Marathon runners, subjected to intense training regimens and prolonged, exhaustive exercises, often experience a compromised immune response. Probiotic supplementation has emerged as a potential remedy to mitigate the impact of prolonged exercise on athletes. Consequently, this study sought to assess the influence of probiotic supplementation on monocyte functionality both before and after the official marathon race. Twenty-seven runners were randomly and double-blindly assigned to two groups: placebo (n 13) and probiotic (PRO) (n 14). Over 30 d, both groups received supplements – placebo sachets containing maltodextrin (5 g/d) and PRO sachets containing 1 × 1010 colony-forming unit Lactobacillus acidophilus and 1 × 1010 colony-forming unit Bifidobacterium bifidum subsp. lactis. Blood samples were collected, and immunological assays, including phagocytosis, hydrogen peroxide production, cytokine levels and monocyte immunophenotyping, were conducted at four different intervals: baseline (start of supplementation/30 d pre-marathon), 24 h-before (1 d pre-marathon), 1 h-after (1 h post-marathon) and 5 d-after (5 d post-marathon). Monocyte populations remained consistent throughout the study. A notable increase in phagocytosis was observed in the PRO group after 30 d of supplementation. Upon lipopolysaccharide stimulation, both PRO and placebo groups exhibited decreased IL-8 production. However, after the marathon race, IL-15 stimulation demonstrated increased levels of 5 d-after, while IL-1-β, IL-8, IL-10, IL-15 and TNF-α varied across different intervals, specifically within the PRO group. Probiotic supplementation notably enhanced the phagocytic capacity of monocytes. However, these effects were not sustained post-marathon.

Factors that are potentially important in the pulmonary pathogenesis of asbestos and other mineral particles are: 1) morphology, 2) Fe-content, 3) solubility under intraphagosomal conditions, 4) value and sign of the surface potential of the particle, 5) hydrophobicity or hydrophilicity, 6) capacity to activate phagocytic leukocytes, and 7) duration of exposure to the particles. The order of importance of these factors in causing severe or fatal pulmonary pathogenicity is estimated to be: 1 > 3 > 7 > 6 ≫ 5 > 4 > 2. The order of pathogenicity of the minerals is estimated as: amphibole asbestos: crocidolite, tremolite, amosite > erionite > serpentine asbestos: chrysotile > talc > silica > simple metal oxides. Particle length, duration of exposure to the particles, and pre-treatment of the particles may however enhance the pathogenic potential of any of the lower-ranked particles.

Leishmaniases affect 12 million people worldwide. They are caused by Leishmania spp., protozoan parasites transmitted to mammals by female phlebotomine flies. During the life cycle, promastigote forms of the parasite live in the gut of infected sandflies and convert into amastigotes inside the vertebrate macrophages. The parasite evades macrophage's microbicidal responses due to virulence factors that affect parasite phagocytosis, survival and/or proliferation. The interaction between Leishmania and macrophage molecules is essential to phagocytosis and parasite survival. Proteins containing leucine-rich repeats (LRRs) are common in several organisms, and these motifs are usually involved in protein–protein interactions. We have identified the LRR17 gene, which encodes a protein with 6 LRR domains, in the genomes of several Leishmania species. We show here that promastigotes of Leishmania (L.) amazonensis overexpressing LaLRR17 are more infective in vitro. We produced recombinant LaLRR17 protein and identified macrophage 78 kDa glucose-regulated protein (GRP78) as a ligand for LaLRR17 employing affinity chromatography followed by mass spectrometry. We showed that GRP78 binds to LaLRR17 and that its blocking precludes the increase of infection conferred by LaLRR17. Our results are the first to report LRR17 gene and protein, and we hope they stimulate further studies on how this protein increases phagocytosis of Leishmania.

Cathepsin-D (Cat-D) is a major proteolytic enzyme in phagocytic cells. In the retinal pigment epithelium (RPE), it is responsible for the daily degradation of photoreceptor outer segments (POSs) to maintain retinal homeostasis. Melanoregulin (MREG)-mediated loss of phagocytic capacity has been linked to diminished intracellular Cat-D activity. Here, we demonstrate that loss of MREG enhances the secretion of intermediate Cat-D (48 kDa), resulting in a net enhancement of extracellular Cat-D activity. These results suggest that MREG is required to maintain Cat-D homeostasis in the RPE and likely plays a protective role in retinal health. In this regard, in the Mregdsu/dsu mouse, we observe increased basal laminin. Loss of the Mregdsu allele is not lethal and therefore leads to slow age-dependent changes in the RPE. Thus, we propose that this model will allow us to study potential dysregulatory functions of Cat-D in retinal disease.

Prospective studies have indicated an age-related impairment of the immune response. Carotenoids have been hypothesised to enhance immune cell function. The aim of the present study was to describe the age-related effects and the impact of in vivo dietary carotenoid depletion and repletion on specific and non-specific immunity. A total of ninety-eight healthy male subjects (aged 20–75 years) received a carotenoid-depleted diet for 3 weeks and were then supplemented daily for 5 weeks with 30 mg β-carotene, 15 mg lycopene and 9 mg lutein. Blood samples were collected at study entry, after depletion and supplementation, and biomarkers of immune status were determined. We found that serum IgA levels were positively correlated with ageing. Lymphocyte phenotyping indicated an increase with age in the memory T-helper cell subpopulation (CD4+CD45RO+) concomitantly with a decrease in naive T-helper cells (CD4+CD45RA+). A significant increase in the natural killer cells subpopulation and a small decrease in B lymphocytes were also observed, especially for the oldest volunteers. From ex vivo cell function exploration, a positive correlation was observed between age and IL-2 production of phytohaemagglutinin-stimulated lymphocytes. Neutrophils' bactericidal activity was significantly impaired with age (from 50 years) and was modulated by carotenoid status. An age effect was found on neutrophils' spontaneous migration but not on directed migration. Immune response in healthy human subjects is mostly affected by age rather than by dietary carotenoid depletion and repletion. Even in carefully selected healthy volunteers, some age-related immune changes occur predominantly from 50 years onwards. This immunosenescence could generate a loss in the immune system adjustment capacity.

Life-cycle stages of the microsporidia Nosema bombycis, the pathogen causing silkworm pebrine, were separated and purified by an improved method of Percoll-gradient centrifugation. Soluble protein fractions of late sporoblasts (spore precursor cells) and mature spores were analysed by two-dimensional polyacrylamide gel electrophoresis (2D-PAGE). Protein spots were recovered from gels and analysed by mass spectrometry. The most abundant differential protein spot was identified by database search to be a hypothetical spore wall protein. Using immunoelectron microscopy, we demonstrated that HSWP5 is localized to the exospore of mature spores and renamed it as spore wall protein 5 (NbSWP5). Further spore phagocytosis assays indicated that NbSWP5 can protect spores from phagocytic uptake by cultured insect cells. This spore wall protein may function both for structural integrity and in modulating host cell invasion.

Microglia, the brain's innate immune cell type, are cells of mesodermal origin that populate the central nervous system (CNS) during development. Undifferentiated microglia, also called ameboid microglia, have the ability to proliferate, phagocytose apoptotic cells and migrate long distances toward their final destinations throughout all CNS regions, where they acquire a mature ramified morphological phenotype. Recent studies indicate that ameboid microglial cells not only have a scavenger role during development but can also promote the death of some neuronal populations. In the mature CNS, adult microglia have highly motile processes to scan their territorial domains, and they display a panoply of effects on neurons that range from sustaining their survival and differentiation contributing to their elimination. Hence, the fine tuning of these effects results in protection of the nervous tissue, whereas perturbations in the microglial response, such as the exacerbation of microglial activation or lack of microglial response, generate adverse situations for the organization and function of the CNS. This review discusses some aspects of the relationship between microglial cells and neuronal death/survival both during normal development and during the response to injury in adulthood.

Antarctic fishes survive freezing through the secretion of antifreeze glycoproteins (AFGPs), which bind to ice crystals to inhibit their growth. This mode of action implies that ice crystals must be present internally for AFGPs to function. The entry and internal accumulation of ice is likely to be lethal, however, so how do fishes survive in its presence? We propose a novel function for the interaction between internal ice and AFGPs, namely the promotion of ice uptake by splenic phagocytes. We show here that i) external mucus of Antarctic notothenioids contains AFGPs and thus has a potential protective role against ice entry, ii) AFGPs are distributed widely through the extracellular space ensuring that they are likely to come into immediate contact with ice that penetrates their protective barriers, and iii) using AFGP-coated nanoparticles as a proxy for AFGP adsorbed onto ice, we suggest that internal ice crystals are removed from the circulation through phagocytosis, primarily in the spleen. We argue that intracellular sequestration in the spleen minimizes the risks associated with circulating ice and enables the fish to store the ice until it can be dealt with at a later date, possibly by melting during a seasonal warming event.

This study aimed to determine whether orally administered ovine serum immunoglobulin (Ig) modulates aspects of immunity such as phagocytosis, lymphocyte proliferation, cytokine production, intestinal and plasma Ig concentrations in growing rats. Forty-five male Sprague–Dawley rats (n = 15/group) were used in the 21-day study, and fed a basal control diet (BD; no Ig) or two test diets: freeze-dried ovine Ig (FDOI) and inactivated ovine Ig (IOI). Phagocytic activity of peripheral blood leukocytes and lymphocyte proliferation in the presence of the mitogen concanavalin A (ConA) was greater (P < 0.05) for the FDOI-fed rats than for the BD- and IOI-fed groups. ConA-stimulated and unstimulated spleen cell culture produced higher (P < 0.05) interferon-γ and interleukin-4, respectively, from rats fed the FDOI than rats fed the BD diet. In the jejunum, ileum and plasma, rats fed FDOI produced higher (P < 0.05) concentrations of secretory IgA (sIgA) than rats fed IOI or BD. Rats fed the FDOI diet had greater jejunal (P = 0.037) and lower plasma (P = 0.025) rat IgG concentrations than rats fed either BD or IOI. In conclusion, an ovine Ig fraction selectively modulated various indices of immune function.

This study reports that the cytopathic effect of Trichomonas vaginalis, an important human parasite of the urogenital tract, occurs due to mechanical stress and subsequent phagocytosis of the necrotic cells. The investigation was done using a primary culture of bovine oviduct epithelial cells (BOECs), grown either in monolayers or as floating cells. Trophozoites displaying different virulence levels were co-incubated with BOECs for times varying between 1 min and 48 h. Analyses were performed using videomicroscopy, scanning and transmission electron microscopy, colourimetric assays and cytochemistry. Injury was observed as early as 1 h after incubation, while after 12 h the host cells were severely damaged when a fresh trichomonad isolate was used. Trichomonads attack the host cells by clustering around them. Mechanical stress on the microvilli of the host cells was observed and appeared to induce plasma membrane damage and cell death. After membrane injury and lysis, fragments of the necrotic cells were ingested by trichomonads. Phagocytosis occurred by trichomonads avidly eating large portions of epithelial cells containing the nucleus and other organelles, but living or intact cells were not ingested. Necrotic fragments were rapidly digested in lysosomes, as shown by acid phosphatase and ruthenium red assays where only the BOECs were labelled. The lytic capacity of the trichomonads was more pronounced in host cell suspensions.

We have examined the development of microglia in the rat retina, using a peroxidase-conjugated lectin derived from Griffonia simplicifolia. Retinas were studied from animals aged from E(embryonic day)12, just after the invagination of the optic cup and prior to the closure of the optic fissure, to adulthood. The lectin also proved a sensitive label for the endothelial cells of the developing retina. Our results provide some support for the view that microglia are derived from the monocyte-macrophage series of blood cells. At E12, most labeled cells were found at the vitreal surface, suggesting that they had come from the hyaloid circulation, while some had entered the retina and appeared to be migrating towards its ventricular surface. From E14 to early postnatal ages, most labeled cells had processes and resembled the amoeboid microglial cells described in silver carbonate staining studies (Ling, 1982). The number of labeled cells rose from about 700 to E14 to a peak of about 27,000 at P(postnatal day)7, and fell to about 19,600 by P12. As early as E16, a regularity was apparent in the distribution of microglial cells over the surface of the retina, the cells tending to avoid each other. Microglial cells are found throughout the thickness of the very young retina, but as the layers of the retina differentiate, they are increasingly restricted to the inner half of the retina. Our findings indicate that microglia enter the retina well before the period of neuronal death, making it unlikely that they invade the retina solely in response to cell death. Our results confirm however that, once in the retina, microglia become associated with, and appear to phagocytose, the pyknotic debris which appears during the period of neuronal death. They also become closely associated with the retinal vasculature. In the adult, the intensity of the labeling of microglia was much reduced. Those cells which were labeled appeared more differentiated, resembling the “resting microglia” described in earlier studies.

The transmission-blocking monoclonal antibody 13.1, which recognizes the ookinete surface antigen Pbs21 of Plasmodium berghei, and an IgG2a isotype control antibody 26.37 were purified by caprylic acid and ammonium sulphate precipitation. Fab fragments were prepared by papain digestion. IgG but not Fab from antibody 13.1 reduced ookinete formation by P. berghei in culture by as much as 94% at a concentration of 100 μg/ml. There was little difference in antibody efficacy in the range 6·25–400 μg/ml in this assay. The parasite was most sensitive to antibody activity in the first 6–9 h of culture, i.e. the gamete/zygote and early retort stages. Peripheral blood leucocytes (PBL) were essential to achieve maximal inhibition by mAb 13.1 (activity was abrogated totally if PBL were removed). Together the data suggest that one of the mechanisms of action of this antibody is antibody-mediated PBL killing. Phagocytosis of parasites was noted in these experiments in all cultures. We have not attempted in this study to distinguish between Fc-mediated opsonization, as opposed to antibody-dependent cellular cytotoxicity.

Human neutrophils were infected with tachyzoites of Toxoplasma gondii in vitro. Infection rates after 1 h of incubation were 38·6–52·0% and 16·0–25·8% in the presence or absence of specific antibody respectively. Apparently, neutrophils killed tachyzoites 18 h after infection in the presence of antibody, whereas tachyzoites proliferated in 25·0–35·0% of infected neutrophils in the absence of antibody. Total tachyzoite counts 18 h after infection were approximately 4 times as high as those 1 h after infection in the absence of antibody and complement. Complement also enhanced phagocytosis and killing of tachyzoites by neutrophils, but the effects were less than that of antibody. Superoxide anion was produced intensively within 1 h after infection, depending on the presence of antibody or complement. Neutrophils seem to be responsible for dissemination of tachyzoites in the host during the initial phase of Toxoplasma infection until antibody is produced.

The balanced polymorphism of glucose-6-phosphate dehydrogenase deficiency (G6PD-) is believed to have evolved through the selective pressure of malaria combined with consumption of fava beans. The implicated fava bean constituents are the hydroxypyrimidine glucosides vicine and convicine, which upon hydrolysis of their β-O-glucosidic bond, become potent pro-oxidants. In this work we show that the glucosides inhibit the growth of Plasmodium falciparum, increase the hexose-monophosphate shunt activity and the phagocytosis of malaria-infected erythrocytes. These activities are exacerbated in the presence of β-glucosidase, implicating their pro-oxidant aglycones in the toxic effect, and are more pronounced in infected G6PD- erythrocytes. These results suggest that G6PD- infected erythrocytes are more susceptible to phagocytic cells, and that fava bean pro-oxidants are more efficiently suppressing parasite propagation in G6PD- erythrocytes, either by directly affecting parasite growth, or by means of enhanced phagocytic elimination of infected cells. The present findings could account for the relative resistance of G6PD- bearers to falciparum malaria, and establish a link between dietary habits and malaria in the selection of the G6PD- genotype.

Effects of infection with the avian schistosome Trichobilharzia ocellata on the activity of the internal defence system of the intermediate snail host Lymnaea stagnalis were studied, utilizing an in vitro phagocytosis assay for determining haemocyte activity. A distinction was made between plasma- and cell-associated effects. The period immediately after penetration of the parasite into the snail host (1.5–72 h post-exposure (p.e.)) was extensively studied. In addition, several time-points coinciding with the later–successive–stages of parasite development (2, 4, 6, 8 and 10 weeks p.e.) were investigated. Plasma-associated enhancement of defence activity was found between 1.5 and 6 h p.e., followed by plasma-associated suppression between 12 and 72 h p.e. A cell-associated activation was found between 1.5 and 6 h p.e. and also at 8 and 10 weeks p.e. How these effects on the defence system may be related to phenomena observed in infected snails at these time-points is discussed.

In the present study, the effects of trans-MUFA, elaidic acid (EA; 18 : 1-9t) and vaccenic acid (VA; 18 : 1-11t) on rat neutrophil functions were compared with those of cis-monounsaturated oleic acid (OA) (18 : 1-9c) and saturated stearic acid (SA; 18 : 0) (10–150 μm). Trans-fatty acids enhanced neutrophil phagocytic capacity, superoxide (O2∙-) and hydrogen peroxide production, and candidacidal activity. The same effects were observed for OA. Cells treated with trans-MUFA showed reduced production of NO∙, whereas those treated with OA showed an increase in production. Treatment with SA did not provoke significant effect on the parameters investigated. The increase in O2∙- production induced by MUFA was not observed when diphenyleneiodonium, an NADPH oxidase inhibitor, was added to the medium. This finding suggests that MUFA stimulate neutrophil NADPH oxidase activity. The addition of 3-[1-[3-(dimethylamino)propyl]-1H-indol-3-yl]-4-(1H-indol-3-yl)-1H-pyrrole-2,5-dione, a protein kinase C (PKC) inhibitor, and wortmannin, a phosphatidylinositol-3 kinase (PI3K) inhibitor, did not affect O2∙- production induced by MUFA. Therefore, the mechanisms by which MUFA stimulate NADPH oxidase are not dependent on PKC and do not seem to involve PI3K. Experiments using Zn2+, an inhibitor of NADPH oxidase H+ channel, indicated that MUFA activate the NADPH oxidase complex in rat neutrophil due to opening of H+ channel.

A 4 base pair deletion in a splice donor site of the Mfrp (membrane-type frizzled-related protein) gene, herein referred to as Mfrprd6/rd6, is predicted to lead to the skipping of exon 4 and photoreceptor degeneration in retinal degeneration 6 (rd6) mutant mice. Little, however, is known about the function of the protein or how the mutation causes the degenerative retinal phenotype. Here we examine ultrastructural changes in the retina of Mfrprd6/rd6 mice to determine the earliest effects of the mutation. We also extend the reported observations of the expression pattern of the dicistronic Mfrp/C1qtnf5 message and the localization of these and other retinal pigment epithelium (RPE) and retinal proteins during development and assess the ability of RPE cells to phagocytize outer segments (OSs) in mutant and wild-type (WT) mice. At the ultrastructural level, OSs do not develop normally in Mfrprd6/rd6 mutants. They are disorganized and become progressively shorter as mutant mice age. Additionally, there are focal areas in which there is a reduction of apical RPE microvilli. At P25, the rod electroretinogram (ERG) a-wave of Mfrprd6/rd6 mice is reduced in amplitude by ~50% as are ERG components generated by the RPE. Examination of β-catenin localization and Fos and Tcf-1 expression, intermediates of the canonical Wnt pathway, showed that they were not different between mutant and WT mice, suggesting that MFRP may operate through an alternative pathway. Finally, impaired OS phagocytosis was observed in Mfrprd6/rd6 mice both in standard ambient lighting conditions and with bright light exposure when compared to WT controls.

Parameters of hemocyte populations have been considered as relevant indicators of bivalve health and are currently used in immunotoxicological studies. Hemocytes in hemolymph can be collected by puncturing either the pericardial cavity or the adductor muscle sinus with a syringe. Flow cytometry is a methodological approach that is increasingly being used in laboratories for the study of hemocyte parameters in aquatic invertebrates. However, various protocols for hemocyte processing in laboratories equipped with different types of cytometers have been published. In this context, two flow cytometers (EPICS XL4®, Beckman Coulter and FacsCalibur®, Becton Dickinson) and two sites of hemocyte collection (pericardial cavity and adductor muscle sinus) were compared for the analysis of hemocyte parameters in the Pacific oyster, Crassostrea gigas. Hemolymph cells were analyzed in terms of their number and organelle contents. Cell mortality, phagocytosis, non specific esterase, extension of the lysosomal compartment and production of reactive oxygen species were quantified. The results showed that the phagocytic index was higher for hemocytes obtained in the muscle sinus hemolymph. The results are discussed with respect to the potential use of flow cytometry as a tool for hemocyte studies in bivalves.

Glial cells provide support and protection for neurons in the embryonic and adult brain, mediated in part through the phagocytic activity of glia. Glial cells engulf apoptotic cells and pruned neurites from the developing nervous system, and also clear degenerating neuronal debris from the adult brain after neural trauma. Studies indicate that Drosophila melanogaster is an ideal model system to elucidate the mechanisms of engulfment by glia. The recent studies reviewed here show that many features of glial engulfment are conserved across species and argue that work in Drosophila will provide valuable cellular and molecular insight into glial engulfment activity in mammals.