Lead induced modulation of splenic macrophage responses on humoral and cell mediated immunity

The heavy metal lead is an environmental toxic material that can induce pathophysiological changes in many organ systems. Previous studies have shown the effects of lead exposure on immune cells in different experimental animals, however, the mechanism of their influence on the immune system is unclear. We reported that in vivo lead exposure inhibits phagocytosis, nitric oxide release, induces DNA fragmentation suggesting the apoptotic death of the target cell. We have also presented evidence that inhibition of macrophage functional responses implicated alteration of humoral and cell mediated immunity. In vivo exposure to lead acetate alters the phagocytic capacity of splenic macrophages as evident from the reduction of phagocytic index of control from 19,792+/-1385.69 to 8893+/-893 in the treated group. The amount of nitric oxide released by the control cell 2.25+/-0.125 microM is also reduced to 1.9375+/-0.0625 microM upon in vivo lead treatment. Functional integrity of the target cell is also decreased after lead exposure as obtained from the percentage of DNA fragmentation. Control group shows 33.29+/-0.11% of fragmented DNA, which is enhanced to 42.43+/-0.725% following the lead treatment. A greater percentage of DNA fragmentation upon lead treatment probably indicating that the heavy metal induces apoptosis. The humoral immune response is also altered after lead exposure as indicated by the decrease of the antibody titre in control group from 1:2048 to 1:128 in the treated group. From the DTH reaction, it was observed that the mean diameter of swollen foot pad of control mice is 0.329+/-0.15 cm and that of lead treated mice is 0.274+/-0.056 cm. It can, therefore, be suggested that lead inhibits normal functional activities of splenic leukocytes, particularly phagocytosis and also affects the functional integrity of cells by inducing DNA fragmentation. The study may demonstrate the usefulness of investigation of humoral immune system and leukocyte functions as sensitive parameters in detecting the effects of lead toxicity.     

Metabolic and immunological responses associated with in vivo glucocorticoid depletion by adrenalectomy in mature Swiss albino rats

The study is undertaken to determine the effect of adrenal corticosteroid depletion after adrenalectomy on carbohydrate, protein and fat metabolism as well as maturation and functional efficacy of the immunocompetent cells. Beside biochemical and hematological parameters, whether in vivo glucocorticoid depletion has any modulatory effects on splenic macrophage responses to bacterial challenge with regards to intracellular killing, nitric oxide release and cellular integrity, were determined. Major findings of our study indicate that blood glucose, urea and total inorganic phosphate levels showed a time dependent increase in adrenalectomized rats compared to control. Total glycogen content in liver was decreased gradually due to adrenal corticosteroid insufficiency. Hematological parameters like hemoglobin concentration, hematocrit value, total leukocyte count and differential count were also found to increase in the adrenalectomized group with respect to intact group. From the functional study of immunocompetent cells, intracellular killing capacity of splenic macrophages recovered from control and adrenalectomized rats after 10 and 20 days of adrenalectomy showed no significant alteration; however, the function of splenic macrophages recovered from rats after 30 days of adrenalectomy showed altered response. Nitric oxide released from splenic macrophages of adrenalectomized rats was less than that of control animal even after stimulation with lipopolysaccharide. DNA fragmentation assay showed a lesser degree of fragmentation of splenic macrophages obtained from adrenalectomized rats indicating, apoptotic death of cells in this group decreases. Adrenal corticosteroid insufficiency due to adrenalectomy interferes with metabolic and hematopoietic functions and modulates the development and maintenance of normal immunitary status, which in turn influences the inflammatory response.     

Effect of ghrelin administration on phagocytic activity in acute cold-restraint stress exposed rats

Ghrelin, an endogenous ligand for growth hormone secretagogue receptor, was identified in the rat stomach. This peptide acts through nitric oxide (NO) by expressing endothelial nitric oxide synthase (eNOS) and down regulating inducible nitric oxide synthase (iNOS) at its gastroproprotective effect against restraint stress induced damage. Recently the ghrelin receptor has also been detected in peripheral systems including immune tissue. We have investigated the possible effect of ghrelin on phagocytic activity of peritoneal macrophages in acute cold-restraint stress (ACRS) exposed rats. The rats were divided into control, stress and ghrelin groups. In ghrelin groups, single dose and three days consecutive dose of ghrelin (20 microg/kg. i.p.) were applied to rats that were exposed to ACRS for 4 h. 1 ml of saline was injected i.p. after ACRS for 3 consecutive days to the rats of the stress groups. Ghrelin administration reduced the increased phagocytic activity induced by ACRS. We conclude that ghrelin exerts an important role at macrophage phagocytic activity in ACRS exposed rats.     

Chitosan induces different L-arginine metabolic pathways in resting and inflammatory macrophages

Chitosan is a linear polymer of N-acetyl-D-glucosamine and deacetylated glucosamine widely used as a wound-healing accelerator in clinical and veterinary medicine. Chitosan enhances the functions of inflammatory cells such as macrophages (Mphi), inducing the production of cytokines as well as the expression of activation markers, Fc receptors and mannose receptor. In this work we studied the effects of chitosan on the arginine metabolic pathways of both resident and inflammatory (proteose-peptone elicited) rat Mphi. Our results show that low molecular weight (LMW) chitosan activated moderately both the inducible nitric oxide synthase (iNOS) and arginase pathways in resident Mphi. In inflammatory Mphi treated with chitosan instead, the arginase activity was strongly enhanced. Supernatants of chitosan-stimulated Mphi enhanced the proliferation of the rat cell line C6. These findings suggest that the healing activity of chitosan could rely on the enhanced arginase activity observed in a wound-associated inflammatory milieu.     

Surfactant-associated protein A inhibits LPS-induced cytokine and nitric oxide production in vivo

The role of surfactant-associated protein (SP) A in the mediation of pulmonary responses to bacterial lipopolysaccharide (LPS) was assessed in vivo with SP-A gene-targeted [SP-deficient; SP-A(-/-)] and wild-type [SP-A(+/+)] mice. Concentrations of tumor necrosis factor (TNF)-alpha, macrophage inflammatory protein-2, and nitric oxide were determined in recovered bronchoalveolar lavage fluid after intratracheal administration of LPS. SP-A(-/-) mice produced significantly more TNF-alpha and nitric oxide than SP-A(+/+) mice after LPS treatment. Intratracheal administration of human SP-A (1 mg/kg) to SP-A(-/-) mice restored regulation of TNF-alpha, macrophage inflammatory protein-2, and nitric oxide production to that of SP-A(+/+) mice. Other markers of lung injury including bronchoalveolar fluid protein, phospholipid content, and neutrophil numbers were not influenced by SP-A. Data from experiments designed to test possible mechanisms of SP-A-mediated suppression suggest that neither binding of LPS by SP-A nor enhanced LPS clearance are the primary means of inhibition. Our data and others suggest that SP-A acts directly on immune cells to suppress LPS-induced inflammation. These results demonstrate that endogenous or exogenous SP-A inhibits pulmonary LPS-induced cytokine and nitric oxide production in vivo.     

Curcumin enhances non-inflammatory phagocytic activity of RAW264.7 cells

Present study was performed to assess the effect of curcumin treatment on macrophage functions using RAW264.7 cells, a murine macrophage cell line. Phagocytic activity of RAW264.7 cells was enhanced by the treatment with curcumin for 48 h while the nitric oxide synthesis from RAW264.7 cells following lipopolysaccharide exposure was blocked. The incubation of RAW264.7 cells with curcumin dose-dependently inhibited the stimulatory responses of macrophage triggered by lipopolysaccharide; the enhanced secretion of inflammatory cytokines such as TNF-α and IL-1β and the up-regulated expression of surface antigens like CD14 and CD40. Curcumin alone, however, was able to increase the basal level of TNF-α secretion and elevated markedly the expression of CD14 and slightly CD40. The marked enhancement of both phagocytic activity and CD14 was detectable as early as 75 min after curcumin treatment which is the minimum time period required for the phagocytosis and CD14 measurement, suggesting a signaling pathway distinct from that triggered by apoptotic cells. In conclusion, this study elucidates that curcumin treatment enhances the phagocytic activity with blocking nitric oxide synthesis, a scavenger function of macrophages in non-inflammatory condition. In addition, this enhancement of phagocytic activity is triggered directly by the signals from curcumin itself not by apoptotic cells.     

Differential effects of the novel non-peptidic opioid 4-tyrosylamido-6-benzyl-1,2,3,4 tetrahydroquinoline (CGPM-9) on in vitro rat t lymphocyte and macrophage functions

Opioid receptors have been reported on immune cells of several species and shown to subserve effector functions of these cell types. Mu-selective opioid agonists such as morphine are immunosuppressive, whereas certain delta-opioid receptor-selective agonists have been associated with immunopotentiation. We have previously shown that intracerebroventricular administration of the non-peptidic delta-opioid receptor agonists did not alter certain parameters of immunocompetence. In this study, we evaluated the in vitro effects of the novel non-peptidic opioid 4-tyrosylamido-6-benzyl-1,2,3,4 tetrahydroquinoline (CGPM-9) on lymphocyte and macrophage functions. We demonstrated that CGPM-9 enhanced rat thymic lymphocyte proliferative response to concanavalin A (2.85- to 5.5-fold increases), and suppressed LPS-induced nitric oxide (67 to 72 percent reduction) and TNF-alpha production (46 percent reduction) by peritoneal macrophages, compared with untreated control. The mu-opioid receptor selective antagonist CTOP used at equimolar doses, significantly suppressed the effect of CGPM-9 on lymphocyte and macrophage functions (CTOP alone did not show any effect on lymphocyte or macrophage functions). In summary, CGPM-9 activated thymic lymphocyte proliferation and suppressed macrophage functions by acting at mu-opioid receptors. This suggests that opioid receptors on immunocytes may be coupled to different signaling pathways depending on the cell type and effector function being analyzed. The mechanism (s) associated with the differential effect of CGPM-9 on these immune cells remains to be elucidated. The pharmacotherapeutic potential for compounds such as CGPM-9 which potentiate T lymphocyte proliferation and suppress production of macrophage-derived inflammatory cytokines is substantial in research and clinical medicine.     

Trikatu,an herbal compound as immunomodulatory and anti-inflammatory agent in the treatment of rheumatoid arthritis – An experimental study

In the present study, trikatu, an herbal compound was evaluated for its immunomodulatory and anti-inflammatory properties with reference to cell mediated immune responses (delayed type hypersensitivity reaction), humoral immune response (haemagglutination titer and plaque forming assay), macrophage phagocytic index, circulating immune complex and inflammatory mediators in rats. For comparison purposes, indomethacin was used as a reference drug for anti-inflammatory studies. The results obtained in our study showed a significant decrease in cell mediated immune responses, humoral immune responses (haemagglutination titre and plaque forming assay) and macrophage phagocytic index in trikatu treated rats (1000 mg/kg/b.wt.) compared to control animals implying its immunosuppressive property. In addition, significant anti-inflammatory effects were observed in trikatu treated adjuvant induced arthritic rats by a reduction in the levels of circulating immune complexes and inflammatory mediators (TNF-alpha and Interleukin-1beta). Thus, in conclusion, our data suggest that trikatu could be considered as a potential anti-inflammatory agent for treating autoimmune inflammatory disorders like rheumatoid arthritis with immunosuppressive property.     

Radiation effects on oxidative metabolism in young and aged rat alveolar macrophages.

The effect of ionizing radiation on metabolic functions of alveolar macrophages (AM) have been well studied. However, variations associated to age have not been established yet. The aim of this work was to perform a comparative study on irradiated alveolar macrophages from young and aged rat lungs. Cell viability and occurrence of apoptosis as well as production of nitric oxide (NO), generation of superoxide anion (O2*-) and total antioxidant capacity were analyzed in vitro after exposure to gamma-irradiation with 10, 25, 50 and 75 Gy. Cell viability decreased only in the aged population at the higher doses. Morphological features of apoptosis were clearly evidenced in irradiated alveolar macrophages from aged animals although the DNA fragmentation assay for apoptosis showed no differences for either of the populations studied. NO production and total reactive antioxidant potential (TRAP) levels showed a dose-dependent modulation. Low radiation doses inhibited the production of NO and decreased TRAP levels whereas higher doses enhanced the NO production and increased the TRAP levels in both macrophage populations. Generation of O2*- was always higher in the aged population for all the doses assayed. We conclude that in vitro young alveolar macrophages exhibited higher radioresistance over the whole range of doses as compared to the aged macrophage population. Our results show that the aging process markedly affects the radioresistance of phagocytic cells. Therefore, immune defense and inflammatory response of lungs from aged patients should be considered when planning radiotherapy protocols.     

Severe acute respiratory syndrome and the innate immune responses: modulation of effector cell function without productive infection

Severe acute respiratory syndrome (SARS) caused by a novel human coronavirus (CoV), designated SARS-CoV, is a highly contagious respiratory disease with the lungs as a major target. Although the exact mechanism of SARS-CoV pathogenesis remains unknown, an intense, ill-regulated local inflammatory response has been suggested as partially responsible for the devastating lung pathology. We investigated the interaction of SARS-CoV with human macrophages (Mphi) and dendritic cells (DC), two key innate immune cells of the host immune system, by focusing on their susceptibility to viral infection and subsequent responses (e.g., phenotypic maturation, T cell-priming activity, phagocytosis, and cytokine production). We found neither cell to be permissive for SARS-CoV replication. However, incubation of Mphi and DC with live, but not gamma irradiation-inactivated, viruses appeared to better sustain their viability. Also, exposure to infectious SARS-CoV led to the phenotypic and functional maturation of DC, with regard to MHC class II and costimulatory molecule expression, T cell-stimulatory capacity, and cytokine production, respectively. Cytokine production was also observed for Mphi, which were refractory to cell surface phenotypic changes. Strikingly, live SARS-CoV could further prime cell types to respond to a suboptimal dose of bacterial LPS (100 ng/ml), resulting in massive release of IL-6 and IL-12. However, the endocytic capacity (e.g., Ag capture) of Mphi was significantly compromised upon exposure to infectious SARS-CoV. This study is the first demonstration that although SARS-CoV does not productively infect human Mphi or DC, it appears to exert differential effects on Mphi and DC maturation and functions, which might contribute to SARS pathogenesis.     

Macrophage biology in the Anx-A1-/- mouse

Historical data suggested that a soluble protein, since identified as annexin-A1 (Anx-A1) was released from macrophages following glucocorticoid stimulation and could modulate eicosanoid production and other functions of these cells. Here, we review some recent findings using a line of Anx-A1(-/-) mice to explore the impact of Anx-A1 gene deletion on macrophage biology. The absence of Anx-A1 selectively alters phagocytic capacity of rodent resident peritoneal macrophages apparently through changes in surface adhesion molecule expression. Anx-A1 is also apparently important in the tonic down-regulation of other macrophage functions such as COX-2 induction, PGE(2) release and the production of reactive oxygen species.     

细虫草胞外多糖对小鼠腹腔巨噬细胞免疫功能研究

本实验在体外条件下,以人工发酵培养的细虫草胞外多糖OgE、OgE-F1和OgE-F2作用于小鼠腹腔巨噬细胞RAW264.7,通过测定其对巨噬细胞的增殖率、代谢MTT活力、NO分泌和吞噬能力的影响,评价细虫草胞外多糖的免疫调节活性。结果表明,细虫草多糖对巨噬细胞无细胞毒性,且能促进巨噬细胞代谢MTT活力;在0.2mg/mL^1.0mg/mL浓度范围内,多糖呈剂量依赖性的促进巨噬细胞分泌NO水平和吞噬能力。本研究表明,细虫草多糖能有效地增强小鼠巨噬细胞的活性,潜在地可改善小鼠的先天性免疫调节。     

T-cell glucocorticoid receptor is required to suppress COX-2-mediated lethal immune activation

Glucocorticoids, acting through the glucocorticoid receptor, potently modulate immune function and are a mainstay of therapy for treatment of inflammatory conditions, autoimmune diseases, leukemias and lymphomas. Moreover, removal of systemic glucocorticoids, by adrenalectomy in animal models or adrenal insufficiency in humans, has shown that endogenous glucocorticoid production is required for regulation of physiologic immune responses. These effects have been attributed to suppression of cytokines, although the crucial cellular and molecular targets remain unknown. In addition, considerable controversy remains as to whether glucocorticoids are required for thymocyte development. To assess the role of the glucocorticoid receptor in immune system development and function, we generated T-cell-specific glucocorticoid receptor knockout mice. Here we show that the T-cell is a critical cellular target of glucocorticoid receptor signaling, as immune activation in these mice resulted in significant mortality. This lethal activation is rescued by cyclooxygenase-2 (COX-2) inhibition but not steroid administration or cytokine neutralization. These studies indicate that glucocorticoid receptor suppression of COX-2 is crucial for curtailing lethal immune activation, and suggest new therapeutic approaches for regulation of T-cell-mediated inflammatory diseases.     

Age-dependent change in reactive oxygen species and nitric oxide generation by rat alveolar macrophages

Immunosenescence is an age-associated dysregulation of the immune function, which contributes to increased susceptibility to disease in the elderly. Alveolar macrophages (AM) are known phagocytes that generate reactive oxygen species (ROS) and nitric oxide (NO), essential mediators for host defence. We studied phagocytosis, ROS and NO production in AM obtained from young, adult and senescent rats (1-2, 9-12 and 18-24 months old, respectively) after exposure to lipopolysaccharide (LPS, 0.1-10 microg mL(-1)), 12-O-tetradecanoylphorbol 13-acetate (TPA, 0.1 microg mL(-1)) or LPS + TPA in culture. Phagocytosis was significantly lower in control AM from adult rats than in AM from young animals. Nevertheless, AM from adult animals pretreated with LPS exhibited higher phagocytic capacity than AM from younger animals. ROS was identified by the NBT test at single cell level and quantified by automated image analysis. When TPA was added to all three populations, AM from adult and senescent animals responded more than AM from young animals. All LPS-stimulated AM produce more NO than controls. However, NO production increased three-, four- and two-fold in young, adult and senescent animals, respectively. Our results demonstrate that AM from young, adult and senescent animals display differential responsiveness to inflammatory mediators. Therefore, aging processes markedly affect AM metabolic functions and may further compromise the lung immune defence response, increasing adverse long-term health effects.     

Critical roles of the WASP N-terminal domain and Btk in LPS-induced inflammatory response in macrophages

While Wiskott-Aldrich syndrome protein (WASP) plays critical roles in TCR signaling as an adaptor molecule, how it transduces innate immune signals remains to be elucidated. To investigate the roles of WASP in innate immune cells, we established bone marrow-derived macrophage (BMDM) cell lines from WASP15 transgenic (Tg) mice overexpressing the WASP N-terminal region (exons 1-5). Upon LPS stimulation, WASP15 Tg BMDM cell lines produce lower levels of inflammatory cytokines, such as TNF-α, IL-6, and IL-12p40 than the wild-type BMDM cell line. In addition, the production of nitric oxide by WASP15 Tg BMDM cells in response to LPS and IFN-γ was significantly impaired. Furthermore, we uncovered that the WASP N-terminal domain associates with the Src homology (SH) 3 domain of Bruton's tyrosine kinase (Btk). Overexpression of the WASP N-terminal domain diminishes the extent of tyrosine phosphorylation of endogenous WASP in WASP15 Tg BMDM cells, possibly by interfering with the specific binding between endogenous WASP and Btk during LPS signaling. These observations strongly suggest that the interaction between WASP N-terminal domain and Btk plays important roles in the LPS signaling cascade in innate immunity.     

Failure of monocytes of trauma patients to convert to immature dendritic cells is related to preferential macrophage-colony-stimulating factor-driven macrophage differentiation

Following trauma, increased inflammatory monokine activation and depressed APC function can occur simultaneously. These contradictory monocyte (Mphi) dysfunctions could result if postinjury Mphi differentiation preferentially favored inflammatory macrophage (Mac) differentiation over development into the most potent APC, dendritic cells (DC). In this report, Mphi of trauma patients with a depressed MLR induction capacity are, for the first time, shown to be unable to differentiate in vitro to immature CD1a(+) DC under the influence of GM-CSF and IL-4. Trauma patient Mphi that retained MLR-inducing capacity had a nonsignificant reduction in DC differentiation capacity. Only patient Mphi populations with depressed differentiation to immature DC (iDC) demonstrated depressed IL-12 and IL-15 production and a continued reduced MLR induction capacity. Neither increased IL-10 production nor decreased CD11c(+) DC precursor numbers correlated with depressed Mphi-to-DC differentiation. Instead, these patients' APC-dysfunctional Mphi populations had increased expression of inflammatory Mac phenotypes (CD64(+), CD86(low), HLA-DR(low)) and up-regulated secretion of M-CSF. M-CSF combined with IL-6 inhibits Mphi-to-iDC differentiation and promotes Mphi-to-Mac differentiation by down-regulating GM-CSFR expression and increasing DC apoptosis. Both depressed GM-CSFR expression and increased Mphi iDC apoptosis, as well as increased expression of CD126 (IL-6R) and CD115 (M-CSFR), were detected in APC-defective patient Mphi. In vitro addition of anti-M-CSF enhanced the IL-4 plus GM-CSF-induced Mphi-to-DC differentiation of these patients. This suggests that, in trauma patients, enhanced Mphi-to-Mac differentiation with concomitant inhibited iDC development is partially due to increased circulating Mphi sensitivity to and production of M-CSF and contributes to postinjury immunoaberrations.     

Endogenous Thrombospondin-1 Regulates Leukocyte Recruitment and Activation and Accelerates Death from Systemic Candidiasis

Disseminated Candida albicans infection results in high morbidity and mortality despite treatment with existing antifungal drugs. Recent studies suggest that modulating the host immune response can improve survival, but specific host targets for accomplishing this goal remain to be identified. The extracellular matrix protein thrombospondin-1 is released at sites of tissue injury and modulates several immune functions, but its role in C. albicans pathogenesis has not been investigated. Here, we show that mice lacking thrombospondin-1 have an advantage in surviving disseminated candidiasis and more efficiently clear the initial colonization from kidneys despite exhibiting fewer infiltrating leukocytes. By examining local and systemic cytokine responses to C. albicans and other standard inflammatory stimuli, we identify a crucial function of phagocytes in this enhanced resistance. Subcutaneous air pouch and systemic candidiasis models demonstrated that endogenous thrombospondin-1 enhances the early innate immune response against C. albicans and promotes activation of inflammatory macrophages (inducible nitric oxide synthase⁺, IL-6^high, TNF-α^high, IL-10^low), release of the chemokines MIP-2, JE, MIP-1α, and RANTES, and CXCR2-driven polymorphonuclear leukocytes recruitment. However, thrombospondin-1 inhibited the phagocytic capacity of inflammatory leukocytes in vivo and in vitro, resulting in increased fungal burden in the kidney and increased mortality in wild type mice. Thus, thrombospondin-1 enhances the pathogenesis of disseminated candidiasis by creating an imbalance in the host immune response that ultimately leads to reduced phagocytic function, impaired fungal clearance, and increased mortality. Conversely, inhibitors of thrombospondin-1 may be useful drugs to improve patient recovery from disseminated candidiasis.     

Target cells for the activity of a synthetic adjuvant: muramyl dipeptide.

Muramyl dipeptide (MDP), a synthetic adjuvant, increased the primary response of CBA mice to sheep red blood cells (SRBC). In reconstituted irradiated recipients, cooperation between T and B lymphocytes was required for the expression of adjuvant activity and MDP increased the efficiency of SRBC-educated T cells. The role of T-derived lymphocytes in mediating the MDP adjuvant activity was also demonstrated in irradiated mice and in mice reconstituted with various splenic cellular types of donors which had received SRBC and MDP 24 hr earlier. In our experiments, the macrophage did not seem to be involved, since MDP did not increase the phagocytic capacity of peritoneal exudate cells and MDP- and SRBC-pretreated macrophages had no increased ability to induce an anti-SRBC immune response. These results demonstrate the importance of T lymphocytes as mediators of the adjuvant activity of MDP.     

The beta-glucan receptor,dectin-1, is predominantly expressed on the surface of cells of the monocyte/macrophage and neutrophil lineages

We recently identified dectin-1 (betaGR) as a major beta-glucan receptor on leukocytes and demonstrated that it played a significant role in the non-opsonic recognition of soluble and particulate beta-glucans. Using a novel mAb (2A11) raised against betaGR, we show here that the receptor is not dendritic cell-restricted as first reported, but is broadly expressed, with highest surface expression on populations of myeloid cells (monocyte/macrophage (Mphi) and neutrophil lineages). Dendritic cells and a subpopulation of T cells also expressed the betaGR, but at lower levels. Alveolar Mphi, like inflammatory Mphi, exhibited the highest surface expression of betaGR, indicative of a role for this receptor in immune surveillance. In contrast, resident peritoneal Mphi expressed much lower levels of betaGR on the cell surface. Characterization of the nonopsonic recognition of zymosan by resident peritoneal Mphi suggested the existence of an additional beta-glucan-independent mechanism of zymosan binding that was not observed on elicited or bone marrow-derived Mphi. Although this recognition could be inhibited by mannan, we were able to exclude involvement of the Mphi mannose receptor and complement receptor 3 in this process. These observations imply the existence of an additional mannan-dependent receptor involved in the recognition of zymosan by resident peritoneal Mphi.     

Dynamics of neuron-glia interplay upon exposure to unconjugated bilirubin

Microglia are the main players of the brain immune response. They act as active sensors that rapidly respond to injurious insults by shifting into different activated states. Elevated levels of unconjugated bilirubin (UCB) induce cell death, immunostimulation and oxidative stress in both neurons and astrocytes. We recently reported that microglial phagocytic phenotype precedes the release of pro-inflammatory cytokines upon UCB exposure. We investigated whether and how microglia microenvironment influences the response to UCB. Our findings revealed that conditioned media derived from UCB-treated astrocytes reduce microglial inflammatory reaction and cell death, suggesting an attempt to curtail microglial over activation. Conditioned medium from UCB-challenged neurons, although down-regulating tumor necrosis factor-α and interleukin-1β promoted the release of interleukin-6 and nitric oxide, the activation of matrix metalloproteinase-9, and cell death, as compared with UCB-direct effects on microglia. Moreover, soluble factors released by UCB-treated neurons intensified the phagocytic properties manifested by microglia under direct exposure to UCB. Results from neuron-microglia mixed cultures incubated with UCB evidenced that sensitized microglia were able to prevent neurite outgrowth impairment and cell death. In conclusion, our data indicate that stressed neurons signal microglial clearance functions, but also overstimulate its inflammatory potential ultimately leading to microglia demise.