Cytotoxic T Cells in the Lungs of Mice Infected with an Influenza A Virus

Cytotoxic T cells are present in the lungs and the bronchoalveolar washings of mice infected intravenously (i.v.) or intranasally (i.n.) with live influenza A/WSN virus. After i.v. injection, cytotoxic T cell activity in both spleens and lungs reaches a peak at 6 days when the level of infectious virus recovered from the lungs falls sharply and the mice do not die. If a lethal dose of virus is given intranasally, very high levels of virus appear rapidly in the lungs, and the development of lung consolidation follows slightly behind the appearance of cytotoxic T cells there. When a non-lethal dose of virus is given intranasally, lower levels of virus are found in the lung and the appearance of cytotoxic T cells is delayed. These results suggest that the cytotoxic T cells play a protective role if the level of virus in the lungs does not reach very high levels. After injection of antithymocyte serum, the subsequent level of cytotoxic T cell activity in the lungs was greatly reduced, suggesting that the T cells recovered in lungs had at an earlier stage been circulating cells. However, splenectomized mice develop high levels of cytotoxic T cell activity, after intranasal infection of mice, indicating that the spleen did not contribute substantially to the T cells recovered in the lungs.     

Cells Mediating Delayed-Type Hypersensitivity in the Lungs of Mice Infected with an Influenza A Virus

Effector cells that demonstrate delayed-type hypersensitivity (DTH) on transfer with antigen to naive mice can he recovered from the lungs of mice inoculated intranasally 6 days earlier with a lethal dose (usually 5 × 10⁴EID₅₀) of influenza A virus. The activity recovered was proportional to the dose of virus instilled intranasally and the extent of lung consolidation observed. Active cells could also be recovered from the draining lymph nodes and from the peripheral blood. The effector cells were identified as T lymphocytes of Ly I phenotype and required I-region sharing between donor and recipient for activity to be elicited. They were cross-reactive within the A group of influenza viruses. Two experiments are reported in which immune cell preparations that expressed DTH activity but had very little cytotoxic T cell activity were transferred to mice inoculated 1 or 2 days earlier with a lethal dose of virus. The mice were not protected from death, and in both experiments, the recipient mice died more rapidly than the controls. These results contrast with earlier results in which cell preparations with high cytotoxic T-cell activity were shown to protect recipient infected mice from death.     

Structural Changes in the Brain of Mice Infected with Influenza A/H5N1 Virus

Structural changes in the brain of outbred mice were studied after infection with infuenza A/H5N1 strain isolated in the Novosibirsk region. High mortality was observed after intranasal infection. Examination of brain specimens revealed vasculopathies with thrombosis of the microcirculatory vessels, pericellular and perivascular edema with multifocal ischemic necrosis, hyperplasia of glial cells, caspase-dependent apoptosis of neurons caused by the cytopathic effect of the virus, and hypercytokinemia.     

Hemopoietic Colony-Forming Cell Responses in Mice Infected with Ectromelia Virus

The responses of in vitro colony-forming cells (granulocyte-macrophage progenitor cells) were studied in the bone marrow, spleen, and blood of mice after intraperitoneal injection of 10(3) plaque-forming units of ectromelia virus. This study showed that a colony-forming cell response occurred during infection and was accompanied by an increase in the serum level of colony-stimulating factor and, at a later stage, colony-inhibiting factor. Changes in the proportions of colony-forming cells were not due to relative changes in other cell populations. The relationships between colony-forming cell responses and levels of infection were complex, higher levels of infection being associated with good colony-forming cell responses in bone marrow and apparently poor colony-forming cell responses in blood and spleen.     

Effect of Methionine Enkephalin on Natural Killer Cell and Cytotoxic T Lymphocyte Activity in Mice Infected with Influenza a Virus

Methionine enkephalin (Met-Enk) was evaluated for efficacy as an immune activator and potential therapeutic agent in influenza A/NWS/33 (H1N1) viral infections in female BALB/C mice. Influenza infection was induced intranasally with an approximate 90% lethal dose of virus and mice were treated intraperitoneally with doses of 10, 3 and 1 mg/kg/day, with treatments given 24 h pre-, 24 h post-and 72 h post-virus exposure. Splenocytes were assayed for natural killer cell (NK) and cytotoxic T lymphocyte (CTL) activity at time periods 76, 96 and 120 h post virus exposure. the 10 mg/kg dosage level significantly increased both CTL and NK activity at all time periods assayed. Other treatment schedules included single doses of 20, 10 and 3 mg/kg/day Met-Enk at either 24 h post-or 72 h post-virus exposure, with highly significant increases in NK and CTL activity noted after the latter treatment. the results of this study demonstrate the immunomodulatory effects of Met-Enk on NK and CTL in influenza infected mice and suggest a potential for therapeutic applications.     

Changes in the Specificity of Antibodies in Mice Infected with Lactate Dehydrogenase-Elevating Virus

Infection with lactate dehydrogenase-elevating virus (LDV) modifies the isotypic distribution of antibodies (Ab) directed to several antigenic proteins with a preferential production of IgG2a. Because it was not known whether the virus could also affect the Ab specificity, the authors addressed this point using human growth hormone (hGH) as a model antigen. Anti-hGH monoclonal antibodies (MoAb) were used as probes to study the occurrence of Ab to three native hGH epitopes (3C11, F11 and 10D6) in sera from LDV-infected CBA/Ht and BALB/c mice immunized with hGH. Competition ELISA was used to determine the extent of Ab directed to cryptic hGH epitopes, i.e. antigenic determinants hidden in the native hormone. Results indicated that in LDV-infected CBA/Ht mice the titres of anti-hGH Ab were lower than in controls, although a consistent isotypic shift to IgG2a subclass was observed. Concurrently, the presence of Ab to epitopes 3C11, F11 and/or 10D6 were markedly reduced in infected animals and most anti-hGH Ab were directed to hGH cryptic epitopes. By contrast, LDV infection increased the amount of anti-KLH Ab elicited by CBA/Ht mice and did not affect Ab specificity, whilst control and LDV-infected BALB/c mice showed similar concentrations of anti-hGH Ab. Furthermore, the proportion of Ab to cryptic hGH epitopes did not change in infected animals even though an important shift to IgG2a was detected. Thus, data presented herein suggest that LDV infection modifies Ab specificity depending on the mice genetic background and on the antigenic characteristics of the immunogen.     

Structural and Functional Changes in Pulmonary Macrophages and Lungs of Mice Infected with Influenza Virus A/H5N1 A/goose/Krasnoozerskoye/627/05

C57Bl/6 mice were intranasally infected with influenza virus A/H5N1 A/goose/Krasnoozerskoye/627/05. The mortality rate of animals reached 70% on day 14 of the disease. The lungs of animals were characterized by necroses, destruction of vessels, hemorrhagic and thrombotic complications, edematous syndrome, and early fibrosis of the interstitium. On days 6-10 after infection, fibrosis was found in the zones of postnecrotic inflammatory infiltration. The expression of lysozyme and myeloperoxidase by pulmonary macrophages was initially increased, but decreased on day 10 of the study. The number of cathepsin D-expressing macrophages was elevated up to the 10th day of examination.     

Macrophage Turnover Kinetics in the Lungs of Mice Infected with Streptococcus pneumoniae

Streptococcus pneumoniae is the most prevalent cause of community-acquired pneumonia and is known to induce apoptosis and necrosis in macrophages in vivo. We analyzed the kinetics of alveolar and lung parenchymal macrophage replacement by newly recruited exudate macrophages in vehicle-treated and S. pneumoniae-challenged bone marrow chimeric CD45.1 mice. After lethal irradiation, CD45.1 alloantigen-expressing recipient mice were transplanted with bone marrow cells from CD45.2 alloantigen-expressing donor mice. After only 24 hours of low-dose S. pneumoniae infection, approximately 60% of CD45.1(pos) recipient-type alveolar macrophages (AM) were replaced by CD45.2(pos) donor-type exudate AM in bronchoalveolar lavage fluid, and this increased to more than 80% on Day 7 of infection. In contrast, lung parenchymal macrophages of S. pneumoniae-infected chimeric CD45.1 mice were replaced by only about 10% by 24 hours, although this increased to over 80% by Days 3 to 7 of infection. This dramatic macrophage turnover was accompanied by early induction of apoptosis/necrosis in donor-type exudate AM peaking at 6 hours after infection, whereas peak apoptosis/necrosis induction in recipient-type AM was delayed until Day 7. Collectively, these data for the first time demonstrate that S. pneumoniae infection of the lung triggers a brisk turnover of both resident and recruited mononuclear phagocyte subsets, and suggest an important role of exudate but not resident macrophages in re-establishing alveolar and lung homeostasis.     

Appearance of Rosette-Forming Macrophages in the Lungs of Influenza Virus-Infected Mice

Approximately one fifth of the macrophages obtained from the lungs of mice infected 2 to 5 days with influenza A/HK virus were found to rosette well with either unmodified human, chicken, or guinea pig erythrocytes, but not with erythrocytes from hamsters, sheep, or mice. Rosette-forming macrophages were seldom seen in suspensions from uninfected mice (3+/-3%) or mice infected 24 h previously (3+/-3%). Rosette formation was not due to virus hemadsorption, as indicated by the failure of specific antiserum to influenza virus to block rosette formation; by the induction of comparable levels of rosette-forming macrophages in the lungs of mice infected with herpes simplex virus type 2, a nonhemadsorbing virus; and by the inhibition of rosette formation at 4 degrees C. Instead, rosette formation appeared to be directly related to macrophage elicitation or activation since nonstimulated macrophage populations such as peripheral blood monocytes, macrophages from uninfected lungs, or noninduced peritoneal macrophages were not observed to rosette to any significant extent. Furthermore, peritoneal macrophages induced with filter-sterilized normal horse serum rosetted at levels comparable to that observed with cells from infected lungs. These results indicate that hemadsorption alone can not be used as a criterion of virus infection of macrophages. However, rosette formation may serve to identify macrophage subpopulations which are active in host defense against viral infections.     

Influenza Virus-Specific T Cells Fail to Reduce Lung Virus Titres in Cyclosporin-Treated, Infected Mice

Cyclosporin A (CsA) inhibited the function(s) of transferred influenza-specific K,D-restricted cytotoxic T cells, which led to clearance of virus in the lungs of influenza virus-infected mice. CsA had no effect on the migration of the transferred cells to the lungs. The pattern of migration and the number of cells recovered from the lungs were similar when cells were transferred into normal, untreated, infected or CsA-treated, infected mice. CsA had no effect on the in vitro expression of cytotoxic activity by the K,D-restricted cytotoxic T cells. These findings strongly suggest that the in vivo clearance of influenza virus by K,D-restricted cytotoxic T cells involves a lymphokine mechanism.     

Hepatitis in Mice Infected with Coxsackie Virus B1

The livers of mice of different ages were readily damaged by Coxsackie virus B₁ infection. The severity of liver damage decreased as the age of the mice increased. Coxsackie B₁ viral crystals were not found in the damaged liver cells in spite of severe pathological changes of the liver, both histologically and electron microscopically, and even though characteristic crystal formation was observed in the pancreas of three of these same animals. Nevertheless, the hepatic damage was considered to be due to direct viral invasion of the hepatic cells. This injury was followed by a variety of degenerative and necrotic processes displaying somewhat characteristic morphological manifestations. The severe hepatic infection produced in the newborn mice resulted in their death from a rather fulminating illness, whereas in the older mice there was recovery from mild to moderate hepatic injury with cellular regeneration by the fourth day after viral inoculation. The experimental preparation used here provides an excellent means for the study of the processes of injury and healing of the liver infected with a virus that is also infectious for man.     

Cellular and Cytokine Characterization of Vascular Inflammation in CBA/J Mice Chronically Infected with Trypanosoma cruzi

Continuing our characterization of the immunopathological events occurring during experimental murine Chagas' disease, an immunohistological examination was conducted of the aortas of chronically infected CBA/J mice. Compared with non-infected mice of identical age, Trypanosoma cruzi -infected mice exhibited a marked vasculitis, with significant infiltration of inflammatory cells into the adventitial layer, including CD4⁺, CD8⁺ T cells and macrophages. Production of interleukin-6 (IL-6) and tumour necrosis factor-α (TNF-α) was evident in the inflammatory infiltrate in the endothelial and smooth muscle layers. Vasculitis was most apparent in proximity to the heart, but extended along the aorta. Such an inflammation could lead to an alteration of the endothelium, altering the protective properties of this layer and further contributing to the focal pathology characteristic of this stage of infection.     

The Recovery of Mice from Influenza A Virus Infection: Adoptive Transfer of Immunity with Influenza Virus-specific Cytotoxic T Lymphocytes Recognizing a Common Virion Antigen

Mice inoculated intranasally with infectious influenza virus of a given A strain were adoptively transferred 24 h later with preparations of secondary influenza virus-immune T cells generated either in vitro or entirely in vivo. The immune cells were raised during infection with homologous or heterologous A strain influenza viruses or with a type B virus. The greatest antiviral effect, measured by reduction in lung virus level of recipient mice, occurred if homologous viruses were used. Sharing of haemagglutinin specificity was shown to be important, but significant antiviral activity was still expressed if neither haemagglutinin nor neuraminidase antigenic specificities were shared. The antiviral effect was type-specific. Adoptive transfer of type A influenza immune T cells did not express antiviral activity against type B virus, and vice versa. On the basis of earlier work, the effector population in the transferred cells was cytotoxic T cells (Tc). Intranasal reinfection of mice with a heterologous type A virus sharing neither haemagglutinin nor neuraminidase antigenic specificity with the first infecting virus induced enhanced and earlier production of cross-reactive Tc against type A influenza viruses. This was paralleled by significantly lower virus levels in the lungs. The results of this work demonstrate heterotypic cell-mediated immunity in influenza virus infection in mice.     

Defects in the Immune System of Mice Infected with Lymphocytic Choriomeningitis Virus

Acute lymphocytic choriomeningitis virus infection of adult mice is associated with general immunosuppression, which develops during the 2nd week of the infection and persists for a period of 2 to 3 months. Studies of some cellular events in the immune system of infected mice brought to light a number of findings which seemed relevant to this immunosuppressive effect. Colony-forming stem cells, which may act as the precursors of the lymphoid cells, were temporarily inhibited during the first period of the infection. Presumably this inhibition also affected the thymus cells, which decreased dramatically at the same time. At a later stage of infection, defects developed within the population of immunocompetent cells, and this was most probably a consequence of the preceding suppression of the precursor cells. The defects in the immunocompetent cells were temporally related to the immunosuppression and seemed to be the ultimate cause of this phenomenon. At all events, antibody-forming cells were not damaged by the virus. In studies of neonatally infected baby mice, it was found that the development of immunological responsiveness was completely abolished for the first 2 weeks of life. It is therefore probable that the generation of immunocompetent cells was also affected in the babies. Evidence was obtained supporting the hypothesis that this effect played an important role for the induction of tolerance to the virus in the neonatally infected mice.     

Experimental Pneumococcal Infection in Embryonated Eggs Previously Infected with Influenza C Virus

Cumulative mortality rates for embryonated eggs, compiled for a variety of pneumococcal strains, indicated a uniformly high degree of virulence. Based on median lethal dose values for combined infection with influenza C virus and single infection with pneumococcus, it is difficult to conclude that pneumococcal virulence is enhanced in combined infection. However, there was a significant increase in mortality during the first 24 hr following combined infection. A significantly greater in vitro growth rate for pneumococcus was shown in amniotic fluid from virus-infected embryonated eggs. Microscopy study of selected embryos revealed the absence of a discernible inflammatory reaction during the first 24 hr. From 48 through 96 hr after inoculation, inflammatory reactions were restricted to the upper and lower respiratory tract and consisted of a polymorphonuclear leukocyte infiltration in response to the localization of pneumococci in the paranasal sinuses, bronchi, alveolar ducts, and alveolar spaces. Extensive, severe inflammatory reactions were more frequently encountered in the combined infection. The various phases from early exudation to eventual resolution, which characterize pneumococcal pneumonia in the human host, were in clear evidence following both the single and combined infection of the chick embryo.     

肾综合征出血热病毒气溶胶感染乳小鼠抗原定位和病理的研究

用肾综合征出血热病毒（ＨＦＲＳＶ）气溶胶对乳小鼠全身暴露感染３０ｍｉｎ，每只乳小鼠吸入了０．０７４空斑形成单位（ＰＦＵ）的病毒。经检测发现，肺从４ｈ～２１ｄ、脑和肾从２～２１ｄ、脾从３～２１ｄ、肝和胸腺从５～２１ｄ、心从７～２１ｄ为阳性。提示，ＨＦＲＳＶ经呼吸道感染后第７ｄ就可造成全身弥散性感染，脑、肾和牌中的病毒来源可能是吸入病毒经血播散而至，肝、胸腺和心中的病毒来源可能是肺、脑、肾和脾中复制的新生病毒经血播散而至。病毒抗原主要定位于器官组织的实质性细胞和血管内皮细胞。病毒定位引起了器官组织细胞的不同程度的变性和坏死，且有出血、淤血和淋巴细胞侵润的现象。病毒的定位和直接作用可能是组织病理变化的一个重要始动因素。