Biphasic production of IL-8 in lipopolysaccharide (LPS)-stimulated human whole blood. Separation of LPS- and cytokine-stimulated components using anti-tumor necrosis factor and anti-IL-1 antibodies.

TNF, IL-1, and IL-6 are integral components of the cytokine cascade released in the response to inflammatory stimuli such as LPS. IL-8 is produced both in response to LPS as well as TNF and IL-1. The early, local production of TNF and IL-1 may therefore contribute to the subsequent expression of IL-8. This hypothesis was tested using LPS-stimulated human whole blood as an ex vivo model of local cytokine production. The production of TNF, IL-1 alpha, IL-1 beta, IL-6, and IL-8 was found to be responsive to a wide range of LPS concentrations (0.1 ng/ml-10 micrograms/ml). These cytokines were first detected between 1 to 4 h post-LPS stimulation, and reached plateau levels after 6 to 12 h. IL-8, however, also displayed a secondary wave of production, with the levels again increasing between 12 to 24 h. The IL-8 present in the plasma after LPS stimulation was biologically active, as assessed by neutrophil chemotaxis. In further studies, addition of anti-TNF and anti-IL-1 neutralizing antibodies, alone and in combination, to LPS-stimulated blood resulted in nearly complete ablation of the secondary phase of IL-8 synthesis at both the levels of protein and mRNA, while leaving the first, LPS-mediated phase of IL-8 synthesis unaffected. This model of cytokine production in human whole blood may reflect the sequence of events in a localized environment of inflammation where both a primary stimulus and the induced early cytokine mediators may serve to elicit multiple, temporally distinct phases of IL-8 production.     

Early response cytokines and innate immunity: essential roles for TNF receptor 1 and type I IL-1 receptor during Escherichia coli pneumonia in mice

The early response cytokines, TNF and IL-1, have overlapping biologic effects that may function to propagate, amplify, and coordinate host responses to microbial challenges. To determine whether signaling from these early response cytokines is essential to orchestrating innate immune responses to intrapulmonary bacteria, the early inflammatory events induced by instillation of Escherichia coli into the lungs were compared in wild-type (WT) mice and mice deficient in both TNF receptor 1 (TNFR1) and the type I IL-1 receptor (IL1R1). Neutrophil emigration and edema accumulation induced by E. coli were significantly compromised by TNFR1/IL1R1 deficiency. Neutrophil numbers in the circulation and within alveolar septae did not differ between WT and TNFR1/IL1R1 mice, suggesting that decreased neutrophil emigration did not result from decreased sequestration or delivery of intravascular neutrophils. The nuclear translocation of NF-kappa B and the expression of the chemokine macrophage inflammatory protein-2 did not differ between WT and TNFR1/IL1R1 lungs. However, the concentration of the chemokine KC was significantly decreased in the bronchoalveolar lavage fluids of TNFR1/IL1R1 mice compared with that in WT mice. Thus, while many of the molecular and cellular responses to E. coli in the lungs did not require signaling by either TNFR1 or IL1R1, early response cytokine signaling was critical to KC expression in the pulmonary air spaces and neutrophil emigration from the alveolar septae.     

Ciliary neurotrophic factor inhibits brain and peripheral tumor necrosis factor production and, when coadministered with its soluble receptor, protects mice from lipopolysaccharide toxicity.

BACKGROUND: The receptor of ciliary neurotrophic factor (CNTF) contains the signal transduction protein gp130, which is also a component of the receptors of cytokines such as interleukin (IL)-6, leukemia-inhibitory factor (LIF), IL-11, and oncostatin M. This suggests that these cytokines might share common signaling pathways. We previously reported that CNTF augments the levels of corticosterone (CS) and of IL-6 induced by IL-1 and induces the production of the acute-phase protein serum amyloid A (SAA). Since the elevation of serum CS is an important feedback mechanism to limit the synthesis of proinflammatory cytokines, particularly tumor necrosis factor (TNF), we have investigated the effect of CNTF on both TNF production and lipopolysaccharide (LPS) toxicity. MATERIALS AND METHODS: To induce serum TNF levels, LPS was administered to mice at 30 mg/kg i.p. and CNTF was administered as a single dose of 10 micrograms/mouse i.v., either alone or in combination with its soluble receptor sCNTFR alpha at 20 micrograms/mouse. Serum TNF levels were the measured by cytotoxicity on L929 cells. In order to measure the effects of CNTF on LPS-induced TNF production in the brain, mice were injected intracerebroventricularly (i.c.v.) with 2.5 micrograms/kg LPS. Mouse spleen cells cultured for 4 hr with 1 microgram LPS/ml, with or without 10 micrograms CNTF/ml, were also analyzed for TNF production. RESULTS: CNTF, administered either alone or in combination with its soluble receptor, inhibited the induction of serum TNF levels by LPS. This inhibition was also observed in the brain when CNTF and LPS were administered centrally. In vitro, CNTF only marginally affected TNF production by LPS-stimulated mouse splenocytes, but it acted synergistically with dexamethasone (DEX) in inhibiting TNF production. Most importantly, CNTF administered together with sCNTFR alpha protected mice against LPS-induced mortality. CONCLUSIONS: These data suggest that CNTF might act as a protective cytokine against TNF-mediated pathologies both in the brain and in the periphery.     

Regulation of bronchoalveolar macrophage proinflammatory cytokine production by dexamethasone and granulocyte-macrophage colony-stimulating factor after stimulation by Aspergillus conidia or lipopolysaccharide

There is substantial evidence that local production of proinflammatory cytokines are very important in host resistance to aspergillosis. Dexamethasone (DEX) down-regulates production of these cytokines by stimulated bronchoalveolar macrophages (BAM) and constitutes a risk factor for aspergillosis. Granulocyte-macrophage colony-stimulating factor (GM-CSF) antagonizes DEX suppression of antifungal activity by BAM. Here we investigated the possibility that GM-CSF could antagonize DEX down-regulation of interleukin (IL)-1alpha and tumour necrosis factor (TNF)-alpha production by stimulated BAM. Control BAM responded to increasing numbers of conidia of Aspergillus fumigatus with increasing production of IL-1 and TNF. DEX (10(-7)M) significantly suppressed IL-1 and TNF production by BAM+conidia. Although GM-CSF did not enhance IL-1 or TNF production by BAM+conidia, GM-CSF significantly antagonized DEX suppression of IL-1 cytokine production. For comparative purposes, lipopolysaccharide (LPS, 1 microg/ml) was used to stimulate BAM in experiments similar to the above. In contrast to the findings with conidia, GM-CSF enhanced the production of IL-1 (5-fold) and TNF (1.5-fold) by LPS treated BAM. DEX suppression of cytokine production by BAM+LPS was modestly but significantly antagonized by GM-CSF. Moreover, differences between regulation of IL-1 and TNF production by BAM+conidia or LPS and peritoneal macrophages (PM)+conidia or LPS were documented. Finally, the anti-inflammatory cytokine IL-10 was minimally produced by BAM + conidia or LPS, but IL-10 was produced by PM + conidia or LPS. In summary, these data indicate that the risk factor for aspergillosis associated with DEX could be lessened in the pulmonary compartment with GM-CSF. On the other hand, desired effects of DEX could be maintained in other compartments.     

Induction of C3 expression in astrocytes is regulated by cytokines and Newcastle disease virus.

Synthesis of complement proteins and their regulation in resident cells of the central nervous system are important pathophysiologic factors that can affect the outcome of inflammatory central nervous system diseases. Primary cultures of rat astrocytes constitutively express C3 mRNA and produce C3 protein; both of them were enhanced by LPS or by a live as well as inactivated Newcastle disease virus, a neurotropic paramixovirus. TNF, IL-1 beta, and IL-8 also increased the levels of C3 mRNA and protein whereas IL-1 alpha and IL-6 had no effect, although all of these cytokines are inducible by LPS. LPS stimulation in the presence of cycloheximide decreased the LPS-mediated C3 mRNA induction by 60%. These data suggest that LPS effect on C3 regulation is mediated directly by LPS as well as by LPS-induced cytokines. Interestingly, C3 mRNA induced by Newcastle disease virus or inactivated Newcastle disease virus was inhibited by protein kinase inhibitors, H-7 and staurosporine, whereas these inhibitors had no effect on C3 induction mediated by LPS or cytokines, indicating the existence of different signal transduction pathways.     

IL-8 in septic shock, endotoxemia, and after IL-1 administration

Much effort has been directed toward elucidating the host response to sepsis and inflammation, resulting in the definition of a cascade of endogenous mediators that direct metabolic and immunological responses. Here we report that IL-8, a novel cytokine produced by a variety of cells in vitro in response to stimulation with bacterial LPS and the proinflammatory cytokines, appears in the circulation of primates in vivo during septic shock, sublethal endotoxemia, and after the administration of IL-1 alpha. The magnitude of the IL-8 response correlates with the severity of the insult, and levels of IL-8 peak relatively late, after those of TNF-alpha and IL-1 beta, and simultaneously with those of IL-6. IL-8 has been primarily defined as a selective activator and chemoattractant of neutrophils, and we demonstrate that after LPS or IL-1 alpha infusion, circulating neutrophil numbers rapidly recover from an initial neutropenia while IL-8 concentrations are maximal, supporting the hypothesis that IL-8 influences circulating leukocyte populations in vivo. We conclude that IL-8 is another participant in the cytokine cascade elicited by sepsis and inflammation and, as such, may play a significant role in host defense and disease.     

Different respiratory syncytial virus and Quillaja saponin formulations induce murine peritoneal cells to express different proinflammatory cytokine profiles

The recognition of a pathogen or a vaccine antigen formulation by cells in the innate immune system leads to production of proinflammatory cytokines, which will determine the ensuing acquired immune response quantitatively and qualitatively. Tumour necrosis factor (TNF)-alpha, interleukin (IL)-1 and IL-6 are the first set of cytokines produced upon such an encounter, which have roles both in protective immunity and immunopathogenesis evident with respiratory syncytial virus (RSV). RSV antigens in different physical adjuvant-vaccine formulations were analysed for their capacity to provoke cultured murine peritoneal cells to produce these three proinflammatory cytokines. RSV immunostimulating complex (ISCOM), i.e. both antigen and adjuvant are incorporated in the same particle, induced high levels of IL-1alpha being of the same magnitude or higher than those of live RSV and lipopolysaccharide (LPS). Live virus and LPS induced higher levels of IL-6 and TNF-alpha than ISCOM and so did non-adjuvanted UV-inactivated RSV but only at high doses. ISCOM-Matrix, i.e. ISCOM without antigens, admixed as a separate entity to inactivated RSV, downregulated or blocked the cytokine response to the inactivated RSV in contrast to ISCOM. Kinetic studies showed that ISCOM induced cytokine production first detected at hours 1, 2, 4 for TNF-alpha, IL-6 and IL-1alpha respectively, which was earlier than for the other antigen formulations containing corresponding doses of antigen and/or Quillaja adjuvant. Peak values for production of TNF-alpha and IL-6 were at 8 h and for IL-1alpha at 72 h following stimulation with ISCOM. The delayed appearance of IL-1alpha may reflect the cell-bound nature of this cytokine.     

Native LDL potentiate TNF alpha and IL-8 production by human mononuclear cells

Native LDL (nLDL) increases expression of adhesion molecules on endothelial cells through induction of Ca(2+) mobilization. Ca(2+) mobilization is also involved in the induction of proinflammatory cytokines, important mediators involved in atherogenesis. The aim of the study was to evaluate the capacity of nLDL to affect spontaneous and lipopolysaccharide (LPS)-stimulated cytokine production. Preincubation of human peripheral blood mononuclear cells (PBMC) with nLDL for 24 h did not influence spontaneous production of tumor necrosis factor alpha (TNF alpha) or interleukin-8 (IL-8), but significantly potentiated LPS-induced production of these cytokines. nLDL preincubation of PBMC did not increase the expression of the LPS receptors Toll-like receptor-4, CD14, or CD11c/CD18. Potentiation of cytokine production by nLDL was mediated through induction of Ca(2+) mobilization, because: a) nLDL induced a sustained pattern of repetitive Ca(2+) transients in human PBMC; b) the Ca(2+) chelator fura 2-acetoxymethyl ester, 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid, an intracellular Ca(2+) chelator, inhibited the potentiating effect of nLDL on LPS-induced cytokine synthesis; c) induction of Ca(2+) mobilization by thapsigargin potentiated LPS-induced cytokine production. nLDL are able to potentiate LPS-induced production of cytokines by human PBMC, and this effect is probably mediated through induction of Ca(2+) mobilization. This may represent an important pathogenetic mechanism in atherogenesis induced by hyperlipoproteinemia.     

Interleukin-1beta and tumor necrosis factor-alpha mediation of endotoxin action on growth hormone

In humans and sheep, endotoxin (LPS) administration results in increased growth hormone (GH) concentrations. To determine the role of cytokines in the effect of LPS on GH, sheep were challenged with IL-1beta or TNF-alpha. GH data were compared with results with LH, where the major effects of LPS are known to act via the hypothalamus. Intracerebroventricular (icv) administration of IL-1beta or TNF-alpha did not alter plasma concentrations of GH. Endotoxin was then administered intravenously (iv) in combination with icv injection of IL-1 receptor antagonist (IL-1RA), TNF antagonist (sTNF-R1), or saline. Administration of LPS increased GH (P < 0.0001), although coadministration of IL-1ra or sTNF-R1 icv did not alter GH response to LPS. In contrast, plasma concentrations of LH were profoundly inhibited by icv administration of either cytokine (P < 0.03), but the LH response to LPS was not altered by cytokine antagonists. Intravenous administration of either IL-1beta or TNF-alpha increased plasma concentrations of GH (P < 0.0001). Administration of IL-1RA and sTNF-R1 iv prevented LPS-induced increases in GH. Although LH was suppressed by high iv doses of IL-1beta (P = 0.0063), the antagonists did not alter the LH response to LPS. To determine whether LPS might directly activate GH release, confocal microscopy revealed colocalization of CD14, the LPS receptor, with GH and, to a lesser extent, LH and some prolactin (PRL)-containing cells, but not ACTH or TSH. These data are consistent with the effects of LPS on GH secretion originating through peripheral cytokine presentation to the pituitary, as well as a potential to act directly on selective populations of pituitary cells via CD14.     

Differential effects of hemorrhage and LPS on tissue TNF-alpha, IL-1 and associate neuro-hormonal and opioid alterations

LPS administration and hemorrhage are frequently used models for the in vivo study of the stress response. Both challenges stimulate cytokine production as well as activate opiate and neuro-endocrine pathways; which in turn modulate the inflammatory process. Differences in the magnitude and tissue specificity of the proinflammatory cytokine and neuro-hormonal responses to these stressors are not well established. We contrasted the tissue specificity and magnitude of the increase in circulating and tissue cytokine (TNF-alpha, IL-1alpha and IL-1beta) content in response to either fixed-pressure hemorrhage (approximately 40 mm Hg) followed by fluid resuscitation (HEM) or lipopolysaccharide (LPS; 100 microg/100 g BW) administration. LPS and HEM elevated circulating levels of TNF-alpha, while neither stress altered circulating IL-1-alpha and IL-beta. LPS-induced increases in TNF-alpha content were greater than those elicited by HEM in all tissues studied except for the lung, where both stressors produced similar increases. Tissue (lung, spleen and heart) content of IL-1alpha was increased by HEM but was not affected by LPS. Tissue (lung, spleen, and heart) content of IL-1beta was increased by LPS but was not affected by HEM. HEM produced greater increases than LPS in epinephrine (16- vs. 4-fold) and norepinephrine (4-fold vs. 60%) levels and similar elevations in beta-endorphin. LPS produced greater elevation in corticosterone levels (2-fold) than HEM (50%). These results suggest differential tissue cytokine modulation to HEM and LPS, both with respect to target tissue and cytokine type. The hormonal milieu to HEM is characterized by marked catecholaminergic and moderate glucocorticoid while that of LPS is characterized by marked glucocorticoid with moderate catecholaminergic influence.     

Endotoxin tolerance in rats: expression of TNF-alpha, IL-6, IL-10, VCAM-1 AND HSP 70 in lung and liver during endotoxin shock.

Endotoxin can induce a state of tolerance against its own pathological effects, commonly referred to as endotoxin tolerance. This phenomenon has been found to be associated with reduced serum levels of cytokines such as TNF-alpha, IL-1, IL-6 and IL-10. In the present study the expression of TNF-alpha, IL-6, IL-10, the adhesion molecule VCAM-1 and the heat shock protein 70 was determined in vivo in lung and liver of LPS-tolerant and naive rats by means of semiquantitative RT-PCR after i.v. LPS injection. TNFalpha, IL-6, IL-10, HSP 70 and VCAM-1 were induced in lung and liver after LPS injection. In liver and lung of endotoxin-tolerant rats TNF-alpha and IL-6 were induced to a lower degree after LPS treatment when compared to non-tolerant controls. The LPS-induced IL-10 expression was also slightly attenuated in the lung of tolerant rats, but in the liver no differences between tolerant and non-tolerant animals were observed. HSP 70 and VCAM-1 were expressed after systemic LPS treatment in liver and lung. The degree of induction, however, was the same in tolerant and untreated controls. The presented data show that endotoxin tolerance is reflected by a reduced cytokine expression in lung and liver in vivo. On the other hand, levels of expression of the adhesion molecule VCAM-1 and the stress protein HSP 70 do not appear to be changed by endotoxin tolerance.     

Effectiveness of a cytokine restraining agent (CRA (TM)) in attenuating disuse deconditioning induced by hindlimb unloading in rats

Spaceflight alters many immune responses and among the regulatory components of an organisms response system that have been to be affected by spaceflight is the cytokine network. Spaceflight, as well as ground-based model systems of spaceflight, have been shown to affect the production and activation of various cytokines including interleukins (IL) and tumor necrosis factor (TNF). Levels of urinary IL-2 are elevated on the first day of spaceflight and again after returning from space. Most results from ground-based studies in rodents indicate either no alterations in cytokines or decreased levels. Results from this experiment indicate that HP 228, a potent cytokine restraining agent (CRA (TM)) was effective in attentuating many of the disuse deconditioning changes induced by the ground-based hindlimb suspension model that simulates weightlessness in rats. HP 228 is a novel heptapeptide with unnatural amino acids and can effectively restrain lipopolysaccharide (LPS)-induced increased levels of several key cytokines, including plasma TNF alpha, IL-1 beta and IL-6. HP 228 has also been shown to be effective in several rodent models of pain, inflammation and LPS-induced lethality, as well as in reducing inducible nitric oxide synthase.     

Synergistic interactions of interleukin 1, interferon-beta, and tumor necrosis factor in terminally differentiating a mouse myeloid leukemic cell line (M1). Evidence that interferon-beta is an autocrine differentiating factor

The effect was investigated of combinations of cytokines known to be cytostatic for some tumor cells, namely interleukin 1 alpha (IL-1 alpha), interferon-beta (IFN-beta), and tumor necrosis factor (TNF), on the growth and differentiation of the mouse myeloid leukemic cell line, M1, cells. IL-1 alpha, IFN-beta, and TNF by themselves are antiproliferative for M1 cells. Treatment of cells with a mixture of any two of the three cytokines resulted in at least additive growth inhibition. None of these cytokines by themselves induced differentiation of M1 cells as assessed by increased expression of Fc receptors (FcR), stimulation of phagocytic activity and by morphologic criteria. However, as little as 1 U/ml IL-1 alpha in conjunction with IFN-beta or TNF increased FcR expression, phagocytic activity and morphologic changes in addition to inhibiting the growth of M1 cells. The combination of IFN-beta and TNF did not induce differentiation, although the growth of the cells was markedly inhibited. Both TNF and lipopolysaccharide (LPS) induced the in vitro production of IFN activity by M1 cells. Furthermore, the induction of differentiation of M1 cells by a combination of IL-1 alpha with either IFN-beta, TNF, or LPS was inhibited by antibody against mouse IFN-beta. Therefore, it appears that IFN-beta provides one of the two required signals for differentiation of M1 cells by these combinations of stimulants, the other being IL-1. Furthermore, the cytostatic effect of TNF by itself on M1 cells was also partly blocked by anti-IFN-beta antibody, suggesting that IFN-beta is also involved in the growth inhibitory effect of TNF for M1 cells. In contrast, the cytostatic effect of IL-1 on M1 cells was not blocked by anti-IFN-beta antibody. In conclusion, both the cytostatic and differentiative effect of TNF appear to be mediated by IFN-beta. Thus, the combination of IL-1 and IFN-beta or inducers of IFN-beta resulted in terminal differentiation of M1 cells. Northern blot analysis using cDNAs for murine IFN-beta1 or human IFN-beta2 showed an increased expression of mRNA for IFN-beta1 but not for IFN-beta2 by stimulation with TNF or LPS, strongly suggesting that IFN-beta 1 rather than IFN-beta 2 is responsible for TNF or LPS effects.     

Depressed IL-1 production by chronic GVHD dermal fibroblasts

Chronic Graft-versus-Host disease (GVHD) is characterized by overt immunosuppression. In addition, the skin is a major anatomical site affected in chronic GVHD for reasons not yet known. Increased collagen deposition, a mononuclear cell infiltrate in the dermis as well as loss of fat and appendages, are observed in the skin. The inflammatory cytokine IL-1 was shown to affect fibroblast proliferation and secretory activities. In the present study, IL-1 generation by dermal fibroblasts, of chronic GVHD or control mice, was assessed. It was shown that two sequential signals are needed for IL-1 generation by dermal fibroblasts; priming by lymphokines/cytokines followed by a challenge with LPS. A variety of recombinant lymphokines and cytokines (G/M-CSF, IL-2, TNF, IL-1 beta and IFNs alpha, beta and gamma) were shown to be efficient in priming dermal fibroblasts for IL-1 generation. IL-1 activity in dermal fibroblasts, most probably of the IL-1 alpha species, was located in frozen-thawed cell lysates or associated to the cell membrane, though not secreted into the culture fluids. Dermal fibroblasts from chronic GVHD mice manifested a pronounced depression in IL-1 generation upon stimulation with exogenous lymphokines/cytokines and LPS. This was observed over a wide range of concentrations of lymphokines/cytokines and LPS. The depressed ability of chronic GVHD fibroblasts to generate IL-1 was pronounced even after few passages of the cells in vitro, and upon stimulation in culture outside the suppressive milieu of the animal.(ABSTRACT TRUNCATED AT 250 WORDS)     

Migration of neutrophils across endothelial monolayers is stimulated by treatment of the monolayers with interleukin-1 or tumor necrosis factor-alpha

To study the effects of the cytokines IL-1 and TNF-alpha on the transendothelial migration of neutrophils, human umbilical vein endothelial cells (HUVEC) were grown to confluence on connective tissue prepared from human amniotic membrane. Pretreatment of HUVEC-amnion cultures with rIL-1 beta (7.5 ng/ml) or rTNF-alpha (5 ng/ml) for 4 h resulted in rapid migration of from 20 to 50% of subsequently added neutrophils across the endothelial monolayer. In contrast, only 3 +/- 3% of added neutrophils penetrated the HUVEC monolayer in the absence of any stimulus. The number of neutrophils that migrated across cytokine-treated HUVEC was similar to the number that traversed untreated monolayers in response to gradients of FMLP; in addition, it was only 35% less than the number of neutrophils that migrated in response to leukotriene B4. No consistent additive effect was seen when migration was induced by both cytokine pretreatment of the HUVEC and a chemotactic gradient. The number of neutrophils that migrated across IL-1-treated cultures was proportional to the number added over the range of 2.5 x 10(5) to 4 x 10(6) neutrophils. When used at optimal concentrations, IL-1 and TNF-alpha were equally effective in stimulating neutrophil migration; no additive effect was seen when HUVEC were pretreated with optimal doses of both cytokines together. Direct addition of IL-1 or TNF-alpha to a 1-h migration assay had no effect on neutrophil adhesion to or migration across HUVEC, either in the presence or absence of a chemotactic gradient. Stimulation of neutrophil transendothelial migration in this system did not appear to be caused by adsorption of cytokine by the amniotic tissue, nor was it due to contamination of the cytokine preparations by LPS. These results suggest that IL-1 and TNF-alpha, generated at sites of inflammation, may act upon the endothelium to promote emigration of neutrophils from the vasculature.     

Involvement of class II MHC molecules in the LPS-induction of IL-1/TNF secretions by human monocytes. Quantitative differences at the polymorphic level

Anti-class II ag mAb (DR and DQ) inhibited, in a dose-dependent manner, LPS-induced IL-1 and TNF secretions from human monocytes (34 to 95% inhibition). The potentiating effect of IFN-gamma on LPS-induced TNF secretion (15.3 +/- 0.7 to 44 +/- 0.6 ng/ml) was also blocked by anti-class II ag mAb (44 +/- 0.6 to 0.3 +/- 0.03 ng/ml). We also report a relationship between interindividual differences in monocyte IL-1 and TNF secretions and the HLA-D-encoded genetic polymorphism. Heterozygotes were, in general, higher secretors of those cytokines than homozygotes. Analysis of these secretions in heterozygotes demonstrated a differential effect of certain haplotype combinations (i.e., DR2-DR4 vs DR2-DR3) that could be arbitrarily characterized as being "low" or "high" secretors (6,230 +/- 2,950 vs 13,029 +/- 6,541 cpm for IL-1, and 12 +/- 10 vs 25 +/- 15 ng/ml for TNF, p = 0.006 and 0.048). DR-associated Dw subtypes appeared to account for differences within certain haplotype combinations (Dw18 vs Dw19 in DRw13/DR4) (11,227 +/- 3,648 vs 17,166 +/- 3,176 cpm for IL-1, and 13 +/- 9 vs 25 +/- 10 ng/ml for TNF, p = 0.02 and 0.047). Interindividual differences were better explained by differences in LPS sensitivity than by differences in the kinetics of secretion and related not to the secretory process itself but to the rate of cytokine synthesis. Finally, there were no relationships between high secretor genotypes and IDD high risk genotypes. Thus, we conclude that, a) LPS-induced IL-1 and TNF secretions are, at least in part, regulated by class II MHC molecules, b) that HLA-D region-encoded genetic polymorphism accounts for interindividual differences in these secretions, and c) that the HLA-associated risk to develop IDD is not explained by these cytokine secretory differences as previously proposed.