Immunological mechanisms giving rise to latency of herpes simplex virus in the spinal ganglia of the mouse

In the model of genital herpes simplex virus (HSV)-infection of mice, early latency could be induced by passive immunization with HSV-specific antibodies and, to a lesser degree, by adoptive transfer of immune lymphocytes prepared from spleen and draining lymph nodes of genitally infected syngeneic mice. Conversely, spontaneously occurring latency was inhibited by treatment of the animals with cyclophosphamide (Cph) and, to a lesser degree, with cyclosporin A (CyA). Whereas the effect of CyA could be compensated by passively administered HSV-specific antibodies, that of Cph could not. Apparently specific antibodies cooperate with a non-specific proliferating cell type, probably macrophages and/or NK-cells, as could be demonstrated by significantly reduced antibody effect in silica-treated mice. Moreover, F(ab)2 fragments, in contrast to complete antibody molecules, were inactive. HSV-specific antibodies and also immune lymphocytes had little effect on virus production in the mucous membranes, immune lymphocytes being at least as active as antibodies. It is therefore not probable that latency is induced by attenuation of the peripheral disease. It can rather be concluded that the neuron itself is the target for the action of specific antibodies, cooperating in turn with macrophages and/or NK cells.     

Inhibition of thein vitro-reactivation of latent herpes simplex virus infection in spinal ganglia: Comparison of various immune factors

Summary In order to study whether the latency of herpes simplex virus (HSV) is immunologically controlled, the influence of different immune mechanisms on thein vitro-reactivation of the virus in latently infected lumbosacral ganglia of mice was investigated.Combined addition of macrophages and antibodies to cultures of ganglionic tissue proved most effective in delaying virus reactivation. This was achieved to a lesser degree when applying antibodies only, whereas macrophages alone were not effective, nor were immune lymphocytes, nor was interferon from L-cells or from the peritoneal cavity of mice.With 1 Figure     

Target cell-dependent T cell-mediated lysis of vaccinia virus-infected cells

Vaccinia virus specific cytotoxicity against infected target cells was observed in vitro. Spleen lymphocytes from normal and immunized mice of the inbred strains C3H and DBA/2 were incubated with vaccinia virus-infected and non-infected ⁵¹Cr-labeled mastocytoma P-815-X2 cells and L-929 fibroblasts, which were used as targets. Cytotoxic lymphocytes could be isolated from the mice as early as 2 days after infection with vaccinia virus. The highest cytotoxic effect was obtained with lymphocytes taken 6 days after infection. The degree of lysi was correlated with the ratio of immune lymphocytes to target cells. Specific blocking of target cell lysis resulted after addition of anti-vaccinia antibody from different sources. The effector cells could be characterized as T cells by elimination of macrophages and B cells. Target cell killing was only possible in a syngeneic system; allogeneic infected target cells were not lysed significantly.     

The activity of T and B lymphocytes in immunity and tolerance to the lymphocytic choriomeningitis virus in mice.

Treatment with anti-theta serum and the Wigzell column technique for cell separation was employed to study the separate functions of the B and T lymphocytes in the late states of immunity to the LCM virus in mice. The cell preparations examined were mixtures of spleen and lymph node cells from immune mice. The results revealed that the anti-viral effect of such cells after transfer to virus carriers was unimpaired in T cell-enriched and B cell-deprived cell preparations. The anti-viral effect was also retained in cell preparations deprived so much of B cells that no antibody was produced in the virus carrier mice receiving transplants of these cells. The results strongly indicate that the anti-viral effect of late immune cells is not only T cell-dependent but that it is also mediated solely by T cells and, moreover, that antibodies have no or very little influence on the virus elimination. The observation that antibody production could be caused neither by column-passed cells nor by anti-theta serum-treated cells, but was obtained by mixtures of these cells, demonstrates that co-operation between T and B cells is crucial for the LCM antibody response. Accordingly, the convincing demonstration of the absence in the persistent virus carriers of cells which, in respect of antibody production, are able to co-operate either with column-passed or with anti-theta serum-treated immune cells, implies that such animals are extremely deficient as regards immune function of both B and T LCM-primed lymphocytes.     

活化淋巴细胞及其染色质诱导抗核抗体生成

目的诱导抗双链ＤＮＡ抗体和其它抗核抗体生成的免疫原是什么？迄今不明。本研究试图阐明不同原因活化的淋巴细胞是诱导抗核抗体生成的免疫原。方法用丝裂原（ＣｏｎＡ／ＬＰＳ）或ＩＬ－２活化淋巴细胞及其染色质免疫同系ＢＡＬＢ／ｃ小鼠，用ＥＬＩＳＡ测定ＩｇＧ抗ｄｓＤＮＡ抗体，用免疫荧光法检测抗核抗体核型和免疫复合物沉积，用免疫印迹法测定抗Ｓｍ和抗ＲＮＰ等抗核抗体。结果上述原因活化的淋巴细胞及其活性染色质均能诱导抗双链ＤＮＡ抗体和其它抗核抗体生成，免疫小鼠的肾脏有显著的免疫复合物沉着。结论据本文结果推测活化淋巴细胞及其细胞核和染色质是诱发抗ｄｓＤＮＡ抗体和其它抗核抗体生成的免疫原。     

Immune protection against HSV-2 in B-cell-deficient mice

The role of antibody in protection of the vaginal mucosa and sensory ganglia against HSV-2 infection was examined using HSV- immune, B-cell-deficient muMT mice. Significantly higher virus titers were detected in the vaginal mucosae of immune muMT mice compared to immune C57BL/6J mice 24 h after HSV-2 rechallenge. However, virus was rapidly cleared in immune muMT mice, and the infection was resolved with only a 2-day delay. Passive transfer of immune serum to immune muMT mice prior to rechallenge resulted in HSV-specific vaginal IgG levels comparable to those of immune C57BL/6J mice. Although transferred antibody failed to prevent reinfection of the majority of recipients, vaginal virus titers at 24 h and clearance kinetics were similar to those of immune C57BL/6J controls. Following vaginal rechallenge, HSV-2 did not spread to the sensory ganglia of immune C57BL/6J mice nor was the rechallenge virus detected in the ganglia of the majority of immune muMT mice. However, protection was severely compromised by T-cell depletion of immune C57BL/6J mice. These results suggest that HSV-specific antibody limits, but does not prevent, infection of the genital epithelia. Further, prevention of virus spread to the sensory ganglia in immune animals requires vigorous T-cell immune responses.     

Kinetics and genetics of herpes simplex virus-induced antibody formation in mice.

The kinetics of antibody synthesis was investigated after intraperitoneal, subcutaneous, and footpad infection of various strains of mice with herpes simplex virus. Immunoglobulin M antibodies appeared 5 days after and immunoglobulin G antibodies appeared 10 to 12 days after intraperitoneal infection with herpes simplex virus type 1. The major histocompatibility complex and the background genome of inbred mice were not found to have a systematical influence on antibody synthesis. Female mice, however, consistently produced more antibodies than did male if the infection was done intraperitoneally, but not if it was done subcutaneously or into footpads. Castration considerably increased the amount of antibodies produced by male mice. The difference in antibody formation between females and males could be abolished by injection of silica; moreover, antibody titers were enhanced by this treatment. This has also been found by immunization with a Formalin-inactivated herpes simplex virus vaccine. The effect of silica in enhancing antibody formation could be observed up to 12 days after infection. Infectious virus could be detected up to 2 days after infection, and herpes simplex virus type 1 antibody-stimulating antigens could be detected up to 4 days in ultrasonicates of macrophages. The assumption is made that androgen-sensitive cell populations, including macrophages and their soluble products, are involved in antibody-depressing mechanisms.     

Apoptosis-inducing protein derived from hepatocyte selectively induces apoptosis in lymphocytes

The liver is where lymphocytes undergo activation-induced cell death (AICD) at the resolution phase of an immune response, which is crucial for homeostasis of the immune system and prevention of autoimmunity. Exploring the machinery of AICD in the liver, we found that a primary culture supernatant of murine hepatocytes had an antiproliferative effect on antigen-stimulated T clone and T lymphoma cells. Biological study showed that the antiproliferation was due to induction of apoptosis in a caspase-dependent manner. The apoptosis-inducing potential was sensitive to trypsin, heat (> 70 degrees ) and acid (< pH 5) treatment but could not be neutralized by anti-tumour necrosis factor-alpha, anti-Fas ligand, or anti-transforming growth factor-beta antibodies. Biochemical study of the isolated and purified apoptosis-inducing component from the supernatant showed that it was a protein with a molecular mass of about 68,000-70,000. It induced apoptotic change in murine T and B cells, and to a lesser degree, in human lymphoid cells, but not in macrophages. Biochemical and biological characteristics distinguish this protein from others that have been reported to induce apoptosis of lymphocytes. The identification of an apoptosis-inducing protein derived from murine hepatocytes, which selectively induces apoptosis in lymphocytes, suggests one possible mechanism for immune suppression in the liver.     

A genetic analysis of macrophage activation and specific antibodies in relation to the resistance of heterogeneous mouse populations to MHV3 infection

Summary The genetically selected high antibody responder mice (H_III) are susceptible and the low antibody responder mice (L_III) are resistant to the experimental infection with Mouse Hepatitis Virus 3 (MHV3). The mortality rates of the F₁ hybrids and of the F₂ segregants showed the codominance of the susceptible and resistant characters. The direct individual intrapopulation correlation between the induction of antiviral state in macrophages activated by IFN gamma and the resistance to the virus infection, showed that an antiviral state could be induced in resistant mouse macrophages, whereas in susceptible mouse macrophages no restriction of virus replication could be observed. A direct inter- and intrapopulation correlation of pre-existing antibody titres against MHV3 with the mortality and a direct interpopulation correlation of those titres with the mean survival time of susceptible animals was shown. The data indicate, among the mechanisms of resistance against the virus infection, a role of IFN gamma macrophage-activation and of antibodies against MHV3 which may delay the mean survival time in susceptible animals.     

Dengue virus-induced thymus-derived suppressor cells in the spleen of mice.

Adoptive transfer of spleen cells obtained from mice given three weekly i.p. doses of dengue type 2 virus (DV) suppressed DV antigen-specific antibody secretion as detected by the Jerne plaque technique. This suppression was produced by non-glass-adherent cells but not by glass-adherent cells. Immune spleen cells depleted of macrophages by carbonyl iron treatment had higher suppressor activity. Immune spleen cell homogenate could transfer the activity equally well. The immune spleen cells were separated into T and B lymphocytes by a nylon wool column. B lymphocytes had no suppressor activity; almost all the suppressor activity was present in T lymphocytes. Thus, macrophages and B lymphocytes had no suppressor activity; it was mediated by T lymphocytes through soluble factors.     

Role of phagocytes and antibodies in freeing the body from influenza virus]

Persistence of influenza virus in normal and immune mice was studied by determinations of the virus infectivity and by the immunofluorescence procedures. After intraperitoneal inoculation the virus could be found in macrophages of the peritoneal exudate in both normal and immune animals. However, in macrophages of normal mice synthesis of V- and S-antigens of influenza virus occurred indicating the possibility of virus reproduction in them, whereas in macrophages of immune animals there was practically no synthesis of S-antigen despite a greater number of cells with V-antigen of influenza virus in their cytoplasma. The assumption that elimination of influenza virus from the body is due to specific antibody and immunologically changed macrophage elements is substantiated.     

Cell-mediated cytotoxicity against Sendai-virus-infected cells.

After injection of Sendai virus, a parainfluenza virus type I, mice generate cytotoxic lymphocytes which lyse specifically Sendai-virus-infected target cells in vitro. Their action is not inhibited by specific antibody in vitro. Killer cell activity appears 4 days after infection, reaches a maximum on the 7th day and disappears on the 14th to 16th day. Decrease of cytotoxic cell activity is correlated with an increase of haemagglutinating antibodies. The cytotoxic effector cell could be characterized as a thymus-derived cell, there is no specific activity in antibody-dependent cell-mediated cytolysis (ADCC). The degree of cytotoxic effector cell activity is only slightly influenced by the dose of injected infective virus. Using different syngeneic Sendai-virus-infected cells as targets for cell-mediated cytotoxicity, a tumor line was not lysed by cytotoxic lymphocytes in spite of viral surface antigens. Preliminary experiments were performed to demonstrate the H-2 gene restriction of the cytotoxic interaction. Using macrophages and tumor cells as targets only syngeneic infected target cells were lysed.     

Glucocorticoid regulation of lactate dehydrogenase-elevating virus replication in macrophages

Lactate dehydrogenase-elevating virus (LDV) is a macrophage-tropic arterivirus which generally causes a persistent viremic infection in mice. LDV replication in vivo seems to be primarily regulated by the extent and dynamics of a virus-permissive macrophage population. Previous studies have shown that glucocorticoid treatment of chronically LDV-infected mice transiently increases viremia 10-100-fold, apparently by increasing the productive infection of macrophages. We have further investigated this phenomenon by comparing the effect of dexamethasone on the in vivo and in vitro replication of two LDV quasispecies that differ in sensitivity to immune control by the host. The single neutralizing epitope of LDV-P is flanked by two N-glycans that impair its immunogenicity and render LDV-P resistant to antibody neutralization. In contrast, replication of the neuropathogenic mutant LDV-C is suppressed by antibody neutralization because its epitope lacks the two protective N-glycans. Dexamethasone treatment of mice 16 h prior to LDV-P infection, or of chronically LDV-P infected mice, stimulated viremia 10-100-fold, which correlated with an increase of LDV permissive macrophages in the peritoneum and increased LDV infected cells in the spleen, respectively. The increase in viremia occurred in the absence of changes in total anti-LDV and neutralizing antibodies. The results indicate that increased viremia was due to increased availability of LDV permissive macrophages, and that during a chronic LDV-P infection virus replication is strictly limited by the rate of regeneration of permissive macrophages. In contrast, dexamethasone treatment had no significant effect on the level of viremia in chronically LDV-C infected mice, consistent with the view that LDV-C replication is primarily restricted by antibody neutralization and not by a lack of permissive macrophages. beta-Glucan, the receptor of which is induced on macrophages by dexamethasone treatment, had no effect on the LDV permissiveness of macrophages.     

Lymphocyte Reactivity Contributes to Protection Conferred by Specific Antibody Passively Transferred to Herpes Simplex Virus-Infected Mice

Passively acquired immunity to herpes simplex virus (HSV) was studied in antithymocyte serum (ATS)-treated mice and athymic nude mice to determine whether immunocompetent lymphocytes contribute to the protection observed after transfer of HSV-specific antibody to infected animals. Mice were given three intraperitoneal injections of 0.1 ml of ATS at 24-h intervals. This treatment reduced concanavalin A and lipopolysaccharide stimulation of lymphocytes harvested from these animals by 90% when compared with the stimulation of lymphocytes harvested from untreated animals. It was found that intraperitoneal injection of 0.5 ml of specific antibody 8 h after corneal HSV type 1 infection or subcutaneous HSV type 2 infection did not protect ATS-treated animals from virus infection. Specific antibody passively transferred to ATS-treated animals 8 and 120 h postinfection also failed to protect lymphocyte-depleted animals from HSV. However, ATS-treated animals were protected from HSV infection by passively acquired antibody when lymphocytes harvested from these animals regained 80% of their ability to be stimulated with concanavalin A and lipopolysaccharide. It was also found that specific antibody conferred protection to nude mice infected with HSV only if they were first reconstituted with syngeneic thymus cells 48 h before infection. The results suggest that both antiviral antibody and thymus-derived lymphocytes contribute to the recovery of HSV-infected hosts after passive immunization.     

The mode of action of splenic suppressor cells in murine plasmacytoma.

Polyclonal IgM secretion after LPS stimulation and antigen-dependent antibody production by splenic cells of BALB/c mice with plasmacytoma were found to be markedly reduced. The former occurred without mediation of suppressor cells, probably as the result of a quantitative reduction of mitogen-responsive B lymphocytes. On the other hand, the reduction in the generation of antibodies to sheep erythrocytes in vitro, as determined by the number of PFC, was found to be dependent on the activity of suppressor cells which had the functional characteristics of macrophages. The targets of these suppressor cells were found to be normal macrophages, B and T lymphocytes which were all independently suppressed. A diffusible factor appeared to be responsible for the suppressive effect of the suppressor macrophages interfering mainly with those early stages of the immune response involving antigen-triggered B cell activation.     

DX5+ NKT cells induce the death of colitis-associated cells: involvement of programmed death ligand-1

NKT cells are activated by CD1d and show an immune regulating function. Here, we investigated whether DX5+ NKT cells could be used to reduce colitis in a chronic colitis mouse model and studied the potential immunological mechanisms involved. Chronic colitis was induced either by transfer of enriched CD62L+ CD4+ T cells to severe-combined-immunodeficient mice or by feeding dextran sodium sulfate to immune competent mice. DX5+ NKT cells were transferred to mice with chronic colitis. Co-transfer of DX5+ NKT cells, but not CD8+ control cells, prevented the onset of colitis, and the immune regulatory effect of DX5+ NKT cells was completely abrogated by injecting CD1d blocking antibody. Moreover, DX5+ NKT cells reduced established colitis in both chronic colitis models. In vitro, DX5+ NKT cells induced cell death of colon-infiltrating lymphocytes isolated from diseased mice. This effect was inhibited in the presence of either anti-CD1d or anti-programmed death ligand-1 (PD-L1) blocking antibodies. The specific potency of DX5+ NKT cells in regulating chronic colitis in two mouse models is demonstrated. In vitro testing suggests that DX5+ NKT cells activated by CD1d induce cell death of colitis-inducing lymphocytes, which is mediated through PD-L1. Therefore, DX5+ NKT cells could be important in the regulation of immune responses associated with chronic colitis.     

Humoral immune response elicited by highly attenuated variants of vaccinia virus and by an attenuated recombinant expressing HIV-1 envelope protein

Attenuated variants of vaccinia virus have excellent potential for the construction of safe recombinant live vaccines. In this investigation, highly attenuated variants of vaccinia virus with several genetic markers and a variant recombinant were tested in Balb/c mice for their ability to induce humoral immune response. Mice primed with variants that had an 8-MDa deletion at the left end of the viral genome induced similar levels of circulating anti-vaccinia antibodies as the wild-type virus. However, mice primed with variants that had several genetic lesions (deletions and point mutations) induced lower levels of circulating anti-vaccinia antibodies. Mice primed and boosted with a recombinant variant with several genetic lesions, and containing the complete envelope gene of the human immunodeficiency virus (HIV) and the bacterial beta-galactosidase (beta-gal) gene, induced significant antibody response to gp 160 and beta-gal. The antibody response to gp 160 was markedly increased by successive inoculations with the recombinant variant. Our findings provide evidence that the extent of activation of the immune system by vaccinia variants can be modulated by the nature of the virus genetic lesion. In addition, when these variants are used as recombinant vaccines, it is possible to induce low levels of circulating anti-vaccinia antibodies after priming and yet achieve significant antibody response to virus-expressed foreign antigens, even after repeated boosters. Such variants could be useful in the design of live recombinant viruses as safe vaccines.     

Properties of mouse leukemia viruses. XVI. Suppression of spontaneous fetal leukemias in AKR mice by treatment with broadly reacting antibody against the viral glycoprotein gp 71.

AKR mice were treated early in life with antibodies prepared in a goat against the major glycoprotein gp71 of Friend murine leukemia virus (FL V) and evaluated over a period of up to 2 years for various parameters associated with AKR thymoma/lymphoma. If both the mothers and offspring were given the antibody, suppression of AKR disease was observed to the degree that the 50% incidence of leukemia was delayed by about 1 year. A lesser effect was found if the treatment was initiated at an age of 3 days and application of the antibody to either mothers alone, or treatment of the mice at later times (39 days) was not successful. Thus, it was concluded that the critical period for treatment is between birth and the first few days of life. Postmortem examination of animals that were successfully treated during this early period revealed a significant proportion exhibiting a unique leukemic pattern. In addition, several died from nonleukemic neoplasms as well as other causes. Levels of virus or antiviral antibodies were also determined at various times in individual mice which had been segregated in family groups. The results demonstrated that overall, successful antibody treatment resulted in (1) suppression of virus and (2) development of significant levels of antiviral antibodies. In contrast, control mice exhibited the opposite pattern: high levels of virus and no detectable antibody. Moreover, it could be shown that antibody treatment reduced the incidence of MCF-like recombinant virus isolation. A hypothesis is presented which suggests that the main function of the administered antibody is to disturb a key event occurring during the early stages of life of the AKR mouse, which is of critical importance for the development of leukemia from 6 months of age onwards.     

Deviation of immune response to Chlamydia psittaci outer membrane protein in lipopolysaccharide-hyporesponsive mice.

The outcome of infection is determined by both the quantity and the quality of an induced immune response. In particular, it has been demonstrated for selected pathogens that induction of TH1 or TH2 type helper T-cell subsets determines whether an immune response gives rise to protective immunity or disease-associated immunopathology. The nature of the antigen and the type of antigen-presenting cells recruited in the induction of a response are critical factors that influence the quality of the immune response. Of particular interest in this respect is the immune response to bacterial particles and the impact of cell wall-associated lipopolysaccharide (LPS) on that response. Nonspecific activation of macrophages and B lymphocytes by LPS could skew the phenotype of activated antigen-presenting cells and selectively alter the immunoglobulin isotypes and helper T-cell subsets that are induced following infection. In an initial attempt to detect immune deviation associated with LPS stimulation, we have compared the immunoglobulin isotypes of antibodies specific for the cysteine-rich outer membrane protein Omp2 induced in normal and LPS-hyporesponsive mice following immunization with Chlamydia psittaci strain guinea pig inclusion conjunctivitis whole elementary bodies. We report that there is a dramatic shift of Omp2-specific antibody from predominantly immunoglobulin G2a (IgG2a) isotype in LPS-hyporesponsive mice to high levels of IgG1 isotype in LPS-responder strains. The dependence of the IgG1 isotype shift on the LPS responder status is linked to the structure of the antigen and its natural processing pathway since LPS-hyporesponsive mice are not, in general, deficient in IgG1 antibody production. In particular, the antibody response to purified recombinant Omp2 is predominantly of the IgG1 isotype even in LPS-hyporesponsive mice.