Heparin-binding protein (CAP37) is internalized in monocytes and increases LPS-induced monocyte activation

Previous studies have shown that the neutrophil-derived heparin-binding protein (HBP), also known as CAP37 or azurocidin, potentiates the LPS-induced release of proinflammatory cytokines (TNF-alpha, IL-1, and IL-6) from isolated human monocytes. To date, the mechanisms by which HBP enhances LPS-induced monocyte activation have not been elucidated, and it is not known whether HBP also increases the LPS-induced production of other bioactive substances. We studied human monocytes activated by recombinant human HBP and LPS and their interaction with the LPS receptor CD14. We hypothesized that the stimulatory effect of HBP on the LPS-induced release of proinflammatory mediators from monocytes was mediated by specific binding of HBP to monocytes, which resulted in an up-regulation of CD14. Our results demonstrated that HBP alone (10 microg/ml) stimulated the production of TNF-alpha from isolated monocytes. In addition, HBP had an additive effect on LPS-induced production of TNF-alpha and PGE2, suggesting a generalized monocyte activation. We used flow cytometry to demonstrate that HBP had a high affinity to monocytes but not to the LPS receptor CD14, and experiments performed at 4 degrees C indicated an energy-dependent step in this process. Confocal microscopy showed that monocytes internalize HBP within 30 min. These data suggest that mechanisms other than increased CD14 expression are responsible for the enhanced release of TNF-alpha or PGE2 in response to HBP and LPS.     

Inhibition of IL-10 expression by IFN-gamma up-regulates transcription of TNF-alpha in human monocytes

Stimulation of human monocytes with LPS induces expression of multiple cytokines, including TNF-alpha, IL-1 beta, IL-6, and IL-10, IL-10 expression is delayed relative to that of TNF-alpha, IL-1 beta, and IL-6. Furthermore, IL-10 feedback inhibits expression of TNF-alpha, IL-1 beta, and IL-6, thus providing an efficient autocrine mechanism for controlling proinflammatory cytokine production in monocytes. The Th1-type lymphokine, IFN-gamma, markedly up-regulates TNF-alpha production in monocytes. However, the precise mechanism by which IFN-gamma mediates this effect is unknown. We examined the effects of IFN-gamma on IL-10 expression in LPS-stimulated monocytes, and the relationship between IL-10 and TNF-alpha production in these cells. LPS stimulation induced rapid, ordered expression of multiple cytokines. Steady-state mRNA levels for TNF-alpha increased rapidly, reached maximal levels by 2 to 3 h poststimulation, and then declined sharply. IL-1 beta and IL-6 mRNA levels also increased markedly following stimulation with LPS, but decreased more slowly than did TNF-alpha. Down-regulation of mRNA for TNF-alpha, IL-1 beta, and IL-6 coincided with a delayed and more gradual increase in IL-10 mRNA levels. Furthermore, neutralization of IL-10 with anti-IL-10 Abs prolonged TNF-alpha mRNA expression, and significantly increased net TNF-alpha production. IFN-gamma suppressed expression of IL-10 mRNA and protein in a dose-dependent manner. Moreover, inhibition of IL-10 production correlated with a marked increase in both the magnitude and duration of TNF-alpha expression. Thus, potentiation of TNF-alpha production by IFN-gamma in monocytes is coupled to inhibition of endogenous IL-10 expression.     

T cell activation signals up-regulate p38 mitogen-activated protein kinase activity and induce TNF-alpha production in a manner distinct from LPS activation of monocytes

p38 mitogen-activated protein kinase (MAPK) (p38) is involved in various cellular responses, including LPS stimulation of monocytes, resulting in production of proinflammatory cytokines such as TNF-alpha. However, the function of p38 during antigenic stimulation of T cells is largely unknown. Stimulation of the human Th cell clone HA-1.70 with either the superantigen staphylococcal enterotoxin B (SEB) or with a specific antigenic peptide resulted in p38 activation and the release of TNF-alpha. MAPK-activated protein kinase-2 (MAPKAPK-2), an in vivo substrate for p38, was also activated by T cell signaling. SB 203580, a selective inhibitor of p38, blocked p38 and MAPKAPK-2 activation in the T cell clone but did not completely inhibit TNF-alpha release. PD 098059, a selective inhibitor of MAPK kinase 1 (MEK1), blocked activation of extracellular signal-regulated kinase (ERK) and partially blocked TNF-alpha production by the clone. In human peripheral T cells, p38 was not activated by SEB, but rather by CD28 cross-linking, whereas in the human leukemic T cell line Jurkat, p38 was activated by CD3 and CD28 cross-linking in an additive fashion. TNF-alpha production by peripheral T cells in response to SEB and anti-CD28 mAb correlated more closely with ERK activity than with p38 activity. Therefore, various forms of T cell stimulation can activate the p38 pathway depending on the cells examined. Furthermore, unlike LPS-stimulated monocytes, TNF-alpha production by T cells is only partially p38-dependent.     

Regulation of monocyte IL-10 synthesis by endogenous IL-1 and TNF-alpha: role of the p38 and p42/44 mitogen-activated protein kinases

IL-10 is an anti-inflammatory cytokine with potent immunomodulatory effects, including inhibition of cytokine production. However, regulation of monocyte IL-10 production is poorly understood. In this report we have investigated the mechanisms of LPS-induced IL-10 production by human peripheral blood monocytes and demonstrate that IL-10 synthesis is uniquely dependent on the endogenous proinflammatory cytokines IL-1 and/or TNF-alpha. LPS signal transduction in monocytes has been shown to involve activation of the p38 and p42 mitogen-activated protein kinase (MAPK) cascades. The results in this paper indicate that inhibition of p38 MAPK potently inhibited the production of IL-10, IL-1beta, and TNF-alpha, whereas blockade of the p42/44 MAPK pathway, while partially inhibiting TNF-alpha and IL-1beta production, had no effect on monocyte secretion of IL-10. Furthermore, neither the inhibition of monocyte TNF-alpha induced by IL-10 nor the stimulation of soluble TNF receptor production was affected by inhibition of the p42/44 MAPK pathway, suggesting that this signaling event is not involved in either monocyte production of or anti-inflammatory responses to IL-10. These data raise the interesting possibility that proinflammatory TNF-alpha-mediated effects may be selectively blocked without modulating the induction or the response to IL-10, whereas the signaling events associated with the anti-inflammatory events induced by IL-10 remain to be elucidated.     

Regulation of monocyte IL-12 production: augmentation by lymphocyte contact and acute ethanol treatment, inhibition by elevated intracellular cAMP

IL-12, a monocyte-derived cytokine, is pivotal in activation of cellular immune response and inflammation. Both inflammatory response and cellular immunity are impaired by acute ethanol consumption. Here, we found that in vitro acute ethanol treatment (25-100 mM) results in a dose-dependent and significant increase of IL-12 in IFN-gamma (100 U/ml) plus Staphylococcal enterotoxin B (SEB; 1 microg/ml) stimulated monocytes and mononuclear cells but not in unstimulated cells from non-alcoholic blood donors. There was significantly greater IL-12 production in the MNC population compared to isolated Mphi (P < 0.001). Prevention of monocyte surface contact with either purified T lymphocytes or monocyte-depleted MNC resulted in a significant, 65+/-20%, decrease in IL-12 production regardless of IFN-gamma, SEB or ethanol stimulation suggesting that Mphi T-cell surface contact provides an additional signal for IL-12 production. In addition to cell surface contact, soluble mediators, particularly IL-10 and PGE2 may regulate IL-12 production. The cyclooxygenase inhibitor, Indomethacin (10(-6)M), augmented both IL-12 and IL-10 levels in isolated monocytes and mononuclear cells whether induced by medium, SEB or SEB plus 25 mM ethanol suggesting that regulation of IL-12 production via the cyclooxygenase pathway is independent of IL-10. Finally, elevation of intracellular cAMP levels by dbcAMP treatment consistently inhibited IL-12 as well as IL-10 production in monocytes induced by IFN-gamma or IFN-gamma plus 25 mM ethanol. These data suggest that augmentation of monocyte IL-12 by acute ethanol is not mediated via the cAMP pathway.     

Effects of proinflammatory cytokines and bacterial toxins on neutrophil rheologic properties

OBJECTIVE: To examine the changes in neutrophil deformability, aggregation, and adherence in response to stimulation with proinflammatory cytokines and bacterial toxins. DESIGN: Prospective, randomized trial. SETTING: Research laboratory. SUBJECTS: Neutrophils isolated from healthy volunteers. INTERVENTIONS: Neutrophils were exposed to tumor necrosis factor (TNF)-alpha, interleukin (IL)-1beta, IL-8, their combination, endotoxin (LPS), lipoteichoic acid (LTA), and staphyloccocal enterotoxin B (SEB). Neutrophil deformability was measured as percent neutrophils filtered through 5-microm diameter filters. Aggregation was measured using a platelet aggregometer. Adherence was determined by examining the binding of neutrophils to albumin-coated latex beads. MEASUREMENTS AND MAIN RESULTS: Exposure to TNF-alpha and IL-1beta led to significant decreases in neutrophil filterability, which was attenuated by cytochalasin D pretreatment. LPS and LTA also decreased deformability, suggesting that these toxins directly stimulated neutrophils independent of cytokines. IL-8 and SEB did not significantly affect neutrophil deformability. TNF-alpha and LPS were associated with significant neutrophil aggregation, which was inhibited by pretreatment with anti-CD18 antibodies. Neutrophil aggregation was not affected by IL-1beta, LTA, or SEB. TNF-alpha, IL-8, and LPS increased neutrophil adherence, which also was attenuated by pretreatment with anti-CD18 antibodies. IL-1beta, LTA, and SEB did not significantly affect neutrophil adherence. CONCLUSIONS: Cytokines and bacterial toxins differ in their effects on neutrophil deformability, aggregation, and adherence. Of the cytokines examined, TNF-alpha appears to have the greatest direct effects on neutrophil rheology. Similarly, endotoxin appears to have greater direct effects on neutrophil rheology than the Gram-positive bacterial toxins, LTA, and staphylococcal enterotoxins.     

Interleukin-18 enhances lipopolysaccharide-induced interferon-gamma production in human whole blood cultures

Interleukin-18 (IL-18) is a newly described cytokine, formerly called interferon-gamma (IFN-gamma)-inducing factor. In a simple 24-h human whole blood culture, IFN-gamma was produced by the combination of lipopolysaccharide (LPS) plus IL-18. To liberate cytokines in the leukocyte and red cell compartments, the detergent Triton X-100 was added to the entire blood culture. The combination of low concentrations of LPS plus IL-18 induced a 3- to 5-fold greater production of IFN-gamma than did either stimulant alone. Tumor necrosis factor-alpha (TNF-alpha), IL-6, and IL-8 were also produced. The presence of IL-10 completely suppressed the production of IFN-gamma and reduced that of TNF-alpha, IL-6, and IL-8. Thus, IFN-gamma, TNF-alpha, IL-8, and IL-6 are produced in a single whole blood culture, making correlations in the synthesis of a T helper type 1 cytokine and proinflammatory cytokines with disease activity possible in a single culture.     

Erratum to "Immunocytochemical detection of cytokines and chemokines in Langerhans cells and in vitro derived dendritic cells"

We have developed a direct immunocytochemical technique to identify cytokine and chemokine production in epidermal Langerhans cells (LC) and in vitro derived CD14-, CD1a+, CD83+, CD40+ dendritic cells (DC) at the single cell level. Formaldehyde fixation combined with saponin permeabilization preserved cellular morphology and generated a characteristic juxtanuclear staining signal due to the accumulation of cytokine to the Golgi organelle. This approach was used for the assessment of TNF-alpha, IL-6, IL-8, IL-10, IL-12, GM-CSF, MIP-1alpha, MIP-1beta and RANTES producing cells. In contrast, a diffuse cytoplasmic staining was evident for IL-1ra, IL-1alpha and IL-1beta production. IL-1ra and IL-1alpha were expressed in 10-25% of unstimulated cultured cells, while all the other cytokines were undetectable. IL-1ra, IL-1alpha and IL-1beta were also the dominating cytokines, expressed in up to 85% of the DC, after 3 h of LPS stimulation. A significantly lower number of cells (0-5%) synthesized TNF-alpha, IL-6, IL-10, IL-12 and GM-CSF. The incidence of chemokine producing cells (IL-8, RANTES, MIP-1alpha, MIP-1beta) peaked 10 h after LPS stimulation in up to 60% of the DC. Both immature CD83- and mature CD83+ DC as well as LC had a similar cytokine production pattern. Thus, in comparison to monocytes, LPS stimulation of DC generated a lower incidence of TNF-alpha, IL-6, IL-10 and IL-12 producing cells while IL-1 was expressed in a comparable number of cells.     

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.     

Resistance to lipopolysaccharide mediated by the Yersinia pestis V antigen-polyhistidine fusion peptide: amplification of interleukin-10

We previously showed that injection of homogenous staphylococcal protein A-V antigen fusion peptide into mice delayed allograft rejection and suppressed the major proinflammatory cytokines tumor necrosis factor alpha (TNF-alpha) and gamma interferon (IFN-gamma) associated with generation of protective granulomas. This study was undertaken to determine if V antigen could prevent endotoxic shock, known to be mediated by excessive production of certain proinflammatory cytokines. After treatment with 50 microg of homogeneous V antigen-polyhistidine fusion peptide (Vh), the 50% lethal dose of purified lipopolysaccharide (LPS) in BALB/c mice immediately rose from 63 microg (normal controls) to 318 microg, fell to near baseline (71 microg) in 6 h, and then slowly rose to a maximum of 566 microg at 48 h before again returning to normal. Injected Vh alone (50 microg) promptly induced the anti-inflammatory cytokine interleukin-10 (IL-10) as well as modest levels of TNF-alpha (an inducer of IL-10) in spleen. Concomitant injection of Vh and an otherwise lethal dose of LPS (200 microg) dramatically decreased levels of TNF-alpha and IFN-gamma in the spleen and peritoneal lavage fluid as compared to values determined for LPS alone. These results would be expected if V antigen directly up-regulated IL-10 that is reported to generally down-regulate proinflammatory cytokines. Mice receiving 200 microg of LPS 48 h after injection of Vh exhibited patterns of cytokine synthesis similar to those observed in endotoxin-tolerant mice, a condition also reported to be mediated by IL-10. These findings suggest that V antigen serves as a virulence factor by amplifying IL-10, thereby repressing proinflammatory cytokines required for expression of cell-mediated immunity.     

The regulation and functional consequence of proinflammatory cytokine binding on human intestinal epithelial cells

Products of an activated immune system may affect cells within the immune system as well as nonlymphoid cells in the local environment. Given the immunologically activated state of the intestinal tract, it is conceivable that locally produced cytokines could regulate epithelial cell function. To assess whether epithelial cells are targets for particular cytokines, we initiated studies on the binding of a panel of proinflammatory cytokines in freshly isolated epithelial cells from normal and inflammatory bowel disease (IBD) patients as well as in cell lines. Isolated intestinal epithelial cells (IEC) were stained with phycoerythrin-conjugated or biotinylated cytokines to determine the expression and density of receptors for IL-1beta, IL-6, granulocyte-macrophage CSF (GM-CSF), and TNF-alpha. Receptors for IL-1beta, IL-6, and GM-CSF were readily detectable in all epithelial cell preparations at levels equal to (GM-CSFR) or lower than those seen on monocytes. However TNFalpha-R were not detectable on freshly isolated IECs. Receptor density was greater in surface vs crypt epithelial cells, but no significant differences were seen between normal and IBD epithelial cells. Expression of IL-1R and IL-6R was enhanced by LPS and IFN-gamma. Functionally, IL-1beta enhanced proliferation of the IEC cell line, DLD1, whereas GM-CSF treatment of de-differentiated crypt-like DLD1 and HT29 cells resulted in enhanced expression of ICAM-1. Furthermore, TNF-alpha treatment enhanced the secretion of IL-8 and GRO-alpha in HT29 cells, but not in freshly isolated IEC cultures. The differential binding and function of proinflammatory cytokines on IEC support the hypothesis that these cytokines may be involved in normal physiological processes as well as in regulating mucosal immune responses.     

Aberrant cytokine expression and autocrine regulation characterize macrophages from young MRL+/+ and NZB/W F1 lupus-prone mice

We investigated whether macrophages (Mphi) from young, lupus-prone MRL+/+ and NZB/W F1 mice expressed common defects in immunoregulatory cytokine production. Endotoxin-activated Mphi from both strains, obtained well before disease signs, had a markedly reduced capacity to maintain IL-1 production compared with Mphi from normal strains (BALB/c, A/J, and C57BL/6). Mphi from lupus-prone mice showed similar defects in IL-6 and TNF-alpha production, which preceded the IL-1 defect. In fact, defective TNF-alpha production appeared to be responsible for aberrant expression of the other cytokines because this defect was the first to be expressed, and treatment with exogenous TNF-alpha reduced the extent of defective IL-1 and IL-6. These "proinflammatory" cytokine defects appeared to be selective because the anti-inflammatory cytokine IL-10 was not expressed aberrantly in the lupus-prone strains. For this reason, and because anti-IL-10 mAb treatment did not correct defective proinflammatory cytokine production, IL-10 did not appear to be responsible for these defects. IFN-gamma was able to normalize TNF-alpha production in Mphi from lupus-prone mice, demonstrating a stimulus-specific induction of the proinflammatory defects. These studies also revealed that Mphi from the three normal strains studied here maintain a precise inverse relationship between levels of TNF-alpha and IL-10, a relationship not seen in Mphi from lupus-prone strains. These findings reveal shared elements of cytokine dysregulation in the two principal animal models of multigenic lupus, and suggest that the study of Mphi (and perhaps other cells of the innate immune system) may provide valuable insights into intrinsic functional defects associated with systemic autoimmunity.     

Evaluation of the hairless rat as a model for in vivo percutaneous absorption

Interleukin-10 (IL-10) is known to inhibit T cell-mediated responses. IL-10 has also been shown to play an important pathogenetic role in allergic diseases. Glucocorticoid is known to inhibit the production and gene expression of many cytokines which induce inflammatory reactions. We examined the effect of dexamethasone on the gene expression and production of IL-10 by human peripheral blood mononuclear cells (PBMCs) and monocytes. PBMCs and monocytes from 5 healthy volunteers were incubated with or without dexamethasone for 1 h, then stimulated with 5 micrograms/ml lipopolysaccharide (LPS). Gene expression and production of IL-10 by human PBMCs were detected without stimulation and increased by LPS stimulation. Dexamethasone suppressed the gene expression and production of IL-10 by LPS-stimulated PBMCs in a dose-dependent manner by 41.6 and 61.1% at 10(-6) M, respectively. Also in monocytes, the gene expression and production of IL-10 were detected without stimulation, increased by LPS stimulation, and significantly suppressed by dexamethasone by 53.1 and 61.2% at 10(-6) M, respectively. This suppressive effect on IL-10 gene expression was not so potent compared with its effect on cytokines such as IL-5. The suppression of IL-10 production by glucocorticoid is suggested to be one of the important mechanisms by which glucocorticoids suppress allergic inflammation in the treatment of allergic diseases.     

Immunomodulatory activities of oat beta-glucan in vitro and in vivo

Previous studies have shown that beta-glucans extracted from yeast or fungi potentiate immune responses. In the present study, the immunomodulatory activities of beta-(1-->3,1-->4)-glucan, derived from oats, were investigated. The ability of oat beta-glucan (ObetaG) to stimulate IL-1 and TNF-alpha release from murine peritoneal macrophages and the murine macrophage cell line P338D1, was assessed. In vitro stimulation of macrophages with ObetaG resulted in the production of IL-1 in a dose and time-dependent manner, whereas only small amounts of TNF-alpha could be detected in the culture supernatants. ObetaG also induced the production of IL-2, IFN-gamma and IL-4 secretion in a dose-dependent manner in cultured spleen cells. The intraperitoneal administration of ObetaG in mice resulted in the accumulation of leucocytes, predominantly macrophages, in the peritoneal cavity. Furthermore, ObetaG was tested for its ability to enhance non-specific resistance to a bacterial challenge in mice. Survival of mice challenged with Staphylococcus aureus was enhanced by a single intraperitoneal administration of 500 microg of ObetaG 3 days prior to bacterial challenge. In conclusion, these studies demonstrated that ObetaG possesses immunomodulatory activities capable of stimulating immune functions both in vitro and in vivo.     

Somitogenesis: segmenting a vertebrate

OBJECTIVE: To evaluate the effect of gliclazide administration to NIDDM patients on 1) monocyte adhesion to cultured endothelial cells, 2) plasma cytokine and lipid peroxide levels, and 3) monocyte cytokine production. RESEARCH DESIGN AND METHODS: Poorly controlled glyburide-treated diabetic patients (n = 8) and healthy control subjects (n = 8) were recruited. At the beginning of the study, glyburide was replaced by an equivalent hypoglycemic dose of gliclazide. Serum and monocytes were isolated from blood obtained from control and diabetic subjects before and after 3 months of treatment with gliclazide. RESULTS: Plasma lipid peroxide levels and monocyte adhesion to endothelial cells are enhanced in NIDDM patients, and gliclazide administration totally reverses these abnormalities. Before gliclazide treatment, serum levels of cytokines did not differ in the control and the diabetic groups, with the exception of an enhancement of tumor necrosis factor-alpha (TNF-alpha) and interleukin-6 (IL)-6 in NIDDM subjects. Basal and lipopolysaccharide (LPS)-stimulated monocyte production of interleukin-1 beta, IL-6, and IL-8 did not differ between the two groups. Furthermore, basal monocyte production of TNF-alpha was similar in the control and the diabetic groups, whereas a marked increase in the LPS-stimulated monocyte production of TNF-alpha was observed in the NIDDM group. Gliclazide treatment lowered LPS-stimulated TNF-alpha production by diabetic monocytes to levels similar to those observed in control subjects. CONCLUSIONS: Gliclazide administration to NIDDM patients inhibits the increased adhesiveness of diabetic monocytes to endothelial cells and reduces the production of TNF-alpha by these cells. These results suggest that treatment of NIDDM subjects with gliclazide may be beneficial in the prevention of atherosclerosis associated with NIDDM.     

Effects of tumor necrosis factor-alpha and interleukin-6 on fluid-phase permeability and ammonia diffusion in CNS-derived endothelial cells

BACKGROUND: Proinflammatory cytokines tumor necrosis factor-alpha (TNF-alpha) and interleukin-6 (IL-6) play an important role in the blood-brain barrier breakdown present in several neurological diseases including multiple sclerosis and AIDS. However, the specific effects of these cytokines on central nervous system-derived endothelial cells (CNS-EC) is not fully understood. In this study the effects of TNF-alpha and IL-6 were tested on different permeability mechanisms of CNS-EC. METHODS: Central nervous system endothelial cells were isolated from human brain and retina and cultured in vitro in a transwell system. Fluid-phase endocytosis and transcytosis, absorptive-mediated endocytosis, and ammonia diffusion were measured with specific methods. Endothelial cells were studied with electron microscopy for the ultrastructural effects of cytokine stimulation. RESULTS: Fluid-phase endocytosis and transcytosis were significantly increased by TNF-alpha and IL-6. This effect was dose dependent and reversible. The ammonia diffusion rate was also significantly increased by TNF-alpha. Absorptive-mediated endocytosis was not enhanced by TNF-alpha. Ultrastructural analysis of cytokine-treated CNS-EC confirmed the alterations in permeability showing an increase in endocytotic activity and a decrease in tight junctions. CONCLUSIONS: The proinflammatory cytokines IL-6 and TNF-alpha induce specific changes in the morphology and permeability of CNS-EC. These alterations can be important in many diseases characterized by increased cytokine production.