Many pathogenic enveloped viruses, including HIV-1, escape complement-mediated virolysis by incorporating host cell regulators of complement activation into their own viral envelope. The presence of complement regulators including CD59 on the external surface of the viral envelope confers resistance to complement-mediated virolysis, which may explain why human pathogenic viruses such as HIV-1 are not neutralized by complement in human fluids, even in the presence of high Ab titers against the viral surface proteins. In this study, we report the development of a recombinant form of the fourth domain of the bacterial toxin intermedilysin (the recombinant domain 4 of intermedilysin [rILYd4]), a 114 aa protein that inhibits human CD59 function with high affinity and specificity. In the presence of rILYd4, HIV-1 virions derived from either cell lines or peripheral blood mononuclear cells of HIV-1–infected patients became highly sensitive to complement-mediated lysis activated by either anti–HIV-1 gp120 Abs or by viral infection-induced Abs present in the plasma of HIV-1–infected individuals. We also demonstrated that rILYd4 together with serum or plasma from HIV-1–infected patients as a source of anti–HIV-1 Abs and complement did not mediate complement-mediated lysis of either erythrocytes or peripheral blood mononuclear cells. These results indicate that rILYd4 may represent a novel therapeutic agent against HIV-1/AIDS

Alcoholic hepatitis (AH) develops only in small proportion of heavy drinkers. To better understand the mechanisms underlying this disparity, we conducted a study to define the relationship between AH development and dysregulated immune responses that might be ameliorated by alcohol abstinence. Sixty-eight AH patients, 65 heavy drinking controls without liver disease (HDC), and 20 healthy controls (HC) were enrolled and followed up to 12 months. At the baseline, HDC and HC had no significant differences in their plasma levels of 38 inflammatory cytokines/chemokines measured using multiplex immunoassays. However, compared to HDC, AH patients had higher baseline levels of 11 cytokines/chemokines (TNF-α, IL-6, IL-8, IP10, IL-4, IL-9, IL-10, FGF-2, IL-7, IL-15, and TGF-α), but lower levels of the anti-inflammatory macrophage-derived chemokine (MDC). AH patients also had more activated, yet dysfunctional immune cells as monocytes, T cells, and B cells expressed higher levels of CD38 and CD69, but low levels of HLA-DR, CD80, and CD86 at baseline. In addition, CD4 T cells produced less IFN-γ in response to T cell stimulation. Upregulated IL-6, IL-8, CD38, and CD69 and downregulated MDC, HLA-DR, CD86, and CD80 correlated positively and negatively, respectively, with disease severity. Longitudinal analysis indicated that levels of IL-6, IL-8, CD38, and CD69 were reduced, whereas levels of MDC, HLA-DR, CD80, and CD86 were increased in abstinent AH patients. All of the cellular immune abnormalities were reversed by day 360 in abstinent AH patients; however, plasma levels of TNF-α, IL-8, IL-10, FGF-2, and IL-7 remained higher. Conclusion: AH patients were in a highly immune-dysregulated state, whereas HDC showed little evidence of immune activation. Alcohol abstinence reversed most, but not all, of the immunological abnormalities.

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Several enveloped viruses including HIV-1, CMV, HSV-1, Ebola virus, vaccinia virus, and influenza virus have been found to incorporate host regulators of complement activation (RCA) into their viral envelopes and, as a result, escape antibody-dependent complement-mediated lysis (ADCML). HCV is an enveloped virus of the family Flaviviridae and incorporates more than 10 host lipoproteins. Patients chronically infected with HCV develop high-titer and cross-reactive neutralizing antibodies (nAbs) yet fail to clear the virus, raising the possibility that HCV may also use the similar strategy of RCA incorporation to escape ADCML. The current study was therefore undertaken to determine whether HCV virions incorporate biologically functional CD59, a key member of RCA. Our experiments provided several lines of evidence demonstrating that CD59 was associated with the external membrane of HCV particles derived from either Huh7.5.1 cells or plasma samples from HCV-infected patients. First, HCV particles were captured by CD59-specific Abs. Second, CD59 was detected in purified HCV particles by immunoblot analysis and in the cell-free supernatant from HCV-infected Huh7.5.1 cells, but not from uninfected or Ad5 (a nonenveloped cytolytic virus)-infected Huh7.5.1 cells by ELISA. Last, abrogation of CD59 function with its blockers increased the sensitivity of HCV virions to ADCML, resulting in a significant reduction of HCV infectivity. Additionally, direct addition of CD59 blockers into plasma samples from HCV-infected patients increased autologous virolysis. Conclusion our study, for the first time, demonstrates that CD59 is incorporated into both cell line-derived and plasma primary HCV virions at levels that protect against ADCML. This is also the first report to show that direct addition of RCA blockers into plasma from HCV-infected patients renders endogenous plasma virions sensitive to ADCML.

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Latently HIV-1-infected cells are recognized as the last barrier towards viral eradication and cure. To purge these cells, we combined a provirus stimulant with a blocker of human CD59, a key member of the regulators of complement activation (RCA), to trigger antibody-dependent complement-mediated lysis (ADCML). Provirus stimulants including prostratin and histone deacetylase inhibitors (HDACi) such as romidepsin (RMD) and suberoylanilide hydroxamic acid (SAHA) activated proviruses in the latently HIV-1-infected T cell line ACH-2 as virion production and viral protein expression on the cell surface were induced. RMD was the most attractive provirus stimulant as it effectively activated proviruses at nanomolar concentrations that can be achieved clinically. Antiretroviral drugs including two protease inhibitors (atazanavir and darunavir) and a reverse transcriptase inhibitor (emtricitabine) did not affect the activity of provirus stimulants in the activation of proviruses. However, saquinavir (a protease inhibitor) markedly suppressed virus production, although did not affect the percentage of cells expressing viral Env on the cell surface. Provirus-activated ACH-2 cells expressed HIV-1 Env that colocalized with CD59 in lipid rafts on the cell surface, facilitating direct interaction between them. Blockage of CD59 rendered provirus-activated ACH-2 cells and primary human CD4+ T cells that were latently infected with HIV-1 sensitive to ADCML by anti-HIV-1 polyclonal antibodies or plasma from HIV-1-infected patients. Therefore, a combination of provirus stimulants with RCA blockers represents a novel approach to eliminate HIV-1.

Bone marrow stromal cell antigen 2 (BST-2, also known as tetherin, CD317, or HM1.24) has recently been identified as a host restriction factor against diverse families of enveloped viruses. However, the effects of BST-2 on the life cycle of hepatitis C virus (HCV), an enveloped RNA virus, remain unclear and controversial. Here we demonstrated that human hepatocytes including Huh7.5.1 cells, primary human hepatocytes (PHHs), and HepG2 cells constitutively expressed low to moderate levels of endogenous BST-2 on the cell surface, which could be robustly up-regulated by all three types of interferons (IFNs) such as IFN-α, IFN-γ, and IFN-λ. IFN-α and IFN-γ showed a synergistic effect in induction of BST-2 expression on human hepatocytes. Over-expression of BST-2 by BST-2-expressing vector transfection or up-regulation of BST-2 by IFN stimulation markedly suppressed HCV production, whereas shRNA-mediated depletion of endogenous BST-2 significantly enhanced HCV production in infected Huh7.5.1 cells. IFN-mediated anti-HCV activity was partially but significantly diminished by shRNA-mediated knockdown of BST-2 expression, indicating that BST- 2 upregulation is directly involved in IFN-mediated inhibition of HCV production. We also found that both BST-2 and HCV core co-localized with intracellular lipid droplets (LDs), suggesting that BST-2-HCV interaction may take place around LDs as LDs constitute an important intracellular organelle for HCV assembly and replication. Taken together, our data suggest that BST-2 is a host restriction factor against HCV, and induction of BST-2 in hepatocytes could be one of the mechanisms by which current HCV standard therapy (IFN-α plus ribavirin) achieves a sustained virological response (SVR).

The vast majority of people living with human immunodeficiency virus type 1 (HIV-1) have pain syndrome, which has a significant impact on their quality of life. The underlying causes of HIV-1-associated pain are not likely attributable to direct viral infection of the nervous system due to the lack of evidence of neuronal infection by HIV-1. However, HIV-1 proteins are possibly involved as they have been implicated in neuronal damage and death. The current study assesses the direct effects of HIV-1 Tat, one of potent neurotoxic viral proteins released from HIV-1-infected cells, on the excitability and survival of rat primary dorsal root ganglion (DRG) neurons. We demonstrated that HIV-1 Tat triggered rapid and sustained enhancement of the excitability of small-diameter rat primary DRG neurons, which was accompanied by marked reductions in the rheobase and resting membrane potential (RMP), and an increase in the resistance at threshold (RTh). Such Tat-induced DRG hyperexcitability may be a consequence of the inhibition of cyclin-dependent kinase 5 (Cdk5) activity. Tat rapidly inhibited Cdk5 kinase activity and mRNA production, and roscovitine, a well-known Cdk5 inhibitor, induced a very similar pattern of DRG hyperexcitability. Indeed, pre-application of Tat prevented roscovitine from having additional effects on the RMP and action potentials (APs) of DRGs. However, Tat-mediated actions on the rheobase and RTh were accelerated by roscovitine. These results suggest that Tat-mediated changes in DRG excitability are partly facilitated by Cdk5 inhibition. In addition, Cdk5 is most abundant in DRG neurons and participates in the regulation of pain signaling. We also demonstrated that HIV-1 Tat markedly induced apoptosis of primary DRG neurons after exposure for longer than 48 h. Together, this work indicates that HIV-1 proteins are capable of producing pain signaling through direct actions on excitability and survival of sensory neurons.