scholarly journals Vaccinia Virus Extracellular Enveloped Virion Neutralization In Vitro and Protection In Vivo Depend on Complement

2008 ◽  
Vol 83 (3) ◽  
pp. 1201-1215 ◽  
Author(s):  
Mohammed Rafii-El-Idrissi Benhnia ◽  
Megan M. McCausland ◽  
Juan Moyron ◽  
John Laudenslager ◽  
Steven Granger ◽  
...  
Keyword(s):  
Vaccinia Virus ◽  
Protective Mechanism ◽  
Humoral Immune ◽  
Humoral Immune Responses ◽  
Rapid Control ◽  
Infected Cells ◽  
Sterilizing Immunity ◽  
Powerful Mechanism

ABSTRACT Antibody neutralization is an important component of protective immunity against vaccinia virus (VACV). Two distinct virion forms, mature virion and enveloped virion (MV and EV, respectively), possess separate functions and nonoverlapping immunological properties. In this study we examined the mechanics of EV neutralization, focusing on EV protein B5 (also called B5R). We show that neutralization of EV is predominantly complement dependent. From a panel of high-affinity anti-B5 monoclonal antibodies (MAbs), the only potent neutralizer in vitro (90% at 535 ng/ml) was an immunoglobulin G2a (IgG2a), and neutralization was complement mediated. This MAb was the most protective in vivo against lethal intranasal VACV challenge. Further studies demonstrated that in vivo depletion of complement caused a >50% loss of anti-B5 IgG2a protection, directly establishing the importance of complement for protection against the EV form. However, the mechanism of protection is not sterilizing immunity via elimination of the inoculum as the viral inoculum consisted of a purified MV form. The prevention of illness in vivo indicated rapid control of infection. We further demonstrate that antibody-mediated killing of VACV-infected cells expressing surface B5 is a second protective mechanism provided by complement-fixing anti-B5 IgG. Cell killing was very efficient, and this effector function was highly isotype specific. These results indicate that anti-B5 antibody-directed cell lysis via complement is a powerful mechanism for clearance of infected cells, keeping poxvirus-infected cells from being invisible to humoral immune responses. These findings highlight the importance of multiple mechanisms of antibody-mediated protection against VACV and point to key immunobiological differences between MVs and EVs that impact the outcome of infection.

scholarly journals Vaccinia Virus Vectors with an Inactivated Gamma Interferon Receptor Homolog Gene (B8R) Are Attenuated In Vivo without a Concomitant Reduction in Immunogenicity

2001 ◽  
Vol 75 (1) ◽  
pp. 11-18 ◽  
Author(s):  
Paulo H. Verardi ◽  
Leslie A. Jones ◽  
Fatema H. Aziz ◽  
Shabbir Ahmad ◽  
Tilahun D. Yilma
Keyword(s):  
Antiviral Activity ◽  
Vaccinia Virus ◽  
Gamma Interferon ◽  
Surprising Result ◽  
Humoral Immune ◽  
Humoral Immune Responses ◽  
Concomitant Reduction ◽  
Ifn Γ

ABSTRACT The vaccinia virus (VV) B8R gene encodes a secreted protein with homology to the gamma interferon (IFN-γ) receptor. In vitro, the B8R protein binds to and neutralizes the antiviral activity of several species of IFN-γ, including human and rat IFN-γ; it does not, however, bind significantly to murine IFN-γ. Here we report on the construction and characterization of recombinant VVs (rVVs) lacking the B8R gene. While the deletion of this gene had no effect on virus replication in vitro, rVVs lacking the B8R gene were attenuated for mice. There was a significant decrease in weight loss and mortality in normal mice, and nude mice survived significantly longer than did controls inoculated with parental virus. This is a surprising result considering the minimal binding of the B8R protein to murine IFN-γ and its failure to block the antiviral activity of this cytokine in vitro. Such reduction in virulence could not be determined in rats, since they are considerably more resistant to VV infection than are mice. Finally, deletion of the B8R gene had no detectable effects on humoral immune responses. Mice and rats vaccinated with the rVVs showed identical humoral responses to both homologous and heterologous genes expressed by VV. This study demonstrates that the deletion of the VV B8R gene leads to enhanced safety without a concomitant reduction in immunogenicity.

scholarly journals By Binding CD80 and CD86, the Vaccinia Virus M2 Protein Blocks Their Interactions with both CD28 and CTLA4 and Potentiates CD80 Binding to PD-L1

2019 ◽  
Vol 93 (11) ◽  
Author(s):  
Patricia Kleinpeter ◽  
Christelle Remy-Ziller ◽  
Eline Winter ◽  
Murielle Gantzer ◽  
Virginie Nourtier ◽  
...  
Keyword(s):  
Immune Response ◽  
Vaccinia Virus ◽  
Cytotoxic T Lymphocyte ◽  
Myxoma Virus ◽  
Parental Virus ◽  
M2 Protein ◽  
First Results ◽  
Infected Cells

ABSTRACTIn this article we report that the M2 protein encoded by the vaccinia virus is secreted as a homo-oligomer by infected cells and binds two central costimulation molecules, CD80 (B7-1) and CD86 (B7-2). These interactions block the ligation of the two B7 proteins to both soluble CD28 and soluble cytotoxic T-lymphocyte associated protein 4 (CTLA4) but favor the binding of soluble PD-L1 to soluble CD80. M2L gene orthologues are found in several other poxviruses, and the B7-CD28/CTLA4 blocking activity has been identified for several culture supernatants of orthopoxvirus-infected cells and for a recombinant myxoma virus M2 protein homolog (i.e., Gp120-like protein, or Gp120LP). Overall, these data indicate that the M2 poxvirus family of proteins may be involved in immunosuppressive activities broader than the NF-κB inhibition already reported (R. Gedey, X. L. Jin, O. Hinthong, and J. L. Shisler, J Virol 80:8676–8685, 2006, https://doi.org/10.1128/JVI.00935-06). A Copenhagen vaccinia virus with a deletion of the nonessential M2L locus was generated and compared with its parental virus. This M2L-deleted vaccinia virus, unlike the parental virus, does not generate interference with the B7-CD28/CTLA4/PD-L1 interactions. Moreover, this deletion did not affect any key features of the virus (in vitroreplication, oncolytic activitiesin vitroandin vivo,and intratumoral expression of a transgene in an immunocompetent murine model). Altogether, these first results suggest that the M2 protein has the potential to be used as a new immunosuppressive biotherapeutic and that the M2L-deleted vaccinia virus represents an attractive new oncolytic platform with an improved immunological profile.IMPORTANCEThe vaccinia virus harbors in its genome several genes dedicated to the inhibition of the host immune response. Among them, M2L was reported to inhibit the intracellular NF-κB pathway. We report here several new putative immunosuppressive activities of M2 protein. M2 protein is secreted and binds cornerstone costimulatory molecules (CD80/CD86). M2 binding to CD80/CD86 blocks their interaction with soluble CD28/CTLA4 but also favors the soluble PD-L1-CD80 association. These findings open the way for new investigations deciphering the immune system effects of soluble M2 protein. Moreover, a vaccinia virus with a deletion of its M2L has been generated and characterized as a new oncolytic platform. The replication and oncolytic activities of the M2L-deleted vaccinia virus are indistinguishable from those of the parental virus. More investigations are needed to characterize in detail the immune response triggered against both the tumor and the virus by this M2-defective vaccinia virus.

scholarly journals MATURATION OF THE HUMORAL IMMUNE RESPONSE IN MICE

1974 ◽  
Vol 139 (2) ◽  
pp. 249-263 ◽  
Author(s):  
Patricia G. Spear ◽  
Gerald M. Edelman
Keyword(s):  
T Cell ◽  
B Cells ◽  
Cell Function ◽  
Spleen Cells ◽  
Con A ◽  
Humoral Immune ◽  
Humoral Immune Responses ◽  
Humoral Antibodies

In spite of the prenatal appearance of immunoglobulin-bearing lymphocytes and θ-positive lymphocytes in the spleens of Swiss-L mice, these mice are not able to produce detectable levels of humoral antibodies in response to antigen until after 1 wk of age. Adult levels of response are not achieved until 4–8 wk of age. In the presence of bacterial lipopolysaccharides, which can substitute for or enhance T-cell function, the B cells from young Swiss-L mice were found to be indistinguishable in function from adult B cells, both with respect to the numbers of plaque-forming cells (PFC) produced in vitro in response to antigen and with respect to the kinetics of PFC induction. The spleen cells from young Swiss-L mice are significantly less sensitive than adult spleen cells, however, to stimulation by the T cell mitogens, concanavalin A (Con A) and phytohemagglutinin (PHA). Very few Con A-responsive cells could be detected at birth but the numbers increased sharply with age until 3 wk after birth. On the other hand, PHA-responsive cells could not be detected in the spleen until about 3 wk of age. The latter cells were found to respond also to Con A, but at a lower dose (1 µg/ml) than that required for the bulk of the Con A-responsive cells (3 µg/ml). The cells that respond both to PHA and to Con A appear in the spleen at about the time that Swiss-L mice acquire the ability to produce humoral antibodies, and these cells can be depleted from the spleen by the in vivo administration of antithymocyte serum. The development of humoral immune responses in these mice therefore appears to be correlated with the appearance of recirculating T lymphocytes that are responsive both to PHA and to Con A.

Chondroitin sulfate activates B cells in vitro, expands CD138+cells in vivo, and interferes with established humoral immune responses

2014 ◽  
Vol 96 (1) ◽  
pp. 65-72 ◽  
Author(s):  
Hilke Brühl ◽  
Josef Cihak ◽  
Nicole Goebel ◽  
Yvonne Talke ◽  
Kerstin Renner ◽  
...  
Keyword(s):  
B Cells ◽  
Immune Responses ◽  
Chondroitin Sulfate ◽  
Humoral Immune ◽  
Humoral Immune Responses

scholarly journals Augmentation of in vitro humoral immune responses in the mouse by an antibody to IgD.

1980 ◽  
Vol 152 (3) ◽  
pp. 493-506 ◽  
Author(s):  
F D Finkelham ◽  
V L Woods ◽  
S B Wilburn ◽  
J J Mond ◽  
K E Stein ◽  
...  
Keyword(s):  
T Cell ◽  
Immune Responses ◽  
Total Serum ◽  
Adjuvant Effect ◽  
Humoral Immune ◽  
Humoral Immune Responses ◽  
Almost All ◽  
T Cell Help

Heterologous anti-delta-chain antibodies have an adjuvant effect on specific in vivo humoral immune responses to simultaneously, or subsequently, injected antigens in the rat and rhesus monkey. We have used a hybridoma-secreted antibody that binds murine delta-chain of the allotype (4.22aM delta a) to study this phenomenon in the mouse and to investigate the mechanism of this effect. Injection of 4.22aM delta a into BALB/c mice removes almost all surface IgD (sIgD) from splenic B lymphocites. sIgD does not reappear until the serum level of 4.22aM delta a decreased 5-7 d after injection. 4.22aM delta a fails to induce detectable proliferation or to raise total serum Ig levels substantially above control values. However, 4.22aM dalta a injected 24 h before antigen elicits an approximately twofold enhancement of serum IgM and a 3- to 10-fold enhancement of serum IgG anti-trintriphenyl (TNP) antibodies in response to immunization with optimal doses of TNP-Ficoll or TNP-sheep red blood cells (TNP-SRBC). 4.22aM delta a injected 1 wk before or 3 d after TNP-SRBC, however, has no effect on IgG anti-TNP levels. The adjuvant effect of anti-delta-chain antibody was markedly decreased when suboptimal antigen doses were used. Furthermore, even in the case of TNP-Ficoll, a relatively T-independent antigen, the ability of 4.22aM dalta a to enhance the anti-TNP antibody response was T cell dependent. Our data suggest that the binding of anti-delta-chain antibody to cell sIgD may partially activate B lymphocytes and make them more capable of differentiating into antibody-secreting cells when stimulated by antigen-specific T cell help.

scholarly journals The collaborative phenotype of secondary B cells is determined by T lymphocytes during in vivo immunization.

1982 ◽  
Vol 155 (2) ◽  
pp. 574-586 ◽  
Author(s):  
N A Speck ◽  
S K Pierce
Keyword(s):  
T Cells ◽  
B Cells ◽  
B Cell ◽  
Cell Populations ◽  
Antibody Synthesis ◽  
Humoral Immune ◽  
Igg1 Antibody ◽  
Humoral Immune Responses

Previous studies have demonstrated that the B cells in immune and nonimmune mice manifest different major histocompatibility complex (MHC) collaborative phenotypes with antigen-specific T cells. Immune, or secondary B cells require syngeneic-like MHC recognition by collaborating T cells, and in its absence fail to be stimulated. Primary B cells manifest a much less stringent requisite for MHC recognition by T cells, and under conditions in which secondary B cells fail to be stimulated, primary B cells are stimulated to secrete IgM antibody. Experiments were conducted to determine whether the acquisition of the secondary B cells' MHC collaborative phenotype was dependent on the presence of T cells during in vivo immunization. B cell populations from T dependently and T independently immunized conventional BALB/c and athymic BALB/c nu/nu mice were compared in their ability to collaborate with allogeneic T cells. Although antigen alone promotes the differentiation of several secondary B cell characteristics, including an increase in the frequency of antigen-specific B cells and a preference for IgG1 antibody synthesis in vitro, the acquisition of the secondary B cells' MHC collaborative phenotype was dependent on the presence of T cells during in vivo immunization. B cell populations from T dependently and T independently immunized conventional BALB/c and athymic BALB/c nu/nu mice were compared in their ability to collaborate with allogeneic T cells. Although antigen alone promotes the differentiation of several secondary B cell characteristics, including an increase in the frequency of antigen-specific B cells and a preference of IgG1 antibody synthesis in vitro, the acquisition of the secondary B cells' MHC collaborative phenotype was found to be dependent on the presence of T cells during in vivo immunization. Thus, the restriction imposed on T cell-B-cell-collaborative interactions in secondary humoral immune responses appears to be the result of T dependent antigen-driven events.

scholarly journals Functional Analysis of Vaccinia Virus B5R Protein: Essential Role in Virus Envelopment Is Independent of a Large Portion of the Extracellular Domain

1998 ◽  
Vol 72 (1) ◽  
pp. 294-302 ◽  
Author(s):  
Elizabeth Herrera ◽  
María del Mar Lorenzo ◽  
Rafael Blasco ◽  
Stuart N. Isaacs
Keyword(s):  
Vaccinia Virus ◽  
Gradient Centrifugation ◽  
Extracellular Domain ◽  
Gene Encoding ◽  
Reading Frame ◽  
Enveloped Virus ◽  
Vaccinia Viruses ◽  
Infected Cells

ABSTRACT Vaccinia virus has two forms of infectious virions: the intracellular mature virus and the extracellular enveloped virus (EEV). EEV is critical for cell-to-cell and long-range spread of the virus. The B5R open reading frame (ORF) encodes a membrane protein that is essential for EEV formation. Deletion of the B5R ORF results in a dramatic reduction of EEV, and as a consequence, the virus produces small plaques in vitro and is highly attenuated in vivo. The extracellular portion of B5R is composed mainly of four domains that are similar to the short consensus repeats (SCRs) present in complement regulatory proteins. To determine the contribution of these putative SCR domains to EEV formation, we constructed recombinant vaccinia viruses that replaced the wild-type B5R gene with a mutated gene encoding a B5R protein lacking the SCRs. The resulting recombinant viruses produced large plaques, indicating efficient cell-to-cell spread in vitro, and gradient centrifugation of supernatants from infected cells confirmed that EEV was formed. In contrast, phalloidin staining of infected cells showed that the virus lacking the SCR domains was deficient in the induction of thick actin bundles. Thus, the highly conserved SCR domains present in the extracellular portion of the B5R protein are dispensable for EEV formation. This indicates that the B5R protein is a key viral protein with multiple functions in the process of virus envelopment and release. In addition, given the similarity of the extracellular domain to complement control proteins, the B5R protein may be involved in viral evasion from host immune responses.

Delivery of mRNA into Dendritic Cells Using Novel Transfer Peptides Leads to Prolonged Antigen Expression and Potent Immune Responses In Vivo.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 4884-4884
Author(s):  
Karrune Woan ◽  
Axel Heiser ◽  
Philipp Dahm ◽  
Johannes Vieweg ◽  
Zhen Su
Keyword(s):  
Dendritic Cells ◽  
Immune Responses ◽  
Rna Binding ◽  
Antigen Expression ◽  
Human Monocyte ◽  
Humoral Immune ◽  
Humoral Immune Responses ◽  
Effective Delivery

Abstract We have previously shown that vaccination with RNA-transfected DC is a potent strategy to stimulate CTL and antitumor immunity in cancer patients. In this study, we investigated whether novel transfer peptides derived from the RNA-binding region of the HIV-1 nucleocapsid protein could be utilized for effective delivery of mRNA into human monocyte-derived dendritic cells (DC). Here we show that both peptide-mediated mRNA delivery and electroporation of DC with mRNA resulted in efficient gene transfer. However, the use of transfer peptides led to prolonged antigen expression and did not negatively affect the viability of DC, the migratory capacity of matured DC, and the production of cytokines by these cells in vitro. In murine studies, DC loaded with transfer peptide-mRNA complexes were clearly superior, compared to mRNA-electroporated DC, in stimulating antigen-specific CTL, CD4+ T cell, and antibody responses. Importantly, no transfer peptide-specific cellular or humoral immune responses were detected in vaccinated mice. Our data suggest that vaccination with transfer peptide-mRNA-loaded DC may represent a promising strategy to stimulate potent anti-tumor immune responses in a vaccination setting.

scholarly journals Vaccinia Virus G7L Protein Interacts with the A30L Protein and Is Required for Association of Viral Membranes with Dense Viroplasm To Form Immature Virions

2003 ◽  
Vol 77 (6) ◽  
pp. 3418-3429 ◽  
Author(s):  
Patricia Szajner ◽  
Howard Jaffe ◽  
Andrea S. Weisberg ◽  
Bernard Moss
Keyword(s):  
Vaccinia Virus ◽  
Protein A ◽  
Direct Interaction ◽  
Uncharacterized Protein ◽  
Lethal Mutant ◽  
C Terminus ◽  
Infected Cells ◽  
The Stability

ABSTRACT The vaccinia virus A30L protein is required for the association of electron-dense, granular, proteinaceous material with the concave surfaces of crescent membranes, an early step in viral morphogenesis. For the identification of additional proteins involved in this process, we used an antibody to the A30L protein, or to an epitope appended to its C terminus, to capture complexes from infected cells. A prominent 42-kDa protein was resolved and identified by mass spectrometry as the vaccinia virus G7L protein. This previously uncharacterized protein was expressed late in infection and was associated with immature virions and the cores of mature particles. In order to study the role of the G7L protein, a conditional lethal mutant was made by replacing the G7L gene with an inducible copy. Expression of G7L and formation of infectious virus was dependent on the addition of inducer. Under nonpermissive conditions, morphogenesis was blocked and viral crescent membranes and immature virions containing tubular elements were separated from the electron-dense granular viroplasm, which accumulated in large spherical masses. This phenotype was identical to that previously obtained with an inducible, conditional lethal A30L mutant. Additional in vivo and in vitro experiments provided evidence for the direct interaction of the A30L and G7L proteins and demonstrated that the stability of each one was dependent on its association with the other.

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