Several Types Of Leukocytes Research Articles

Hematological Evaluation of Three Common Teleosts in Relation to The Environmental Changes from Trang Province, Thailand.

Hematological evaluation of fish is essential to the assessment of their physiological status. This study describes the morphometric analysis and comparison of blood cell characteristics in Zanarchopterus sp., Gerres filamentosus Cuvier, 1829 and Leiognathus decorus (De Vis, 1884). The species were collected at two locations off the coast of Trang Province, Thailand. A comparative hematological evaluation was made to assess the effects of environmental conditions on the blood of the fish. Ten individuals of each species were collected from a seagrass bed at Libong Island, where human activities are increasing, and from a secluded sandy beach. Their blood samples were analysed using the blood smear technique. Erythrocytes of all the studied fishes were either elliptical or oval. The morphometric data from both locations showed that erythrocytes were of similar size, except for those of Zanarchopterus sp. Fish from both stations showed several types of leukocytes, including neutrophils and lymphocytes.The highest proportion of leukocytes was made up of lymphocytes, followed by neutrophils. However, monocytes were only observed in fish from Libong Island and the erythrocytic nuclei of fish collected from Libong Island were both reniform and lobate. Our results show the potential of hematological evaluation as an early warning signal of environmental impacts on aquatic animals. The determination of baseline parameters could provide a tool for the monitoring of environmental quality.

Semaphorin 4A as a Potential Biomarker for Diagnosis of Systemic Lupus Erythematosus

ABSTRACT Background Semaphorin 4A (Sema4A) is an immunoregulatory molecule that is closely related to the pathogenesis of some autoimmune diseases. However, the relationship between Sema4A and systemic lupus erythematosus (SLE) remains unknown. We therefore aimed to investigate the expression and clinical value of Sema4A in SLE patients. Methods Patients with SLE, rheumatoid arthritis (RA), and healthy controls (HC) were enrolled. The whole blood samples were collected from SLE (83), RA (29) and HC (85), and the expression of Sema4A on several types of leukocytes in peripheral blood was detected by flow cytometry. The serum samples were collected from SLE(77), RA (23) and HC (63), and the concentrations of soluble Sema4A in plasma were detected by ELISA. The diagnostic value of membrane-bound and soluble Sema4A in SLE patients was evaluated using a receiver operating characteristic (ROC) curve. Results The concentration of soluble Sema4A was significantly higher in plasma from SLE patients compared to that from HC and RA patients. In SLE patients, the ratio of CD4+CD11c+ myeloid dendritic cells (mDCs) expressing Sema4A increased significantly, and the levels of soluble Sema4A and membrane-bound Sema4A were negatively correlated with the levels of C3 and C4, respectively. The same result was observed for membrane-bound Sema4A on CD4+CD11c+ mDCs cells. In addition, the level of soluble Sema4A negatively correlated with the concentration of hemoglobin (Hb). Importantly, the expression ratio of membrane-bound Sema4A on CD4+CD11c+ mDCs was positively correlated with systemic lupus erythematosus disease activity index (SLEDAI). Finally, we revealed that soluble and membrane- bound Sema4A had high sensitivity and specificity for diagnosis of SLE, and had a greater ability to distinguish between SLE and RA. Conclusion Sema4A has potential as a new diagnostic biomarker for SLE, and is promising for distinguishing between SLE and RA.

Clothianidin alters leukocyte profiles and elevates measures of oxidative stress in tadpoles of the amphibian, Rana pipiens

Neonicotinoid pesticide use is widespread and highly debated, as evidenced by recent attention received from the public, academics and pesticide regulatory agencies. However, relatively little is known about the physiological effects of neonicotinoid insecticides on aquatic vertebrates. Amphibians (larval stages in particular) are excellent vertebrate bioindicators in aquatic systems due to their risk of exposure and sensitivity to environmental stressors. Previous work with wood frog (Rana sylvatica) tadpoles exposed to formulated products containing thiamethoxam or clothianidin in outdoor mesocosms found significant shifts in leukocyte profiles, suggesting the tadpoles were physiologically stressed. The main objective of the present study was to characterize this stress response further using complementary measures of stress after exposure to clothianidin on northern leopard frogs (Rana pipiens) during their aquatic larval stages. Laboratory static-renewal exposures were conducted over eight weeks with the technical product clothianidin at 0, 0.23, 1, 10 and 100 μg/L, and diquat dibromide at 532 μg/L was used as a positive control. We assessed tadpole leukocyte profiles and measures of oxidative stress as these sub-lethal alterations could affect amphibian fitness. We found changes in several types of leukocytes at 1 and 10 μg/L, suggesting that these tadpoles exhibited signs of mild physiological stress. Clothianidin also induced an oxidative stress response at 0.23, 1 and 100 μg/L. However, we found no differences in survival, growth, development time or hepatosomatic index in frogs exposed to clothianidin. Our study indicates that tadpoles chronically exposed to clothianidin have increased stress responses, but in the absence of concentration-response relationships and effects on whole-organism endpoints, the implications on the overall health and fitness of these changes are unclear.

157 IL-10+ regulatory B cell migration into inflamed skin limits cutaneous inflammation

To maintain skin homeostasis several types of leukocytes exist in or are recruited into the skin. While most of these leukocytes are well characterized, the roles of skin-associated B cells remain largely unknown. Our lab previously demonstrated that a subset of IL-10+ regulatory B cells preferentially migrates into the inflamed skin of mice, a process that required α4β1 integrin. Thus, we hypothesized that B regulatory cells suppress skin inflammation and that impaired migration of these cells into the skin exacerbates skin inflammation. To test this hypothesis, we used two mouse models of skin inflammation: (i) IL-17-dominated psoriasiform skin inflammation induced by imiquimod (IMQ) cream and (ii) IFN-γ-mediated cutaneous delayed contact hypersensitivity elicited by dinitrofluorobenzene (DNFB). Furthermore, we employed a mouse model with a B cell-specific tamoxifen-inducible deletion of α4-integrin (Itga4). Here, we found that inducing the deletion of Itga4 in B cells led to a significant decreased in skin-associated IL-10+ B cells in inflamed skin and included both conventional B2 (CD19+B220highCD43-) and innate-like B1 (CD19+B220-/lowCD43+) cells. The decrease accumulation of IL-10+ regulatory B cells was associated with a significant increase in the clinical and histopathological parameters of skin inflammation in both skin inflammation models. Thus, our data show a crucial function of skin-homing IL-10+ regulatory B cells in the suppression of IL-17- and IFN-γ-dominated skin inflammation; supporting the notion that B regulatory cells are critical players in the cutaneous environment during inflammatory skin diseases.

Production of transgenic cattle by somatic cell nuclear transfer (SCNT) with the human granulocyte colony-stimulation factor (hG-CSF).

The hG-CSF (human Granulocyte Colony-Stimulating Factor) is a growth and stimulation factor capable of inducing the proliferation of bone marrow cells, several types of leukocytes, among other hematopoietic tissue cells. hG-CSF is used in used to treat anomalies that reder a small number of circulating white blood cells, which may compromise the immune defenses of the affected person. For these reasons, the production of hG-CSF in a bioreactor system using the mammary gland of genetic modified animals is a possibility of adding value to the bovine genetic material and reducing the costs of hG-CSF production in pharmaceutical industry. In this study, we aimed the production of transgenic hG-CSF bovine through the lipofection of bovine primary fibroblasts with an hG-CSF expression cassette and cloning these fibroblasts by the somatic cell nuclear transfer (SCNT) technique. The bovine fibroblasts transfected with the hG-CSF cassette presented a stable insertion of this construct into their genome and were efficiently synchronized to G0/G1 cell cycle stage. The transgenic fibroblasts were cloned by SCNT and produced 103 transferred embryos and 2 pregnancies, one of which reached 7 months of gestation.

Leukocyte-Derived Prolactin

The polypeptide hormone prolactin (PRL) is produced by the anterior pituitary and is best known for its role in the development of the mammary gland and lactation. Whereas the pituitary is the most important source of PRL in serum, PRL is also produced elsewhere by for instance the decidua and leukocytes. Although PRL does not seem to be crucial for the normal function of the immune system in mice, it has been proposed that PRL may exert immunomodulating effects under stress or pathological conditions. We summarize the expression of leukocyte subsets expressing PRL and focus on the regulation of PRL expression under normal circumstances and in several disorders. The literature contains plenty of evidence that several types of leukocytes produce bioactive PRL. Furthermore, it is clear that regulation of PRL in leukocytes differs from that in the pituitary and PRL production in leukocytes can be regulated by cytokines and other stimulators of leukocytes. Several groups showed that PRL expression in leukocytes can be increased under pathological conditions and it has been speculated that leukocyte-derived PRL may contribute to hyperprolactinemia and exert immunomodulating effects.

Eosinophil granules function extracellularly as receptor-mediated secretory organelles

Intracellular granules in several types of leukocytes contain preformed proteins whose secretions contribute to immune and inflammatory functions of leukocytes, including eosinophils, cells notably associated with asthma, allergic inflammation, and helminthic infections. Cytokines and chemokines typically elicit extracellular secretion of granule proteins by engaging receptors expressed externally on the plasma membranes of cells, including eosinophils. Eosinophil granules, in addition to being intracellular organelles, are found as intact membrane-bound structures extracellularly in tissue sites of eosinophil-associated diseases. Neither the secretory capacities of cell-free eosinophil granules nor the presence of functional cytokine and chemokine receptors on membranes of leukocyte granules have been recognized. Here, we show that granules of human eosinophils express membrane receptors for a cytokine, IFN-gamma, and G protein-coupled membrane receptors for a chemokine, eotaxin, and that these receptors function by activating signal-transducing pathways within granules to elicit secretion from within granules. Capacities of intracellular granule organelles to function autonomously outside of eosinophils as independent, ligand-responsive, secretion-competent structures constitute a novel postcytolytic mechanism for regulated secretion of eosinophil granule proteins that may contribute to eosinophil-mediated inflammation and immunomodulation.

Heparan Sulfate Proteoglycan–Involving Immunomodulation by Cathelicidin Antimicrobial Peptides LL-37 and PR-39

Antimicrobial peptides, which are categorized by homologous structural motifs, are effector molecules of innate immunity with direct antimicrobial activity and multiple other functions. The most prominent families are defensins and cathelicidins. These peptides are expressed in the granules of several types of leukocytes and in a wide variety of tissue types[1]. Peptide antibiotics of the cathelicidin family are characterized by a highly conserved signal sequence and proregions (“cathelin” = cathepsin L inhibitor), but show substantial heterogeneity in the carboxy-terminal domain that encodes the mature peptide[2,3,4]. Based on amino acid sequences, cathelicidin peptides are categorized in (a) linear, a-helical peptides without cysteines, e.g., LL-37/human cathelicidin antimicrobial peptide (hCAP)-18 from human; (b) peptides with an even number of cysteines linked by disulfide bridges, e.g., protegrins (porcine cathelicidin peptides); and (c) peptides with an unusually high proportion of one or two amino acids, e.g., PR-39 from porcine leukocytes (Table 1).

Immune Regulatory Mechanisms Influence Early Pathology in Spinal Cord Injury and in Spontaneous Autoimmune Encephalomyelitis

Injuries to the central nervous system (CNS) trigger an inflammatory reaction with potentially devastating consequences. In this report we compared the characteristics of the inflammatory response on spinal cord injury (SCI) caused by a stab wound between the T7 and T9 vertebrae and spontaneous experimental autoimmune encephalomyelitis (EAE). SCI and EAE were compared in two types of myelin basic protein Ac1-11-specific T-cell receptor transgenic mice: T/R+ mice harbor regulatory T cells, and T/R- mice lack regulatory T cells. Our results show that 8 days after SCI, T/R- mice developed a strong T-cell infiltrate in the spinal cord, with remarkable down-modulation of CD4 expression that was accompanied by a local increase in Mac-3+ and F4/80+ reactivity and diffuse local and distal astrogliosis. In contrast, T/R+ mice exhibited a modest increase in CD4+ cells localized to the site of injury, without CD4 down-modulation; focal astrogliosis was restricted to the site of the lesion, although Mac-3+ and F4/80+ cells were also present. Similarly to T/R- mice that underwent SCI, T cells displaying down-modulated CD4 expression were found in the CNS of older T/R- mice afflicted by spontaneous EAE. Overall, our results suggest that common mechanisms regulate T-cell accumulation in CNS lesions of different causes, such as mechanic lesion or autoimmune-mediated damage.

Immunology of the mammary gland

The mammary gland is an organ of specific structure whose elementary task is to supply offspring with nutritive and other biologically active substances during the first weeks, or, depending on the species, the first months of life. This prolongs the period of close contact between the mother and her young, which is necessary for their regular growth. Most mammal offspring are born with physiological agammaglobulinaemia, because of the specific structure of the placenta, so that they receive the first specific protection against pathogenic microorganisms through colostrum. Furthermore, this gland is in direct contact with the outer environment through the secretary ducts, so that there are great possibilities for the occurrence of infections. It is therefore necessary to secure protective mechanisms which would prevent such infections. It is clear that there is a distinct connection between the immunological system and the mammary gland, and that link is the central topic of this paper. It presents the basic mechanisms of mammary gland defense which are divided into two categories: nonspecific (innate) and specific immune response. The mammary gland secretion contains several types of leukocytes, such as lymphocytes, macrophages, and neutrophiles, as well as 2% epithelial cells. On the average, there are 0.2 x 106 somatic cells in one mililiter of milk. Macrophages account for most of these (58%), as well as lymphocytes (28%), while a smaller number of somatic cells (12%) are polymorphonuclears (PMN). The paper considers the characteristics and main functions of these cell types.

The immunologic basis for the treatment of psoriasis with new biologic agents

Psoriasis vulgaris is the most prevalent T-cell-mediated inflammatory disease in humans. The pathogenesis of psoriasis is linked to activation of several types of leukocytes that control cellular immunity and to a T-cell-dependent inflammatory process in skin that accelerates the growth of epidermal and vascular cells in psoriasis lesions. Critical steps in immunologic activation include Langerhans cell maturation (activation), T-cell activation, differentiation and expansion of type 1 T cells, selective trafficking of activated T cells to skin, and induction of an inflammatory cytokine and chemokine cascade in skin lesions. In turn, each of these steps offers an opportunity for intervention with engineered biologic therapeutics. (J Am Acad Dermatol 2002;46:1-23.) Learning objective: By reading this article, the participant will be able to understand molecules involved in regulating immune responses and the ways in which rationally designed biologic agents may be used to intercept pathologic immune activation.

Follicular dendritic cells and dissemination of Creutzfeldt-Jakob disease.

The contribution of immune system cells to the propagation of transmissible encephalopathies is not well understood. To determine how follicular dendritic cells (FDC) may act, we challenged lymphotoxin beta null and wild-type (wt) controls with a Creutzfeldt-Jakob disease (CJD) agent. There was only a small difference in incubation time to clinical disease even after peripheral challenge with low infectious doses (31 in a total of 410 days). Brain pathology with extensive microglial infiltration, identified by keratan sulfate, as well as astrocytic hypertrophy, was also equivalent in all groups despite the fact that null mice had neither FDC nor splenic metallophilic macrophages that filter particulate antigen. Because FDC accumulate pathologic prion protein (PrP) in infected but not wt mice, we studied the cellular distribution of PrP by confocal microscopy. The majority of pathologic PrP collected on the plasma membrane of FDC, as identified by the Ca(+2)-binding protein S100A. This surface distribution suggested that agent aggregated with pathologic PrP might spread by cell-to-cell contacts. While several types of leukocytes may be involved in agent dissemination, cells of myeloid lineage were found to be infectious. Moreover, perivascular tracks of microglia and abnormal PrP after intraperitoneal inoculation were consistent with hematogenous spread. In summary, FDC are not required for CJD agent spread from the periphery, although FDC may enhance spread through surface accumulation of pathologic PrP. While it is still not clear where the infectious agent replicates, macrophages can sequester appreciable levels of infectivity and hence act as reservoirs for prolonged latent infection.

Use of hydroethidine and flow cytometry to assess the effects of leukocytes on the malarial parasite Plasmodium falciparum.

Flow cytometry was evaluated as a method of assessing in vitro the effects of leukocytes on blood-stage Plasmodium falciparum. Hydroethidine is converted by metabolizing cells to ethidium, a nucleic acid fluorochrome. After incubation with hydroethidine, viable and dead leukocytes and parasitized and uninfected erthrocytes could all be identified on the basis of fluorescence intensity and size. Leukocytes can therefore be eliminated from further analysis; this allows assessment, at any parasite developmental stage, of the level of parasitemia within erythrocytes in the presence of any of several types of leukocytes. Whether leukocytes actually kill intraerythrocytic parasites can therefore be determined and the level of cytotoxicity can be assessed. The ability of leukocytes to prevent merozoites from invading new erythrocytes, i.e., inhibition of parasite invasion, can also be assessed by this method. When erythrocytes containing schizont-stage parasites were cocultured with different leukocyte populations and the level of parasitemia was determined after merozoite release and invasion, only cultures containing gamma delta T cells inhibited parasite invasion. The different blood-stage forms of the parasite vary in nucleic acid content, which allows each of the developmental stages to be distinguished by flow cytometry; this permits assessment of changes in parasite development in the presence of leukocytes. Monocyte-derived macrophages (MDMs) appeared to have an effect on parasite development. In this instance, when erythrocytes containing ring-form parasites were cocultured with MDMs and harvested 24 h later, the parasites in cultures containing MDMs were at the late schizont stage, whereas parasites in control cultures were early trophozoites; this finding suggests that MDMs accelerate parasite development. Together, these results indicate that flow cytometry is potentially useful for measuring the following effects mediated by leukocytes: (i) level of cytotoxicity, (ii) changes in parasite development, and (iii) inhibition of parasite invasion.

Gorging Response of Culicine Mosquitoes (Diptera: Culicidae) to Blood Fractions

Although anopheline mosquitoes will ingest plasma without blood cells, Culex spp. and Aedes spp. require the phagostimulatory effect of blood cells; this effect can be duplicated by the addition of adenine-nucleotides to plasma. Because activation of platelets released ADP and ATP into the plasma, they were suspected as the major source of the phagostimulant. This paper describes quantitatively the role of platelets in ingestion by Aedes aegypti (L.) and Culex univittatus Theobald. We found that about 10(6)/mm3 inactivated platelets are required to induce engorgement by 80-90% of the mosquitoes of both species. Thrombin activation of the platelets reduced the effective dose to < 2 x 10(4)/mm3. Other blood fractions also were tested as possible sources of stimulation. A series of washed red blood cells (RBC) dilutions was tested; 5 x 10(5) RBC/mm3 were required to induce 90% engorgement. Several types of leukocytes derived from blood by standard methods also induced engorgement at their physiological concentrations. Macrophages and cultured lymphocytes that do not contain any platelets induced gorging in Cx. univittatus, but not in Ae. aegypti. Because RBC and leukocytes do not release nucleotides unless broken, we suggest that their phagostimulatory effect is due to platelet contamination, which invariably occurs during standard methods of blood fractionation.

Activation of mechanical responses in leukocytes

Different kinds of leukocytes undergo cytoskeleton-dependent mechanical responses associated with their specific physiological functions. We have investigated cellular stiffening of several types of leukocytes using a method which measures the force resisting cellular indentation. We have found that lymphocytes stiffen in response to crosslinking cell surface antigens in a process associated with the much studied capping and patching processes. Further studies of myosin-deficient mutants of the ameba Dictyostelium discoideum suggest that this stiffening process results from a myosin dependent contractile process. Rat basophilic leukemia cells and pancreatic islet cells stiffen when triggered to secrete. The function of these cytoskeleton dependent processes is now unknown, but, at least in the islet cells, may be related to a regulation of the rate of secretion. Primary neutrophils stiffen in response to the chemotactic agent, fMet-Leu-Phe. This stiffening may be responsible for retention of these cells in the pulmonary microcirculation during response to inflammation. These observations pose the challenge of determining the structural basis, mechanism, and physiological function of each of these cellular responses.

Regulation of human leukocyte function by lipoxygenase products of arachidonic acid.

Oxygenation of arachidonic acid via the 5- and 15-lipoxygenase pathways yields a variety of products, including monohydroxyeicosatetraenoic acids (HETEs) and more complex products termed leukotrienes. Of these products leukotriene B4 (LTB4) is the most potent modulator of PMN function. Nanomolar concentrations of LTB4 elicit PMN chemotaxis both in vitro and in vivo, and 3- to 10-fold higher concentrations of LTB4 promote lysosomal degranulation, PMN leukocyte aggregation, and adherence, in addition to enhancing the expression of neutrophil and eosinophil C3b receptors. The PMN-directed effects of LTB4 are mediated by binding to a distinct subset of PMN receptors. LTB4 is bound stereospecifically by 26,000-40,000 receptors per PMN, and the receptors have a KD of 10.8-13.9 nM. The receptor for LTB4 may also mediate PMN activation by other 5-lipoxygenase products, as the capacity of these products to activate PMN leukocyte function correlates with their capacity to compete with LTB4 for binding by the receptor. Several observations suggest that LTB4 may function both as an extracellular mediator of PMN function and as a critical intracellular intermediate in cellular activation by other agents. These observations include the localization of the LTB4 receptor in neutrophil granules, the dependence of PMN activation by different stimuli on the lipoxygenation of arachidonic acid, and the capacity of LTB4 to promote calcium flux in liposomes. Arachidonic acid oxygenation products have complex effects on T-lymphocyte function. LTB4 enhances T-lymphocyte chemokinesis and inhibits the generation of lymphokines and cellular transformation. LTC4 inhibits the generation of lymphokines without affecting transformation or chemokinesis. 5-HETE enhances and 15-HETE inhibits chemokinesis, whereas these products have no effects on the other lymphocyte functions. Thus, the lipoxygenase products of arachidonic acid are potent modulators of T-lymphocyte and PMN-function; these products may function both as extracellular mediators and as intracellular intermediates in cellular activation.

Antitumor effect of tuftsin.

Tuftsin, a physiological tetrapeptide derived from the Fc region of leukophilic IgG possesses a variety of immunopotentiating properties including the ability to act as an immunotherapeutic agent against the experimental tumors, L1210 leukemia and Cloudman S-91 melanoma. Although the mechanism of action of tuftsin in vivo is not known, several types of leukocytes have been shown to become cytotoxic effector cells following activation with tuftsin. These cells presently include macrophages, natural killer cells, and granulocytes. The possibility that tuftsin can also activate other types of effector cells have not been ruled out. We feel this small peptide has a high potential (largely unrecognized) as an antitumor immunopotentiating agent. It is naturally occurring in man and appears to be relatively non-toxic. Its exact sequence (Thr-lys-Pro-Arg) is known and it can be chemically synthesized. Methods are also available to monitor the levels of tuftsin in body fluids. These properties along with its ability to control infectious disease make this agent one of the more promising immunopotentiators.