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E L Cooper

Publications and source records attributed to E L Cooper.

At least 19 recordsLinked to original sources
PubMed2005

Still waiting for the toll?

Multicellular organisms including invertebrates and vertebrates live in various habitats that may be aquatic or terrestrial where they are constantly exposed to deleterious pathogens. These include viruses, bacteria, fungi, and parasites. They have evolved various immunodefense mechanisms that may protect them from infection by these microorganisms. These include cellular and humoral responses and the level of differentiation of the response parallels the evolutionary development of the species. The first line of innate immunity in earthworms is the body wall that prevents the entrance of microbes into the coelomic cavity that contains fluid in which there are numerous leukocyte effectors of immune responses. When this first barrier is broken, a series of host responses is set into motion activating the leukocytes and the coelomic fluid. The responses are classified as innate, natural, non-specific, non-anticipatory, non-clonal (germ line) in contrast to the vertebrate capacity that is considered adaptive, induced, specific, anticipatory and clonal (somatic). Specific memory is associated with the vertebrate response and there is information that the innate response of invertebrates may under certain conditions possess specific memory. The invertebrate system when challenged affects phagocytosis, encapsulation, agglutination, opsonization, clotting and lysis. At least two major leukocytes, small and large mediate lytic reactions against several tumor cell targets. Destruction of tumor cells in vitro shows that phagocytosis and natural killer cell responses are distinct properties of these leukocytes. This has prompted newer searches for immune function and regulation in other systems. The innate immune system of the earthworm has been analyzed for more than 40 years with every aspect examined. However, there are no known entire sequences of the earthworm as exists in these other invertebrates. Because the earthworm lives in soil and has been utilized as a successful monitor for pollution, there are studies that reveal up and down regulation of responses in the immune system after exposure to a variety of environmental pollutants. Moreover, there are partial sequences that appear in earthworms after exposure to environmental pollutants such as cadmium and copper. There are now attempts to define the AHR receptor crucial for intracellular signaling after exposure to pollutants, but without linking the signals to changes in the immune system. There are several pathways for signal transduction, including JAK/STAT, TOLL, TRAF PIP3, known in invertebrates and vertebrates. For resistance to pathogens, conserved signal transduction components are required and these include a Toll/IL-1 receptor domain adaptor protein that functions upstream of a conserved p38 MAP kinase pathway. This pathway may be an ancestral innate immune signaling pathway found in a putative common ancestor of nematodes, arthropods and even vertebrates. It could also help us to link pollution, innate immunity and transduction in earthworms.

Animals↗
PubMed2004

Anticipating innate immunity without a Toll.

Earthworm innate immunity depends upon small and large leukocytes (coelomocytes) that synthesize and secrete humoral antimicrobial molecules (e.g. lysenin, fetidin, eiseniapore, coelomic cytolytic factor [CCF]; Lumbricin I). Small coelomocytes (cytotoxic) are positive (CD11a, CD45RA, CD45RO, CDw49b, CD54, beta(2)-m and Thy-1 [CD90]; CD24; TNF-alpha) but negative using other mammalian markers. Large coelomocytes (phagocytic) are uniformly negative. Specific earthworm anti-EFCC 1, 2, 3, 4 mAbs are negative for Drosophila melanogaster hemocytes and mammalian cells but positive those of earthworms. Coelomocytes contain several lysosomal enzymes involved in phagocytosis and a pattern recognition molecule (CCF) that may trigger the prophenoloxidase cascade a crucial innate immune response. Earthworms and other invertebrates possess natural, non-specific, non-clonal, and non-anticipatory immune response governed by germ line genes. Toll and Toll-like receptor signaling is essential for phagocytosis and antimicrobial peptide synthesis and secretion in insects and vertebrates but has not yet been shown to be essential in earthworm innate responses.

Animals↗
PubMed2004

Earthworm leukocytes kill HeLa, HEp-2, PC-12 and PA317 cells in vitro.

Earthworm coelomic fluid contains biologically active molecules and leukocytes that participate in phagocytosis, encapsulation. Presumably they synthesize and secrete several effector modulators of innate immune responses such as antibacterial molecules, cytotoxic proteins and cytokines. Several lytic molecules have been detected in coelomic fluid previously but it is not yet clear which are actually released from the coelomocytes. Our aim was to analyze the cytotoxic effects of coelomocytes on mammalian target cells and to provide evidence that the lytic factors originate from coelomocytes. Cell-free coelomic fluid, supernatants of short-term cultured coelomocytes, and lysates from coelomocytes--derived by mechanical and detergent extraction--were used in cytotoxicity assays performed on different mammalian standard tumor cell lines and mouse fibroblasts. We used native and denaturized (using proteinase K, and trypsin digestions, or heat-inactivation) coelomocyte lysates (CCL). The viability controls of targeted cells were made by measuring photometrically and analyzing by inverted microscopy. According to our results the coelomic fluid, the supernatant of cultured coelomocytes, and the CCL significantly decreased ratios of living cells compared to controls in a dose-dependent manner. Our experiments performed with CCLs suggest that coelomocytes are responsible for the productions of cytotoxic components presumably proteins.

Animals↗
PubMed2004

Earthworm leukocyte populations specifically harbor lysosomal enzymes that may respond to bacterial challenge.

Earthworm leukocytes (coelomocytes) are responsible for innate cellular immune functions such as phagocytosis and encapsulation against parasites and pathogens. Microbial killing results from the combined action of the phagocytic process with humoral immune factors such as agglutinins (e.g., lectins), lysosomal enzymes (e.g., acid phosphatase, lysozyme), and various cytotoxic and antimicrobial molecules. There is also evidence of weak adaptive immune responses against foreign transplants. This study focused on aspects of the innate immune response. First, anti-human acid phosphatase (anti-AcP) polyclonal antibody characterized different acid hydrolase patterns in coelomocytes. Second, flow cytometry identified a strongly immunoreactive coelomocyte population. Third, ultrastructural and cytochemical analyses revealed acid phosphatase in discrete granules (lysosomes) of effector hyaline and granular coelomocytes but not in mature chloragocytes. Coelomocytes were exposed to bacteria to assess how phagocytosis influences: (a) the production of acid phosphatase using Western blot, and (b) release of acid phosphatase using ELISA from cell-free coelomic fluid. Fourth, after phagocytosis, acid phosphatase levels differed between controls and experimentals. Fifth, we found a 39-kDa molecule that reacted intensely with anti-AcP. Our results suggest that effector earthworm coelomocytes may not eliminate pathogens only by phagocytosis but also by extracellular lysis.

Acid Anhydride Hydrolases↗
PubMed2004

Immune response to dietary proteins, gliadin and cerebellar peptides in children with autism.

The mechanisms behind autoimmune reaction to nervous system antigens in autism are not understood. We assessed the reactivity of sera from 50 autism patients and 50 healthy controls to specific peptides from gliadin and the cerebellum. A significant percentage of autism patients showed elevations in antibodies against gliadin and cerebellar peptides simultaneously. For examining cross-reaction between dietary proteins and cerebellar antigens, antibodies were prepared in rabbits, and binding of rabbit anti-gliadin, anti-cerebellar peptides, anti-MBP, anti-milk, anti-egg, anti-soy and anti-corn to either gliadin- or cerebellar-antigen-coated wells was measured. In comparison to anti-gliadin peptide binding to gliadin peptide at 100%, the reaction of anti-cerebellar peptide to gliadin peptide was 22%, whereas the binding of anti-myelin basic protein (MBP), anti-milk, anti-egg and anti-soy to gliadin was less than 10%. Further examination of rabbit anti-gliadin (EQVPLVQQ) and anti-cerebellar (EDVPLLED) 8 amino acid (AA) peptides with human serum albumin (HSA) and an unrelated peptide showed no binding, but the reaction of these antibodies with both the cerebellar and gliadin peptides was greater than 60%. This cross-reaction was further confirmed by DOT-immunoblot and inhibition studies. We conclude that a subgroup of patients with autism produce antibodies against Purkinje cells and gliadin peptides, which may be responsible for some of the neurological symptoms in autism.

Adolescent↗
PubMed2001

UVB irradiation suppresses cytokine production and innate cellular immune functions in mice.

We examined whether ultraviolet-B (UVB) irradiation (6 kJ/m2) alters cytokine production and other innate immune reactions by murine peritoneal macrophages and peripheral neutrophils. Along with these experiments, serum IgG levels were also assessed. In addition, using scanning electron microscopy (SEM) we observed macrophages that had been exposed to UVB in vitro. Results showed that UVB irradiation: (1) decreased IL-12 production while increasing IL-1alpha secretion from macrophages, but had no effect on IL-1alpha from neutrophils; (2) suppressed phagocytosis of macrophages but not of neutrophils; (3) diminished active oxygen production of macrophages but not of neutrophils; (4) had no effect on serum IgG levels; and (5) caused significant cell destruction of macrophages in vitro. These results suggested: (1) that UVB irradiation could induce characteristic suppression of innate immunity; (2) that innate cellular immunity was more susceptible to the effects of UVB irradiation than humoral immunity.

Animals↗
PubMed2001

Evidence for perforin-like activity associated with earthworm leukocytes.

Earthworm (Eisenia fetida) coelomic fluid contains several leukocytes (coelomocytes): basophils, acidophils and neutrophils as well as chloragocytes. Small coelomocytes and coelomocyte lysate are cytotoxic for the tumor cell target K562. The expression of a lytic factor was investigated by immunocytochemistry using light and transmission electron microscopy. A rat-anti-mouse-perforin-mAb labeled mainly small coelomocytes (nearly 20%) as visualized by light microscopy. TEM analysis using immunogold showed a homogenous labeling in the cytoplasm of small coelomocytes. The highest number of immunogold particles was estimated in coelomocytes with many small cytoplasmic granules. Coelomocytes with large lysosomal granules were also labeled but less intensely. No antibody binding was observed for chloragocytes either in light or electron microscopy. This suggests that the perforin-like activity is associated with only one cell type and that chloragocytes are responsible for other lytic activities. MALDI-MS revealed calreticulin usually associated with perforin in mammalian cells that mediate lysis (e.g. NK, CTL). Together, results strongly suggest the presence of putative perforin in earthworms. This in turn supports the hypothesis that perforin is a conserved component important in immune defense during evolution.

Journal Article↗
PubMed2000

Different types of response to foreign antigens by leech leukocytes.

We used morphological and immunocytochemical approaches to characterize and to show the behavior of cells involved in leech inflammatory responses. Leeches were injected with bacterial lipopolysaccharide, fluoresceinated yeasts, sulfate spheres and ciliates (Protozoa). Shortly after injection, migrating cells appeared in the area of injection. The response of the cells occurred in relation to the injected micro or macro antigens. Each injection first provoked a migration of cells towards the non-self material. Afterwards, different responses (degranulation, phagocytosis, encapsulation, melanization) occurred. The migrating cells involved in these series of processes have a similar behavior and are characterized by CD markers of macrophages, NK cells and granulocytes, which are typical of many invertebrates and vertebrates.

Animals↗
PubMed2000

Lipopolysaccharide-dependent induction of leech leukocytes that cross-react with vertebrate cellular differentiation markers.

We have designed experiments to characterise leech leukocytes that mediate inflammatory responses. Shortly after inflicting injury to the body wall in the presence of lipopolysaccharides, many cells resembling macrophages, NK cells and granulocytes of vertebrates and many invertebrates migrated to the lesioned area. Nuclei of migrating cells incorporated bromodeoxyuridine. Using human monoclonal antibodies, macrophage-like cells were positive for CD25, CD14, CD61, CD68, CD11b and CD11c. NK-like cells were positive for CD25, CD56, CD57 and CD16, and granulocytes were positive for CD11b and CD11c. In blots of leech extracts, the CD25 monoclonal antibody recognised a band of about 55 kD; the CD56 monoclonal antibody, two bands of about 140 and 210 kD; the CD57 monoclonal antibody, two bands of about 106 and 70 kD; the CD14 monoclonal antibody, a band of about 50 kD; the CD16 monoclonal antibody, a band of about 60 kD. CD61 and CD68 both recognised a band of about 110 kD; CD11b recognised a band of 200 kD, and CD11c, a band of 180 kD.

Animals↗
PubMed2000

Ontogeny recapitulates phylogeny: comparative immunology in Germany.

The innate immune system is in the spot light of modern immunology. Whenever protists, invertebrates and vertebrates are threatened by pathogens, they rapidly activate highly effective antimicrobial defense reactions. Because this young field develops very dynamically, it is important to ask what we really know about the mechanisms governing the innate immune defense system. This was the topic of a recent meeting entitled 'The Evolution of the Immune System', held at the Friedrich Schiller University in Jena, Germany. Leading scientists in the field of innate immunity presented their latest data in a historical and friendly setting.

Allergy and Immunology↗
PubMed1999

Cell adhesion and the immune system: a case study using earthworms.

In the earthworm's immune system, cell adhesion, which occurs by putative receptors on leukocytes, is essential after recognition of self vs. non-self. Confrontation with foreign antigens is a normal event in the environment, replete with microbial pathogens that pose a threat to survival. To better understand what happens when an effector cell first recognizes a foreign target followed by its adhesion to it, isolated leukocytes, in sufficient quantities to be subjected to various analyses, have been extremely beneficial. In vitro approaches when accompanied by biochemical, immunological, and molecular technologies, have opened up new vistas concerning the immune response of earthworms and other invertebrates. The most recent discovery includes the preliminary identification of cell differentiation (CD) markers that play vital roles in recognitive and adhesive events. Certain leukocyte effectors show characteristics of natural killer (NK) cells that may act differently depending upon their source, whether autogeneic, allogeneic, xenogeneic, or expressed under normal or varying environmental conditions including exposure to xenobiotics. At the level of earthworm evolution, there is apparently a dissociation of phagocytosis from the process of killing by NK-like effectors. There are at least three future challenges. First, it is essential to determine the precise nature of the CD markers with respect to their molecular structure. Second, once their molecular and biochemical characteristics have been defined, the role of these markers in cellular and humoral mechanisms must be clarified in order to define effector cell products and resulting immune responses. Third, there is a need to differentiate between the several lytic factors that have been found in earthworms with respect to molecular structure, and biochemical and functional characterization.

Animals↗
PubMed1999

Histopathological changes after induced injury in leeches.

Cells involved in leech inflammatory responses have been characterized by morphological, histochemical, and immunohistochemical methods. Macrophage-like cells, NK-like cells, and granulocytes migrated shortly after injury by pricking with bacterial lipopolysaccharide. Inflammatory responses increased progressively and provoked cell migration to the body wall and then to wound surfaces. Macrophages, NK cells, and granulocytes display similar features and behavior traits in invertebrates and vertebrates.

Animals↗
PubMed1999

Biochemical characteristics of Eiseniapore, a pore-forming protein in the coelomic fluid of earthworms.

The cytolytic protein Eiseniapore (38 kDa) from coelomic fluid of the earthworm Eisenia fetida functionally requires sphingomyelin as revealed by using mammalian erythrocytes and phospholipid vesicles. The effects of ions, glycoproteins and phospholipids were investigated for the two-step Eiseniapore action mode, binding and pore formation in different assays. Eiseniapore lysis is activated by thiol groups but inhibited by metal ions. Eiseniapore binding to target membranes is inhibited by Eiseniapore-regulating factor, vitronectin, heparin and lysophosphatidylcholine. Ca2+ and Mg2+ were found to be not necessary for membrane binding or lytic activity. Sphingomyelin was essential for Eiseniapore-induced leakage of liposomes. We describe a cytolytic protein/toxin in Eiseniapore which differs from the established classification; it can be activated by thiol groups and is inhibited by sphingomyelin. Electron microscopy of erythrocyte membranes confirmed ring-shaped structures (pores) with a central channel with outer (10 nm) and inner (3 nm) diameters as shown previously [Lange, S., Nüssler, F., Kauschke, E., Lutsch, G., Cooper, E.L. & Herrmann, A. (1997) J. Biol. Chem. 272, 20 884-20 892] using artificial membranes. Functional evidence of pore formation by Eiseniapore was revealed as protection of lysis by carbohydrates occurred at an effective diameter above 3 nm. From these results, we suggest a plausible explanation for the mechanism by which components of the earthworm's immune system destroy non-self components.

Animals↗
PubMed1998

Chemotactic responses of tunicate (Urochordata, Ascidiacea) hemocytes in vitro.

A number of molecules were found to alter the motility of tunicate hemocytes. Bacterial lipopolysaccharide (LPS) significantly enhanced cell mobility relative to non-stimulated controls. Responses to LPS were not directional and so represented chemokinesis. In contrast, checkerboard analyses indicated that two tunicate hemolymph proteins, tunIL1-alpha and -beta, stimulated truly directional chemotaxis by hemocytes. The data suggest that tunIL1 proteins may contribute to defense by altering the localization of immunocompetent cells.

Animals↗
PubMed1997

Participation of a sialic acid-specific lectin from freshwater prawn Macrobrachium rosenbergii hemocytes in the recognition of non-self cells.

Phagocytic activity of circulating hemocytes from freshwater prawns is mediated by a small group of granulocytes. Recognition of nonself cells by these cells seems to be mediated by two independent mechanisms: specific, via O-acetylsialic acid, as well as N-acetylated sugars on recognized cells and a nonspecific one. Both mechanisms show differences in their optimal temperature and time differences for activity. Hemocytes with phagocytic activity interact positively with rabbit IgG raised against the serum lectin. Attempts to elucidate the nature of the membrane-lectin on hemocytes performed by electroimmunotransfer blot assays on cell lysates indicates the presence of two major proteins with a molecular weight of 68-72 kDa. Electron microscopy revealed a regular distribution of lectin on hemocyte plasma membranes. Our results suggest the active participation of a membrane lectin in the recognition of nonself.

Animals↗
PubMed1997

Interaction of earthworm hemolysin with lipid membranes requires sphingolipids.

Lytic activity in the coelomic fluid of earthworm (Eisenia fetida fetida) has been ascribed to eiseniapore, a hemolytic protein of 38 kDa. Since receptors for eiseniapore on target cell membranes are not known, we used lipid vesicles of various composition to determine whether specific lipids may serve as receptors. Lytic activity of eiseniapore was probed by the relief of fluorescence dequenching from the fluorophore 8-aminonaphthalene-1,3, 6-trisulfonic acid originally incorporated into the vesicle lumen as a complex with p-xylene-bis-pyridinium bromide. Hemolysin binds to and disturbs the lipid bilayer only when distinct sphingolipids consisting of a hydrophilic head group as phosphorylcholine or galactosyl as well as the ceramide backbone, e.g. sphingomyelin, are present. Cholesterol enhances eiseniapore lytic activity toward sphingomyelin-containing vesicles probably due to interaction with sphingomyelin. Leakage of vesicles was most efficient when the lipid composition resembled that of the outer leaflet of human erythrocytes. Presumably, an oligomeric protein pore formed by six monomers is responsible for leakage of sphingomyelin-containing vesicles. The secondary structure of eiseniapore did not change upon binding to lipid membranes. The lytic activity of eiseniapore was completely abolished after its denaturation or after preincubation with polyclonal antibodies. Our results suggest that the presence of specific sphingolipids is sufficient to mediate lytic activity of eiseniapore. This action contributes to our understanding of earthworm immune responses.

Animals↗