IL-1beta mRNA Expression Research Articles

Oxidized low density lipoprotein inhibits lipopolysaccharide-induced binding of nuclear factor-kappaB to DNA and the subsequent expression of tumor necrosis factor-alpha and interleukin-1beta in macrophages.

A large body of evidence suggests that oxidized LDL (oxLDL) has a role in atherogenesis. One effect is the impact on macrophage function. We have studied the effects of oxLDL and oxysterols on the binding of the transcription factors nuclear factor (NF)-kappaB and AP-1 to DNA. These transcription factors are involved in the regulation of several genes and expressed during activation of macrophages, for example by endotoxin (LPS). OxLDL did not induce binding of NF-kappaB. However, the LPS-induced response to NF-kappaB was substantially reduced after preincubation with oxLDL. Medium and highly oxidized LDL also decreased the constitutive DNA-binding of AP-1. Similar effects on AP-1-binding were seen with the oxysterols, 7beta-hydroxycholesterol, 24- hydroxy-, 25-hydroxy-, and 27-hydroxy-cholesterol. Our data therefore suggest an effect of oxLDL on the DNA-binding of AP-1, which might be mediated by the oxysterol content of oxLDL. A decreased LPS-induced TNF-alpha and IL-1beta mRNA and protein expression were found in macrophages incubated with oxLDL before LPS-exposure. These observations suggest that macrophages that internalize extensively oxidized LDL are suppressed in their response to inflammatory stimulation.

Early and Persistent Alterations in the Expression of Interleukin-1α, Interleukin-1β and Tumor Necrosis Factor α mRNA Levels in Fibrosis-Resistant and Sensitive Mice after Thoracic Irradiation

Fibrosis, characterized by the accumulation of collagen, is a consequence of a chronic inflammatory response. The purpose of this study was to determine if tumor necrosis factor alpha (TNF-alpha), interleukin-1 alpha (IL-1 alpha) and IL-1 beta mRNA expression are altered acutely after irradiation, during the so-called "latent" phase of pulmonary injury, and to examine if these alterations persist through the development of pneumonitis and fibrosis. Further, we wished to determine if these changes differ between two strains of mice which vary in their sensitivity to radiation. Fibrosis-sensitive (C57BL/6) and fibrosis-resistant (C3H/HeJ) mice were irradiated with a single dose of 5 or 12.5 Gy to the thorax. Total lung RNA was prepared and immobilized by slot blotting and hybridized with radiolabeled cDNA probes encoding for TNF-alpha, IL-1 alpha and IL-1 beta. Autoradiographic data were quantified by video densitometry and results normalized to a control probe encoding for glyceraldehyde-3-phosphate dehydrogenase. It was found that TNF-alpha mRNA levels were increased in C57BL/6 mice at days 1 and 7 postirradiation after 5 Gy and day 14 postirradiation after both 5 and 12.5 Gy, and IL-1 alpha mRNA levels were increased in C57BL/6 mice at days 56, 112 and 182 postirradiation after both 5 and 12.5 Gy, and IL-1 beta mRNA levels in the C3H/HeJ mice were increased at days 56 and 182 postirradiation after 12.5 Gy. In summary, these studies demonstrated early and persistent alterations in TNF-alpha, IL-1 alpha and IL-1 beta mRNA levels even at the lower dose (5 Gy). The temporal relationship between the elevation of these cytokines and the strain-dependent variation in fibrosis response suggests that IL-1 alpha and TNF-alpha contribute to the radiation-induced component of pulmonary fibrosis, whereas IL-1 beta may have a protective function.

Hypothermic response of mice to ornithine-containing lipids and to endotoxin

The hypothermic response of mice to ornithine-containing lipids (Orn-Ls) of the form alpha-N-(3-acyloxyacyl)-ornithine and to endotoxin (Escherichia coli 0111:B4 lipopolysaccharide [LPS]) was studied. After the administration of Orn-L or LPS to C3H/HeSlc mice, body temperature decreases were determined at 30-min intervals by inserting a thermistor into the rectum of each mouse. When Orn-L (750 microg) or LPS (70 microg) was injected into the mice, body temperature decreases of 0.8 and 2.0 degrees C, respectively, occurred 1.8 to 2.0 h later. These body temperature decreases were completely suppressed by the preadministration of indomethacin. When anti-tumor necrosis factor alpha (TNF-alpha) antibody was administered before the administration of Orn-L or LPS, only the body temperature decrease by LPS was suppressed. The body temperature decrease by Orn-L was suppressed by anti-interleukin-1beta (IL-1beta) antibody preadministration. Next, in order to study IL-1beta and TNF-alpha mRNA expression in macrophages, peritoneal macrophages were collected 40 min after the administration of Orn-L or LPS to mice. The expression of IL-1beta mRNA by stimulation with Orn-L was as strong as that by stimulation with LPS, but the expression of TNF-alpha mRNA by stimulation with Orn-L was very weak. Our previous studies of in vitro macrophage activation by Orn-L proved that strong induction of IL-1 and prostaglandin E2 generation by Orn-L occurred (Y. Kawai and K. Akagawa, Infect. Immun. 57:2086-2091, 1989). From these experiments, the weak body temperature decrease in mice caused by Orn-L was found to be mediated by cytokines different from those which mediate the strong body temperature decrease caused by LPS. Namely, it was caused by prostaglandin E2 being mediated by IL-1 but not by TNF-alpha.

In situ hybridization analysis of synovial and systemic cytokine messenger RNA expression in superantigen-mediated Staphylococcus aureus arthritis.

To investigate patterns of synovial and systemic cytokine messenger RNA (mRNA) expression in mice with superantigen-mediated Staphylococcus aureus arthritis. Mice were inoculated intravenously with 1 x 10(7) colony-forming units of toxic shock syndrome toxin-1-producing S aureus LS-1. Synovial tissues and spleens were obtained at varying time intervals after bacterial inoculation, and examined for mRNA expression of interleukin-1beta (IL-1beta), IL-4, IL-10, IL-12, tumor necrosis factor alpha (TNFalpha), TNFbeta, interferon-gamma (IFN-gamma), transforming growth factor beta, and perforin, by an in situ hybridization technique. In situ hybridization revealed early synovial up-regulation of TNFalpha and IL-1 beta mRNA expression. Peak frequencies of these proinflammatory cytokines were observed at the second and third week of the infection. Expression of T cell-derived cytokine mRNAs was detected later, and in a relatively low frequency. Notably, induction and peak numbers of Th2 cytokine (IL-4 and IL-10) mRNA expression preceded Th1 cytokine (IFNgamma and TNFbeta) mRNAs. In comparison with synovial tissues, peak spleen cytokine mRNA expression of IL-1beta, TNFalpha, TNFbeta, IL-12, and IFNgamma occurred earlier, but displayed a clearly lower magnitude of expression. These findings demonstrate synovial and systemic up-regulation of cytokine mRNA expression during S aureus arthritis, indicating that both monocyte/macrophage and T cell-derived products are involved in the pathogenesis of this disease.

Prostaglandin E1 inhibits collagen expression in anti-thymocyte antibody-induced glomerulonephritis: Possible role of TGFβ

To test whether or not prostaglandins mediate extracellular matrix formation in immune-mediated glomerular disease, rats with anti-thymocyte antibody-induced glomerulonephritis were treated with prostaglandin E1 (PGE1) (250 micrograms/twice daily/s.c.). Glomerular expression of collagen types III and IV was assessed by Northern blotting, immunohistology and Western blotting. Proliferation of glomerular cells was evaluated by staining for the proliferating cell nuclear antigen (PCNA) and consecutive cell counting. At day five after induction of the disease, glomerular mRNA levels of collagen types III and IV were three- to fivefold higher compared with non-nephritic controls. Similarly glomerular deposition of these collagens was markedly increased when assessed by immunohistology. The treatment of nephritic rats with PGE1 reduced the increased glomerular mRNA levels as well as the protein concentration and the deposition of extracellular collagens. The number of PCNA positive cells which was significantly higher in nephritic rats when compared with control animals (24 hr, nephritis 2.53 +/- 0.33 and Control 0.26 +/- 0.06, P = 0.011; 5 days, nephritis 5.10 +/- 1.13 and Control 0.75 +/- 0.08, cells per glomerular cross section, P = 0.03) was reduced by PGE1 (24 hr, nephritis+PGE1 0.44 +/- 0.30, P = 0.0001; 5 days, nephritis +/- PGE1 1.91 +/- 1.84 cells per glomerular cross section, P = 0.001). Prostaglandin E1 also ameliorated the glomerular infiltration of monocytes at 24 hours (nephritis 4.36 +/- 2.82, nephritis + PGE1 2.20 +/- 1.82, cells per glomerular cross section) and five days (nephritis 1.51 +/- 0.58, nephritis+PGE1 1.12 +/- 0.61, cells per glomerular cross section). To further characterize possible mechanisms by which PGE1 reduces extracellular matrix deposition, the glomerular expression of transforming growth factor (TGF-beta), and interleukin 1 beta (IL-1 beta) was assessed by Northern blotting. Nephritic glomeruli showed increased mRNA levels of TGF-beta at day 5 and IL-1 beta at 24 hours when compared with control kidneys. Treatment of the animals with PGE1 inhibited the mRNA expression of TGF-beta and IL-1 beta. These data demonstrate that PGE1 reduces the glomerular expression of extracellular matrix proteins in anti-thymocyte antibody-induced glomerulonephritis, suggesting a beneficial role of prostaglandins in this proliferative glomerular immune injury. The effects of PGE1 might be mediated by inhibition of TGF-beta and IL-1 beta production.

Human immunodeficiency virus envelope glycoprotein 120 alters sleep and induces cytokine mRNA expression in rats [published errata appear in Am J Physiol 1996 Aug;271(2 Pt 2):section R following table of contents and 1996 Dec;271(6 Pt 3):section R following table of contents

Sleep is altered during the course of viral infection, including that in which the human immunodeficiency virus (HIV) is the etiologic agent. Alterations in the sleep of HIV-infected individuals occur early in the course of infection, prior to the onset of AIDS. The mechanisms for such alterations in sleep are not known. The HIV envelope glycoprotein 120 (gp120) induces the synthesis and secretion of cytokines that enhance [e.g., interleukin (IL)-1 and tumor necrosis factor] and suppress (e.g., IL-10 and IL-1 receptor antagonist) sleep. We used a well-defined rat model to test the hypothesis that the HIV gp120 alters sleep. Recombinant HIV-1IIIB gp120 was injected intracerebroventricularly (20- 500 ng) into rats prior to dark onset. Sleep-wake behavior was not altered after the 20-ng dose, whereas both non-rapid eye movement sleep (NREMS) and rapid eye movement sleep (REMS) were initially enhanced and subsequently suppressed after the 100-ng dose. NREMS was enhanced for 8 h after the 500-ng dose; REMS was not affected by this dose. Brain temperature was not altered by any of the gp120 doses used in this study. In addition, mRNA expression for IL-1 beta and IL-10 was induced in the hypothalamus by gp120; this brain region is crucial for the regulation of sleep. These new data support the hypothesis that altered cytokine concentrations within the central nervous system play a pivotal role in the complex alterations in sleep observed during HIV infection.

Transcriptional and post-transcriptional regulation of GM-CSF-induced IL-1 beta gene expression in PMN.

Polymorphonuclear leukocytes (PMN) play an important role in inflammation, immune responses, and tissue repair by secreting interleukin- 1 beta (IL-1 beta). We investigated the regulation of IL-1 beta gene expression in human PMN treated with granulocyte-macrophage colony-stimulating factor (GM-CSF). GM-CSF induced IL-1 beta mRNA accumulation at 0.1 ng/ml and maximal induction was observed at 1 ng/ml. IL- 1 beta mRNA levels reached a maximum with 1-2 h after stimulation with GM-CSF and returned to baseline levels by 4-6 h. The time course of IL-1 beta mRNA induction by GM-CSF was more protracted than previously reported for PMN stimulated with tumor necrosis factor-alpha (TNF-alpha, 10 ng/ml). Nuclear run-on analysis indicated that GM-CSF, like TNF, increases IL-1 beta transcription. Kinetic studies with the RNA synthesis inhibitor, actinomycin D, showed that GM-CSF induces stable IL-1B mRNA. Cycloheximide enhanced the IL-1 beta mRNA accumulation by GM-CSF at the level of mRNA stabilization, but blocked IL-1 beta mRNA expression by TNF. Thus, GM-CSF increases IL-1 beta message accumulation in PMN at both the transcriptional and post-transcriptional levels by mechanisms that are different from TNF induction of IL-1 beta gene expression.

Effects of immunoglobulin and gamma-interferon on the production of tumour necrosis factor-alpha and interleukin-1 beta by peripheral blood monocytes in the acute phase of Kawasaki disease.

In order to study the in vitro effects of intact immunoglobulin (Ig) and gamma-interferon (INF-gamma) in patients with Kawasaki disease, the production of tumour necrosis factor-alpha (TNF-alpha) and interleukin-1 beta (IL-1 beta) was measured in peripheral blood monocytes (PBM) both before and after intravenous immunoglobulin (IVIG) therapy. Spontaneous production of TNF-alpha and IL-1 beta both before and after IVIG therapy was significantly higher than in healthy controls. Intact Ig enhanced in vitro the production of TNF-alpha and IL-1 beta both before and after IVIG therapy approximately 3-4 times as compared to the spontaneous production. INF-gamma did not affect the production of the two cytokines. Ig enhanced IL-1 beta mRNA expression in PBM of KD by 3-8 times more than that of spontaneous production.

Interleukin 1 beta, interleukin 6, tumor necrosis factor alpha, and interleukin 10 responses in peripheral blood mononuclear cells of cynomolgus macaques during acute infection with SIVmac251.

HIV infection ultimately leads to AIDS, despite the immune responses elicited soon after infection. In addition to the observed changes in lymphoid cell subsets, alteration of the cytokine network most likely accompanies and/or contributes to the lack of protective immune responses. In an attempt to delineate the early events in the immune response to lentivirus infection, we have sequentially monitored levels of proinflammatory (IL-1 beta, IL-6, and TNF-alpha) and antiinflammatory (IL-10) cytokine mRNAs in PBMCs of cynomolgus macaques during primary SIVmac infection. Eight monkeys were infected i.v. with 4 AID50 of cell-free SIVmac251. All monkeys seroconverted between days 16 and 21 postinfection (p.i.), and had detectable peripheral viremia. The viral load peaked between days 12 and 16 p.i., and fell sharply thereafter. A marked increase in the expression of IL-6 mRNA was observed in all macaques during the first weeks following infection. An increase in the levels of expression of IL-1 beta, TNF-alpha, and IL-10 mRNA was also determined in six, six, and five of the eight monkeys, respectively. While IL-6, TNF-alpha, and IL-10 increased transiently, increased levels of IL-1 beta mRNA expression were sustained over 44 days in most monkeys.

Intravitreal transforming growth factor-β2 decreases cellular infiltration in endotoxin-induced ocular inflammation in rabbits

Transforming growth factor-beta (TGF-beta), a multifunctional cytokine which has been identified in normal and inflamed ocular fluids, may play a role in the evolution of inflammatory ocular lesions. In this study we utilized a rabbit model of LPS-induced uveitis to determine if exogenous TGF-beta 2 could alter its course. Recombinant TGF-beta 2 (1-2000 ng), LPS (10 or 20 ng), or TGF-beta 2 (100 ng) plus LPS (10 ng) were injected intravitreally in one eye of a New Zealand white rabbit and the contralateral eye served as a paired control which received an equal volume of vehicle. The uveitic response was assessed by biomicroscopic examination of the anterior uvea and analysis of protein and cells in the aqueous humor. Ocular tissues were processed for histologic, immunohistochemical and in situ hybridization analyses. Rabbits injected with doses of TGF-beta 2 > or = 500 ng developed a mild uveitic response, compared to LPS alone, accompanied by expression of IL-1 beta mRNA and protein in the anterior uvea. Interestingly, rabbits coinjected with LPS (10 ng) and a nonuveitic dose (100 ng) of TGF-beta 2 exhibited a similar increase in ocular vascular permeability, but a decrease in inflammatory cell infiltration into the anterior uvea and aqueous humor (1185 +/- 117 versus 2465 +/- 176; p < 0.05). No evidence of inflammation was observed in eyes injected with 100 ng TGF-beta 2 alone. Similar to other models of inflammation, TGF-beta may interrupt the cascade of events leading to ocular inflammation, thereby suggesting therapeutic potential.

Impact of interferon-gamma receptor deficiency on experimental Staphylococcus aureus septicemia and arthritis.

The role of IFN-gamma in the regulation of host resistance of Staphylococcus aureus was studied using IFN-gamma receptor-deficient (IFN-gamma R-/-) mice in a model of S. aureus-induced septicemia and arthritis. IFN-gamma R-/- mice and wild-type controls were inoculated intravenously with a toxic shock syndrome toxin-1-producing S. aureus LS-1 strain. IFN-gamma R-/- mice displayed significantly more frequent and more severe arthritis compared with wild-type littermates (p < 0.01) throughout the course of infection. Notably, IFN-gamma R-/- mice developed severe sepsis with high mortality early after the inoculation with staphylococci. However, the mortality of wild-type mice became significantly higher at later stages of the disease compared with IFN-gamma R-/- mice (p < 0.05). This differential outcome of sepsis-related mortality was associated with deficiencies of bacterial elimination from blood and parenchymatous organs and correlated well to serum levels of IL-6 and spleen IL-1 beta and TNF-beta mRNA expression. Thus, bacterial growth and proinflammatory cytokines IL-1 beta, TNF-beta, and IL-6 were higher at the early stage of infection in IFN-gamma-/- mice but increased at the later stage in wild-type littermates. Our data indicate that the absence of IFN-gamma R leads to harmful as well as beneficial effects in S. aureus infection, depending on the stage of the disease and the localization of the infection.

T cell-dependent secretion of IL-1 beta by a dendritic cell line (XS52) derived from murine epidermis.

IL-1 beta has been reported to play an essential role in the induction of T cell-mediated immune responses in skin, and Langerhans cells are considered to be the primary source of IL-1 beta in epidermis. We have established recently a long-term dendritic cell line (XS52) from mouse epidermis. This line resembles resident epidermal Langerhans cells in many respects, including the potent capacity to present a protein Ag (KLH) to a CD4+ Th1 clone (HDK-1) and the expression of IL-1 beta mRNA. We sought to determine whether XS52 cells secrete IL-1 beta upon Ag-dependent interaction with T cells and, if so, to elucidate the mechanism. Despite constitutive expression of mRNA for both IL-1 beta and the IL-1 beta-converting enzyme (ICE), XS52 cells secreted no detectable IL-1 beta spontaneously. When they were cultured with HDK-1 T cells and KLH, relatively large amounts of IL-1 beta (17.5-kDa form) were detected in the culture supernatant. IL-1 beta was secreted by LPS-stimulated XS52 cells, but not by LPS- or Con A-stimulated HDK-1 cells, suggesting that IL-1 beta is secreted primarily by XS52 cells in the coculture system. Incubation with HDK-1 cells alone or with KLH alone caused no IL-1 beta secretion, indicating the requirement for both T cells and Ag. IL-1 beta secretion was associated with a striking up-regulation of IL-1 beta mRNA and a modest up-regulation of ICE mRNA and enzymatic activity. IL-1 beta secretion was blocked by Ac-YVAD-CHO (a peptide inhibitor of ICE), CTLA4-Ig fusion protein, or anti-Ia mAb. IL-1 beta secretion was triggered in a T cell-independent manner by either CTLA4-Ig or anti-Ia mAb in immobilized forms. Thus, the XS52 dendritic cell line secretes, by an ICE-dependent mechanism, biologically relevant amounts of IL-1 beta upon Ag-dependent interaction with T cells, with both Ia molecules and B7-related molecules playing essential roles.

Mice with the xid B cell defect are less susceptible to developing Staphylococcus aureus-induced arthritis.

To investigate the role of B cells in the development of experimental Staphylococcus aureus-induced arthritis, we used X-linked immunodeficiency (xid) mice that carry a Bruton's tyrosine kinase mutation affecting the function of B cells. NFR/N.xid and congenic NFR/N mice were inoculated i.v. with a toxic syndrome toxin-1 producing S. aureus LS-1 strain. B cell-deficient NFR/N.xid mice developed less frequent (p < 0.01) and less severe (p < 0.01) arthritis than NFR/N mice did. These clinical findings were corroborated by histopathologic evaluation, indicating that NFR/N.xid mice had significantly lower (p < 0.01) erosivity of the disease. Interestingly, infected NFR/N.xid mice showed decreased bacterial burden in blood, joints, and other organs compared with the control mice. Serologic studies displayed poor B cell responses to staphylococcal cell walls, toxic shock syndrome toxin-1, and ssDNA, accompanied by a low level of Igs in infected NFR/N.xid mice. More importantly, xid defect affected cytokine profile. The in vitro experiments showed that the lymphocytes from NFR/N.xid mice had low IL-6, but high IFN-gamma production upon stimulation with staphylococcal cell walls compared with NFR/N mice. Furthermore, the in situ hybridization technique revealed the relative increase of IFN-gamma, but marked decrease of IL-1 beta mRNA expression in spleens of infected NFR/N.xid mice. No significant difference in IL-4, IL-10, and TNF-alpha mRNA expression was found between both strains. Our findings demonstrate that B cells may, directly or indirectly, contribute to the pathogenesis of septic arthritis. The results indicate that increased IFN-gamma production along with low IL-6 and IL-1 beta synthesis found in xid mice may provide a more favorable outcome of S. aureus arthritis.

Long-term intracerebroventricular corticotropin-releasing hormone administration induces distinct changes in rat splenocyte activation and cytokine expression.

The effects of long-term corticotropin-releasing hormone (CRH) infusion in the lateral ventricle of the rat on hypothalamic-pituitary-adrenocortical (HPA) axis parameters and on the immune system function were studied. Compared with infusion of vehicle, the CRH treatment produced a sustained overactivity of the HPA axis, as evidenced by elevated plasma ACTH and corticosterone levels, increased anterior pituitary POMC messenger RNA (mRNA) expression, and adrenal enlargement. Long-term CRH treatment also inhibited body weight gain and reduced thymus and spleen weight. In the CRH-treated animals, both Concanavalin A (Con A)-induced T lymphocyte proliferation and lipopolysaccharide (LPS)-induced B lymphocyte mitogenesis was largely suppressed. Surprisingly, interleukin-2 (IL-2) levels were higher in supernatants of splenocyte cultures from CRH-treated rats than in those of control animals. However, IL-2 receptor alpha chain (IL-2R alpha) mRNA expression after Con A stimulation was highly suppressed in the CRH-treated animals. In addition, Northern blot analysis of RNA from splenocytes isolated from spleens of CRH-treated rats revealed a marked expression of IL-1 beta mRNA, in contrast to the barely detectable levels of this cytokine in control animals. Moreover, incubation of total splenocytes and spleen macrophages with LPS resulted in an enhanced induction of IL-1 beta mRNA in cells of CRH-treated rats compared with that of control animals. When adrenalectomized rats were treated with CRH or vehicle, the effects of the CRH treatment on T and B cell proliferation, IL-2 production, and IL-1 beta mRNA expression were abolished. Thus, a continuously increased HPA axis drive results in disparate changes in immune system function. Whether the observed changes in cytokine expression should be regarded as physiologically adaptive adjustments in support of immune function or as potentially pathological anomalies remains to be elucidated.

Long-term intracerebroventricular corticotropin-releasing hormone administration induces distinct changes in rat splenocyte activation and cytokine expression

The effects of long-term corticotropin-releasing hormone (CRH) infusion in the lateral ventricle of the rat on hypothalamic-pituitary-adrenocortical (HPA) axis parameters and on the immune system function were studied. Compared with infusion of vehicle, the CRH treatment produced a sustained overactivity of the HPA axis, as evidenced by elevated plasma ACTH and corticosterone levels, increased anterior pituitary POMC messenger RNA (mRNA) expression, and adrenal enlargement. Long-term CRH treatment also inhibited body weight gain and reduced thymus and spleen weight. In the CRH-treated animals, both Concanavalin A (Con A)-induced T lymphocyte proliferation and lipopolysaccharide (LPS)-induced B lymphocyte mitogenesis was largely suppressed. Surprisingly, interleukin-2 (IL-2) levels were higher in supernatants of splenocyte cultures from CRH-treated rats than in those of control animals. However, IL-2 receptor alpha chain (IL-2R alpha) mRNA expression after Con A stimulation was highly suppressed in the CRH-treated animals. In addition, Northern blot analysis of RNA from splenocytes isolated from spleens of CRH-treated rats revealed a marked expression of IL-1 beta mRNA, in contrast to the barely detectable levels of this cytokine in control animals. Moreover, incubation of total splenocytes and spleen macrophages with LPS resulted in an enhanced induction of IL-1 beta mRNA in cells of CRH-treated rats compared with that of control animals. When adrenalectomized rats were treated with CRH or vehicle, the effects of the CRH treatment on T and B cell proliferation, IL-2 production, and IL-1 beta mRNA expression were abolished. Thus, a continuously increased HPA axis drive results in disparate changes in immune system function. Whether the observed changes in cytokine expression should be regarded as physiologically adaptive adjustments in support of immune function or as potentially pathological anomalies remains to be elucidated.

Products of enteropathogenic Escherichia coli inhibit lymphocyte activation and lymphokine production.

The aim of this study was to determine whether products of enteric bacteria are able to regulate lymphocyte activation and cytokine production. Whole bacteria and bacterial lysates from different strains of Escherichia coli were tested for their ability to inhibit cytokine production by peripheral blood mononuclear cells as determined by reverse transcription-PCR, Northern (RNA) blotting of cellular RNA, or enzyme-linked immunosorbent assay for cytokine protein. Lysates from two pathogenic strains of E. coli, enteropathogenic E. coli (EPEC) and enterohemorrhagic E. coli, inhibited mitogen-stimulated expression of interleukin-2 (IL-2), IL-4, IL-5, and gamma interferon. IL-1 beta, IL-6, IL-8, IL-10, IL-12, and Rantes mRNA expression was not affected. The inhibitory activity was dose dependent, protease and heat sensitive, nondialyzable, and not due to cellular toxicity. The inhibitory activity remained in EPEC strains having mutations in known virulence factors. Nonpathogenic E. coli HB101 transformed with a 22-kb cosmid clone derived from EPEC chromosomal DNA expressed the inhibitory activity. Thus, certain strains of pathogenic E. coli express a protein or proteins encoded by chromosomal genes that selectively inhibit lymphocyte activation and lymphokine production. Therefore, immunosuppressive factors produced by pathogenic bacteria could be important in modifying gastrointestinal immune responses in enteric bacterial infections or gastrointestinal autoimmune diseases.

Subdiaphragmatic vagotomy blocks induction of IL-1 beta mRNA in mice brain in response to peripheral LPS

To test the possibility that the vagus nerve is involved in the communication between the immune system and the brain, we injected sham-operated and vagotomized mice with physiological saline or lipopolysaccharide (LPS; 400 micrograms/kg ip). Vagotomy attenuated LPS-induced depression of general activity measured 2 h after treatment but did not alter the increase in plasma levels of IL-1 beta in response to LPS. In addition, vagotomy abrogated the LPS-induced increase in the levels of transcripts for IL-1 beta, as determined by semiquantitative polymerase chain reaction after reverse transcription, in the hypothalamus and hippocampus, but not in the pituitary of vagotomized mice. This relationship between the effects of vagotomy on the behavioral effects of LPS and the LPS-induced brain expression of IL-1 beta mRNA indicates that vagal afferent fibers play a prominent role in the pathways of communication between the immune system and the brain.

Complement and cytokine gene expression in cultured microglia derived from postmortem human brains

Microglia were successfully cultured from human brain tissue from normal and neurologically diseased cases obtained 3.5-10 hours postmortem. Final cell preparations were more than 99% pure as judged by latex bead phagocytosis, expression of microglial phenotypic markers, and absence of astrocytic markers. The expression of complement genes C1qB, C3, and C4 as well as genes for interleukin-(IL-)1 alpha, IL-1 beta, IL-6, tumor necrosis factor (TNF)alpha, IL-1 receptor antagonist, and transforming growth factor beta, but not inducible nitric oxide synthase, by these cells was detected by polymerase chain reaction (PCR) analysis. The pattern of gene expression was evaluated following stimulation of the cells with lipopolysaccharide, phorbol myristate acetate, gamma interferon, and beta amyloid peptide. There was considerable variation in gene response to these activating agents. However, it was of interest that beta-amyloid peptide (1-40) increased the expression of IL-1 beta mRNA in these cells. The number of cases in this study was too small to permit evaluation of microglial response according to the disease state, but the results demonstrate the potential for such studies in the future.

Downregulation of Proinflammatory Cytokine Release in Whole Blood From Septic Patients

Using animal models or healthy volunteers, injection of lipopolysaccharide (LPS) or bacteria causes activation of macrophages with excessive synthesis and secretion of proinflammatory cytokines. Although these models mimic the effects of LPS in the host, they may represent more of an experimental expression of endotoxemia than natural infection itself. Therefore, as an ex vivo model of sepsis, whole blood from 15 patients with severe sepsis and 20 control patients without infection was stimulated with LPS to study the kinetics of mRNA expression and release of proinflammatory cytokines, tumor necrosis factor (TNF)-alpha, interleukin (IL)-1 beta, and IL-6. Stimulation of whole blood with 1 microgram/mL LPS resulted in a maximum increase of cytokine secretion in the control group, while a marked (P < .01) depression of TNF-alpha, IL-1 beta, and IL-6 release was observed in the septic group, which persisted up to 10 days after study enrollment. While IL-1 beta mRNA expression was similar in peripheral blood mononuclear cells (PBMCs) harvested from LPS-stimulated whole blood in septic and control patients, the half-life and consequently the expression of TNF-alpha and IL-6 mRNA were strongly reduced in the septic group. These data indicate a downregulatory mechanism of cytokine release in whole blood from patients with severe sepsis that occurs on different levels. Although excessive secretion of proinflammatory cytokines has been considered deleterious for the host, the reduced capacity of PBMCs in whole blood from septic patients to synthesize and secrete proinflammatory cytokines to an inflammatory stimulus may result in immunodeficiency, because these cytokines in low concentrations are involved in the upregulation of essential cellular and humoral immune functions.

Exaggerated messenger RNA expression of inflammatory cytokines in human T-cell lymphotropic virus type I-associated myelopathy.

To investigate the expression of inflammatory cytokine messenger RNA (mRNA) in peripheral blood mononuclear cells of patients with human T-cell lymphotropic virus type I (HTLV-I)-associated myelopathy (HAM). Seventeen patients with HAM, 18 HTLV-I-seropositive carriers, and 10 seronegative individuals were studied. We compared the expression of tumor necrosis factor alpha (TNF-alpha), granulocyte-macrophage colony-stimulating factor (GM-CSF), interferon alpha (IFN-alpha), IFN-beta, and IFN-gamma, and interleukin 1 alpha (IL-1 alpha) and IL-1 beta by reverse transcriptase-polymerase chain reaction. In patients with HAM, the reverse transcriptase-polymerase chain reaction products of TNF-alpha, GM-CSF, IFN-gamma, and IL-1 alpha were detected in significantly higher incidences than in HTLV-I-seropositive carriers and seronegative controls. Furthermore, simultaneous mRNA expression of three or more of these four cytokines was detected in all patients with HAM compared with only 21.4% of HTLV-I-seropositive carriers. By contrast, there was no significant difference in mRNA expression of IFN-alpha, IFN-beta, and IL-1 beta among patients with HAM, HTLV-I-seropositive carriers, and HTLV-I-seronegative controls. An exaggerated mRNA expression of several inflammatory cytokines, including TNF-alpha, GM-CSF, IFN-gamma, and IL-1 alpha, was demonstrated in peripheral blood mononuclear cells of patients with HAM. Moreover, transcripts of these cytokines were simultaneously up-regulated in patients with HAM, suggesting that an inflammatory state in the central nervous system may be related to the pathogenesis of HAM.