Characteristics of infection of B and T lymphocytes from mice after inoculation with cytomegalovirus.

Viremia was produced by inoculating intravenously BALB/c mice with murine cytomegalovirus. Virus was detected in plasma and granulocytes only during the first 8 days after infection. Lymphocyte-associated viremia, detectable by cocultivation on syngeneic or allogeneic fibroblasts, persisted for at least 4 weeks. Eight to 10 days after infection, sonicated lymphocytes had no demonstrable free virus. When whole lymphocytes with no demonstrable free virus were enclosed in a Millipore chamber and placed on a fibroblastic feeder layer, T cells produced free virus but B cells did not. Compared to normal calf serum, specific hyperimmune serum reduced B cell-associated infectious centers by 74% and T cell-associated infectious centers by only 38%. Normal mouse sera reduced by 36% and 30% infectious center production by B cells and T cells, respectively. Lymphocytes enriched with Fc receptor-positive cells produced significantly more infectious centers than receptor-negative cells.     

Selective induction of murine oncornavirus gene expression in somatic cell hybrids between mouse peritoneal macrophages and SV-40-transformed human cells

Somatic cell hybrids of BALB/c and C57BL/6 mouse peritoneal macrophages and human Lesch-Nyhan fibroblasts transformed by infection with simian virus 40 have been examined by radioimmunoassay for synthesis of the viral envelope and major core proteins of the murine endogenous type-C oncornavirus. Little or no detectable viral proteins (approximately 1 ng/mg of protein) were present in extracts of the parental cells. In contrast, hybrid clones of BALB/c cells containing human chromosome 7 with the simian virus 40 genome synthesized extremely high concentrations of the murine type-C virus core protein (210 to 6000 ng/mg) and also the envelope glycoprotein (83 to 250 ng/mg). The highest concentrations were in cells containing human chromosomes 5 and 11 in addition to 7. There was little or no change in the concentration of viral proteins in the C57BL/6-human cell hybrids. These results suggest that human-cell gene products can be active in the induction of the murine oncornavirus genome.     

Expression of type C virus p30 in mouse cells infected with herpes simplex virus.

Expression of type C virus p30 in BALB/c and NIH Swiss mouse cells infected with live (productive infection) or uv-irradiated herpes simplex virus (uv-HSV) types 1 and 2 was studied by use of either immunofluorescent (FA) or radioimmunoassay (RIA) procedures. No evidence for enhanced expression of p30 was obtained with either of these procedures although activation of type C virus in uv-HSV-infected BALB/c cells was readily demonstrated at low frequencies (～10^?4) by infectious center assay. While FA staining with rabbit anti-murine leukemia virus (MuLV) p30 serum was observed in mouse cells productively infected with HSV or infected with uv-HSV, this was shown to be nonspecific. It was concluded that activation of type C virus in uv-HSV-infected BALB/c cells does not occur in significantly more cells than detected by infectious center assay.     

Immunological properties of murine thymus-dependent lymphocyte surface glycoproteins.

The immunological properties of two murine thymus-dependent (T) lymphocyte surface glycoproteins, T200 and T25, were investigated. T200 is a lymphocyte-specific antigen with a high degree of species specificity. It shares antigenic determinants with molecules present on thymus-independent (B) lymphocytes. T25 has antigenic determinants which cross-react with antigens on mouse brain, rat thymocytes and rat brain. An antiserum against a purified rat brain glycoprotein which carries Thy-1.1 reacts with T25. Absorption of this antiserum with BALB/c thymocytes or brain homogenate produces a Thy-1.1 specific serum which reacts with T25 from AKR/J thymocytes but not with T25 from AKR/Cum thymocytes. These results confirm that T25 is the molecule on the surface of mouse T cells which carries the Thy-1 antigen. T25 also carries antigenic determinants, recognized by anti-thymocyte serum (ATS), which were found on secondary mouse embryo fibroblasts and untransformed fibroblast cell lines but which were not detected on fibroblast cell lines transformed with murine sarcoma virus (MSV) or with Simian virus 40 (SV40).     

Viral RNA of murine sarcoma virus produced by a hamster-mouse somatic cell hybrid

The viral RNA genome of murine sarcoma virus synthesized by a hybrid cell, THR-23, which was isolated by Sendai virus fusion of a hamster cell (HT-1) carrying the defective Moloney sarcoma genome and Swiss mouse cells shedding Rauscher leukemia virus was characterized. The THR-23 clone produces a 500-fold excess of sarcoma over leukemia virus. The high molecular weight (HMW) RNA from the virion of THR-23 gave a single peak in sucrose gradients with a sedimentation coefficient of 60 S, smaller than that of Rauscher murine leukemia virus (70 S). The 60 S HMW RNA of THR-23 virus dissociates to a 30 S subunit RNA after heat denaturation. The size is different from 35 S subunit RNA of 70 S RLV RNA. 30 S intracellular virus-specific subunit RNA was detected in HT-1 and THR-23 cells by hybridization with DNA product made in vitro from Moloney murine sarcoma and leukemia virus. These results suggest that the defective genome (30 S RNA) of the HT-1 cell is synthesized in the THR-23 hybrid cell, aggregated to 60 S viral HMW RNA and incorporated into the virion. In contrast to the avian system, the murine sarcoma genome is smaller by sedimentation than the murine leukemia genome.     

Spontaneous expression of endogenous type C RNA virus by BALB/c splenic B lymphocytes in continuous culture.

B-lymphocyte cultures were established from spleens of BALB/c, C57B1/6, NIH Swiss, and SWR mice of various age groups. Spontaneous, consistent, and thus predictable release of a B-tropic mouse endogenous virus occurred from the very first passage in cultured lymphocytes derived from BALB/c mice 6 months old or older but not from similar lymphocytes derived from BALB/c mice of 1.5 or 3 months of age. C57Bl/6, NIH Swiss, and SWR mice belonging to various age groups ranging between 1.5 and 18 months failed to exhibit such a spontaneous release of viral particles. We conclude that in BALB/c splenic B lymphocytes a breakdown of cellular control mechanisms occurs in older animals leading to viral production while such a phenomenon is absent in C57Bl/6, NIH Swiss, and SWR mice.     

Short- and long-term studies on chemical carcinogenesis in BALB/Mo mice

To study the interactions between chemical carcinogens and oncogenic retroviruses, BALB/Mo mice which carry the Moloney murine leukemia virus (M-MuLV) as an endogenous virus, and conventional (M-MuLV-free) BALB/c mice, as well as their Bc1 (M-MuLV+ or M-MuLV-) hybrids were injected neonatally with a single dose of urethane. BALB/Mo and V+ Bc1 mice showed accelerated lymphoma development; similar results were obtained in BALB/Mo mice receiving one or two doses of urethane transplacentally. Lung adenomas developed with shorter latency and higher incidence in BALB/Mo mice given urethane at birth; however, significant differences in the incidence of lung adenomas in BALB/Mo mice were found only in two experiments. Additional short-term experiments were carried out to investigate the mechanism of the higher susceptibility to sister chromatid exchange induction observed in BALB/Mo lymphocytes. It was found that BALB/Mo spleen lymphocytes incubated with cordycepin, an antiviral antibiotic, with or without mitomycin C treatment, showed reduction in both M-MuLV synthesis and sister chromatid exchange frequency, and the latter values were similar to those seen in control cultures. These data suggest that the integration of M-MuLV proviral DNA into the host genome is per se not sufficient to increase the susceptibility to carcinogenic stimuli, but that other events, such as viral gene expression and amplification, are most likely required for the chemical-viral synergistic effect to occur.     

Analysis of the inhibitory effect of peritoneal macrophages on the spread of herpes simplex virus.

Peritoneal macrophages obtained from mice after an intraperitoneal injection of tryptose peptone inhibited the development of herpes simplex virus type 2 plaques in syngeneic mouse embryonic fibroblasts. In contrast, peritoneal macrophages, spleen cells, and thymocytes from untreated mice showed only a minimal inhibitory effect on the development of viral plaques. The effect was age dependent. Macrophages from 2 and 3-week-old mice showed weaker functions, requiring a larger number of cells for an equivalent reduction of plaques and virus yield than those from adult mice. When macrophages were treated with procaine, their phagocytic activity was completely abolished. However the procaine-treated macrophages still could inhibit the development of viral plaques. Peritoneal macrophages did not show any increased cytotoxicity against herpes simplex virus-infected cells; plaque inhibition might rather be attributable to their cytostatic effects on target cells.     

Activation/proliferation and apoptosis of bystander goat lymphocytes induced by a macrophage-tropic chimeric caprine arthritis encephalitis virus expressing SIV Nef

Caprine arthritis encephalitis virus (CAEV) is the natural lentivirus of goats, well known for its tropism for macrophages and its inability to cause infection in lymphocytes. The viral genome lacks nef, tat, vpu and vpx coding sequences. To test the hypothesis that when nef is expressed by the viral genome, the virus became toxic for lymphocytes during replication in macrophages, we inserted the SIVsmm PBj14 nef coding sequences into the genome of CAEV thereby generating CAEV-nef. This recombinant virus is not infectious for lymphocytes but is fully replication competent in goat macrophages in which it constitutively expresses the SIV Nef. We found that goat lymphocytes cocultured with CAEV-nef-infected macrophages became activated, showing increased expression of the interleukin-2 receptor (IL-2R). Activation correlated with increased proliferation of the cells. Interestingly, a dual effect in terms of apoptosis regulation was observed in exposed goat lymphocytes. Nef was found first to induce a protection of lymphocytes from apoptosis during the first few days following exposure to infected macrophages, but later it induced increased apoptosis in the activated lymphocytes. This new recombinant virus provides a model to study the functions of Nef in the context of infection of macrophages, but in absence of infection of T lymphocytes and brings new insights into the biological effects of Nef on lymphocytes.     

Differences in mouse interferons according to cell source and mode of induction.

Mouse interferon induced by ultraviolet-irradiated Newcastle disease virus or polyriboinosinic-polyribocytidylic acid in T lymphocytes, B lymphocytes, macrophages, and primary mouse embryonic cell culture was studied. Irrespective of the inducer, interferons produced by T or B lymphocytes were relatively heat stable and of low antigenicity when reacted with antiserum against L-cell interferon (ALI), whereas interferons produced by macrophages and mouse embryo cells were heat labile and of high antigenicity against ALI. Mouse interferons induced by ultraviolet-irradiated Newcastle disease virus were separated into three components by chromatography on CH-Sepharose 4B. Interferons produced by T and B lymphocytes consisted primarily of component 1 (unbound fraction), whereas interferons produced by macrophages or mouse embryo cells consisted primarily of component 3 (eluted by 0.5 M NaCl). Component 1 was heat stable and of low antigenicity against ALI, properties characteristic of T- and B-cell interferon. Components 2 and 3 were heat labile and of high antigenicity against ALI, properties characteristic of macrophage and mouse embryo cell interferon. In contrast, interferon induced in mice sensitized with BCG differed from these interferons induced in B cells, T cells, macrophages, and fibroblasts in being extremely acid labile and nonreactive against ALI.     

Pathogenesis of murine cytomegalovirus infection: the macrophage as a permissive cell for cytomegalovirus infection, replication and latency.

Macrophages harvested from the peritoneal cavities of mice of several strains were permissive to infection with murine cytomegalovirus (MCMV). Macrophages from six mouse strains released equivalent amounts of plaque-forming virus into the culture fluids and cells from mouse strains scored similarly in numbers of infectious centres. Twenty to 50% of the infected macrophages obtained after thioglycollate activation formed infectious centres. When studied by in situ hybridization, more than 82% of infected macrophages (with or without thioglycollate activation) contained MCMV DNA. Macrophages obtained from latently infected mice were examined for their content of MCMV. Using co-cultivation assays, MCMV was frequently recovered from thioglycollate activated macrophages harvested from latently infected mice but only rarely recovered from non-activated macrophages. MCMV DNA--mouse DNA hybridization assays revealed four to seven virus genome DNA copies per 100 cells. These studies indicate that macrophages harvested from mice susceptible (BALB/cSt) or resistant (C3H) to MCMV infection replicated virus equivalently and that macrophages are a reservoir of MCMV during latent and chronic infections. Activation of macrophages may be one of the important steps leading to the exacerbation of in vivo latent infections.     

Differential expression of murine CD81 highlighted by new anti-mouse CD81 monoclonal antibodies

We describe the use of a soluble CD81-Fc fusion protein to screen for novel monoclonal antibody (MAb) reactive with the extracellular loops of murine CD81 (TAPA-1). Two such MAbs, Eat1 and Eat2 (for Extracellular Anti-TAPA1), were used to assess the expression and function of CD81 on murine lymphocytes. Although CD81 is expressed uniformly on all human lymphocytes, murine CD81 was found to be expressed at much higher levels on resting B cells than on resting T cells. This was particularly evident when staining with the new MAbs, Eat1 and Eat2. The molecule is also functionally active on B cells, as Eat1 and Eat2 induce homotypic adhesion of B lymphocytes. Stimulated B cells undergo early apoptotic events in the presence of Eat2, as shown by binding of Annexin V-fluorescein isothiocyanate (FITC). Polyclonal activation of murine T cells also induces higher level CD81 expression, and many immortalized murine T-cell lines express high levels of the protein. In contrast to human CD81, which is expressed equally on all thymocytes, murine CD81 is induced during thymic development, being expressed at high levels on CD4+CD8+ thymocytes, in contrast to other subsets of thymocytes. Finally, murine dendritic cells, splenic macrophages, and non-killer (NK) cells all express high levels of CD81. We conclude that CD81 is differentially expressed in the murine immune system, and is involved in regulating the adhesion and activation of murine B cells.     

An anti-mouse macrophage monoclonal antibody reacting with T-derived leukaemic cells.

Hybridomas secreting anti-mouse macrophage antibodies were obtained by fusing a murine plasmacytoma with lymphocytes of a rat immunized against mouse macrophages. An IgM, monoclonal antibody (3 A 35) reacted with mouse monocytes, macrophages and polymorphonuclear leucocytes. It did not bind appreciably to erythrocytes, platelets and unstimulated T or B lymphocytes. However, 3 A 35 bound to various murine T-derived leukaemic cells and to a small proportion of Con A-stimulated thymocytes. Cross absorption experiments confirmed the existence of a common antigenic determinant on macrophages and leukaemic cells. The possibility that 3 A 35 identified a previously described antigen common to macrophages and normal or leukaemic T cells was investigated. The antibody was tested against thymocytes and macrophages of various mouse strains, some congenic for H-2 or T1a. The 3 A 35-detected antigen was found to be different from Ly5, Tla and Qa and did not represent a I-J encoded allotypic specificity.     

柯萨奇B3病毒诱导小鼠单核—巨噬细胞产生肿瘤坏死…

观察柯萨奇Ｂ３病毒对小鼠单核－巨噬细胞的感染，及诱导白细胞介素６和肿瘤坏死因子的产生。发现ＣｏｘＢ３能够感染小鼠腹腔巨噬细胞，并释放感染性病毒颗粒；一定感染量病毒的感染，对Ｍｏ／Ｍψ的存活率无明显影响。     

Increased neoplasm development due to immunosuppressive treatment with FK-506 in BALB/C mice persistently infected with the mouse herpesvirus (MHV-72).

BALB/c mice were infected with the lymphotropic mouse gammaherpesvirus (MHV-72). At late (7-12 months) post-infection intervals the latent virus was detected in the cells of lymphatic system (peripheral blood, lymphocytes and macrophages, thymocytes, lymph nodes, bone marrow, and peritoneal macrophages,) and in the spleen, lungs, liver, and kidney by cocultivation as well as by explantation. The MHV-72 infected mice, in which latency had been established, were treated with the immunosuppressive (IS) drug FK-506 (2 mg/kg/mouse for 30 days). This treatment increased the probability of virus reactivation by over two-fold. During the post-treatment observation period of 19 months, the incidence of lymphomas and the development of MHV-related lymphoproliferative and hemoblastic disorders raised to nearly five-fold in the drug treated mice as compared to untreated animals.