Mimotopes selected with neutralizing antibodies against multiple subtypes of influenza A

<p>Abstract</p> <p>Background</p> <p>The mimotopes of viruses are considered as the good targets for vaccine design. We prepared mimotopes against multiple subtypes of influenza A and evaluate their immune responses in flu virus challenged Balb/c mice.</p> <p>Methods</p> <p>The mimotopes of influenza A including pandemic H1N1, H3N2, H2N2 and H1N1 swine-origin influenza virus were screened by peptide phage display libraries, respectively. These mimotopes were engineered in one protein as multi- epitopes in Escherichia coli (E. coli) and purified. Balb/c mice were immunized using the multi-mimotopes protein and specific antibody responses were analyzed using hemagglutination inhibition (HI) assay and enzyme-linked immunosorbent assay (ELISA). The lung inflammation level was evaluated by hematoxylin and eosin (HE).</p> <p>Results</p> <p>Linear heptopeptide and dodecapeptide mimotopes were obtained for these influenza virus. The recombinant multi-mimotopes protein was a 73 kDa fusion protein. Comparing immunized infected groups with unimmunized infected subsets, significant differences were observed in the body weight loss and survival rate. The antiserum contained higher HI Ab titer against H1N1 virus and the lung inflammation level were significantly decreased in immunized infected groups.</p> <p>Conclusions</p> <p>Phage-displayed mimotopes against multiple subtypes of influenza A were accessible to the mouse immune system and triggered a humoral response to above virus.</p

HIV and HCV Co-Culture Promotes Profibrogenic Gene Expression through an Epimorphin-Mediated ERK Signaling Pathway in Hepatic Stellate Cells.

Accelerated fibrosis in patients co-infected with hepatitis C virus (HCV) and human immunodeficiency virus (HIV) has been a major cause of mortality in the highly active anti-retroviral therapy (HAART) era. However, the role of co-infection in accelerating the progression of liver fibrosis, particularly with regard to the effects of co-infection on hepatic stellate cells (HSCs), remains unclear. We hypothesized that HIV and HCV induce liver fibrosis synergistically by altering the regulation of epimorphin production, and thereby indirectly alter HSC function. Here, we examined the effects of epimorphin on HSC proliferation and invasion, and the changes in fibrogenesis-related gene activity in HSCs (LX2) in the presence of inactivated CXCR4-tropic HIV and HCV (JFH1). The combination of HIV and HCV significantly increased epimorphin expression, which increased the proliferation and invasion capabilities of HSCs. Epimorphin also induced the expression of profibrogenic tissue inhibitor of metalloproteinase 1 (TIMP1) in an extracellular signal-regulated kinase (ERK)-dependent manner. These data indicated that the effects of HIV/HCV co-infection on hepatic fibrosis might be mediated in part by EPM. Strategies to limit the expression of EPM might represent a novel therapeutic approach to prevent the progression of hepatic fibrosis during HIV/HCV co-infection

HIV-1 infection depletes human CD34⁺CD38^- hematopoietic progenitor cells via pDC-dependent mechanisms

<div><p>Chronic human immunodeficiency virus-1 (HIV-1) infection in patients leads to multi-lineage hematopoietic abnormalities or pancytopenia. The deficiency in hematopoietic progenitor cells (HPCs) induced by HIV-1 infection has been proposed, but the relevant mechanisms are poorly understood. We report here that both human CD34⁺CD38^- early and CD34⁺CD38⁺ intermediate HPCs were maintained in the bone marrow (BM) of humanized mice. Chronic HIV-1 infection preferentially depleted CD34⁺CD38^- early HPCs in the BM and reduced their proliferation potential <i>in vivo</i> in both HIV-1-infected patients and humanized mice, while CD34⁺CD38⁺ intermediate HSCs were relatively unaffected. Strikingly, depletion of plasmacytoid dendritic cells (pDCs) prevented human CD34⁺CD38^- early HPCs from HIV-1 infection-induced depletion and functional impairment and restored the gene expression profile of purified CD34⁺ HPCs in humanized mice. These findings suggest that pDCs contribute to the early hematopoietic suppression induced by chronic HIV-1 infection and provide a novel therapeutic target for the hematopoiesis suppression in HIV-1 patients.</p></div

Chronic HIV-1 infection impairs proliferation of early CD34⁺CD38^- HPCs <i>in vivo</i> of humanized mice.

<p>(A) Representative histograms showing BrdU labeling of CD34⁺ HPC subsets from BM of humanized mice. Numbers indicate percentages of BrdU⁺ within the total CD34⁺ HPCs, early CD34⁺CD38^- HPCs and intermediate CD34⁺CD38⁺ HPCs. (B-E) Pooled data indicating percentages of BrdU⁺ cells within early CD34⁺CD38^- HPCs (B) and intermediate CD34⁺CD38⁺ HPCs (C); and absolute cell numbers of BrdU⁺ early CD34⁺CD38^- HPCs (D) and intermediate CD34⁺CD38⁺ HPCs (E) in BM of mock (n = 4) and HIV-1-infected (n = 6) humanized mice. Data are shown as the mean ± s.e.m. *<i>P</i> < 0.05 (two-tailed unpaired Student’s t-test).</p

Chronic HIV-1 infection preferentially depletes early CD34⁺CD38^- HPCs in humanized mice <i>in vivo</i>.

<p>(A) Representative histograms showing p24 expression on CD3⁺ T cells and CD34⁺ HPCs from BM of humanized mice with chronic HIV-1 infection. Numbers indicate percentages of p24-expressing CD3⁺ T cells and CD34⁺ HPCs. Data are representative of three independent experiments with mice reconstituted with two to three donors. (B) Representative dot plots showing chronic HIV-1 infection depleting early CD34⁺CD38^- HPCs from BM of humanized mice. Numbers indicate percentages of early CD38^-CD34⁺ and intermediate CD34⁺CD38⁺ HPCs. (C) Pooled data indicating mean percentages of early CD34⁺CD38^- HPCs and intermediate CD34⁺CD38⁺ HPCs in BM of humanized mice with mock and HIV-1 infection shown in the stacked bar graph. <i>P</i> values are shown. (D-E) Pooled data indicating absolute cell numbers of early CD34⁺CD38^- HPCs (D) and intermediate CD34⁺CD38⁺ HPCs (E) in BM of humanized mice with mock (n = 7) or chronic HIV-1 infection (n = 5). (F) Representative dot plots showing chronic HIV-1 infection depleting early CD34⁺CD38^- HPCs from BM of HIV-1-infected patients compared to that of healthy control (HC) donors. Numbers indicate percentages of early CD38^-CD34⁺ and intermediate CD34⁺CD38⁺ HPCs. (G-H) Pooled data indicating percentages of early CD34⁺CD38^- HPCs (G) and intermediate CD34⁺CD38⁺ HPCs (H) in BM of HC donors (n = 6) and HIV-1-infected patients (n = 5). (D, E, G, H) Data are shown as the mean ± s.e.m. *<i>P</i> < 0.05 and ***<i>P</i> < 0.001 (two-tailed unpaired Student’s t-test).</p

Depletion of pDCs during chronic HIV-1 infection rescues early CD34⁺CD38^- HPCs in humanized mice.

<p>(A) Representative dot plots showing the recovery of early CD34⁺CD38^- HPCs in BM of chronic HIV-1-infected humanized mice with pDC depletion. Numbers indicate percentages of early CD34⁺CD38^- HPCs and intermediate CD34⁺CD38⁺ HPCs in various groups of mice. (B and C) Summary data of percentages (B) and absolute cell numbers (C) of early CD34⁺CD38^- HPCs and intermediate CD34⁺CD38⁺ HPCs from BM in mock (n = 11), HIV-1-infected (n = 12) and HIV-1-infected humanized mice with pDC depletion (n = 13). Each dot represents one mouse. *<i>P</i> < 0.05 and ***<i>P</i> < 0.001 (two-tailed unpaired Student’s t-test). (D) Summary data of CFU that developed from CD34⁺ HPCs of mock-infected mice (n = 4), HIV-1-infected mice (n = 5) and HIV-1-infected mice with pDC depletion (n = 4). *<i>P</i> < 0.05 (two-tailed unpaired Student’s t-test). Error bars, s.e.m</p

Depletion of pDCs reverses the dysregulated gene expression profile of human CD34⁺ HPCs affected by HIV-1 infection.

<p>Humanized mice were infected with HIV-1 JR-CSF and treated with 15B or control IgG at 11 weeks post-infection and terminated at 21 weeks post-infection. huCD45⁺Lin^-CD34⁺ cells from BM of mock, HIV-1/IgG or HIV-1/15B treated mice were purified by flow cytometry (> 95% purity). Total mRNA were isolated and used for the cDNA microarray assay. (A) Heat map showing the relative expression of 6108 significantly up- or down-regulated genes indicated by the color bars in huCD45⁺Lin^-CD34⁺ cells from HIV-1-infected over mock-infected samples (green, suppressed genes; red, induced genes. Fold change ≥ 2). (B) Table showing numbers of genes significantly up-regulated and down-regulated in humanized mice with HIV-1 infection treated with control IgG or 15B antibody for pDC depletion. (C) Pathway maps representing a set of signaling and metabolic maps. Sorting was performed for statistically significant maps. Experimental data are represented as green (JR-CSF + IgG) and red (JR-CSF + 15B) histograms. The height of the histogram corresponds to the relative expression of particular pathways. (D) Fold changes of 88 genes from hematopoietic cell lineage in HIV-1-infected mice treated with control IgG or 15B antibody for pDC depletion in relation to mock controls. (E) Expression levels of genes associated with HPC proliferation, differentiation and cell death in huCD45⁺Lin^-CD34⁺ cells are indicated by the heat-map color bars (green, suppressed genes; red, induced genes. Fold change ≥ 2). (F) Summary data of the expression of HPC-associated genes, including CD38, HES1, Notch-1 and AML-1 in huCD45⁺Lin^-CD34⁺ cells from various groups of mice (n = 4 per group). Data are shown as the mean ± s.e.m. *<i>P</i> < 0.05 (two-tailed unpaired Student’s t-test).</p