012357-Lee1" target="_blank">[10]</a>. The lentiviral vector VRX494 contains 937 bp of antisense (AS) HIV-1 envelope, and eGFP transcriptionally regulated by the HIV-1 LTR <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0012357#pone.0012357-Lu2" target="_blank">[32]</a>. The vector M589 contains an internal SFFV promoter regulating expression of the C46 heptad repeat-anchored with a linker and transmembrane domain:GFP fusion protein (maC46:GFP). The control vectors HJ57 and M420 do not contain an inhibitor cassette.</p

Self-reported risk factors for HCV transmission in the anti-HCV positive population at the Berlin study site.

<p>IVDU, intravenous drug-use.</p><p>Multiple responses were possible. Percentages are given in relation to all anti-HCV positive patients (n = 535).</p

Preferential survival of cells expressing the maC46 fusion inhibitor following HIV-1 infection.

<p>A. Flow cytometric analysis for expression of GFP and HIV-1 p24 Gag at the indicated times post-infection. Non-transduced CEMx174 cells were mixed with transduced cells expressing the maC46 inhibitors such that approximately 25–30% of the cells were GFP⁺. Mixtures of cells were then infected with HIV-1_NL4-3 at a MOI of 10⁻³ TCID₅₀ per cell. Expression of HIV-1 p24 Gag and GFP were determined by flow cytometry on days 5, 9, 14 and 21. B. Survival advantage of cells expressing maC46 following HIV-1 infection. Mixtures of non-transduced CEMx174 cells and transduced cells expressing the shI, maC46, and antisense <i>env</i> inhibitors, or transduced with the HJ57 control vector (GFP) were generated such that approximately 25–30% of the cells were GFP⁺. Cells were then infected with HIV-1_NL4-3 at MOI of 10⁻³ TCID₅₀ per cell or remained uninfected. The percentage of GFP⁺ cells in the absence of HIV-1 infection was stable for each vector examined over the 21 day culture period (data not shown). C. Mixtures of non-transduced CEMx174 cells and cells transduced with vectors expressing either the sh(<i>tat/rev</i>), antisense env (VRX494), or the maC46 fusion inhibitors, were generated at the ratios indicated at time 0 and then infected with HIV-1_NL4-3 at an MOI of 10⁻³ TCID₅₀/cell. Initial mixtures of vector-transduced CEMx174 cells ranged from 1% up to 95% GFP⁺ cells. The percentage of GFP⁺ cells was determined by flow cytometry at the indicated times post-infection.</p

Adsorption of HIV-1 virions to the surface of transduced cells expressing maC46.

<p>A. Localization of virus in PM-1 cells transduced with M589, which expresses maC46 and GFP, was analyzed by confocal microscopy. Mixtures of maC46-GFP transduced and non-transduced cells (approximately 10% GFP⁺) were infected with HIV-1_D117/II. After 10 days, cells were stained for HIV-1 p24 (red) and analyzed for expression of GFP (green). maC46-GFP⁺ cells show p24 signal only on the cell membrane, whereas non-transduced cells have a cytoplasmic p24 staining. B. A3-CCR5 or A3-CCR5-maC46 cells were incubated with HIV-pseudotyped viral particles. After 5 hours cells were fixed, stained for p24, and analyzed for the presence of bound viral particles. C. A3-CCR5 cells expressing maC46 were analyzed for their potential to transfer replication-incompetent lentiviral particles pseudotyped with an HIV envelope to PM-1 cells. A3-CCR5 or A3-CCR5-maC46 cells were incubated with the viral particles for 5 hours and after washing, transferred to PM-1 cells expressing RFP at a final ratio of 1∶1. GFP expression of PM-1 cells was analyzed by flow cytometry after 3 days.</p

Patient characteristics.

a<p>ULN = upper limit of normal; ^bof 42 HCV-RNA negative patients, 3 were still on antiviral treatment at the time of study inclusion.</p

Distribution of anti-HCV and HCV-RNA status for each of the two study sites alone and all patients combined.

<p>Serum samples for HCV-RNA analysis were available from 685 out of 757 anti-HCV positive patients.</p