Effects of macrophage colony-stimulating factor (M-CSF) on lipopolysaccharide (LPS)-induced mediator production from monocytes in vitro

M-CSF is a macrophage-lineage-specific growth factor that causes proliferation and differentiation of progenitor cells in the bone marrow. To investigate the effects of M-CSF on more matured cells, human monocytes were cultured in the presence or absence of M-CSF for 6 days. Addition of M-CSF at more than 10(2) U/ml resulted in higher viability and caused morphological differentiation to large macrophage-like cells. LPS-induced mediator production was also compared between M-CSF-treated and control cell. Monocytes were incubated with or without M-CSF for 3 days, and were stimulated with 1 microgram/ml of LPS for 2 days. IL-1 beta was not detected in the both culture supernatants, and PGE2 production was not influenced by M-CSF. However, amounts of G-CSF, GM-CSF, IL-6, and TNF-alpha produced in response to 1 microgram/ml of LPS were 1.5 to 2 times greater from monocytes treated with 10(4) U/ml of M-CSF than from control cells. The priming effect of M-CSF on LPS-induced cytokine production was found to require 3-day preincubation, and reached a maximum at the concentration of 10(4) U/ml. M-CSF-treated cells responded to a 10 times lower concentration of LPS than control cells in terms of cytokine production. M-CSF was also shown by flowcytometric analysis to influence the expression of CD14, a receptor for LPS, which might render monocytes more sensitive to LPS.     

Induction of interleukin-12 (IL-12) by recombinant glycoprotein gp120 of human immunodeficiency virus type 1 in human monocytes/macrophages: requirement of gamma interferon for IL-12 secretion

We studied the effects of the gp120 glycoprotein of human immunodeficiency virus type 1 on the expression of interleukin-12 (IL-12) in human monocytes and in monocyte-derived macrophages. Induction of the mRNA for both the p35 and p40 subunits of IL-12 was observed in both cell types after gp120 treatment. We then evaluated cytokine secretion by using an enzyme-linked immunosorbent assay which recognizes only the IL-12 heterodimer. No IL-12 was detected in monocytes/macrophages treated with gp120 alone. A consistent IL-12 secretion was found in macrophages primed with gamma interferon (IFN-gamma) and subsequently treated with gp120. Low levels of IL-12 were occasionally observed in IFN-gamma-primed monocytes stimulated with gp120. The greater response of macrophages than of monocytes to the priming effect of IFN-gamma was consistent with the finding that IFN-gamma induced a much stronger antiviral state to vesicular stomatitis virus in macrophages than in monocytes. These data indicate that gp120 is an inducer of IL-12 expression in monocytes/macrophages and that IFN-gamma is an essential cofactor for IL-12 secretion, especially in differentiated macrophages.     

Human macrophages induced in vitro by macrophage colony-stimulating factor are deficient in IL-12 production

IL-12 is important for Th1 differentiation. Myeloid-derived antigen-presenting cells (APC) such as monocytes, macrophages (Mphi) and dendritic cells (DC) are believed to be major sources of IL-12 in vivo. We have compared IL-12 production of fresh monocytes with Mphi differentiated in vitro using macrophage colony-stimulating factor (M-CSF) or human plasma, and in vitro generated dendritic cells, since these differentiated cell types represent APC at sites of antigen challenge. Macrophages stimulated with lipopolysaccharide (LPS) or heat-killed Listeria monocytogenes in the presence or absence of IFN-gamma produced minimal IL-12 p70 by comparison with DC or monocytes, despite comparable production of TNF-alpha. M-CSF-induced Mphi produced low levels of IL-10 constitutively and high levels after stimulation with LPS, but neutralization of IL-10 did not augment Mphi IL-12 production. Exposure of Mphi to TNF-alpha, granulocyte-macrophage CSF or IFN-gamma did not substantially up-regulate IL-12. Therefore M-CSF induces a differentiated Mphi phenotype in which IL-12 production is down-regulated, perhaps irreversibly. This may be the default pathway for monocyte-Mphi development in the absence of inflammation.     

Tacrine inhibits nicotinic secretory and current responses in adrenal chromaffin cells

Previous studies have demonstrated an infiltration of monocytes and increased levels of granulocyte-macrophage colony-stimulating factor (GM-CSF) in the asthmatic lung. To study the possible effects of this cytokine upon the differentiation and function of these newly recruited monocytes, we have developed a model in which monocytes isolated from human peripheral blood were differentiated into macrophages in serum in the presence or absence of GM-CSF. After 7 days, the macrophages increased in size and granularity, had increased phagocytic activity, and expressed various adhesion molecules, CD14 and major histocompatibility complex (MHC) class II. The effects of GM-CSF on antigen presentation by cultured macrophages on the antigen-specific proliferative response of CD4+ T cells to Dermatophagoides pteronyssinus or purified protein derivative of tuberculin and the mitogen phytohaemagglutinin was determined. CD4+ T-cell proliferation was reduced when either antigen was presented by macrophages cultured in serum alone, compared with the values obtained with freshly isolated monocytes. However, CD4+ cell proliferation was comparable to that observed with monocytes when antigen was presented by macrophages which had been pre-cultured with 50 U/ml GM-CSF. CD4+ T-cell proliferation to phytohaemagglutinin was similar when all three populations were used as accessory cells. High numbers of macrophages partially suppressed CD4+ T-cell proliferation in response to antigen presented by monocytes, but there was no significant difference between macrophages cultured in the presence or absence of GM-CSF. This data suggests that GM-CSF directs monocyte differentiation into macrophages with an antigen-presenting, rather than a suppressive, phenotype. Elevated levels of GM-CSF in the asthmatic lung may therefore maintain recently recruited monocytes in an inflammatory and T-cell activating state.     

Synergistic effect of immunoregulatory cytokines on peripheral blood monocytes from patients with inflammatory bowel disease

Active inflammatory bowel disease (IBD) is characterized by increased monocyte secretion of proinflammatory cytokines. Immunoregulatory cytokines such as Interleukin (IL)-4, IL-10, and IL-13 are capable of inhibiting the proinflammatory cytokine response of activated monocytes. The aim of our study was to determine the effect of different antiinflammatory cytokines under various culture conditions and to evaluate combinations of antiinflammatory cytokines in down-regulating monocyte response in IBD. Peripheral monocytes from patients with active IBD were isolated and stimulated with pokeweed mitogen (PWM). IL-4, IL-10, IL-13 and a combination of IL-4/IL-10 and IL-10/IL-13 were added at different concentrations and different times. Secretion of IL-1beta and TNF-alpha was assessed using sandwich ELISA systems. There was a diminished down-regulation of TNF-alpha by IL-4 and IL-13 in IBD when the cytokines were added at the time of stimulation, while there was a significantly higher down-regulation when monocytes were primed with these Th-2 cytokines 24 hr before activation. IL-10 plus IL-4 and IL-10 plus IL-13, respectively, inhibited the proinflammatory cytokine response of monocytes as well as matured macrophages much more than IL-4, IL-10, or IL-13 alone. Even at suboptimal concentrations for each cytokine alone, a combination of cytokines showed synergistic inhibitory effects. In summary, a combination of antiinflammatory cytokines is more effective in down-regulating the response of activated monocytes than using the cytokines alone and thus may have a potential therapeutic benefit for patients with IBD.     

IL-10 prevents the differentiation of monocytes to dendritic cells but promotes their maturation to macrophages

Human monocytes cultured with granulocyte-macrophage colony-stimulating factor (GM-CSF) and IL-13 for 7 days differentiate into cells with the morphology and function of dendritic cells (DC). We have investigated the effect of IL-10 on this differentiation pathway. In the presence of IL-10 cells did not develop DC morphology, did not express CD1a and had lower levels of MHC class II. IL-10 promoted the differentiation of large cells with the morphology, cytochemistry and membrane phenotype of macrophages, including staining for nonspecific esterase and high levels of CD14, CD16 and CD68. The effect of IL-10 was dose dependent and was best appreciated when the cytokine was added at the initiation of the culture, as addition on day 3 was less inhibitory. When added to already differentiated DC on day 6, IL-10 caused only a modest reduction of MHC class II and CD1a expression, and no acquisition of the macrophage markers CD14, CD16 and CD68. Prolonged incubation up to 5 days with IL-10 did not induce a shift of differentiated DC to macrophages. On the other hand, the macrophages obtained by culturing for 7 days with GM-CSF+IL-13+IL-10 did not shift to DC upon removal of IL-10 for up to 3 days. Thus, the effect of IL-10 on monocyte differentiation, occurs only at the precursor level and confers an irreversible phenotype. From a functional point of view, cells cultured in the presence of IL-10 were poor stimulators of allogeneic cord blood T cells in mixed lymphocyte reaction (MLR) and presented tetanus toxin (TT) to specific T cell lines with much less efficiency than control DC. In contrast, IL-10-cultured DC showed 7 times greater endocytosis of FITC-dextran. This increased endocytosis was mostly mediated via the mannose receptor, as demonstrated by blocking with unlabeled mannose. In conclusion, IL-10 inhibits DC differentiation from monocytes and, in a substantial proportion of the cells, promotes the differentiation to mature macrophages. Intriguingly, IL-10 inhibits antigen presentation while it stimulates endocytic activity.     

Focal adhesion kinase upregulated by granulocyte-macrophage colony-stimulating factor but not by interleukin-3 in differentiating myeloid cells

The involvement of focal adhesion kinase (FAK) in myeloid differentiation was investigated in primary murine bone marrow (BM) cells. In unstimulated BM, FAK mRNA was detected in myeloid and lymphoid cells, but not in erythroid precursors. When the BM cells were incubated with granulocyte-macrophage colony-stimulating factor (GM-CSF) or interleukin-3 (IL-3), FAK expression showed a remarkable difference depending on the cytokine. Although FAK was upregulated in the cells stimulated by GM-CSF (GM-treated cells), the kinase was barely detectable in the cells cultured with IL-3 (IL-3-treated cells). Morphology and flow cytometry analysis showed GM-CSF promoted the growth and differentiation of monocyte/macrophage lineage stronger than IL-3. In addition, motility of the cytokine-differentiated cells showed an overt distinction between the cultures, which was closely correlated with FAK expression. After 7 days of stimulation, GM-treated cells showed active migration and chemoattractant-induced morphologic polarization. In contrast, IL-3-treated cells showed minimal migration and polarization. These results suggest an important role of GM-CSF in the terminal differentiation of monocytes/macrophages, and possible involvement of FAK in functional maturity of this lineage.     

Tumors and other lesions composed of adipose tissue. Integrated diagnostic imaging]

Atherosclerosis is characterized as a chronic inflammatory-fibroproliferative disease of the vessel wall. The attachment of monocytes and T-lymphocytes to the injured endothelium followed by their migration into the intima is one of the first and most crucial steps in lesion development. The co-localization of CD4+ T-cells and macrophages in the lesion, the abundant expression of HLA Class II molecules and the co-stimulatory molecule CD40 and its ligand (CD40L) indicate a contribution of cell-mediated immunity to atherogenesis. Transgenic mouse models revealed that dependent on the model T- and B-cells may promote lesion progression, monocytes and macrophages are in contrast essential for the development of atherosclerotic lesions. Apart from the local process in the vessel wall, systemic signs of an inflammatory reaction are also associated with lesion development. Thus plasma levels of C-reactive protein and fibrinogen and the white blood cell count are positively correlated to the risk of cardiovascular disease. Recently, an inflammatory phenotype of circulating peripheral blood monocytes could be demonstrated as a specific cellular correlate to lipid and lipoprotein risk factors. Thus the pool size of LPS receptor (CD14)dim and Fc gamma IIIa receptor (CD16a)+ monocytes positively correlates to plasma cholesterol levels, to triglycerides levels and to the apolipoprotein E4 (apo E4) phenotype in contrast to a negative correlation to the high density lipoprotein (HDL) cholesterol concentration. This CD14dim CD16a+ monocytes are further characterized by a high expression of beta 1- and beta 2-integrins, suggesting a higher capacity for attachment at sites of inflammation. A proinflammatory cytokine pattern and an expansion of these cells in other inflammatory diseases are indicating that these cells promote the inflammatory process during atherogenesis. Surface expression of the activation antigen CD45RA on monocytes in correlation to plasma LDL cholesterol and Lp(a) levels further indicates an inflammatory reaction. Regarding the potential mechanisms of the phenotypic changes of peripheral blood monocytes, in a serum free in vitro differentiation model supplemented with M-CSF monocytes from probands which are homozygous for apo E4 showed a significantly higher increase of CD16a expression compared to apo E3/E3 cells indicating that a genetic polymorphism of a single apolipoprotein gene locus may affect monocyte differentiation. The further characterization of the cellular immunology of monocytes and T-lymphocytes in lesion development will provide new specific diagnostic and therapeutic targets in atherogenesis.     

Induction of interleukin-10 by human immunodeficiency virus type 1 and its gp120 protein in human monocytes/macrophages

In this study, we evaluated the effects of human immunodeficiency virus type 1 (HIV-1) and its gp120 protein on interleukin-10 (IL-10) expression in cultured human monocytes/macrophages. Infection of either 1-day monocytes or 7-day monocyte-derived macrophages with HIV-1 strain Ba-L resulted in clear-cut accumulation of IL-10 mRNA at 4 and 24 h. Likewise, treatment of these cells with recombinant gp120 induced IL-10 mRNA expression and caused a marked increase in IL-10 secretion. Monoclonal antibodies to gp120 strongly inhibited recombinant gp120-induced IL-10 secretion by monocytes/macrophages. Moreover, the addition of IL-10 to monocytes/macrophages resulted in a significant inhibition of HIV-1 replication 7 and 14 days after infection. On the whole, these results indicate that HIV-1 (possibly through its gp120 protein) up-regulates IL-10 expression in monocytes/macrophages. We suggest that in vivo production of IL-10 by HIV-primed monocytes/macrophages can play an important role in the early response to HIV-1 infection.     

Specific activated T cells regulate IL-12 production by human monocytes stimulated with Cryptococcus neoformans

IL-12 production mediated by a T cell-independent and/or T cell-dependent pathway was investigated in human monocytes responding to Cryptococcus neoformans. The data of this study showed that: 1) appreciable levels of IL-12 were observed when freshly isolated monocytes were exposed to acapsular C. neoformans or Candida albicans and secretion occurred within 24-48 h of incubation; 2) monocytes alone were poor producers of IL-12 when stimulated with encapsulated C. neoformans; 3) the presence of specific anti-glucuronoxylomannan mAb favored IL-12 secretion and Fc cross-linking could play a role; 4) monocytes were able to secrete consistent levels of IL-12 when cultured with activated T cells responding to C. neoformans; 5) the maximum secretion of IL-12 was observed at 5-7 days of culture and was strongly regulated by the presence of endogenous IFN-gamma; and 6) the interaction between CD40 on monocytes and CD40 ligand on activated T lymphocytes responding to C. neoformans played a critical role in IL-12 secretion. These data highlight the mechanisms of IL-12 production by human monocytes exposed to C. neoformans, indicating a possible biphasic secretion of IL-12, dependent on the direct effect of fungal insult, and characterized by consistent secretion of IL-12 that is dependent on the interaction of CD40 with the CD40 ligand expressed on activated T cells responding to C. neoformans.     

Interleukin-10 in non-Hodgkin's lymphoma

Interleukin-10 (IL-10) is a pleiotropic cytokine produced by type 2 helper cells (Th2), as well as by monocytes and macrophages, and normal and neoplastic B lymphocytes. It is highly homologous to an open reading frame of Epstein-Barr virus (EBV) called BCRF1, and EBV infection of B-cells up-regulates IL-10. IL-10 production has strong immunosuppressive effects via inhibition of Th1 type cytokines, including interferon gamma and interleukin-2. On B-cells, IL-10 has a potent stimulating effect, inducing proliferation and differentiation. Interestingly, in cell lines derived from B-cell lymphomas, IL-10 production has been found to be up-regulated, and it serves as an autocrine growth factor. In patients with non-Hodgkin's lymphoma (NHL), serum IL-10 levels are significantly increased when compared to normal individuals and NHL patients in remission. The prognostic significance of these increased levels vary according to the assay used. Both human IL-10 and viral IL-10 are increased, and when specific assays for human IL-10 are used, there seems to be no prognostic significance, whereas when the assay cross-reacts with viral IL-10, high levels correlate with poor prognosis. These results suggest that viral IL-10 might have some pathogenic role in NHL.     

Macrophage colony-stimulating factor stimulates synthesis and secretion of a mouse homolog of a human IgE-dependent histamine-releasing factor by macrophages in vitro and in vivo

Treatment of murine resident peritoneal macrophages with macrophage-CSF (M-CSF) up-regulated the synthesis of a discrete set of proteins, including a 26-kDa protein (p26). The sequence of 20 NH2-terminal amino acids of the purified p26 was identical with the mouse homolog of a human IgE-dependent histamine-releasing factor (HRF). Among macrophage activators tested (M-CSF, granulocyte-macrophage-CSF, IL-3, TNF-alpha, IFN-gamma, and LPS), only M-CSF could up-regulate the p26 HRF synthesis by cultured macrophages. M-CSF not only increased the levels of p26 HRF mRNA and protein, but also stimulated the secretion of an N-glycosylated p26 HRF with a m.w. of 30 kDa. Repeated injections of M-CSF into mouse peritoneal cavity for 4 days elicited macrophages expressing abundant p26 HRF. A single i.p. injection of M-CSF failed to increase the p26 HRF level in peritoneal macrophages of thioglycollate-, LPS-, or adjuvant-treated mice, while M-CSF challenge to OVA-immunized mice caused macrophage infiltration and overproduction of p26 HRF, similarly as did OVA challenge. The Ag-specific priming for enhanced synthesis and secretion of p26 HRF by M-CSF was also demonstrated in cultured macrophages prepared from OVA-immunized mice. An i.p. injection of M-CSF or recombinant p26 HRF triggered eosinophil recruitment, even in the absence of the Ag, in the sensitized mice, but not in normal mice. Furthermore, recombinant p26 HRF could induce eosinophilia without marked macrophage and lymphocyte infiltrations. Our results suggest that p26 HRF secreted by M-CSF-stimulated macrophages may be an important mediator for the late phase allergic inflammation.     

Cytokine regulation of granulocyte-macrophage colony stimulating factor and macrophage colony-stimulating factor production in human arterial smooth muscle cells

Smooth muscle cells (SMC) are the major cell type found in the walls of large blood vessels and appear to participate in local immune and inflammatory reactions, as well as in certain vascular diseases. We tested whether human arterial SMC can produce in vitro the colony stimulating factors (CSFs), granulocyte macrophage-CSF (GM-CSF) and macrophage CSF (M-CSF). Untreated internal mammary artery and aortic SMC produced no detectable GM-CSF but constitutively made M-CSF, measured by ELISA and radioimmunoassay, respectively. Interleukin-1 (IL-1) and, to a lesser extent, tumor necrosis factor alpha (TNF alpha) stimulated GM-CSF formation within 3 h; mRNA levels also increased particularly in the presence of the protein synthesis inhibitor, cycloheximide. IL-1, TNF alpha and, in addition, interferon-gamma (IFN-gamma) raised the M-CSF levels within 6 h; cycloheximide potentiated the effects of IL-1 and TNF alpha on mRNA levels. These results suggest that cytokine-stimulated human arterial SMC may be a source of the M-CSF found in atherosclerotic lesions. Since monocytes/macrophages can be activated by GM-CSF and M-CSF, while GM-CSF can also affect granulocyte function, SMC may participate in inflammatory reactions and vascular diseases by releasing these cytokines.     

A combination of osteoclast differentiation factor and macrophage-colony stimulating factor is sufficient for both human and mouse osteoclast formation in vitro

Both human and murine osteoclasts can be derived in vitro from hematopoietic cells or monocytes that are co-cultured with osteoblasts or marrow-derived stromal cells. The osteoclastogenic stimulus provided by murine osteoblasts and marrow-derived stromal cells is now known to be mediated by osteoclast differentiation factor (ODF), a membrane-bound tumor necrosis factor-related ligand. This study demonstrates that mouse spleen cells and monocytes form osteoclasts when cultured in the presence of macrophage-colony stimulating factor (M-CSF) and a soluble form of murine ODF (sODF). Numerous multinucleated osteoclasts expressing tartrate resistant acid phosphatase (TRAP) and calcitonin receptor (CTR) formed within 7 days of culture and engaged in extensive lacunar bone resorption. Osteoclast number and bone resorption area was dependent on sODF concentration. Long-term cultured human monocytes also formed bone resorbing osteoclasts in response to co-stimulation by sODF and M-CSF, although this required more than 11 days in culture. This human osteoclast differentiation was strongly inhibited by granulocyte-macrophage colony stimulating factor. This study further characterises murine osteoclast differentiation caused by sODF and M-CSF co-stimulation in vitro, and shows that the same co-stimulation causes human osteoclast differentiation to occur. We propose that this methodology can be employed to investigate the direct effects of cytokines and other factors on human osteoclast differentiation.     

Phospholipid vesicle binding and aggregation by four novel fish annexins are differently regulated by Ca2+

The aim of this study was to further assess the role of pooled human immunoglobulin (PHIG) on cytokine production from PBMC stimulated with a bacterial superantigen. Human PBMC were cultured with Streptococcus pyrogenic exotoxin A (SPE-A) with or without PHIG and several proinflammatory cytokine levels of culture supernatants were measured. Serum cytokine levels of KD patients before and after PHIG therapy were also examined. PHIG greatly reduced the production of IL-12, interferon-gamma (IFN-gamma) and other cytokines from SPE-A-stimulated PBMC, while exogenous IL-12, but neither IL-1 nor tumour necrosis factor-alpha (TNF-alpha), restored IFN-gamma production inhibited by PHIG. Although PHIG partially adsorbed SPE-A, its inhibitory effect on cytokine production was not played by anti-SPE-A antibody. Although purified CD4+ T cells cultured with human HLA-DR-transfected mouse L cells and SPE-A could not effectively produce IFN-gamma, they produced large amounts of IFN-gamma if exogenous IL-12 was introduced. KD patients in the acute phase had higher levels of serum IFN-gamma than did controls and patients with bacterial infection. Although IL-12 levels of children with or without KD were not significantly different, IL-12 levels of children were much higher than those of adults. However, serum levels of IL-12 of KD patients were transiently but significantly decreased by PHIG therapy and IFN-gamma amounts subsequently reverted to basal levels thereafter. These findings indicate that PHIG inhibits IL-12 production of SPE-A-activated monocytes and thereby decreases IFN-gamma synthesis by T cells and suggest that inhibition of IL-12 and IFN-gamma production is an important part of the mechanisms underlying PHIG therapy on KD.     

Interaction between interleukin 10 and interleukin 6 in human B-cell differentiation

Contrary to their opposing action on human T-lymphocytes and monocytes, both Interleukin (IL-)10 and IL-6 are potent differentiation factors of human B-cells. Both are known to induce immunoglobulin (Ig) production. The precise mechanism of this converging effect of IL-6 and IL-10 remains elusive, however. Here we investigated the role of IL-6 in the IL-10 dependent B-cell differentiation into Ig secreting cells. We found that co-stimulation of SAC-stimulated human peripheral B-lymphocytes with IL-10 and IL-6 exhibited no additive effect on Ig production over stimulation with IL-10 alone, and that IL-6 receptor blockade only mildly inhibited IL-10 induced Ig synthesis. In fact, we could show that stimulation of B-cells with IL-10 somewhat suppressed SAC induced autocrine IL-6 production. Despite this suppression IL-6 levels remained sufficiently high to stimulate its receptor, and IL-6 binding to the B-cell surface was not affected. The failure of IL-6 to exert an additional effect on SAC+IL-10 induced Ig production suggests that IL-10 may recruit components of the IL-6 intracellular pathway for Ig induction. In conclusion we could demonstrate that IL-10 acts on B-cell differentiation independently of autocrine IL-6 and that it had a considerably mild effect on B lymphocytic autocrine IL-6 secretion. This still allows an IL-6 effect in the presence of IL-10 which appears adaptive with a view to the converging effects of these two cytokines on human B lymphocytes. Our study thus adds to the appreciation of the complex cytokine regulation of the immune system.     

Appetitive sexual behavior in male rats: 2. sexual reward and level-changing behavior

The immunodeficiency of patients with chronic renal failure (CRF) is related to multiple and complex alterations of the cytokine network and of its target cells such as T or B lymphocytes, monocytes, fibroblasts or endothelial cells. Chronic activation of monocytic functions is recognized as a key factor in these immunological disorders. Since macrophage colony stimulating factor (M-CSF) is essential for the activation of several functions of monocytes and macrophages and their production of cytokines such as interleukin-1, interleukin-6, and tumor necrosis factor alpha, we investigated its involvement in patients with CRF. When measured by ELISA, M-CSF serum levels were significantly higher in patients with progressive CRF and those on hemodialysis (HD) and continuous ambulatory peritoneal dialysis (CAPD) than in controls. M-CSF serum levels did not correlate with the degree of renal insufficiency and were probably related to complex alterations in its production and/or degradation by the specific M-CSF receptors of macrophages. In HD patients the M-CSF serum concentrations inversely correlated with the number of circulating lymphocytes and were significantly higher in anemic patients requiring treatment with erythropoietin. Our results suggest that M-CSF may play a role in altering the immune system in uremic patients by maintaining in the circulation and tissues permanently primed monocytes and/or macrophages that can then be triggered to an activated state by secondary stimuli such as endotoxins, complement components, other cytokines or contact with foreign surfaces.     

Reduction of fecal contamination of street-vended beverages in Guatemala by a simple system for water purification and storage, handwashing, and beverage storage

CD4+ cells from young (3 months) and old (19 months) mice were stimulated by plate-bound anti-CD3 monoclonal antibody (mAb) alone or also by soluble anti-CD28 mAb. Supernatants were analysed by enzyme-linked immunosorbent assay (ELISA) to determine cytokine concentrations. Total RNA was extracted from cells, reverse transcribed and the cDNA amplified by polymerase chain reaction (PCR) to evaluate the amount of specific mRNA. The results indicate that anti-CD3 alone is not sufficient to induce interleukin-2 (IL-2) production in CD4+ cells from both young and old mice. However, anti-CD28, together with anti-CD3 mAb, induces a much higher production of IL-2 in CD4+ cells from young as compared with old mice. Conversely, interferon-gamma (IFN-gamma) production is also induced by anti-CD3 alone and is higher in CD4+ cells from old as compared with young mice. Upon addition of anti-CD28 mAb, IFN-gamma production increases in both groups, but it remains much higher in old than in young mice. Also the production of IL-4 and IL-10 is induced by anti-CD3 mAb but it is increased by the addition of anti-CD28 mAb. CD4+ cells from old mice produce more IL-4 and IL-10 as compared with cells from young mice. The amounts of cytokine specific mRNA in CD4+ cells from young and old mice parallel the cytokine levels in culture supernatants. Results on the mRNA turnover indicate that when CD4+ cells are stimulated by anti-CD3 or costimulated also by anti-CD28 mAb, the IFN-gamma, IL-4 and IL-10 specific mRNAs are more stable in old than in young mice, suggesting that mRNA stability has a relevant role in the different patterns of cytokine production.     

Cytokine-regulated expression of activated leukocyte cell adhesion molecule (CD166) on monocyte-lineage cells and in rheumatoid arthritis synovium

OBJECTIVE: To determine whether monocyte/macrophage expression of the CD6 ligand, activated leukocyte cell adhesion molecule (ALCAM) (CD166), is regulated by cytokines during inflammation in rheumatoid arthritis (RA). METHODS: We used flow cytometry to test whether cytokines present in rheumatoid synovium could regulate ALCAM cell surface expression on peripheral blood (PB) monocytes and RA synovial fluid (SF) macrophages, and we examined ALCAM expression in situ in RA synovium by immunofluorescence. RESULTS: The monocyte differentiation factors interleukin-3, macrophage colony-stimulating factor (M-CSF), and granulocyte-macrophage colony-stimulating factor augmented ALCAM expression on PB monocytes. ALCAM was expressed on monocyte-lineage cells in situ in inflamed synovium from patients with RA (9 of 9), but not in uninflamed synovium from patients with joint trauma (0 of 3). Furthermore, in vitro culture-induced ALCAM expression on PB monocytes and CD14+ RA SF cells was inhibited by an M-CSF neutralizing antibody. CONCLUSION: ALCAM expression on PB and SF monocytes/macrophages is enhanced by M-CSF.