Intracoelomic parasitic Sporozoa in the burrowing spatangoid echinoidEchinocardium cordatum: coelomocyte reaction and formation of brown bodies

Echinocardium cordatum frequently harbours in its coelomic cavity the protozoan parasiteLithocystis schneideri. Motile stages of this gregarine (trophozoites and gamonts) may be surrounded by echinoid coelomocytes which show a peculiar transformation of their shape (i.e. each cell develops a single spine-like extension giving a pincushion aspect to the parasite). Encysted stages of the gregarine (gametocysts and sporocysts) are found mostly within brown bodies. Brown bodies are particular mesothelium-covered formations occurring usually in the coelomic cavity ofE. cordatum. It is suggested that brown bodies naturally originate from detached fragments of mesenteries.     

Identité immunologique et métabolique entre le liquide coelomique,les coelomocytes et les ovocytes de Perinereis cultrifera (Annélide Polychète)

Immunological study of Perinereis cultrifera reveals the existence of an identical antigen in the coelomic fluid of females (oocyte diameter > 140 (μm), in oocytes, and in coelomocytes. This factor is not found in the body fluid of males or young females.

The elution patterns obtained after Sephadex chromatography shows a similar glycoprotein fraction (fraction I) in the coelomic fluid, in coelomocytes, and in soluble oocyte extracts. This fraction includes the main part of the antigenic components of the coelomic fluid.

Gas chromatography reveals that identical monosaccharides are present, albeit in varying proportions, in samples of fraction I obtained from the different sources.

The metabolic interrelationships of coelomocytes, coelomic fluid and oocytes is discussed. Glycoprotein synthesized by coelomocytes may be discharged into the coelomic fluid and contribute to the development of the cortical alveoli of the oocytes. No evidence of an involvement of this material in yolk synthesis has been obtained.     

Amassin,an olfactomedin protein,mediates the massive intercellular adhesion of sea urchin coelomocytes

Sea urchins have a fluid-filled body cavity, the coelom, containing four types of immunocytes called coelomocytes. Within minutes after coelomic fluid is removed from the body cavity, a massive cell-cell adhesion of coelomocytes occurs. This event is referred to as clotting. Clotting is thought to be a defense mechanism against loss of coelomic fluid if the body wall is punctured, and it may also function in the cellular encapsulation of foreign material and microbes. Here we show that this intercoelomocyte adhesion is mediated by amassin, a coelomic plasma protein with a relative molecular mass (Mr) of 75 kD. Amassin forms large disulfide-bonded aggregates that adhere coelomocytes to each other. One half of the amassin protein comprises an olfactomedin (OLF) domain. Structural predictions show that amassin and other OLF domain-containing vertebrate proteins share a common architecture. This suggests that other proteins of the OLF family may function in intercellular adhesion. These findings are the first to demonstrate a function for a protein of the OLF family.     

Fine structure of the axial complex of Sphaerechinus granularis (Lam.) (Echinodermata: Echinoidea)

Summary Three regions of the axial complex in Sphaerechinus granularis can be distinguished: 1) The axial organ which protrudes from one side of the axial sinus; the sinus septum which separates the sinus from the body cavity and encloses the stone canal; the pulsating vessel which runs along the inside of the axial organ. 2) The blindly-ending terminal sinus in which the pulsating vessel broadens out to the contractile terminal process. 3) The ampulla of the stone canal which connects the axocoel and water vascular system and which opens out through the madreporite.A single-layered, monociliated coelomic epithelium surrounds all regions of the axial complex. This epithelium contains smooth muscle cells at the contractile areas. Canaliculi, surrounded by basal lamina, are formed through infolding of epithelia; they end blindly in the fluid and connective tissue -matrix of the inner structures.The lacunae of the dorso-ventral mesentery connect the periesophageal and the perianal haemal ring with the axial organ. The axial organ contains many coelomocytes rich in pigment and granules. These coelomocytes are separated into compartments by elastic fibres. Phagocytosis of whole cells and transformational stages of coelomocytes suggest storage and degradation functions. An excretory function via the water vascular system is also suggested.     

Ultrastructure of the Coelomocytes in the Tentacular Coelom of Thysanocardia nigra Ikeda, 1904 (Sipuncula)

Free-floating coelomocytes in the tentacular coelomic cavity of the sipunculan Thysanocardia nigra Ikeda, 1904, were studied using light interference contrast microscopy and scanning and transmission electron microscopy. The following coelomocyte types were distinguished: hemerythrocytes, amoebocytes, and two morphological types of granular cells. No clusters of specialized cells that had been reported to occur in the trunk coelom of Th. nigra were found in the tentacular coelom. The corresponding types of coelomocytes from the tentacular and trunk coelomic cavities were shown to differ in size. These two coeloms are completely separated in sipunculans.     

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.     

Biosynthesis and functions of eicosanoids generated by the coelomocytes of the starfish, Asterias rubens

Eicosanoids are a group of oxygenated fatty acid derivatives formed from C20 polyunsaturated fatty acids, including arachidonic and eicosapentaenoic acids. The potential of the coelomocytes of the starfish, Asterias rubens, to generate eicosanoids through the cyclooxygenase (COX) and lipoxygenase (LOX) pathways was investigated using reverse-phase high performance liquid chromatography, enzyme immunoassay and gas chromatography–mass spectrometry. The principal LOX product was identified as 8-hydroxyeicosatetraenoic acid (8-HETE) with 8-hydroxyeicosapentaenoic acid (8-HEPE) synthesised at significantly lower levels. No classical prostaglandins (PG), such as PGE₂ or PGD₂, were found to be generated by ionophore-challenged coelomocytes. Incubation of coelomocytes with lipopolysaccharides from either Escherichia coli or Salmonella abortus failed to induce an increase in generation of LOX products and the presence of 8-HETE (0–25 μM) had no significant effect on the in vitro phagocytic activity of Asterias coelomocytes. Neither indomethacin (a COX inhibitor) or esculetin (a LOX inhibitor) had any effect on the clearance of the bacterium, Vibrio splendidus, from the coelomic cavity of starfish suggesting that products of these enzymes are not involved in such coelomocyte responses to foreign particles.     

Ultrastructure of Microvillar Coelomocytes from the Trunk Coelom of Thysanocardia nigra Ikeda, 1904 (Sipuncula) from the Sea of Japan

This research is part of a study on the ultrastructure of coelomocytes and cellular complexes from the body cavity of sipunculans. New free-swimming elements called microvillar cells in the trunk coelom of Thysanocardia nigra Ikeda, 1904 are examined using transmission electron microscopy. The cell harbors a giant vesicle filled with a fibrous matrix and rosettes of minute osmiophilous granules. The nucleus is peripheral, and a few cell organelles are situated between the cell membrane and the vesicular membrane. The cell membrane bears numerous microvilli with enlarged apical points. Numerous small microvillar vesicles swimming in the coelomic fluid separate from the microvillar cells. The functional morphology of coelomocytes and cellular complexes is discussed.Original Russian Text Copyright © 2005 by Biologiya Morya, Maiorova, Adrianov.     

Cellular Responses of Congenital Immunity in the Starfish <Emphasis Type="Italic">Asterias rubens</Emphasis>

The work deals with analysis of changes of cellular defense factors in the starfish Asterias rubens in response to injection of human erythrocytes (HE). The number of circulating coelomocytes, dynamics of their production of active oxygen forms, activity of peroxidase, and dynamics of elimination of human hemoglobin from coelomic fluid were estimated before immunization with HE as well as at 6–144 h. The number of coelomocytes was counted in Goryaev chamber, production of active oxygen forms was determined in the test of spontaneous and zymosan-induced reduction of Tetrazolium Nitro Blue, peroxidase activity—in a color enzymatic reaction. Time of human hemoglobin elimination from the coelomic fluid was determined by spectrophotometric method by hemoglobin binding with acetone cyanohydrin with formation of a colored product. It is revealed that injection of human erythrocytes into the starfish Asterias rubens leads to a decrease of the number of coelomocytes in 24–96 h and to an increase of their specific production of active oxygen forms in 96–120 h after the HE injection. In coelomic fluid of Asterias rubens the presence of peroxidase activity is established. The circulation time of human hemoglobin released from erythrocytes in coelomic fluid of these animals does not exceed 24 h. It is suggested that the cellular defense reactions are the major factor of the starfish congenital immunity.__________Translated from Zhurnal Evolyutsionnoi Biokhimii i Fiziologii, Vol. 41, No. 2, 2005, pp. 107–113.Original Russian Text Copyright © 2005 by Kudryavtsev, D’yachkov, Kazakov, Kanaikin, Kharazova, Polevshchikov.     

Expression of SpC3, the sea urchin complement component,in response to lipopolysaccharide

The homologue of the vertebrate complement component C3 that is expressed in the coelomocytes of the purple sea urchin, Strongylocentrotus purpuratus, designated SpC3, was investigated for changes in response to immune challenge or injury. Immunoquiescent animals were used in this study because they have reduced or no detectable SpC3 in their coelomocytes or coelomic fluid (CF). Animals were injected with lipopolysaccharide (LPS) or sterile sea water (SSW, injury control). Changes in the amounts of SpC3 in coelomic fluid and in coelomocytes were then followed over time by Western blots and ELISA. Changes in mRNA from the SpC3 gene (Sp064) were also followed by RT-PCR. Although all animals responded to injury with increased levels of SpC3 in the coelomic fluid, those challenged with LPS had greater amounts of SpC3 in both CF and coelomocytes than those receiving SSW. In most of the animals receiving LPS, initial increases in SpC3 were observed within 1 h post-injection, while the earliest response in the animals receiving SSW was 6 h. The appearance of SpC3 in the coelomocytes was delayed compared to its appearance in CF, and was first detected several days after challenge. Changes in mRNA from the Sp064 gene paralleled the appearance of SpC3 in the coelomic fluid. Increases in the number of coelomocytes per milliliter of CF and in the percentage of coelomocytes that were SpC3+ also occurred after challenge with LPS or in response to injury, with a slightly greater increase in response to LPS. Although the changes in SpC3 were not as great as those identified previously for human C3 expressed in macrophages, the kinetics of the response are similar to that of acute-phase reactants in mammals.     

Functional anatomy of the respiratory system of Branchipolynoe species (Polychaeta, Polynoidae), commensal with Bathymodiolus species (Bivalvia, Mytilidae) from deep-sea hydrothermal vents

 The gills of three species of Branchipolynoe have been studied in order to better understand the morphological and anatomical adaptations of their respiratory system. These Polynoidae live commensally inside the pallial cavity of different species of Bathymodiolus (Mytilidae), found clustered near deep-sea hydrothermal vents and cold seeps, and which harbor chemolithoautotrophic bacteria in their gills. As the mussels exploit hydrothermal fluid, the pallial cavity is perfused with a sulfide-rich hydrothermal water. The gills of Branchipolynoe species are well-developed branched outgrows of the body wall, located on the parapodia, and filled with coelomic fluid. They do not contain blood vessels. Living animals are red, due to the presence of extracellular hemoglobins in the coelom. The gill epidermis is made of supporting cells and a few ciliated cells arranged in longitudinal rows along the branches. Myoepithelial and ciliated cells line the interior of the coelomic cavity which contains the respiratory pigments. Coelomic fluid circulation inside the gills and body cavity is probably facilitated by both the cilia and myoepithelial contractions. The cuticle, the epidermis, and the coelomic epithelium are completely devoid of bacteria. The gill surface areas per unit body weight and the minimum diffusion distances, between external milieu and coelomic hemoglobins, have been calculated and compared with data already obtained on vascular gills of littoral or hydrothermal species of Polychaeta. In Branchipolynoe species, the respiratory surface area is very large, similar to that of a free-living hydrothermal species Alvinella pompejana, and the minimum diffusion distance is short, similar to that of the littoral species Arenicola marina. Although the organization of these coelomic gills in Branchipolynoe species is totally different from that of usual vascular gills, their characteristics represent a unique and effective respiratory system in Polynoidae which has adapted to the hypoxic and sulfide-rich micro-habitat which probably holds in the mantle cavity of vent mussels. In the gill epidermis, numerous secondary and large compound lysosomes are present which might be involved in sulfide detoxification. Accepted: 5 August 1998     

Morphological and ultrastructural characterization of sea urchin immune cells

The free circulating coelomocytes in the coelomic cavity of echinoderms are considered to be immune effectors by phagocytosis, encapsulation, cytotoxicity, and by the production of antimicrobial agents. Although echinoderms (especially sea urchin embryo) have been used as a model organisms in biology, no uniform criteria exist for classification of coelomocytes in echinoderms, and few studies have reported about the biological functions of their coelomocytes. Hence, we study the coelomocytes in the echinoid sea urchin, Paracentrotus lividus, and describe their morphological and ultrastructural features using light and transmission electron microscopes. We classify the coelomocytes of P. lividus into red spherule and colorless spherule cells, small cells, vibratile cells, and phagocytic cells; petaloid and filopodial cells. To our knowledge, this is the first report describing ultrastructural details of the coelomocytes of P. lividus. J. Morphol. 276:583–588, 2015. © 2015 Wiley Periodicals, Inc.     

Coelomocyte Morphology in the Holothurians Apostichopus japonicus (Aspidochirota: Stichopodidae) and Cucumaria japonica (Dendrochirota: Cucumariidae)

The cellular composition of the coelomic fluid of the Far Eastern holothurinans Apostichopus japonicus and Cucumaria japonica was studied using light and transmission electron microscopy and histochemistry. In the coelomic fluid of A. japonicus, the following types of coelomocytes were distinguished: progenitor cells; amoebocytes; vacuolated cells; small (or young) morula cells; morula cells of type I, type II, and type III; crystal cells; and vibratile cells. In the coelomic fluid of C. japonicawere found progenitor cells, amoebocytes, vacuolated cells, morula cells of type I and type II, crystal cells, and hemocytes containing a respiratory pigment. The issue of stem cell type, which gives rise to coelomocytes, is discussed.     

Fine Structure of the Hepatic Sacculations of Glossobalanus minutus (Enteropneusta,Hemichordata)

Abstract The hepatic region of Glossobalanus minutus is characterized by deep foldings of the dorsal side of the gut epithelium which affect the neighbouring tissues and structures: coelomic spaces, musculature and epidermis. The following cell types of the gut epithelium are described: vacuolated cells, undifferentiated cells, two types of mucous cells and two types of granular secretory cells. The nature and function of the different cell types are discussed. Data on the general ciliation and subepithelial nerve plexus of the gut epithelium are also given, with special mention of a possible neuroendocrine secretion towards the subjacent blood spaces. A well-developed blood sinus (gut sinus) lies between the gut and the visceral peritoneum. The ultrastructural features of the gut epithelium and its close association with the blood sinus point to an absorptive function. The coelomic cavity is reduced to a narrow space limited by two peritoneal sheets (visceral and parietal) of myoepithelial nature. Amoebocyte-like cells (coelomocytes) occur free in the coelomic fluid, and muscular, unicellular bridges are attached to both peritoneal walls across the coelomic space. The dorsal epidermis follows the gut foldings and is formed by flat, overlapping cells. The present observations are compared with previous histological, histochemical and ultrastructural data.     

Ultrastructure of coelomic epithelium and coelomocytes of the starfish Asterias rubens L. in norm and after wounding

Samples of coelomic epithelium (CE) and coelomocyte suspension of intact and wounded starfish Asterias rubens L. were studied by electron microscopy. The CE was shown to be composed of three types of cells: flagellar (approximately 60%), secretory (approximately 3%), and myoepithelial (approximately 37%); flagellar and secretory cells form the CE apical surface. Secretory cells are represented by two subtypes, i.e., granular and mucous secretory cells. Myoepithelial cells are located in the basal zone of the epithelium. In 4–5% of cases, adjacent flagellar cells are separated by various sizes of intercellular gaps. These gaps seem to be lacunae left by the flagellar cells after their release into the coelomic cavity. The morphological pattern of the conversion of CE flagellar cells into coelomocytes was characterized. After a moderate wounding used in the present study, no significant structural alterations in the CE organization were revealed. In coelomocyte suspension, small rounded young coelomocytes (approximately 3%) and the larger mature coelomocytes (approximately 97%) were found. On the surface of one of the young coelomocytes, a flagellum was revealed. Surface of the mature coelomocytes forms processes of various size and structure; their cytoplasm contains lysosomes and phagocytic vacuoles of different size. After wounding, a coelomocyte activation was found that consisted of a sharp rise in the number and length of filopodia on their surface, as well as the formation of multicellular aggregates. The complex of ultrastructural data allows it to be suggested that the histogenesis of coelomocytes from CE flagellar cells is a process of cell transdifferentiation.     

The fine structure of coelomocytes in the annelid Enchytraeus fragmentosus

Two types of coelomocytes, the mucocyte and the phagocyte, occur in Enchytraeus fragmentosus. Other free cells observed within the coelomic cavity include chloragogen cells, peritoneal cells and some anucleate granular cells. Three forms of mucocytes occur and are believed to represent developmental stages. The first stage is one in which the mucous droplets are forming in the Golgi region. The second stage is a mature form, and the third stage is one in which the mucous droplets are being released. The phagocytes generally are quite large, and inclusions vary from recognizable portions of chloragogen cells to extremely small, electron-dense cytosomes. The origin of the coelomocytes could not be determined. Probable functions of coelomocytes are discussed.     

Activity and immunodetection of lysozyme in earthworm Dendrobaena veneta (Annelida)

In the present study, lysozyme-like activity against Micrococcus luteus was detected in the coelomic fluid, the extract from coelomocytes, intestine and in the homogenates from cocoons of Dendrobaena veneta. Four hours after immunization with Escherichia coli, the lysozyme activity in the coelomic fluid increased about three times and in the extract of coelomocytes - four times, in comparison to the control. In three cases: of the coelomic fluid, the homogenates from cocoons and the extract from coelomocytes, the antibody against HEWL (hen egg white lysozyme) recognized only one protein with a molecular mass of about 14.4 kDa. In the coelomic fluid, apart from the protein with molecular mass of 14.4 kDa the antibody directed against human lysozyme recognized an additional protein of 22 kDa. Using the bioautography technique after electrophoretic resolution of native proteins in acidic polyacrylamide gels, two lytic zones of M. luteus were observed in the case of the coelomic fluid and three after the analysis of the extract of coelomocytes and the egg homogenates. The results indicated the existence of several forms of lysozyme with a different electric charge in the analyzed D. veneta samples. The highest lysozyme activity in the intestine of D. veneta was observed in the midgut. The antibody directed against human lysozyme indicated a strong positive signal in epidermal and midgut cells of earthworm.     

Ultrastructure of the tentacles of Themiste lageniformis (Sipuncula)

Summary An ultrastructural study of the tentacles of Themiste lageniformis (Sipuncula) was conducted as part of a larger study of head metamorphosis in the species.The oral surface of the tentacles is constructed of a multiciliated, pseudostratified, columnar epithelium while the aboral surface is an unciliated, cuboidal epithelium. Intraepidermal mucous cells lie near the junction of the oral and aboral regions. The basal portion of the epidermal cells is embedded in a thick, collagenous extracellular matrix which contains outer circular muscles, inner longitudinal muscles, the main tentacular nerve and its branches. Three tentacular canals are present and are lined by peritoneum. Hemerythrocytes and coelomocytes flow through the lumen of the canals in a regular pattern.Justification for the designation of the tentacular canals as coelomic rather than vascular is discussed.     

Expression of the starfish complement component C3 gene homolog under the influence of bacterial lipopolysaccharide

A fragment of a homolog of the complement component C3 gene was cloned and sequenced from the starfish Asterias rubens. A phylogenetic analysis of the gene, termed ArC3-like, demonstrated its close similarity to the C3 gene homolog of deuterostome invertebrates. A high level of ArC3-like mRNA expression was observed in circulating cells (coelomocytes), a gut derivate (hepatopancreas), and the male gonad, but not in the stomach, female gonad, and rectal gland of A. rubens. ArC3-like gene expression was detected in all types of starfish coelomocytes, including lymphocyte-like cells and granular and nongranular amebocytes. Bacterial lipopolysaccharide (LPS) injected into the coelomic cavity of starfish increased the ArC3-like gene expression in coelomocytes and the hepatopancreas as compared to the control level (sterile sea water injection). The level of ArC3-like gene expression in coelomocytes in response to LPS reached its maximum 6 h after the injection and decreased to the baseline level 24 h after the injection. In the hepatopancreas, the level of ArC3-like gene expression reached its maximum 6–12 h after the LPS injection stimulation and remained high even 48 h after the injection. A long-term upregulation in response to LPS was demonstrated for the ArC3-like gene.     

Sea urchin coelomic fluid agglutinin mediates coelomocyte adhesion.

The sea urchin Paracentrotus lividus coelomic fluid was found to contain agglutinin which agglutinates animal erythrocytes and promotes adhesion of autologous coelomocytes. Hemagglutinating activity depended upon the presence of calcium ions and was relatively heat-stable. Through a combination of methods including ammonium sulfate precipitation and both size exclusion and ion exchange chromatographies, we purified the anti-rabbit agglutinating factor. The intact agglutinin migrates as a single band with an apparent M(r) of over 200,000. Three distinct protein bands with a calculated M(r) of 174,000, 137,000, and 76,000, respectively were observed under reducing conditions. The purified agglutinin strongly promoted the in vitro adhesion of autologous coelomocytes.