T lymphocytes and the transfer of immunity to Trypanosoma musculi in mice.

Trypanosoma musculi produces a resolving infection in mice and the immune response is thymus dependent. Spleen cells from immune and from uninfected mice were transferred to T cell-deprived mice and restored their ability to control the infection, the immune cells being effective most rapidly. Treatment of the cells in vitro with anti-theta serum did not impair their ability to restore immunocompetence and it is proposed that, though T-cell dependent, the immune response is effected by theta- cells.     

A lung model of schistosome immunity in mice.

When mice are challenged intravenously with schistosomula of Schistosoma mansoni, host cell reaction and parasite attrition proceed entirely in the lung, where these events can be followed by quantitative histology and worm recovery. In nonimmune animals the destruction of schistosomula in the lungs proceeds gradually, resulting in the elimination of about 80% of the challenge organisms after 6 days. Cell reaction begins promptly, as evidenced by the appearance of neutrophilic foci around many of the lung schistosomula within 30 minutes after injection, and results in increasing numbers of damaged organisms and residual inflammatory foci 24 hours and 6 days later, respectively. In contrast, when schistosomula are injected into mice immune by virtue of an established S. mansoni infection, parasite destruction is augmented and accelerated, a process already evident by 24 hours. By the sixth day, 98% of the challenge organisms have been eliminated, a substantially greater reduction in parasite survival than that occurring in the normal host. This increased attrition of schistosomula is also reflected in the decreased numbers of parasites recovered from minced lung tissue of immune mice 6 days after challenge. Immune cellular inflammatory reactions to schistosomula are, likewise, greatly intensified and can be readily distinguished from those of normal mice by the proportions of parasites involved and by the large numbers of eosinophils surrounding them. In some instances, degranulation of eosinophils onto the parasite tegument is observed. Schistosomula cultured for 24 or 44 hours in a medium containing mouse red blood cells elicit significantly less cellular reaction and show greater survival in the lungs of immune animals than do freshly derived schistosomula. It would therefore appear that the susceptibility of maturing schistosomes to immune cellular attack is limited to the first day or two after their metamorphosis from cercariae. These observations form the framework of a new in vivo model for analyzing the dynamics of the cellular and humoral processes involved in the immune destruction of a metazoan parasite. The model also lends itself to studies of the immunologic interrelationships between innate and acquired resistance to infection with schistosomes, as well as the mechanisms by which these parasites evade the host immune response.     

Transfer of immunity to cryptococcosis by T-enriched splenic lymphocytes from Cryptococcus neoformans-sensitized mice.

Splenic enriched T-cells and sera were obtained from inbred CBA/J mice injected 7 or 35 days earlier with either 10(3) viable Cryptococcus neoformans or sterile physiological saline. The transfer of enriched T-cells collected 7 days after immunization or of normal enriched T-cells did not transfer immunity to C. neoformans or delayed-type hypersensitivity responsiveness to cryptococcal culture filtrate (CneF) antigen to the recipients. However, enriched T-cells harvested 35 days after immunization, when transferred to recipient mice, were able to confer immunity as indicated by the reduction in numbers of C. neoformans cells in the tissues, and they also transferred delayed-type hypersensitivity responsiveness to CneF antigens. Sera from either sensitized or normal mice were unable to transfer immunity to recipient animals. These results suggested that there was a time requirement for development of the immune response in the donor mice and that T-cells were crucial in the host defense against a cryptococcal infection. Culturing of day-35 C. neoformans-sensitized T-cells in the presence of homologous antigen (CneF) but not in the presence of heterologous antigen (purified protein derivative or 2, 4-dinitro-1-fluorobenzene) induced the production of migration inhibition factor, thus indicating that lymphocytes from C. neoformans-injected mice were specifically sensitized to CneF antigen.     

Sister chromatid exchange analysis in lung and peripheral blood lymphocytes of mice exposed to methyl isocyanate by inhalation

Mice were exposed to 1, 3, or 6 ppm methyl isocyanate (MIC) for 6 hr/day for four consecutive days. Lung cells and peripheral blood lymphocytes (PBLs) were removed and cultured for analysis of sister chromatid exchange (SCE) and cell cycle kinetics. MIC caused a small but significant increase in SCE frequency of cultured lung cells from mice exposed to 1, 3, or 6 ppm MIC. MIC did not significantly increase SCE levels in PBLs of mice exposed to concentrations as high as 6 ppm. In cultured PBLs, MIC had a stimulatory effect on cell cycling rates as measured by the replicative index, and it caused a significant reduction in mononuclear leucocyte counts and the mitotic indices.     

Persistence of SCE-inducing lesions in lymphocytes of mice exposed to diaziquone

Male C57B1/6 mice were injected i.p. with either 1.25 or 5.0 mg/kg diaziquone (AZQ) and killed at various time intervals from 1 to 99 days post treatment for examination of sister chromatid exchange (SCE) persistence in the peripheral blood lymphocytes (PBLs) and splenocytes. SCE frequencies were found to decay steeply during the first week after exposure in both PBLs and splenocytes. This pattern was followed by a slower decline to baseline over the next week. However, high-frequency cell (HFC) analysis indicates that significant numbers of HFCs persist in the PBLs through day 28 and splenocytes at day 99 post exposure. Mathematical modeling of the time-response curves indicates that the average life span of the majority of AZQ-induced SCE-producing lesions in murine PBLs and splenocytes responsive to phytohemagglutinin is between 3 and 5 days.     

The role of antibody affinity and titre in immunity to Schistosoma mansoni following vaccination with highly irradiated cercariae.

Sera from rabbits and rats vaccinated with highly irradiated cercariae of Schistosoma mansoni (VRabS, VRatS) were found to be of substantially higher affinity than sera from CBA mice vaccinated four times (4 X CVMS), single sex sera (SSS) or chronic infection sera (CIS). In contrast, VRabS and SSS appeared to possess the highest titres of antibody, followed by CIS and VRatS, with 4 X CVMS displaying the lowest titre. Two mouse strains selectively bred for high-affinity (HA) or low-affinity (LA) antibody following vaccination were tested for their ability to resist a challenge infection. LA mice, which produce high titres of low-affinity antibody, manifested significantly more resistance than HA mice, which produce low titres of high-affinity antibody. Immunoprecipitation studies demonstrated that sera from vaccinated LA mice (LVMS) recognized 125I-labelled schistosomular surface antigens more intensely than sera from vaccinated HA mice (HVMS). However, peritoneal macrophages from HA and LA mice in the presence of HVMS, LVMS or 4 X CVMS, and naive macrophages activated in vitro with interferon-gamma (IFN-gamma)/lipopolysaccharide (LPS) mediated comparable levels of schistosomula killing in vitro. The experiments described here provide evidence that the titre of antibody rather than its affinity may be a more critical factor in the development of optimal immunity to S. mansoni.     

Suppression of lymphoproliferation by hapten-specific suppressor T lymphocytes from mice exposed to ultraviolet radiation.

Application of a contact-sensitizing agent to the skin of mice previously exposed to UV radiation at a different site results in the induction of hapten-specific suppressor T lymphocytes. When splenic lymphocytes from such mice were cultured with normal lymphocytes and hapten-conjugated splenic adherent cells, the primary proliferative response was suppressed. The cell responsible for the suppression in vitro was a T lymphocyte, and two signals were required for its induction, ultraviolet radiation and hapten sensitization. The T cell suppressing lymphoproliferation was specific for the hapten applied after UV radiation. The UV-induced T suppressor cell inhibited only primary lymphoproliferation; the response of lymphocytes from immunized mice was unaffected. The activity of the UV-induced suppressor cell was not affected by mitomycin C treatment. Thus, suppression of the primary proliferative response of lymphocytes to hapten-modified syngeneic cells in vitro correlates with in vivo suppression of contact hypersensitivity by these UV-induced suppressor cells. This suggests that the suppressor cells act by preventing the proliferation of hapten-specific responder clones. Use of this in vitro assay system should facilitate investigation of the characteristics of these cells and the mechanism by which these regulatory T lymphocytes inhibit contact sensitization.     

Impaired lung homeostasis in neonatal mice exposed to cigarette smoke

In infants, smoke exposure is associated with more respiratory illnesses and decreased lung function. We hypothesized that perinatal lung is particularly susceptible to the damaging effects of cigarette smoke (CS) and that exposure to CS during this period may alter expression of immune response genes and adversely affect lung growth. To test this, we exposed neonatal mice to 14 days of CS. Immediately after exposure to CS, pulmonary gene expression profiling was performed on 2-week-old CS-exposed lung and age-matched control lung. Nitrotyrosine, TUNEL, MAC3, and phospho-SMAD-2 (p-SMAD2) staining was also performed. At 8 weeks of age, lung volume measurements were determined and mean linear intercept measurements were calculated. Pulmonary gene expression profiling revealed that CS exposure significantly inhibited type 1 and type 2 interferon pathway genes in neonatal lung, compared with age-matched control lung. Neonatal CS-exposed lung also had a significant increase in n-tyrosine, TUNEL, and p-SMAD2 staining when compared with adult CS-exposed lung and age-matched control lung. Lung volumes at 8 weeks of age were modestly but significantly decreased in mice exposed to CS in the neonatal period compared with age-matched controls, consistent with impaired lung growth. The results of this study indicate that exposure to CS during the neonatal period inhibits expression of genes involved in innate immunity and mildly impairs postnatal lung growth. These findings may in part explain the increased incidence of respiratory symptoms in infants and children exposed to CS.     

Trichostatin-A enhances adaptive immune responses to DNA vaccination.

BACKGROUND: The efficacy of DNA vaccines to date has overall been disappointing, especially in large animal models and in humans, which might be partially explained by the chromatinization of the administered foreign DNA. Trichostatin-A (TSA) is a specific inhibitor for histone deacetylase (HDAC) and a gene expression enhancer in in vitro DNA viral infection, including herpes simplex type 1 (HSV-1) and cytomegalovirus (CMV). OBJECTIVES: We sought to determine if increasing antigen expression of DNA vaccines through inhibition of HDAC-induced chromatin silencing using TSA would enhance DNA vaccine efficacy in vivo. STUDY DESIGN: A luciferase assay was used to detect effects of TSA on different promoters in vitro. The effects of TSA on DNA vaccination of mice were determined by neutralization assays for antibody production and interleukin staining for detection of specific T cell responses. RESULTS: Mice receiving TSA in combination with a DNA vaccine exhibited higher antibody responses to the vaccine than mice not given TSA. Co-administration of TSA also enhanced specific CD8 T cells response. CONCLUSION: Drugs such as TSA that reduce initial gene silencing by preventing histone deacetylation can increase immune responses to DNA vaccination.     

Severe neutrophil-mediated lung inflammation in myeloperoxidase-deficient mice exposed to zymosan

Objective and design  

This study examines the role of myeloperoxidase (MPO), a major constituent of neutrophils that generates hypochlorous acid, in neutrophil recruitment into the zymosan-exposed lung of mice.     

Evans blue dye adjuvant enhances delayed hypersensitivity while blocking immunity to Mycobacterium tuberculosis in mice.

Evans blue dye functions as an adjuvant with protein antigens in saline to induce cell-mediated immunological responses in mice. But when used to help induce cell-mediated tuberculoimmunity, it decreased mouse resistance to tuberculosis instead of helping induce immunity. This paradox was investigated. As could be expected from previous work with other antigens, the dye did promote induction of delayed hypersensitivity in mice to tuberculoprotein when injected in saline with killed tubercle bacilli. Peritoneal macrophages from mice injected with the dye responded normally to migration inhibition factor. Morphologically, these cells were moderately "activated" compared with similar cells taken from untreated mice. However, such cells incubated with tuberculosis growth inhibition lymphokine in an in vitro test for tuberculoimmunity did not express tuberculoimmunity, whereas macrophages from untreated mice did. Therefore, Evans blue dye did promote induction of cell-mediated immunological responses and tuberculoimmunity in lymphocytes, but under the conditions used in these experiments, it also blocked expression of tuberculoimmunity by macrophages.     

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.     

Gamma-irradiated scrub typhus immunogens: development of cell-mediated immunity after vaccination of inbred mice.

Mice immunized with three injections of gamma-irradiated Karp strain of Rickettsia tsutsugamushi were evaluated for the presence of cell-mediated immunity by using delayed-type hypersensitivity, antigen-induced lymphocyte proliferation, and antigen-induced lymphokine production. These animals also were evaluated for levels of circulating antibody after immunization as well as for the presence of rickettsemia after intraperitoneal challenge with viable Karp rickettsiae. After immunization with irradiated Karp rickettsiae, a demonstrable cell-mediated immunity was present as evidenced by delayed-type hypersensitivity responsiveness, lymphocyte proliferation, and production of migration inhibition factor and interferon by immune spleen lymphocytes. Also, a reduction in circulating rickettsiae was seen in mice immunized with irradiated rickettsiae after challenge with 1,000 50% mouse lethal doses of viable, homologous rickettsiae. All responses except antibody titer and reduction of rickettsemia were similar to the responses noted in mice immunized with viable organisms. Antibody levels were lower in mice immunized with irradiated rickettsiae than in mice immunized with viable rickettsiae. Furthermore, mice that were immunized with viable rickettsiae demonstrated markedly lower levels of rickettsemia after intraperitoneal challenge compared with either mice immunized with irradiated rickettsiae or nonimmunized mice.     

DNA vaccination against tuberculosis: expression of a ubiquitin-conjugated tuberculosis protein enhances antimycobacterial immunity

Genetic immunization is a promising new technology for developing vaccines against tuberculosis that are more effective. In the present study, we evaluated the effects of intracellular turnover of antigens expressed by DNA vaccines on the immune response induced by these vaccines in a mouse model of pulmonary tuberculosis. The mycobacterial culture filtrate protein MPT64 was expressed as a chimeric protein fused to one of three variants of the ubiquitin protein (UbG, UbA, and UbGR) known to differentially affect the intracellular processing of the coexpressed antigens. Immunoblot analysis of cell lysates of in vitro-transfected cells showed substantial differences in the degradation rate of ubiquinated MPT64 (i.e., UbG64 < UbA64 < UbGR64). The specific immune response generated in mice correlated with the stability of the ubiquitin-conjugated antigen. The UbA64 DNA vaccine induced a weak humoral response compared to UbG64, and a mixed population of interleukin-4 (IL-4)- and gamma interferon (IFN-gamma)-secreting cells. Vaccination with the UbGR64 plasmid generated a strong Th1 cell response (high IFN-gamma, low IL-4) in the absence of a detectable humoral response. Aerogenic challenge of vaccinated mice with Mycobacterium tuberculosis indicated that immunization with both the UbA64- and UbGR64-expressing plasmids evoked an enhanced protective response compared to the vector control. The expression of mycobacterial antigens from DNA vaccines as fusion proteins with a destabilizing ubiquitin molecule (UbA or UbGR) shifted the host response toward a stronger Th1-type immunity which was characterized by low specific antibody levels, high numbers of IFN-gamma-secreting cells, and significant resistance to a tuberculous challenge.     

Cytolytic T lymphocytes and antibodies to myocytes in adriamycin-treated BALB/c mice. Evidence for immunity to drug-induced antigens.

Female BALB/c mice were given a single intravenous injection of between 0.1 and 10 mg adriamycin/kg body weight and were killed between 2 and 16 days later. Natural killer (NK) cell activity in the spleen was measured using YAC cell targets. Natural killer cell activity was slightly elevated 2 to 5 days after drug injection and significantly depressed by day 9 compared with spleen cells from untreated animals. Adriamycin-treated mice developed both cytolytic T lymphocytes (CTL) and antibodies to drug-treated myocytes. Peak CTL response occurred between days 9 and 13, whereas antibody reactivity continued to increase throughout the observation period. The effector cell belonged to the CD8+ T lymphocyte subpopulation, because cytolytic activity could be reduced by treating the cells with anti-Lyt 2 antibody and complement, whereas anti-L3T4 (CD4+ cell-specific) treatment either had no effect or increased cytotoxicity. Both CTL and antibody reactivity could be absorbed with adriamycin-treated myocyte monolayers but not by non-drug-treated myocytes. Furthermore CTL reactivity could be only partly removed by adriamycin-treated skin fibroblasts. Adriamycin concentrations in the heart were measured by flourometry and demonstrated only a gradual decrease in the drug over the 16-day period. Immunofluorescent staining of myocardial sections demonstrated increased numbers of both T lymphocytes and macrophages in the hearts of adriamycin-treated mice compared with untreated controls.     

Thymus-dependent lymphocytes of dengue virus-infected mice spleens mediate suppression through prostaglandin.

Our earlier observations indicate that adoptive transfer of spleen cells obtained from dengue type 2 virus (DV)-primed mice suppressed DV antigen-specific antibody secretion as detected by Jerne PFC technique. Findings of this paper indicate that the suppression was produced by non-glass-adherent cells, macrophage-depleted (by carbonyl iron) cells and by T lymphocytes of the spleen but not by the glass-adherent cells and B lymphocytes. The activity of these cells is dependent upon production of prostaglandin as shown by abrogation of their suppressor activity by pre-treatment of cells by indomethacin or aspirin which are known to block synthesis of prostaglandins.     

Feeding enhances fibronectin adherence of quiescent lymphocytes through non-canonical insulin signalling

Nutritional availability during fasting and refeeding affects the temporal redistribution of lymphoid and myeloid immune cells among the circulating and tissue-resident pools. Conversely, nutritional imbalance and impaired glucose metabolism are associated with chronic inflammation, aberrant immunity and anomalous leukocyte trafficking. Despite being exposed to periodic alterations in blood insulin levels upon fasting and feeding, studies exploring the physiological effects of these hormonal changes on quiescent immune cell function and trafficking are scanty. Here, we report that oral glucose load in mice and healthy men enhances the adherence of circulating peripheral blood mononuclear cells (PBMCs) and lymphocytes to fibronectin. Adherence to fibronectin is also observed upon regular intake of breakfast following overnight fasting in healthy subjects. This glucose load-induced phenomenon is abrogated in streptozotocin-injected mice that lack insulin. Intra-vital microscopy in mice demonstrated that oral glucose feeding enhances the homing of PBMCs to injured blood vessels in vivo. Furthermore, employing flow cytometry, Western blotting and adhesion assays for PBMCs and Jurkat-T cells, we elucidate that insulin enhances fibronectin adherence of quiescent lymphocytes through non-canonical signalling involving insulin-like growth factor-1 receptor (IGF-1R) autophosphorylation, phospholipase C gamma-1 (PLCγ-1) Tyr⁷⁸³ phosphorylation and inside-out activation of β-integrins respectively. Our findings uncover the physiological relevance of post-prandial insulin spikes in regulating the adherence and trafficking of circulating quiescent T-cells through fibronectin–integrin interaction.     

Herd immunity to Newcastle disease virus in poultry by vaccination.

Newcastle disease is an economically important disease of poultry for which vaccination is applied as a preventive measure in many countries. Nevertheless, outbreaks have been reported in vaccinated populations. This suggests that either the vaccination coverage level is too low or that vaccination does not provide perfect immunity, allowing the virus to spread in partially vaccinated populations. Here we study the requirements of an epidemiologically effective vaccination program against Newcastle disease in poultry, based on data from experimental transmission studies. The transmission studies indicate that vaccinated birds with low or undetectable antibody titres may be protected against disease and mortality but that infection and transmission may still occur. In fact, our quantitative analyses show that Newcastle disease virus is highly transmissible in poultry with low antibody titres. As a consequence, herd immunity can only be achieved if a high proportion of birds (>85%) have a high antibody titre (log(2) haemagglutination inhibition titre > or =3) after vaccination. We discuss the implications for the control of Newcastle disease in poultry by vaccination.