Role Of Hemocytes Research Articles

Role of neuronal and non-neuronal acetylcholine signaling in Drosophila humoral immunity

Acetylcholine (ACh) is one the major neurotransmitters in insects, whose role in mediating synaptic interactions between neurons in the central nervous system is well characterized. It also plays largely unexplored regulatory functions in non-neuronal tissues. Here we demonstrate that ACh signaling is involved in the modulation of the innate immune response of Drosophila melanogaster. Knockdown of ACh synthesis or ACh vesicular transport in neurons reduced the activation of drosomycin (drs), a gene encoding an antimicrobial peptide, in adult flies infected with a Gram-positive bacterium. drs transcription was similarly affected in Drosophila α7 nicotinic acetylcholine receptor, nAChRalpha7 (Dα7) mutants, as well as in flies expressing in the nervous system a dominant negative form (Dα7DN) of this specific receptor subunit. Interestingly, Dα7DN elicited a comparable response when it was expressed in non-neuronal tissues and even when it was specifically produced in the hemocytes. Consistently, full activation of the drs gene required Dα7 expression in these cells. Moreover, knockdown of ACh synthesis in non-neuronal cells affected drs expression. Overall, these findings uncover neural and non-neural cholinergic signals that modulate insect immune defenses and shed light on the role of hemocytes in the regulation of the humoral immune response.

Characterization and mode of action analysis of black soldier fly (Hermetia illucens) larva-derived hemocytes.

With the growing importance of the black soldier fly (Hermetia illucens) for both sustainable food production and waste management as well as for science, a great demand of understanding its immune system arises. Here, we present the first description of the circulating larval hemocytes with special emphasis on uptake of microorganisms and distinguishing hemocyte types. With histological, zymographic, and cytometric methods and with a set of hemocyte binding lectins and antibodies, the hemocytes of H. illucens are identified as plasmatocytes, crystal cells, and putative prohemocytes. Total hemocyte counts (THC) are determined, and methods for THC determination are compared. Approximately 1100 hemocytes per microliter hemolymph are present in naive animals, while hemocyte density decreases dramatically shortly after wounding, indicating a role of hemocytes in response to wounding (and immune response in general). The determination of the relative abundance of each hemocyte type (differential hemocyte count, DHC) revealed that plasmatocytes are highly abundant, whereas prohemocytes and crystal cells make up only a small percentage of the circulating cells. Plasmatocytes are not only the most abundant but also the professional phagocytes in H. illucens. They rapidly engulf and take up bacteria both in vivo and in vitro, indicating a very potent cellular defense against invading pathogens. Larger bioparticles such as yeasts are also removed from circulation by phagocytosis, but slower than bacteria. This is the first analysis of the potent cellular immune response in the black soldier fly, and a first toolbox that helps to identify hemocyte (types) is presented.

English

Haemocytes in insects mediates the cellular responses like phagocytosis, encapsulation and clotting which signifies the immunological functions of any insect. In the domesticated variety of silkworm B.mori five types of haemocytes have been identified in all the stages. The most abundant cell was found to be plasmatocytes followed by Granular cells. Spherule cells and Oenocytoids were found to less during IV and V instars and least or absent in all other stages. Haemocytes found be to fluctuating before and after spinning. Plasmatocytes and granular cells decreased gradually with respect to number of days in pupa. The total haemocytic count increases gradually in the silkworm larval stages and found to be maximum at the last instar and least during adult stages where the role of haemocytes is not required because they die after laying eggs.

The Role of Hemocytes in the Hawaiian Bobtail Squid, Euprymna scolopes: A Model Organism for Studying Beneficial Host-Microbe Interactions.

Most, if not all, animals engage in associations with bacterial symbionts. Understanding the mechanisms by which host immune systems and beneficial bacteria communicate is a fundamental question in the fields of immunology and symbiosis. The Hawaiian bobtail squid (Euprymna scolopes) engages in two known symbioses; a binary relationship with the light organ symbiont Vibrio fischeri, and a bacterial consortium within a specialized organ of the female reproductive system, the accessory nidamental gland (ANG). E. scolopes has a well-developed circulatory system that allows immune cells (hemocytes) to migrate into tissues, including the light organ and ANG. In the association with V. fischeri, hemocytes are thought to have a number of roles in the management of symbiosis, including the recognition of non-symbiotic bacteria and the contribution of chitin as a nutrient source for V. fischeri. Hemocytes are hypothesized to recognize bacteria through interactions between pattern recognition receptors and microbe-associated molecular patterns. Colonization by V. fischeri has been shown to affect the bacteria-binding behavior, gene expression, and proteome of hemocytes, indicating that the symbiont can modulate host immune function. In the ANG, hemocytes have also been observed interacting with the residing bacterial community. As a model host, E. scolopes offers a unique opportunity to study how the innate immune system interacts with both a binary and consortial symbiosis. This mini review will recapitulate what is known about the role of hemocytes in the light organ association and offer future directions for understanding how these immune cells interact with multiple types of symbioses.

The Role of Hemocytes in Anopheles gambiae Antiplasmodial Immunity

Hemocytes synthesize key components of the mosquito complement-like system, but their role in the activation of antiplasmodial responses has not been established. The effect of activating Toll signaling in hemocytes on Plasmodium survival was investigated by transferring hemocytes or cell-free hemolymph from donor mosquitoes in which the suppressor cactus was silenced. These transfers greatly enhanced antiplasmodial immunity, indicating that hemocytes are active players in the activation of the complement-like system, through an effector/effectors regulated by the Toll pathway. A comparative analysis of hemocyte populations between susceptible G3 and the refractory L3-5 Anopheles gambiae mosquito strains did not reveal significant differences under basal conditions or in response to Plasmodium berghei infection. The response of susceptible mosquitoes to different Plasmodium species revealed similar kinetics following infection with P. berghei,P. yoelii or P. falciparum, but the strength of the priming response was stronger in less compatible mosquito-parasite pairs. The Toll, Imd,STAT or JNK signaling cascades were not essential for the production of the hemocyte differentiation factor (HDF) in response to P. berghei infection, but disruption of Toll, STAT or JNK abolished hemocyte differentiation in response to HDF. We conclude that hemocytes are key mediators of A. gambiae antiplasmodial responses.

<i>In vitro</i> Studies on Cellular Mediated Immune Response in Haemocytes of Crab— <i>Episesarma tetragonum</i>

Studying the biological interaction with the invertebrate immune system is attractive for the advancement of a basic knowledge on invertebrate and vertebrate general immunity, because it offers various possible alternatives for disease management. The invertebrate immune system involves encapsulation, nodule formation and phagocytosis process by haemocytes (cellular mediated). In the present study, we investigated the morphology of the haemocytes of Episesarma tetragonum , their role in cellular mediated immune responses such as encapsulation, phagocytosis and viability assays. Encapsulation response of haemocytes is tested with the CM cellulose, Sepharose CL-6B and DEAE Sepharose 6CLB beads. DEAE Sepharose 6CLB beads showed (80%) good cellular response. Phagocytic activity of haemocytes showed pathogen engulfing activity against the microbial patterns (N-acteyl muramic acid (NAM), N-acetyl glutamic acid (NAG), lipopolysaccharide, β-1,3-glucans, chitins, laminarins, zymosan and peptidoglycans) present on the surface of the pathogens. Phagocytic activities of the haemocytes were decreased when increasing the incubation time. Viability of haemocytes was decreased when the time intervals of absorption of tryphan blue were increased. The present investigation enlightens the role of hemocytes in the cellular mediated immune responses of crustacean crab including all invertebrates.

Immune parameters of QX-resistant and wild caught Saccostrea glomerata hemocytes in relation to Marteilia sydneyi infection

Sydney rock oysters (SRO) Saccostrea glomerata suffer mass mortalities during summer and autumn as a result of infection by a protozoan parasite Marteilia sydneyi (QX disease). Mass selected disease resistant (QXR) lines have been used with some success in affected estuaries in recent years, with resistance attributed to oxidative defense systems. However, the role of hemocytes in resistance to QX by SRO has not been fully explored. In the present study, fifty QXR and fifty wild caught (WC) oysters were collected from a lease at Pimpama River during a QX outbreak in January 2011. Hemocytes characteristics (type, morphology) and functions (mortality, phagocytosis and oxidative activity) from both oyster lines were analyzed by flow cytometry in the context of infection intensity and parasite viability (determined histologically). Amongst the QXR oysters, 20% were diseased containing viable parasite, 74% had killed M. sydneyi and 6% were uninfected. In contrast, 86% of WC oysters were diseased, 2% had killed M. sydneyi and 12% were healthy. Significant differences in hemocyte number and physiology between the two oyster lines were found (ANOVA). Phagocytosis rate and the mean oxidative activity per cell were similar between both oyster lines. Higher numbers of infiltrating and circulating hemocytes, higher percentage of circulating granulocytes, their higher size and complexity in QXR oysters, and the production of reactive oxygen species were associated with the ability to kill the parasite. High abundance of M. sydneyi in the digestive tubule epithelium of both oyster lines implied inability to kill the parasite at the beginning of the infection. However, QXR oysters had the ability to kill M. sydneyi at the stage of sporangiosorae in the epithelium of digestive tubules. The similar phagocytic ability of hemocytes from both oyster lines, the size of the parasite at this infection stage, and its localization suggested that encapsulation is likely to be the main process involved in the eradication of M. sydneyi by QXR oysters.

The role of haemocytes from the crab Carcinus aestuarii (Crustacea, Decapoda) in immune responses: A first survey

For the first time, a functional study of haemocytes from the crab Carcinus aestuarii was performed in order to evaluate their involvement in immune responses. Total haemocyte count (THC), phagocytosis, haemolymph opsonisation properties, hydrolytic and oxidative enzyme activities, and production of intracellular superoxide anion were evaluated. A great variability in THC was recorded among individuals, and haemocyte mean number was 6.4 (×10 6) cells/ml haemolymph. Although only hyalinocytes were able to phagocytose yeast cells or Zymosan, phagocytic index was low (3%) and did not increase significantly (4%) after pre-incubation of yeast and Zymosan in cell-free haemolymph, suggesting that haemolymph did not have opsonising properties. All haemocyte types produced superoxide anion, whereas only granulocytes were positive to the hydrolytic enzymes assayed. In addition, only granulocytes were positive to phenoloxidase activity. Both Petri dish and spectrophotometric assays revealed a very low lysozyme-like activity in cell-free haemolymph (CFH) and haemocyte lysate (HL), although enzyme activity was higher in CFH than in HL. Interestingly, normalisation of data as to total protein content in CFH and HL resulted in an opposite situation, lysozyme-like activity being higher in HL than in CFH. This demonstrated that haemolymph of C. aestuarii has a high quantity of total proteins, functional properties of which need to be better investigated in future studies. Overall, the results obtained in the present study indicated that C. aestuarii haemocytes are not very active phagocytic cells, but they are more active in terms of both hydrolytic and oxidative enzyme activities and superoxide anion production.

The role of hemocytes, serine protease inhibitors and pathogen-associated patterns in prophenoloxidase activation in the desert locust, Schistocerca gregaria

The prophenoloxidase-activating system is an important component of the innate immune response of insects, involved in wound healing and melanotic encapsulation. In this paper we show that in the desert locust, Schistocerca gregaria, hemocytes, challenged with microbial elicitors, are indispensable for the limited proteolytic activation of prophenoloxidase (proPO) in plasma. In addition, we assessed the influence of serine protease inhibitors on the induction of PO-activity in plasma. While soybean Bowman–Birk inhibitor (SBBI) inhibited the PO activation by laminarin-treated hemocytes, the endogenous pacifastin-related inhibitors, SGPI-1 ( S. gregaria pacifastin-related inhibitor-1) and SGPI-2 did not affect the PO-activity under similar conditions. On the other hand, real-time PCR analysis revealed that the transcripts, encoding SGPI-1–3, were more abundant in the fat body of immune challenged animals, as compared to control animals.

Mechanisms of heavy-metal sequestration and detoxification in crustaceans: a review.

This review is an update of information recently obtained about the physiological, cellular, and molecular mechanisms used by crustacean organ systems to regulate and detoxify environmental heavy metals. It uses the American lobster, Homarus americanus, and other decapod crustaceans as model organisms whose cellular detoxification processes may be widespread among both invertebrates and vertebrates alike. The focus of this review is the decapod hepatopancreas and its complement of metallothioneins, membrane metal transport proteins, and vacuolar sequestration mechanisms, although comparative remarks about potential detoxifying roles of gills, integument, and kidneys are included. Information is presented about the individual roles of hepatopancreatic mitochondria, lysosomes, and endoplasmic reticula in metal sequestration and detoxification. Current working models for the involvement of mitochondrial and endoplasmic reticulum calcium-transport proteins in metal removal from the cytoplasm and the inhibitory interactions between the metals and calcium are included. In addition, copper transport proteins and V-ATPases associated with lysosomal membranes are suggested as possible sequestration processes in these organelles. Together with several possible cytoplasmic divalent and trivalent anions such as sulfate, oxalate, or phosphate, accumulations of metals in lysosomes and their complexation into detoxifying precipitation granules may be regulated by variations in lysosomal pH brought about by bafilomycin-sensitive proton ATPases. Efflux processes for metal transport from hepatopancreatic epithelial cells to the hemolymph are described, as are the possible roles of hemocytes as metal sinks. While some of the cellular processes for isolating heavy metals from general circulation occur in the hepatopancreas and are beginning to be understood, very little is currently known about the roles of the gills, integument, and kidneys in metal regulation. Therefore, much remains to be clarified about the organs and mechanisms involved in metal homeostasis in decapod crustaceans.

Preliminary study on haemocyte response to white spot syndrome virus infection in black tiger shrimp Penaeus monodon.

White spot syndrome virus (WSSV) has been a major cause of shrimp mortality in aquaculture in the past decade. In contrast to extensive studies on the morphology and genome structure of the virus, little work has been done on the defence reaction of the host after WSSV infection. Therefore, we examined the haemocyte response to experimental WSSV infection in the black tiger shrimp Penaeus monodon. Haemolymph sampling and histology showed a significant decline in free, circulating haemocytes after WSSV infection. A combination of in situ hybridisation with a specific DNA probe for WSSV and immuno-histochemistry with a specific antibody against haemocyte granules in tissue sections indicated that haemocytes left the circulation and migrated to tissues where many virus-infected cells were present. However, no subsequent haemocyte response to the virus-infected cells was detected. The number of granular cells decreased in the haematopoietic tissue of infected shrimp. In addition, a fibrous-like immuno-reactive layer appears in the outer stromal matrix of tubule walls in the lymphoid organ of infected shrimp. The role of haemocytes in shrimp defence after viral infection is discussed.

NUCLEOPOLYHEDROVIRUS PATHOLOGY IN SPRUCE BUDWORM LARVAE

AbstractChoristoneura fumiferana (Clemens) (Lepidoptera: Tortricidae) multiple nucleopolyhedrovirus (CfMNPV) expressing green fluorescent protein was used to study aspects of nucleopolyhedrovirus infection in the spruce budworm. The temporal and spatial distribution of fluorescence indicated that the virus infected the midgut, entered the tracheal system, and traveled to the epidermis, fat body, and muscles. In contrast to Autographa californica (Speyer) (Lepidoptera: Noctuidae) multiple nucleopolyhedrovirus (AcMNPV) infection, hemocytes from infected C. fumiferana did not exhibit fluorescence until after CfMNPV had passed from the midgut into the tracheae. Therefore the role of hemocytes may be limited during CfMNPV infection. Also the fluorescence pattern spread from the tracheolar cells to tracheal epithelial cells throughout the tracheal system. Our results indicate that the temporal and spatial events involved in CfMNPV infection of C. fumiferana larvae are consistent with those observed in other lepidopteran hosts infected with AcMNPV Minor deviations between these two systems may be attributed to differences in virulence, infection rate, and possibly host range of the virus.

DP 4 Roles of hemocytes in allorejection in Botryllus scalaris and Symplegma reptans (colonial ascidians)

DP 4 Roles of hemocytes in allorejection in Botryllus scalaris and Symplegma reptans (colonial ascidians)

Adoptive transfer of alloimmune memory in the solitary tunicate,Styela plicata

The role of hemocytes in histocompatibility responses of the tunicate, Styela plicata, was tested by adoptive transfer. Hemocytes were transferred to naive hosts from compatible individuals that had been presensitized to allogeneic tissue. Forty-four percent of the hosts that received presensitized hemocytes were capable of eliminating subsequent allografts more rapidly than control tunicates injected with saline, plasma, or hemocytes that had not been sensitized to the original graft donor. This adoptive transfer of alloimmune memory was specific to the presensitizing allogeneic tissue type. The transfer of hemocyte populations enriched in lymphocyte-like cells by density gradient centrifugation also specifically enhanced alloreactivity. Hemocyte fractions depleted of lymphocyte-like cells were incapable of inducing alloimmune memory. These data confirm that allograft rejection in S. plicata is a cell-mediated response involving lymphocyte-like cells. © 1996 Wiley-Liss, Inc.

Role of Hemocytes in the Propagation of a Microsporidian Infection in Larvae of Galleria mellonella

Role of Hemocytes in the Propagation of a Microsporidian Infection in Larvae of Galleria mellonella

Suppression of the prophenoloxidase cascade in the larval haemolymph of the turnip moth, Agrotis segetum by an entomopathogenic nematode, Steinernema carpocapsae and its symbiotic bacterium

To study the suppression of the prophenoloxidase cascade in the larval haemolymph of the turnip moth, Agrotis segetum by Steinernema carpocapsae and its symbiotic bacterium Xenorhabdus nematophilus, living S. carpocapsae, heat-killed S. carpocapsae, living X. nematophilus, and heat-killed X. nematophilus were injected in four different treatments into final-instar larvae of A. segetum. In all of the treatments, the activation of prophenoloxidase cascade (initiated by laminarin in the insect haemolymph which was drawn 1 h after injection) was inhibited. This inhibition occurred in a dose-dependent manner. On the other hand, injection of non-pathogenic and fungus-feeding nematodes, Aphelenchus avenae did not induce a similar inhibition, and the nematodes were found to be entrapped in nodules within a melanized sheath. Co-injection of Aph. avenae with S. carpocapsae or X. nematophilus suppressed the prophenoloxidase cascade, while Aph. avenae survived in the haemolymph without being trapped by haemocytes. In all haemolymph of larvae into which foreign-bodies were injected, the levels of prophenoloxidase were essentially the same as those from control larvae, when determined by α-chymotrypsin treatment. These results indicate that S. carpocapsae and/or X. nematophilus suppress the prophenoloxidase cascade at an early step and also interfere with the role of haemocytes in self-defence.

Roles of hemocytes in defense mechanisms of the ascidian, Halocynthia roretzi: K. Azumi and H. Yokosawa.Department of Biochemistry, Faculty of Pharmaceutical Sciences, Hokkaido University, Sapporo 060, Japan

Roles of hemocytes in defense mechanisms of the ascidian, Halocynthia roretzi: K. Azumi and H. Yokosawa.Department of Biochemistry, Faculty of Pharmaceutical Sciences, Hokkaido University, Sapporo 060, Japan

Role of hemocytes and plasma on lipid transport in the freshwater mollusc Diplodon delodontus

1. 1. Composition of lipids and fatty acids from plasma and hemocytes separated from freshwater bivalve molluscs Diplodon delodontus were studied. In both hemolymph components phospholipids were the major lipids, but quantitative composition of phospholipids and fatty acids was different in hemocytes and plasma indicating specific functions. 2. 2. When molluscs were fed 14C fatty acids or 14C triacylglycerols, dietary fatty acids were principally transported by plasma and to a lesser extent by hemocytes. Labeled triacylglycerol was only found in hemocytes. 3. 3. When isolated blood cells were incubated with different 14C fatty acids or 14C trioleylglycerol, they fixed the majority of the label but were unable to esterify fatty acids or hydrolyze trioleylglycerol. 4. 4. At least five proteins were separated in plasma, the majority having a molecular weight higher than 200 × 10 3 daltons. 5. 5. Three plasma compounds with lipoprotein characteristics were recognized.

The role of haemocytes in the biosynthesis of protocatechuate in the cockroach colleterial system.

The role of haemocytes in the biosynthesis of protocatechuate in the cockroach colleterial system.