In vivo radioprotection by 5-androstenediol: stimulation of the innate immune system

We showed previously that 5-androstenediol stimulates myelopoiesis, increases the numbers of circulating neutrophils and platelets, and enhances resistance to infection in gamma-irradiated mice. We have extended those studies to include monocytes, natural killer (NK) cells, eosinophils and basophils, and we have measured the activation marker CD11b using flow cytometry. Androstenediol (160 mg/kg) was administered subcutaneously to female B6D2F1 mice 24 h before whole-body gamma irradiation. Androstenediol treatments increased the blood levels of neutrophils, monocytes and NK cells in unirradiated animals; decreased the numbers of circulating eosinophils; and ameliorated radiation-induced decreases in neutrophils, monocytes, NK cells, erythrocytes and platelets. The androstenediol treatments had no significant effect on the numbers of circulating B cells or T cells. CD11b labeling intensity on monocytes was decreased slightly after androstenediol treatment. In contrast, radiation or androstenediol alone caused increases in CD11b labeling intensity on NK cells. Androstenediol and radiation combined caused a marked increase in NK cell CD11b. The results indicate that androstenediol increases the numbers of the three major cell types of the innate immune system (neutrophils, monocytes and NK cells), that androstenediol-induced changes in blood elements in irradiated animals persist for at least several weeks, and that there is a significant positive interaction between radiation and administration of androstenediol in the activation of NK cells.     

Chlamydial and poxvirus infections of circulating monocytes of a flap-necked chameleon (Chamaeleo dilepis)

Of blood films examined from 170 specimens of 15 Chamaeleo spp. in Tanzania, three C. dilepis had an intracytoplasmic inclusion within monocytes. One of the lizards was maintained in captivity and was sequentially bled over a 55 day period. At 46 days, a second type of inclusion was occasionally seen within monocytes. The lizard became ill and was euthanatized on day 55. All circulating monocytes were found to have either one or both types of inclusions. Histologic examination of multiple tissues demonstrated similar inclusions within macrophages in the spleen and liver. Transmission electron microscopic examination of monocytes revealed the presence of a chlamydia-like organism and pox-like virus. These pathogens have not been reported previously in chameleons, nor has a combined infection of circulating monocytes with these two pathogens been reported for any animal.     

Replication and persistence of measles virus in defined subpopulations of human leukocytes.

Replication of Edmonston strain measles virus was studied in several human lymphoblast lines, as well as in defined subpopulations of circulating human leukocytes. It was found that measles virus can productively infect T cells, B cells, and monocytes from human blood. These conclusions were derived from infectious center studies on segregated cell populations, as well as from ultrastructural analyses on cells labeled with specific markers. In contrast, mature polymorphonuclear cells failed to synthesize measles virus nucleocapsids even after infection at a relatively high multiplicity of infection. Measles virus replicated more efficiently in lymphocytes stimulated with mitogens than in unstimulated cells. However, both phytohemagglutinin and pokeweed mitogen had a negligible stimulatory effect on viral synthesis in purified populations of monocytes. In all instances the efficiency of measles virus replication by monocytes was appreciably less than that of mitogenically stimulated lymphocytes or of continuously culture lymphoblasts. Under standard conditions of infection, all of the surveyed lymphoblast lines produced equivalent amounts of measles virus regardless of the major histocompatibility (HL-A) haplotype. Hence, no evidence was found that the HL-A3,7 haplotype conferred either an advantage or disadvantage with respect to measles virus synthesis in an immunologically neutral environment. A persistent infection with measles virus could be established in both T and B lymphoblasts. The release of infectious virus from such persistently infected cells was stable over a period of several weeks and was approximately 100-fold less than peak viral titers obtained in each respective line after acute infection.     

The effect of hydrocortisone on neutrophil functional activity

The male (CBA X C57BL) FI mice received 125 mg of hydrocortisone per kg body weight intraperitoneally. The functional activity of neutrophils has been evaluated by means of nitroblue terazolium test (NBT-test) values taken before or after heat-killed S. marcescens cell stimulation in vitro by 2, 12, 24 h 3, 7 or 14 days post hormonal treatment. Throughout the 1st day after hydrocortisone administration the NBT-test values taken prior to as well as post microbial neutrophil stimulation were steadily increased. This effect could be seen as early as 2 h post hormone administration and it was linked with growing leukopenia and total decrease of blood granulocyte content. By the 3rd day the same very values turned up to become normal. The NBT-test values after microbial stimulation of neutrophils were 1.7 or 2.4 lower after hydrocortisone had been added to blood in vitro in a dose 3.5 X 10(-6) M or 7 X 10(-5) M.     

Chronic bacterial osteomyelitis suppression of tumor growth requires innate immune responses

Clinical studies over the past several years have reported that metastasis-free survival times in humans and dogs with osteosarcoma are significantly increased in patients that develop chronic bacterial osteomyelitis at their surgical site. However, the immunological mechanism by which osteomyelitis may suppress tumor growth has not been investigated. Therefore, we used a mouse model of osteomyelitis to assess the effects of bone infection on innate immunity and tumor growth. A chronic Staphylococcal osteomyelitis model was established in C3H mice and the effects of infection on tumor growth of syngeneic DLM8 osteosarcoma were assessed. The effects of infection on tumor angiogenesis and innate immunity, including NK cell and monocyte responses, were assessed. We found that osteomyelitis significantly inhibited the growth of tumors in mice, and that the effect was independent of the infecting bacterial type, tumor type, or mouse strain. Depletion of NK cells or monocytes reversed the antitumor activity elicited by infection. Moreover, infected mice had a significant increase in circulating monocytes and numbers of tumor associated macrophages. Infection suppressed tumor angiogenesis but did not affect the numbers of circulating endothelial cells. Therefore, we concluded that chronic localized bacterial infection could elicit significant systemic antitumor activity dependent on NK cells and macrophages.     

Differential Immune Response Associated to Malaria Outcome Is Detectable in Peripheral Blood following Plasmodium yoelii Infection in Mice

Malaria infection in humans elicits a wide range of immune responses that can be detected in peripheral blood, but we lack detailed long-term follow-up data on the primary and subsequent infections that lead to naturally acquired immunity. Studies on antimalarial immune responses in mice have been based on models yielding homogenous infection profiles. Here, we present a mouse model in which a heterogeneous course of Plasmodium yoelii lethal malaria infection is produced in a non-congenic ICR strain to allow comparison among different immunological and clinical outcomes. Three different disease courses were observed ranging from a fatal outcome, either early or late, to a self-resolved infection that conferred long-term immunity against re-infection. Qualitative and quantitative changes produced in leukocyte subpopulations and cytokine profiles detected in peripheral blood during the first week of infection revealed that monocytes, dendritic cells and immature B cells were the main cell subsets present in highly-parasitized mice dying in the first week after infection. Besides, CD4⁺CD25^high T cells expanded at an earlier time point in early deceased mice than in surviving mice and expressed higher levels of intracellular Foxp3 protein. In contrast, survivors showed a limited increase of cytokines release and stable circulating innate cells. From the second week of infection, mice that would die or survive showed similar immune profiles, although CD4⁺CD25^high T cells number increased earlier in mice with the worst prognosis. In surviving mice the expansion of activated circulating T cell and switched-class B cells with a long-term protective humoral response from the second infection week is remarkable. Our results demonstrate that the follow-up studies of immunological blood parameters during a malaria infection can offer information about the course of the pathological process and the immune response.     

Population kinetic study on the origin of guinea pig monocyte heterogeneity

Population kinetic studies were performed on guinea pig peripheral blood monocyte fractions isolated by counter-flow centrifugation elutriation following a single in vivo pulse of tritiated thymidine. Labeled large monocytes (volume 317 micron3; relative distribution 49%; circulating half-life 5.7 hr; and production rate 17,000 cells/ml blood/hr) accumulated in the circulation more rapidly, had a faster turnover time, and were produced in greater numbers than small monocytes (volume 283 micron3; relative distribution 34%; circulating half-life 10.8 hr; and production rate of 6100 cells/ml blood/hr). The kinetic data do not support a maturational sequence of small into large monocytes. Intermediate monocytes (volume 300 micron3; relative distribution 11%; circulating half-life 18.2 hr) and very large monocytes (volume 354 micron3; relative distribution 6%; circulating half-life 36.5 hr) had production rates, respectively, of only 1200 and 320 cells/ml blood/hr. Maxima in the labeling index curve for small and large monocytes suggested a generation time of 24 hr while grain count analysis revealed that these two cell fractions were derived from a precursor population with similar numbers of reductive divisions. Grain count analysis of intermediate and very large monocytes revealed that these cells differed from both small and large monocytes. Our data support the concept that monocyte subsets exist in guinea pig peripheral blood with different kinetics of production and survival.     

Bidirectional effects of cytokines on the growth of Mycobacterium avium within human monocytes

Certain cytokines including IFN-gamma possess macrophage-activating factor activity that enhances the ability of these effector cells to destroy intracellular pathogens. A panel of recombinant and highly purified human cytokines was screened to detect this effect on the activation of human monocytes to kill Mycobacterium avium in an in vitro model. Peripheral blood monocytes obtained from 15 healthy donors were precultured for 2 days before infection. Monocytes were infected with two strains of M. avium, one AIDS-associated and relatively avirulent strain (86m2096), and the other a non-AIDS-associated isolate that demonstrated consistent and rapid growth in cultured human monocytes (LR114F). The effects of recombinant and purified human cytokines on M. avium infection were assayed by determining CFU of M. avium in lysates of infected monocytes after 0, 4, and 7 days of culture. After infection, monocytes were cultured in medium alone or continuously in the presence of the following cytokines: IL-1 alpha, IL-1 beta, IL-2, IL-3, IL-4, IL-6, IFN-gamma, granulocyte-macrophage-CSF, or macrophage-CSF. In some experiments, cultures were performed in the presence of indomethacin (IM) in addition to cytokines. Culture in the presence of rIFN-gamma was associated with a decrease in mycobacterial growth within human monocytes. The combination of 300 U/ml of IFN-gamma plus 1 micrograms/ml of IM was associated with a 10-fold decrease (p less than 0.01) in intracellular growth of the virulent strain (LR114F) compared with unstimulated cultures. No other cytokine or combination of a cytokine with IM inhibited the intracellular growth of either strain of M. avium in human monocytes. Rather, several cytokines enhanced the intracellular growth of M. avium. IL-3, IL-6, and macrophage-CSF increased the growth of one, and IL-1 alpha of both strains of M. avium tested. IL-1 alpha and IL-6 also induced M. avium growth in tissue culture medium without monocytes. These studies indicate bidirectional effects of cytokines on intracellular parasitism that may influence the outcome of M. avium infection.     

Systemic and brain macrophage infections in relation to the development of simian immunodeficiency virus encephalitis

The brains of individuals with lentiviral-associated encephalitis contain an abundance of infected and activated macrophages. It has been hypothesized that encephalitis develops when increased numbers of infected monocytes traffic into the central nervous system (CNS) during the end stages of immunosuppression. The relationships between the infection of brain and systemic macrophages and circulating monocytes and the development of lentiviral encephalitis are unknown. We longitudinally examined the extent of monocyte/macrophage infection in blood and lymph nodes of pigtailed macaques that did or did not develop simian immunodeficiency virus encephalitis (SIVE). Compared to levels in macaques that did not develop SIVE, more ex vivo virus production was detected from monocyte-derived macrophages and nonadherent peripheral blood mononuclear cells (PBMCs) from macaques that did develop SIVE. Prior to death, there was an increase in the number of circulating PBMCs following a rise in cerebrospinal fluid viral load in macaques that did develop SIVE but not in nonencephalitic macaques. At necropsy, macaques with SIVE had more infected macrophages in peripheral organs, with the exception of lymph nodes. T cells and NK cells with cytotoxic potential were more abundant in brains with encephalitis; however, T-cell and NK-cell infiltration in SIVE and human immunodeficiency virus encephalitis was more modest than that observed in classical acute herpes simplex virus encephalitis. These findings support the hypothesis that inherent differences in host systemic and CNS monocyte/macrophage viral production are associated with the development of encephalitis.     

C5a and C5a(desArg) enhance the susceptibility of monocyte-derived macrophages to HIV infection

Mononuclear phagocytes, which include circulating blood monocytes and differentiated tissue macrophages, are believed to play a central role in the sexual transmission of HIV infection. The ability of HIV to productively infect these cells may be influenced by action of exogenous or host-derived substances at the site of viral entry. Given the potent capacities of inflammatory mediators to stimulate anaphylatoxic and immunomodulatory functions in mucosa, the effects of complement-derived anaphylatoxins on the susceptibility of monocytes and monocyte-derived macrophages (MDM) to HIV-1 infection were examined. In our in vitro system, the susceptibility to infection was up to 40 times increased in MDM that had been exposed to C5a or C5a(desArg), but not to C3a or C3a(desArg), for 2 days before adding of virus. By contrast, the treatment with complement anaphylatoxins did not affect HIV replication in fresh monocytes. Stimulatory effect of C5a and its desArg derivative on HIV infection correlated with the increase of TNF-alpha and IL-6 secretion from MDM. All these functional effects of C5a and C5a(desArg) were reversible by treatment of cells with the mAb that functionally blocks C5aR. Taken together, these results indicate that C5a and C5a(desArg) may increase the susceptibility of MDM to HIV infection through stimulation of TNF-alpha and IL-6 secretion from these cells.     

In Vivo Monocyte Tropism of Pathogenic Feline Immunodeficiency Viruses

Virus-infected monocytes rarely are detected in the bloodstreams of animals or people infected with immunodeficiency-inducing lentiviruses, yet tissue macrophages are thought to be a major reservoir of virus-infected cells in vivo. We have identified feline immunodeficiency virus (FIV) clinical isolates that are pathogenic in cats and readily transmitted vertically. We report here that five of these FIV isolates are highly monocytotropic in vivo. However, while FIV-infected monocytes were numerous in the blood of experimentally infected cats, viral antigen was not detectable in freshly isolated cells. Only after a short-term (at least 12-h) in vitro monocyte culture were FIV antigens detectable (by immunocytochemical analysis or enzyme-linked immunosorbent assay). In vitro experiments suggested that monocyte adherence provided an important trigger for virus antigen expression. In the blood of cats infected with a prototype monocytotropic isolate (FIV subtype B strain 2542), infected monocytes appeared within 2 weeks, correlating with high blood mononuclear-cell-associated viral titers and CD4 cell depletion. By contrast, infected monocytes could not be detected in the blood of cats infected with a less pathogenic FIV strain (FIV subtype A strain Petaluma). We concluded that some strains of FIV are monocytotropic in vivo. Moreover, this property may relate to virus virulence, vertical transmission, and infection of tissue macrophages.     

Regulation of tissue factor gene expression in the monocyte procoagulant response to endotoxin.

Tissue factor is the cellular receptor and cofactor for plasma factor VIIa which initiates the coagulation protease cascade on cell surfaces. Although normally absent from all intravascular cell types, tissue factor can be induced to appear on circulating monocytes and vascular endothelial cells by specific inflammatory or immunological mediators. In this study, we have examined the regulation of endotoxin-induced tissue factor gene expression in peripheral blood monocytes.     

Monocyte Subset Dynamics in Human Atherosclerosis Can Be Profiled with Magnetic Nano-Sensors

Monocytes are circulating macrophage and dendritic cell precursors that populate healthy and diseased tissue. In humans, monocytes consist of at least two subsets whose proportions in the blood fluctuate in response to coronary artery disease, sepsis, and viral infection. Animal studies have shown that specific shifts in the monocyte subset repertoire either exacerbate or attenuate disease, suggesting a role for monocyte subsets as biomarkers and therapeutic targets. Assays are therefore needed that can selectively and rapidly enumerate monocytes and their subsets. This study shows that two major human monocyte subsets express similar levels of the receptor for macrophage colony stimulating factor (MCSFR) but differ in their phagocytic capacity. We exploit these properties and custom-engineer magnetic nanoparticles for ex vivo sensing of monocytes and their subsets. We present a two-dimensional enumerative mathematical model that simultaneously reports number and proportion of monocyte subsets in a small volume of human blood. Using a recently described diagnostic magnetic resonance (DMR) chip with 1 µl sample size and high throughput capabilities, we then show that application of the model accurately quantifies subset fluctuations that occur in patients with atherosclerosis.     

Neuroinvasion of fluorescein-positive monocytes in acute simian immunodeficiency virus infection

Monocytes and macrophages play a central role in the pathogenesis of human immunodeficiency virus (HIV)-associated dementia. They represent prominent targets for HIV infection and are thought to facilitate viral neuroinvasion and neuroinflammatory processes. However, many aspects regarding monocyte brain recruitment in HIV infection remain undefined. The nonhuman primate model of AIDS is uniquely suited for examination of the role of monocytes in the pathogenesis of AIDS-associated encephalitis. Nevertheless, an approach to monitor cell migration from peripheral blood into the central nervous system (CNS) in primates had been lacking. Here, upon autologous transfer of fluorescein dye-labeled leukocytes, we demonstrate the trafficking of dye-positive monocytes into the choroid plexus stromata and perivascular spaces in the cerebra of rhesus macaques acutely infected with simian immunodeficiency virus between days 12 and 14 postinfection (p.i.). Dye-positive cells that had migrated expressed the monocyte activation marker CD16 and the macrophage marker CD68. Monocyte neuroinvasion coincided with the presence of the virus in brain tissue and cerebrospinal fluid and with the induction of the proinflammatory mediators CXCL9/MIG and CCL2/MCP-1 in the CNS. Prior to neuroinfiltration, plasma viral load levels peaked on day 11 p.i. Furthermore, the numbers of peripheral blood monocytes rapidly increased between days 4 and 8 p.i., and circulating monocytes exhibited increased functional capacity to produce CCL2/MCP-1. Our findings demonstrate acute monocyte brain infiltration in an animal model of AIDS. Such studies facilitate future examinations of the migratory profile of CNS-homing monocytes, the role of monocytes in virus import into the brain, and the disruption of blood-cerebrospinal fluid and blood-brain barrier functions in primates.     

Expression of CXCR4 on feline peripheral blood mononuclear cells: effect of feline immunodeficiency virus infection

CXCR4 expression on feline peripheral blood mononuclear cells (PBMC) was analyzed. While monocytes and B lymphocytes expressed CXCR4, no CXCR4 was detected on T lymphocytes, in stark contrast to the expression pattern on T lymphocytes from humans. In spite of the important role that CXCR4 plays in infection with feline immunodeficiency virus, expression on PBMC in vivo was unaffected by infection with either a primary or a cell culture-adapted virus strain.     

The stimulation of sheep monocyte mitosis in vitro by autologous plasma taken after surgery and the implantation of foreign material.

When blood monocytes from sheep are grown upon glass in 100% autologous plasma they divide with low mitotic indices, but if the plasma is taken from the sheep three or four days after the implantation of a small Teflon or stainless steel cylinder, the maximum mitotic indices of the monocytes cultured in that plasma rise. Plasma collected from the sheep in the first two days after operation is often toxic to cultured autologous monocytes. An equivalent sham operation increases the mitogenicity of the plasma, but without a preceding toxic period. No consistent effects upon the numbers of circulating blood monocytes were noted following implantation operations. A hypothesis is presented which suggests that blood monocyte multiplication in vivo and in vitro is dependent upon (a) a lesion which causes cell attachment, and (b) a humoral component of the plasma which stimulates mitosis. Surgery, and the implantation of foreign materials will increase the concentration of such a substance in plasma.     

Cytochemical study of the natural resistance of the body in Trichophyton rubrum-induced mycosis (tinea)

Clinical-cytochemical examination of 27 patients with tinea has permitted estimating nonspecific immune status of an organism. The NBT-test is shown to be highly informative and active. Differences in metabolic activity of phagocytizing subpopulations of peripheral blood are revealed. The results obtained underlie the development of the pathogenetically substantiated therapy method using diucyphon and chlorophyllipt, immunomodulators of different pharmacological groups.     

Tumor necrosis factor alpha induction in human monocytes

The present study was undertaken to assess the presence of tumor necrosis factor (TNF)-alpha mRNA and protein in circulating human blood monocytes and to study the TNF-alpha gene expression in human monocytes isolated by continuous Percoll gradient fractionation. The technique of RNA isolation directly from the blood samples was used to study TNF-alpha mRNA expression in circulating human blood leukocytes. It was shown that human blood leukocytes of healthy donors contained no presynthesized pool of TNF-alpha mRNA as well as no TNF-alpha protein. It was found that early pretreatment with cycloheximide interferes with TNF-alpha mRNA induction by Staphylococcus aureus.     

The monocyte-macrophage-mast cell axis in dengue pathogenesis

Dengue virus, the causative agent of dengue disease which may have hemorrhagic complications, poses a global health threat. Among the numerous target cells for dengue virus in humans are monocytes, macrophages and mast cells which are important regulators of vascular integrity and which undergo dramatic cellular responses after infection by dengue virus. The strategic locations of these three cell types, inside blood vessels (monocytes) or outside blood vessels (macrophages and mast cells) allow them to respond to dengue virus infection with the production of both intracellular and secretory factors which affect virus replication, vascular permeability and/or leukocyte extravasation. Moreover, the expression of Fc receptors on the surface of monocytes, macrophages and mast cells makes them important target cells for antibody-enhanced dengue virus infection which is a major risk factor for severe dengue disease, involving hemorrhage. Collectively, these features of monocytes, macrophages and mast cells contribute to both beneficial and harmful responses of importance to understanding and controlling dengue infection and disease.