Trypanosoma cruzi: peripheral blood monocytes and heart macrophages in the resistance to acute experimental infection in rats

The contribution of mononuclear phagocytes to host resistance against acute Chagas disease has been studied in vivo in a rat model inoculated with Trypanosoma cruzi, Y strain. Acute T. cruzi infection triggered a dramatic increase (937%) in peripheral blood monocyte number at day 12 of infection. At this point, histological analysis of the heart showed high parasitism and diffuse and a moderate to intense mononuclear inflammatory process. Ultrastructural study revealed a large number of macrophages, in addition to lymphocytes and undiffer entiated cells. Clusters of macrophages exhibited different morphologi cal phenotypes, with evident signs of activation (increase in size, surface rufflings, and amount of cytoplasmic organelles). Cell-to-cell contacts involving macrophages and lymphocytes or macrophages and mono cytes were observed. Depletion of macrophages by treatment with silica, a selective cytotoxic agent for these cells, caused a significant increase in the number of amastigote nests in cardiomyocytes. The present findings indicate that the early phase of infection with T. cruzi induces rapid production, maturation, and activation of the monocyte/macrophage system so as to control T. cruzi replication, emphasizing the crucial role for macrophages in the rat resistance to Chagas disease.     

Production of hydrogen peroxide by peripheral blood monocytes and specific macrophages during experimental infection with Trypanosoma cruzi in vivo

Acute Chagas' disease triggers potent inflammatory reaction characterized by great increase of peripheral blood monocyte (PBM) and macrophage numbers. We studied the respiratory burst responses of PBM and peritoneal and splenic macrophages to in vivo infection (rats). The ultrastructure of heart inflammatory macrophages was also investigated. The infection increased the hydrogen peroxide (H2O2) production by PBM and splenic macrophages but not by peritoneal macrophages. Accordingly, the PBM and spleen cell numbers increased but the total number of peritoneal cells was similar to controls. Heart macrophages of infected rats exhibited increase (number and size) and activated morphology in parallel to high cardiomyocyte parasitism. Our data highlight the importance of innate immunity and H2O2production to host resistance during acute phase of T. cruzi infection. A novel finding is that H2O2production seems related to specific types of monocytes/macrophages that are able to release this agent when in presence of high parasite load.     

Roles of macrophages in measles virus infection of genetically modified mice

Knowledge of the mechanisms of virus dissemination in acute measles is cursory, but cells of the monocyte/macrophage (MM) lineage appear to be early targets. We characterized the dissemination of the Edmonston B vaccine strain of measles virus (MV-Ed) in peripheral blood mononuclear cells (PBMC) of two mouse strains expressing the human MV-Ed receptor CD46 with human-like tissue specificity and efficiency. In one strain the alpha/beta interferon receptor is defective, allowing for efficient MV-Ed systemic spread. In both mouse strains the PBMC most efficiently infected were F4/80-positive MMs, regardless of the inoculation route used. Circulating B lymphocytes and CD4-positive T lymphocytes were infected at lower levels, but no infected CD8-positive T lymphocytes were detected. To elucidate the roles of MMs in infection, we depleted these cells by clodronate liposome treatment in vivo. MV-Ed infection of splenic MM-depleted mice caused strong activation and infection of splenic dendritic cells (DC), followed by enhanced virus replication in the spleen. Similarly, depletion of lung macrophages resulted in strong activation and infection of lung DC. Thus, in MV infections of genetically modified mice, blood monocytes and tissue macrophages provide functions beneficial for both the virus and the host: they support virus replication early after infection, but they also contribute to protecting other immune cells from infection. Human MM may have similar roles in acute measles.     

Primary stage of feline immunodeficiency virus infection: viral dissemination and cellular targets.

The objective of this study was to identify cellular and organ targets of acute feline immunodeficiency virus (FIV) infection in vivo. Tissues of FIV-infected cats were studied at eight time points during the first 3 months after experimental infection. FIV nucleic acids were first detected by in situ hybridization 21 days after infection, approximately 1.5 weeks after lymph node enlargement was first observed and 3 weeks before the primary acute flu-like illness. The majority of FIV-infected cells were present in lymphoid organs, though low numbers of infected cells were noted in nonlymphoid organs as well. Germinal centers harbored many of the FIV-infected cells within lymphoid tissues. The thymic cortex was also a major site of early infection. Combined in situ hybridization and immunohistochemistry revealed that T lymphocytes were the primary target of early FIV infection in tissues of cats before the onset of clinical signs of acute illness. An unidentified population of mononuclear cells and a few macrophages were also infected. During the ensuing acute flu-like illness, the proportion of FIV-infected macrophages in tissues increased dramatically. This early shift in the predominant cellular localization of FIV from T lymphocytes to macrophages may be important for establishing viral persistence.     

Ischaemia alters the effects of cardiomyocyte‐derived extracellular vesicles on macrophage activation

Myocardial ischaemia is associated with an exacerbated inflammatory response, as well as with a deregulation of intercellular communication systems. Macrophages have been implicated in the maintenance of heart homeostasis and in the progression and resolution of the ischaemic injury. Nevertheless, the mechanisms underlying the crosstalk between cardiomyocytes and macrophages remain largely underexplored. Extracellular vesicles (EVs) have emerged as key players of cell‐cell communication in cardiac health and disease. Hence, the main objective of this study was to characterize the impact of cardiomyocyte‐derived EVs upon macrophage activation. Results obtained demonstrate that EVs released by H9c2 cells induced a pro‐inflammatory profile in macrophages, via p38MAPK activation and increased expression of iNOS, IL‐1β and IL‐6, being these effects less pronounced with ischaemic EVs. EVs derived from neonatal cardiomyocytes, maintained either in control or ischaemia, induced a similar pattern of p38MAPK activation, expression of iNOS, IL‐1β, IL‐6, IL‐10 and TNFα. Importantly, adhesion of macrophages to fibronectin was enhanced by EVs released by cardiomyocytes under ischaemia, whereas phagocytic capacity and adhesion to cardiomyocytes were higher in macrophages incubated with control EVs. Additionally, serum‐circulating EVs isolated from human controls or acute myocardial infarction patients induce macrophage activation. According to our model, in basal conditions, cardiomyocyte‐derived EVs maintain a macrophage profile that ensure heart homeostasis, whereas during ischaemia, this crosstalk is affected, likely impacting healing and post‐infarction remodelling.     

Trypanosoma evansi: immune response and acetylcholinesterase activity in lymphocytes from infected rats

The existence of cholinergic receptors in the immune system cells is well documented. This study aimed to evaluate the acetylcholinesterase activity (AChE) in lymphocytes from rats infected with Trypanosoma evansi in acute and chronic phase disease. Twenty animals were infected with 10⁶ trypomastigotes forms each and 10 were used as negative controls. The two groups of inoculated rats were formed according to the degree of parasitemia and the period post-infection (PI). Group A: rats with 4 days PI and between 24 and 45 parasites/field (1000×); group B: rats with 30 days PI and parasitemia with jagged peaks between 0 and 1 parasites/field; group C: not-infected animals. At 4 days PI (acute phase) and 30 days PI (chronic phase) the rats were anesthetized to collect blood for hemogram and separation of lymphocytes. After separation, the AChE activity was measured in lymphocytes. It was observed that the number of lymphocytes increased significantly in group A compared to group C. The activity of AChE in lymphocytes significantly increased in acute phase and decreased in chronic phase in the infected rats when compared to not-infected (P < 0.05). Statistical analysis showed a positive correlation between the number of lymphocytes and AChE activity in lymphocytes in 4 days PI (r²: 0.59). Therefore, the infection by T. evansi influences AChE activity in lymphocytes of rats indicating changes in the responses of cholinergic system in acute phase, possibly due to immune functions performed by these enzymes.     

HSP and apoptosis in leukocytes from infected or vaccinated animals by Brucella abortus

The production of hsp and apoptosis of leukocytes in the peripheral blood of animals naturally infected with Brucella spp or treated with the vaccine Brucella abortus 19 have been investigated in this study. Cytokines able to induce phagocytic activity in macrophages of non treated healthy animals were found in the supernatant of bovine leukocytes cultivated in vitro. A long-lasting antibody response against hsp 60 kDa and 27 kDa, which lasts a long time, is induced in naturally infected animals, while in animals vaccinated with B. abortus 19 we detected an antibody response against hsp 60 and 70 kDa which is much shorter, disappearing in two months. During the early phase of infection, lymphocytes and monocytes of naturally infected animals show a delay of apoptosis in vitro compared to the same cells coming from healthy controls and vaccinated animals.     

The mediator of cellular immunity. X. Interaction of macrophages and specifically sensitized lymphocytes.

Specifically sensitized lymphocytes have a focusing influence on mononuclear phagocytes that is expressed in the local accumulation and division of macrophages in bacteria-induced exudates. This was demonstrated by injecting Listeria monocytogenes into the peritoneal cavity of normal rats immediately before the animals were transfused with thoracic duct lymphocytes from either Listeria-immune donors or donors that had been infected with the unrelated parasite, Francisella tularensis. Sensitized lymphocytes originally present in the intravenous inocula were found later in the exudates. The arrival in the inflamed peritoneal cavity of specifically sensitized lymphocytes was associated with an exuberant influx of newly formed host cells and a local proliferative response that involved both immunoblasts and macrophages. These cytokinetic observations provide a plausible explanation of the delayed inflammatory response induced by the parasite, and imply that sensitized lymphocytes contribute to the host's defence by encouraging the prompt and purposeful deployment of monocyte-derived macrophages in centers of infection. In addition to their focusing influence on mononuclear phagocytes, specifically sensitized lymphocytes or their products can enhance the metabolic and microbicidal activity of macrophages. This activation process was revealed in the ability of rats infected with L. monocytogenes or BCG to control the growth of F. tularensis at a challenge site in the testis. But resistance was expressed only when the challenge organisms were injected with killed bacteria against which the recipients had been specifically immunized. The results accord with the view that macrophages are functionally activated by an immunological mechanism, and imply that the process is triggered locally by sensitized lymphocytes which are recruited from the blood.     

Enhanced survival by vitamin A supplementation during a retrovirus infection causing murine AIDS

Infection by LP-BM5 murine leukemia virus (MuLV) produces an AIDS-like condition in mice. The viral infection suppressed the percentage of peripheral blood cells showing surface markers for macrophages, activated macrophages, T lymphocytes and activated lymphoid cells. High dietary vitamin A (retinyl palmitate) caused increased numbers of activated macrophages. It also increased the percentage of cells with markers for Ia+ cells and macrophages in the retrovirally infected mice compared to infected controls. In uninfected mice retinyl palmitate stimulated the percentage of cells with activated lymphocytes bearing IL-2R, and T cytotoxic cells. These were associated with a retarded death rate during infection with LP-BM5 murine leukemia in C57BL/6 mice. By 25 weeks of infection and 20 weeks of retinyl palmitate supplementation 71.3% survived, while 45.0% virally infected controls survived. The mice also had elevated numbers of B cells measured in the blood after 4 and 8 weeks of dietary treatment. Vitamin A stimulation may play a role in the slower death rate for retrovirally infected mice.     

In vivo studies of the early, peritoneal, cellular and free radical response in rats infected with Fasciola hepatica by flow cytometric analysis

Early recruitment of the peritoneal cell population was observed during migration of newly excysted juvenile flukes. The peritoneal lavages were examined for T cells, cytotoxic NK cells (CNK) and free radicals production of rats at an early stage of infection by Fasciola hepatica. Male Sprague–Dawley rats were infected with 50 metacercariae of F. hepatica and non-infected controls were euthanized 2, 4 and 7 days post infection (d.p.i.), respectively. The peritoneal fluid of experimental animals was analyzed by flow cytometry to estimate cell phenotypes. The peritoneal areas were infiltrated by inflammatory cells, particularly from numerous neutrophils, eosinophils and CD4+ lymphocytes, which were significantly higher for infected rats than non-infected. CNK cells dominated in the peritoneal fluid of infected rats as early as 2 d.p.i. However, after 4 d.p.i. there was a decreased level of CNK cells which may indicate a change from a cytotoxic natural killer (NK) to a regulatory NK response. The challenged group generated very high in vivo levels of inducible nitric oxide (NO) from eosinophils. Superoxide expression was very high in macrophages and neutrophils compared to the uninfected control. In conclusion, our studies suggest that early F. hepatica infection could directly affect lymphoid cells and generate a high in vivo NO production by eosinophils in the peritoneal cavity. Moreover juvenile flukes could stimulate the macrophages and neutrophils to generate H₂O₂ radicals. The host parasite interactions resulting from immune response regulation by effector cells and immune evasion are discussed.     

CD4 Depletion in SIV-Infected Macaques Results in Macrophage and Microglia Infection with Rapid Turnover of Infected Cells

In rhesus macaques (RMs), experimental depletion of CD4⁺ T-cells prior to SIV infection results in higher viremia and emergence of CD4-independent SIV-envelopes. In this study we used the rhesus recombinant anti-CD4 antibody CD4R1 to deplete RM CD4⁺ T-cells prior to SIVmac₂₅₁ infection and investigate the sources of the increased viral burden and the lifespan of productively infected cells. CD4-depleted animals showed (i) set-point viral load two-logs higher than controls; (ii) macrophages constituting 80% of all SIV vRNA⁺ cells in lymph node and mucosal tissues; (iii) substantial expansion of pro-inflammatory monocytes; (iv) aberrant activation and infection of microglial cells; and (v) lifespan of productively infected cells significantly longer in comparison to controls, but markedly shorter than previously estimated for macrophages. The net effect of CD4⁺ T-cell depletion is an inability to control SIV replication and a shift in the tropism of infected cells to macrophages, microglia, and, potentially, other CD4-low cells which all appear to have a shortened in vivo lifespan. We believe these findings have important implications for HIV eradication studies.     

Changes in lymphocyte and macrophage subsets due to morphine and ethanol treatment during a retrovirus infection causing murine AIDS

Infection by LP-BM5 murine leukemia virus (MuLV) suppressed significantly the percentage of peripheral blood cells showing surface markers for macrophages, lymphocytes and activated lymphoid cells. Chronic administration of a 7% (36% calories) ethanol diet or injection of 1.9 mg/mouse/day of morphine for a 7 day period were followed by 3 week periods of abstinence and then 1 week periods of consumption of 5% ethanol diets or morphine injection to female C57BL/6 mice resulted in changes in the numbers of macrophages and lymphocyte subsets. The number of lymphocytes of various subsets were not significantly changed by the ethanol exposure except those showing activation markers which were reduced. The percentage of peripheral blood cells showing markers for macrophage functions and their activation were significantly reduced after "binge" use of ethanol. Ethanol retarded suppression of cells by retroviral infection. However by 25 weeks of infection there was a 8.6% survival in the ethanol fed mice infected with retrovirus which was much less than virally infected controls (45.0%). Morphine treatment also increased the percentage of cells with markers for macrophages and activated macrophages in virally infected mice, while suppressing them in uninfected mice. The second and third morphine injection series suppressed lymphocyte T-helper and T-suppressor cells, but not total T cells. However, suppression by morphine was significantly less during retroviral disease than suppression caused by the virus only. At 25 weeks of infection 44.8% of morphine treated, infected mice survived. Morphine treatment also caused deaths such that the survival in morphine treated, retrovirally infected was higher than would have been expected if the death rate in virally infected, and morphine injected animals were combined during combined treatment. Thus these drugs of abuse can modulate peripheral blood lymphoid subsets, suppression caused by retroviral infection, and survival.     

Spontaneous regression of Friend virus-induced erythroleukemia. III. The role of macrophages in regression.

The spontaneous regression of erythroleukemia induced by the RFV strain of Friend virus is a macrophage-dependent process. Functional suppression or elimination of the macrophage population in leukemic mice with silica, carrageenan, anti-macrophage serum, or trypan blue inhibited regression. Prior protection of the macrophages with PVNO allowed regression in silica or carrageenan-treated mice. Macrophage phagocytic activity was inhibited in about half the RFV-induced leukemic mice at 25 to 30 days post virus inoculation. Those animals with normal macrophages regressed, whereas whereas those with inhibited macrophages did not. Progressor mice could be induced to regress by inoculation with normal syngeneic macrophages; other cell types were ineffective. The inhibition of macrophage function in leukemic mice was the result of infection of the macrophages by virus. Removal of the infected cells by cytolysis with anti-virus antiserum and C restored the phagocytic activity of the population. Inhibited macrophages were less capable of responding to immobilized antigen-antibody complexes than normal macrophages, suggesting that the loss of function was due to a change in their Fc receptor.     

A naturally occurring epizootic caused by Sendai virus in breeding and aging rodent colonies. II. Infection in the rat.

Sendai virus infected a hysterectomy derived, barrier maintained breeding colony and a conventional aging rat colony. The virus produced seroconversion in the colonies followed by a 7-month period of decreasing titers. Clinical signs were absent during the months when titers were highest, and there was no increase in mortality, but multifocal interstitial pneumonia with perivascular and peribronchial cuffing by lymphocytes and plasma cells was present in rat lungs examined histologically. Such lesions were absent before the period of seroconversion. During the months of declining titers, the interstitial and perivascular lesions decreased in frequency and severity. The peribronchial lesions did not decrease, however, and were still present in many rats 7 months after the acute infection. Attempts to isolate the virus from weanling rats were unsuccessful.     

巨细胞病毒感染对新生豚鼠胸腺的影响

新生豚鼠皮下接种豚鼠巨细胞病毒(GPCMV)后,导致动物胸腺急性感染。感染豚鼠胸腺在接种后第五天开始出现病毒,第十天达高峰。此外,感染动物胸腺的发育受到抑制,细胞总数和T淋巴细胞数随朐腺中病毒滴度的增高而进行性下降,至接种GPCMV后第十天最显著。由于病毒对T细胞的作用,细胞表面红细胞受体的丧失导致胸腺Null细胞百分比高于对照动物。     

急性巨细胞病毒感染导致新生豚鼠脾细胞亚群的定量改变

新生豚鼠皮下接种豚鼠巨细胞病毒(GPCMV)后,导致脾脏GPCMV急性感染,脾脏肿大,脾细胞增生,计算表明,脾T淋巴细胞,B淋巴细胞、吞噬细胞数显著高于正常动物,接种病毒后第5天即可在脾细胞检出高滴度感染性病毒(达2.5 log10 TCID50/10~7细胞),其中,主要是脾B淋巴细胞、T淋巴细胞和巨噬细胞携带病毒。     

Trypanosoma cruzi-cardiomyocyte interaction: role of fibronectin in the recognition process and extracellular matrix expression in vitro and in vivo

We investigated the involvement of fibronectin (FN) in Trypanosoma cruzi-cardiomyocyte invasion and the extracellular matrix (ECM) components expression during T. cruzi infection in vivo and in vitro. Treatment of trypomastigotes with FN or a synthetic peptide (MRGDS) prior to cardiomyocyte interaction reduced T. cruzi infection, indicating that FN mediates the parasite invasion through its RGD sequence. In murine experimental Chagas' disease, an enhancement of the ECM components was detected in the myocardium during the late acute infection, coinciding with inflammatory infiltrates accumulation. In contrast, highly infected cardiomyocytes displayed a reduction in FN expression in vitro, while laminin spatial distribution was altered. Although it has been demonstrated that cardiomyocytes are able to synthesize cytokines upon T. cruzi infection, our data suggest that matrix remodeling is dependent on cytokines secreted by inflammatory cells recruited in immune response.     

Immunoregulation in experimental schistosomiasis: III. Role of macrophages in soluble egg antigen-induced chronic spleen cell augmentation of baseline lymphocyte reactivity

Spleen cells from mice infected for 20 weeks with Schistosoma mansoni, exposed in vitro to soluble schistosomal egg antigens (SEA), treated with mitomycin C (Mc), and cocultured with syngeneic responder spleen cells increased the baseline proliferation of the otherwise unstimulated responder cells in cocultures. The role of macrophages in this “spontaneous” thymidine incorporation was studied directly by removal of macrophages on Sephadex G-10 columns. Removal of esterase-positive, Sephadex G-10-adherent cells (macrophages) greatly reduced the amount of SEA-induced, chronically infected spleen cell-mediated stimulation observed in cocultures. It also reduced an elevated background of spontaneous DNA synthesis seen with control cultures of spleen cells from infected animals. Depletion of T lymphocytes from chronic spleen cell populations by treatment with anti-Thy 1.2 serum and complement prior to exposure to SEA partially abrogated the augmentation effect. Comparison of these results with mitogen (concanavalin A)induced spleen cell-mediated stimulation (which is elevated, rather than reduced, by macrophage removal) and with known alterations in splenic T- and B-lymphocyte ratios in chronic murine Schistosomiasis suggests that antigen-stimulated, chronically infected splenic macrophage-de-pendent baseline augmentation may depend on specific T-lymphocyte-derived lymphokine induction. These results may reflect a general mechanism whereby animals harboring a persistent, chronic infection can respond quickly to a second or challenge infection or a flareup of the primary infection.     

CD8+ Lymphocytes Control Viral Replication in SIVmac239-Infected Rhesus Macaques without Decreasing the Lifespan of Productively Infected Cells

While CD8+ T cells are clearly important in controlling virus replication during HIV and SIV infections, the mechanisms underlying this antiviral effect remain poorly understood. In this study, we assessed the in vivo effect of CD8+ lymphocyte depletion on the lifespan of productively infected cells during chronic SIVmac239 infection of rhesus macaques. We treated two groups of animals that were either CD8+ lymphocyte-depleted or controls with antiretroviral therapy, and used mathematical modeling to assess the lifespan of infected cells either in the presence or absence of CD8+ lymphocytes. We found that, in both early (day 57 post-SIV) and late (day 177 post-SIV) chronic SIV infection, depletion of CD8+ lymphocytes did not result in a measurable increase in the lifespan of either short- or long-lived productively infected cells in vivo. This result indicates that the presence of CD8+ lymphocytes does not result in a noticeably shorter lifespan of productively SIV-infected cells, and thus that direct cell killing is unlikely to be the main mechanism underlying the antiviral effect of CD8+ T cells in SIV-infected macaques with high virus replication.