单纯疱疹病毒糖蛋白单克隆抗体对致死性腹腔感染BALB/c幼鼠的被动保护作用

用单纯疤疹病毒(HSV)Ⅰ型SM44株和Ⅱ型SaV株分别腹腔感染BALB/c小鼠,于感染前后不同时间经腹腔注射HSV单克隆抗体(McAb)观察6株McAbs对致死性腹腔感染小鼠的被动保护作用。结果4株McAbs(2C5、1A12、Mad-2、1D10)对HSV-Ⅰ感染的小鼠有保护作用,5株McAbs(2C5、1A12、2A8、1D10、CH-A9)对HSV-Ⅱ感染的小鼠有保护作用。体内保护作用与体外的中和活性相关;并分析了McAbs在中和试验中有无补体参与条件下的保护能力。还证实了HSV糖蛋白在急性感染病程中其型特异性和型共同性抗原决定簇在体内的表达。     

单纯疱疹病毒体外诱导特异性抗体应答的实验研究

用灭活的单纯疱疹病毒（Ｈｅｒｐｅｓｓｉｍｐｌｅｘｖｉｒｕｓ，ＨＳＶ１）为致敏原，体外诱导成人外周血淋巴细胞产生特异性抗体应答，特异性抗体诱生水平与血清抗体无相关性（Ｒ＝０．４５，Ｐ＞０．０５），抗体类型为ＩｇＧ。同法致敬新生儿淋巴细胞则不能诱生任何类型的特异抗体，表明此体外抗体应答属继发性免疫应答。特异性抗体应答有明显的ＨＳＶ１抗原剂量依赖性，且需要Ｔ、Ｂ细胞的相互作用。ＨＳＶ１体外致敏的实验研究，为探讨正常和疾病状态下体外特异性抗体应答和免疫调节提供一个有用的实验模型。     

Relationship of Antibody to Outcome in Neonatal Herpes Simplex Virus Infections

Neutralizing antibody titers to herpes simplex virus type 1 (HSV-1) and HSV-2 were measured at birth in normal infants and uninfected infants of mothers with genital HSV infections during pregnancy and at the onset of infection in 5 infants with mild infections and 11 infants with severe infections. Thirty-eight percent of premature and 29% of term infants had neutralization titers of <1:5. High titers ([unk]1:40) were found in 55% of infants of mothers with primary infections during pregnancy and in 76% of infants of mothers with recurrent infections. The mean titers to HSV-1 and -2 in 5 infected infants with mild infections were 1:56 and 1:65 at the time of onset of infection, whereas the mean titers in 11 infants with severe infections were 1:11 and 1:12. Six natally exposed infants who remained asymptomatic were also studied and had a mean titer to HSV-1 of 1:85 and to HSV-2 of 1:69. Therefore, infants with high titers of transplacentally derived antibody had a more favorable outcome than infants with lower titers. Ninety-five percent of the infants of mothers with recurrent infections had a Rawls index of more than 85, suggesting that the antibody response was to HSV-2. However, low levels of antibody with this type specificity failed to protect four infants from infection with HSV-2. Augmentation of the neutralization titer to HSV-2 by the amount of complement present in cord serum was less than twofold. The study suggests that the quantity of antibody derived transplacentally affects the outcome of infection after natal exposure to herpes simplex virus. Complete neutralization of virus by antibody may occur in some infants, and prolongation of the incubation period and modification of the infection may occur in others.     

Natural Killer Cytotoxicity and Antibody-Dependent Cell Cytotoxicity to Herpes Simplex Virus-Infected Target Cells in Murine Pregnancy

ABSTRACT: In vitro natural killer cytotoxicity (NKC) and antibody-dependent cell cytotoxicity (ADCC) activity against herpes simplex virus (HSV)-infected cells were evaluated in a pregnant murine model (C₅₇B1₆inbred strain). Virgin (n = 16) and pregnant (late gestation) mice (n = 15) were infected intraperitoneally with HSV, type 1. After 18 hr, a 0.5-ml aliquot of the peritoneal wash was frozen for virus plaque assay, and the cells were cultured in the ⁵¹chromium release assay for NKC and ADCC. %NKC (mean ± S.E.) to HSV-infected targets was significantly suppressed (P < 0.05) in pregnant mice, 10.3% ± 1.9, compared to that of virgin mice, 32.5% ± 2.5. This suppression was abrogated with HSV-specific antisera (%ADCC); 53.9% ± 4.4 (pregnant) compared to 49.1% ± 3.6 (virgin). The diminished NKC activity in pregnant mice was reflected in an increased mean number of virus particles in the peritoneal wash, 266 + 66 PFU/ml, compared to 38 ± 11 PFU/ml in virgin mice (P < 0.05). We concluded that NKC, but not ADCC, to HSV-infected targets was suppressed and that HSV elimination was impaired in pregnant mice.     

Natural Killer Cell Cytotoxicity and Antibody-Dependent Cellular Cytotoxicity to Herpes Simplex Virus-Infected Cells in Human Pregnancy

ABSTRACT: Natural killer cell (NKC) cytotoxicity and antibody-dependent cellular cytotoxicity (ADCC) represent the ability of human leukocyte effector cells to destroy target cells in the absence and presence of antibody, respectively. Since these immune systems play a pivotal role in the body's primary lines of defense against a variety of pathogens including herpes simplex virus (HSV), a study was undertaken to evaluate the influence of pregnancy on these systems. Eleven uncomplicated gravidas were followed serially through each trimester and compared to 11 nonpregnant female controls. Mononuclear cells were acquired by Ficoll-Hypaque centrifugation of heparinized blood. Chang liver cells infected with HSV-I were utilized as target cells in a ⁵¹Cr release assay. Mean NKC values in the pregnant patients were uniformly lower than in the controls. No similar decreases in ADCC activity were observed in a comparison between the two study populations. These data support previous observations suggesting that pregnancy represents a relatively immunocompromised state. Differences apparently exist between NKC and ADCC effector cell populations with regard to the influence of pregnancy. Although these physiologic alterations in immunoregulation may help support the fetoplacental allograph, detrimental conditions may exist regarding susceptibility to various pathogens such as HSV.     

Sequential Changes in T and B Lymphocyte Responses to Herpes Simplex Virus in Man

Lymphocytes of seven patients with primary herpetic infection, twenty-three patients with recurrent herpes labialis and of nineteen control subjects were separated into T and B enriched cells by the use of nylon wool columns. In the absence of a herpetic infection the thymidine incorporation and macrophage migration inhibition responses to herpes simplex virus (HSV), Candida albicans and PPD, and the thymidine incorporation induced by PHA were functions of T cells. When a herpetic infection was present, the unfractionated lymphocyte response to HSV was increased, as measured by thymidine incorporation, but the T cell response was unchanged. However, T cells did show an increased response to HSV when prepared by elimination of cells forming rosettes with zymosan-complement. T cells of some patients were stimulated by contact with zymosan, and this correlated with the response to C. albicans. It is suggested that lymphocyte responses to HSV in man are mediated by T cells, but that these cells are specifically retained by nylon wool columns at the time of a herpetic infection. This may be associated with acquisition of an Fc receptor by the sensitized T cells.     

Frequency of Cytotoxic T Lymphocyte Precursors to Herpes Simplex Virus Type 1 as Determined by Limiting Dilution Analysis

The conditions for establishing a limiting dilution assay to measure cytotoxic T lymphocyte precursors (CTL-P) against herpes simplex virus type 1 (HSV-1) were determined. Analysis by Poisson statistics demonstrated that the estimated frequency of HSV-1-reactive cells in the spleens of normal mice was less than 1/250,000. In contrast, mice immunized previously with infectious HSV-1 demonstrated a CTL-P frequency between 1/3,500 and 1/15,670. The generation of a maximum cytotoxic T lymphocyte response required that mice be primed in vivo with infectious virus. Immunization with inactivated virus either failed to elicit detectable CTL-P frequencies or gave frequencies markedly less than those induced with infectious virus. To obtain positive cultures, the responder cell population had to be exposed to stimulator splenocytes expressing viral antigens. Normal splenocytes without virus or normal splenocytes with T cell growth factor did not result in significant cytotoxicity. Split culture analysis comparing cytotoxicity against syngeneic and allogeneic virus-infected targets provided evidence for specificity, H-2 restriction, and the T cell nature of the CTL-P. It was determined that precursors were eliminated by treatment with anti-Thy 1, Lyt 2.1, or Lyt 1.1, indicating the CTL-P were Lyt 1(+)2(+) cells. Cytotoxicity was reduced after treatment of the responders with anti-Lyt 2 plus complement, which gave further evidence of the T cell nature of the cytotoxic T lymphocytes. These experiments demonstrated the feasibility of using the limiting dilution approach as a highly sensitive and quantitative means to measure the cell-mediated immune response to HSV-1 antigens.     

An OspC-Specific Monoclonal Antibody Passively Protects Mice from Tick-Transmitted Infection by Borrelia burgdorferi B31

A murine monoclonal antibody directed against Borrelia burgdorferi B31 outer surface protein C (OspC) antigen was generated by a method whereby borreliae were inoculated into the mouse via the natural transmission mode of tick feeding. Passive immunization with this antibody resulted in protection of C3H/HeJ and outbred mice from a tick-transmitted challenge infection. Immunofluorescence staining of borrelia cells indicated surface exposure of the OspC epitope reactive with the monoclonal antibody.     

Macrophage Dependence of Polyriboinosinic Acid-Polyribocytidylic Acid-Induced Resistance to Herpes Simplex Virus Infection in Mice

The relative contributions of macrophages and lymphocytes to the induction of resistance to primary herpes simplex virus type 1 (HSV-1) infection by polyriboinosinic-polyribocytidylic acid complex [poly (I:C)] were investigated in C58 mice. The induction of resistance was found to be strongly dependent on macrophages compared to lymphocytes. Macrophage-deficient (silica-treated) mice produced less interferon and were not as responsive to prophylactic treatment of HSV-1 infections with poly (I:C) as were either normal, lymphocyte-deficient (cyclophosphamide-treated), or T-lymphocyte-deficient (anti-thymocyte serum-treated, adult-thymectomized) mice. Silica and cyclophosphamide treatments reduced the therapeutic activity of poly (I:C), whereas T-cell depletion did not have a significant effect. Similarly, the protection of mice with exogenous interferon was markedly reduced in silica-treated mice and moderately reduced in cyclophosphamide-treated mice, but unaffected in T-cell-deficient mice. Furthermore, suppression of HSV-1 plaque formation was obtained by cocultivation of infected mouse fibroblast monolayers with peritoneal (macrophage-rich) cells, but not with splenic (lymphocyte-rich) cells, from poly (I:C)-treated mice. Peritoneal cells did not protect heterologous (human) fibroblasts, suggesting that the protection of mouse embryo fibroblasts is mediated by interferon. Collectively, the data indicate that macrophages are required for the production of poly (I:C)-induced interferon and that macrophages and perhaps B-lymphocytes are important for mediating the protection against HSV-1 infection after interferon has been produced.     

Characterization of a Type-Common Human Recombinant Monoclonal Antibody to Herpes Simplex Virus with High Therapeutic Potential

We report the characterization of a type-common human recombinant monoclonal antibody previously isolated by antigen selection from a phage-displayed combinatorial antibody library established from a herpes simplex virus (HSV)-seropositive individual. Competition with well-characterized murine monoclonal antibodies and immunodetection of gD truncations revealed that this antibody recognizes the group Ib antigenic site of glycoprotein D, a highly conserved and protective type-common determinant. To our knowledge, this is the first human group Ib monoclonal antibody ever described. The antibody also displayed first-order neutralization kinetics and a high neutralization rate constant, was capable of completely inhibiting syncytium formation by a fusogenic strain of HSV type 1, and efficiently neutralized low-passage clinical isolates of both HSV serotypes. Taken together with our earlier observations of the in vivo antiviral activities of this human recombinant antibody in animal models of HSV infection, the present results support the high therapeutic potential of this antibody.     

Production of Specific Antibodies by Cerebrospinal Fluid Lymphocytes in Patients with Herpes Zoster, Mumps Meningitis and Herpes Simplex Virus Encephalitis

We applied a new method consisting of short-term culture (18 h) of lymphocytes from cerebrospinal fluid (CSF-L) and peripheral blood (PBL) in viral antigen-coated ELISA plates and subsequent measurement of IgG and IgM antibodies bound to antigen. Utilizing mumps virus, herpes simplex virus (HSV), varicella zoster virus (VZV), and measles virus as antigens, we demonstrated production by CSF-L of antibodies against the aetiological agent only in all patients with mumps meningitis and HSV encephalitis and also in all patients with herpes zoster without central nervous system (CNS) symptoms. This might be considered as direct evidence that specific antibodies are produced within the CNS in inflammatory nervous system diseases. CSF-L usually produced higher amounts of antibodies than the corresponding number of PBL. In comparison with concentrations of free antibodies determined in parallel, our method had higher specificity and sensitivity and gave more precise information about the antibody response in infections of the nervous system.     

Immunity to Chlamydial Infections of the Eye II. Studies of Passively Transferred Serum Antibody in Resistance to Infection with Guinea Pig Inclusion Conjunctivitis

Conjunctival infection of guinea pigs by the chlamydial agent of guinea pig inclusion conjunctivitis confers immunity. However, the mechanism of resistance to this intracellular pathogen is not yet defined. In the study reported here, serum immunoglobulin was passively transferred with resultant titers in excess of those known to be associated with immunity. Nonetheless, when the passive transfer recipients were challenged, they acquired infection which was neither delayed nor attenuated. Eye secretion antibody titers appeared and increased only after 11 days of infection in both passive transfer recipients and control groups, suggesting but not proving de novo local synthesis of secretory antibody. This study suggests that cellular or secretory immune mechanisms may predominate in resistance to this infection.     

表达2型单纯疱疹病毒糖蛋白D的重组痘苗病毒对动物的免疫效果

用表达２型单纯疱疹病毒（ＨＳＶ－２）糖蛋白Ｄ的重组痘苗病毒（Ｒ－ｇＤ－Ｖ）免疫ＣＢＡ小鼠，通过间接免疫荧光（ＩＦ）（观察小鼠抗体滴度变化）和四甲基偶氮唑蓝法（ＭＴＴ）动态观察细胞毒性Ｔ细胞活性（ＣＴＬ），均见增高并检测了其产生的抗体对动物的保护作用。同时对Ｒ－ｇＤ－Ｖ和２型单纯疱疹病毒（ＨＳＶ－２）产生的二次ＣＴＬ活性进行了比较，结果表明，Ｒ－ｇＤ－Ｖ产生抗体３周达高峰（１：６４０），７周下降为１：３２０，抗体对病毒攻击动物有保护作用，其中和指数（Ｎ）为６３。Ｒ－ｇＤ－Ｖ在鼠体内产生初次ＣＴＬ８ｄ达高峰，１２ｄ后消失；二次ＣＴＬ持续８周之久，且ＣＴＬ活性明显高于初次。Ｒ－ｇＤ－Ｖ和ＨＳＶ－２相比较产生ＣＴＬ在统计学上有差异（Ｐ＜０．０５）。说明Ｒ－ｇＤ－Ｖ能引起体液免疫和细胞免疫，对动物有保护作用。     

Host Defense Mechanisms Against Herpes Simplex Virus I. Control of Infection In Vitro by Sensitized Spleen Cells and Antibody

Sensitized mouse spleen cells decrease the spread of herpes simplex virus infection in cell culture lines derived from human and murine tissues. These washed, sensitized cells act alone and additively in combination with antibody to diminish the ability of single virus-infected cells to spread infection to contiguous cells. This control of infection is not species specific, unlike interferon, and appears to be distinct from the effect of antibody. Lymphotoxin was not detected in this lymphocyte-mediated response. This control of herpes simplex virus infection in vitro by sensitized lymphoid cells is immunologically specific; spleen cells from donor animals immunized with a heterotypic virus do not cause herpesvirus plaque size reduction. The ratio of spleen cells from immunized animals to target monolayer cells needed to produce this effect is > 4:1. Plaque size reduction of herpes simplex virus by spleen cells requires intact, immune, non-glass-adhering lymphoid cells.     

CD44 Expression by Leucocytes in Rheumatoid Arthritis and Modulation by Specific Antibody: Implications for Lymphocyte Adhesion to Endothelial Cells and Synoviocytes In Vitro

Anti-CD44 MoAb IM7 induced the loss of CD44 from mouse leucocytes thereby inhibiting leucocyte migration and joint inflammation in murine arthritis. Thus, targeting CD44 with MoAb may have potential for the treatment of patients with inflammatory joint diseases. Expression of CD44 by peripheral blood (PB) and synovial fluid (SF) leucocytes from rheumatoid arthritis (RA) patients was compared and the ability of IM7 to modulate this expression determined. RASF lymphocytes showed increased CD44 expression compared with those in PB indicative of an activated phenotype. As inflammatory SF did not up-regulate CD44 expression on PB lymphocytes, the increased CD44 expression by SF lymphocytes was a result of the selective homing of CD44^high cells to the synovium rather than an effect of the synovial environment. RASF granulocytes showed reduced CD44 expression compared with those in PB, again indicative of an activated phenotype. However, this reduction could be induced on PB granulocytes following culture with inflammatory SF and was inhibited by anti-TNF-α MoAb, implying that soluble factors in inflammatory SF such as TNF-α induced granulocyte activation and CD44 loss. IM7 induced the loss of CD44 from lymphocytes (both from PB and SF) and granulocytes in vitro , but was subsequently re-expressed after 24 h culture in the absence of the MoAb. This loss of CD44 was blocked by serine- and metalloprotease inhibitors implying that IM7 induced the proteolytic cleavage of CD44 by a mechanism similar to that reported for the loss of CD44 from PMA-activated granulocytes. Furthermore, IM7-treated CD44^low lymphocytes showed reduced adherence to both an endothelial cell line and RA synovial fibroblasts in vitro . The unique ability of IM7 to reduce CD44 expression by lymphocytes suggests that it could prevent lymphocyte extravasation and synovial infiltration in RA as previously reported in murine arthritis.     

Use of a Single Monoclonal Antibody To Determine the Susceptibilities of Herpes Simplex Virus Type 1 and Type 2 Clinical Isolates to Acyclovir

This report describes a flow cytometry drug susceptibility assay that uses a single fluorochrome-labeled monoclonal antibody to determine the acyclovir susceptibilities of herpes simplex virus (HSV) type 1 or type 2 clinical isolates. This assay yields 50% effective doses (drug concentrations that reduce the number of antigen-positive cells by 50%) for HSV clinical isolates that are equivalent to those obtained with the plaque reduction assay.     

Protection of Mice Against Vaccinia Virus by Bacterial Infection and Sustained Stimulation with Specific Bacterial Antigens

In these experiments, mice which have a strong delayed-type hypersensitivity to mycobacteria were found, when elicited with old tuberculin, to be more resistant to intravenous vaccinia virus challenge than controls. This was manifest as protection from killing when large amounts of virus were injected, or as significantly less tail swelling and damage as well as lower titers of infectious virus when a lesser inoculum was used. Preliminary experiments indicate that animals sensitized with Staphylococcus aureus and elicited with phage lysate of staphylococcus are also more resistant to vaccinia infection.