Regulation Of IL-1β Research Articles

Paradoxically high resistance of natural killer T (NKT) cell-deficient mice to Legionella pneumophila: another aspect of NKT cells for modulation of host responses

In the present study, we examined the roles of natural killer T (NKT) cells in host defence against Legionella pneumophila in a mouse model. The survival rate of NKT cell-deficient Jalpha281 knock-out (KO) mice was significantly higher than that of wild-type mice. There was no bacterial overgrowth in the lungs, but Jalpha281 KO mice showed enhanced pulmonary clearance at a later stage of infection, compared with their wild-type counterparts. The severity of lung injury in L. pneumophila-infected Jalpha281 KO mice was less, as indicated by lung permeability measurements, such as lung weight and bronchoalveolar lavage fluid albumin concentration. Recruitment of inflammatory cells in the lungs was approximately twofold greater in Jalpha281 KO mice on day 3. Interestingly, higher values of interleukin (IL)-1beta and IL-18, and increased caspase-1 activity were noted in the lungs of Jalpha281 KO mice from an early time point (6 h). Exogenous alpha-galactosylceramide, a ligand of NKT cells, induced IL-12 and gamma interferon at 6 h, but suppressed IL-1beta at later time points in wild-type, whereas no effects were evident in Jalpha281 KO mice, as expected. Systemic administration of heat-killed L. pneumophila, but not Escherichia coli LPS, reproduced exaggerated production of IL-1beta in the lungs of Jalpha281 KO mice. These results demonstrate that NKT cells play a role in host defence against L. pneumophila, which is characterized by enhanced lung injury and decreased accumulation of inflammatory cells in the lungs. The regulation of IL-1beta, IL-18 and caspase-1 may be associated with the modulating effect of host responses by NKT cells.

Increased Interleukin (IL)-1β Messenger Ribonucleic Acid Expression in β-Cells of Individuals with Type 2 Diabetes and Regulation of IL-1β in Human Islets by Glucose and Autostimulation

Elevated glucose levels impair islet function and survival, and it has been proposed that intraislet expression of IL-1beta contributes to glucotoxicity. The objective was to investigate IL-1beta mRNA expression in near-pure beta-cells of patients with type 2 diabetes (T2DM) and study the regulation of IL-1beta by glucose in isolated human islets. Laser capture microdissection was performed to isolate beta-cells from pancreas sections of 10 type 2 diabetic donors and nine controls, and IL-1beta mRNA expression was analyzed using gene arrays and PCR. Cultured human islets and fluorescence-activated cell sorter-purified human beta-cells were used to study the regulation of IL-1beta expression by glucose and IL-1beta. Gene array analysis of RNA from beta-cells of individuals with T2DM revealed increased expression of IL-1beta mRNA. Real-time PCR confirmed increased IL-1beta expression in six of 10 T2DM samples, with minimal or no expression in nine control samples. In cultured human islets, IL-1beta mRNA and protein expression was induced by high glucose and IL-1beta autostimulation and decreased by the IL-1 receptor antagonist IL-1Ra. The glucose response was negatively correlated with basal IL-1beta expression levels. Autostimulation was transient and nuclear factor-kappaB dependent. Glucose-induced IL-1beta was biologically active and stimulated IL-8 release. Low picogram per milliliter concentrations of IL-1beta up-regulated inflammatory factors IL-8 and IL-6. Evidence that IL-1beta mRNA expression is up-regulated in beta-cells of patients with T2DM is presented, and glucose-promoted IL-1beta autostimulation may be a possible contributor.

Coordinate Regulation of IL-1β and MMP-13 in Rat Tendons Following Subrupture Fatigue Damage

Mechanical overloading is a major causative factor of tendinopathy; however, its underlying mechanisms are unclear. We hypothesized mechanical overloading would damage tendons and alter genes associated with tendinopathy in a load-dependent manner. To test this hypothesis, we fatigue loaded rat patellar tendons in vivo and measured expression of the matrix-degrading enzyme MMP-13 and the inflammatory cytokine IL-1beta. We also examined these responses in cultured tenocytes exposed to intermittent hydrostatic pressure in vitro. Additionally, we hypothesized load-induced changes in tenocyte MMP-13 expression would be dependent on expression of IL-1beta. In vivo fatigue loading at 1.7% strain caused overt microstructural damage and upregulated expression of MMP-13 and IL-1beta, while 0.6% strain produced only minor changes in matrix microstructure and downregulated expression of both MMP-13 and IL-1beta. Loading of cultured tenocytes at 2.5 and 7.5 MPa produced comparable changes in expression to those of in vivo tendon loading. Blocking IL-1beta expression with siRNA suppressed load-induced both MMP-13 mRNA expression and activity. The data suggest fatigue loading alters expression of MMP-13 and IL-1beta in tendons in vivo and tenocytes in vitro in a load-dependent manner. The data also suggest MMP-13 is regulated by both IL-1beta-dependent and IL-1beta-independent pathways.

Cytosolic Antiviral RNA Recognition Pathway Activates Caspases 1 and 3

During an innate immune response, macrophages recognize viruses by their pattern recognition receptors. In this study, we have studied the role of membrane-associated TLRs and cytoplasmic retinoic acid inducible gene-I (RIG-I)-like receptors (RLR) in regulation of IFN-beta, IL-29, IL-1beta, and IL-18 production and caspases 1 and 3 activation in human macrophages. We provide evidence that TLRs are mainly involved in transcriptional up-regulation of IL-1beta gene expression, whereas cytosolic dsRNA recognition pathway stimulates powerful IFN-beta and IL-29 gene transcription. However, robust IL-1beta secretion occurred only if two TLRs were triggered simultaneously or if a single TLR was activated in conjunction with the RLR pathway. Markedly, TLR activation did not stimulate IL-18 processing or secretion. In contrast, triggering of cytosolic RNA recognition pathway with poly(I:C) transfection or influenza A virus infection resulted in caspase-1- and -3-mediated proteolytic processing of pro-IL-18 and secretion of biologically active IL-18. Furthermore, caspase 3-dependent processing of pro-IL-18 was also observed in human HaCaT keratinocytes, and forced expression of RIG-I and its downstream effector, mitochondrial antiviral signaling protein, activated proteolytic processing of pro-IL-18, caspase-3, and apoptosis in these cells. The present results indicate that in addition to robust IFN-beta, IL-29, IL-1beta, and IL-18 generation, RIG-I/mitochondrial antiviral signaling protein pathway activates caspase-3, suggesting a role for these RIG-I-like receptors beyond the innate cytokine response, hence, in the induction of apoptosis of the virus-infected cell.

Signaling molecules involved in production and regulation of IL-1β by murine peritoneal macrophages in vitro on treatment with Concanavalin A

In the present study we report the activation of murine peritoneal macrophages in vitro on treatment with Concanavalin A (ConA). ConA (10 μg/ml) treatment of macrophages resulted in the transcription of IL-1β gene at 16 h and maximum production of IL-1β at 24 h. To investigate the signaling molecules involved in the production of IL-1β different pharmacological inhibitors were used. It was observed that genestein, wortmannin, H-7, TMB-8, PD98059, SB202190, and tyrophostin (AG490) down regulated the expression of IL-1β. These observations suggested the involvement of tyrosine kinase, PI3 kinase, protein kinase C, p42/44, p38, Ca ++ and JAK2 signaling molecules in ConA induced production of IL-1β by macrophages. Maximum protein tyrosine kinase activity and expression of PI3K in macrophages was seen at 5 min, PKC activity and Ca ++ release was found at 10 min after ConA treatment. Maximum expression of phospho-JAK2 at 2.5–5 min, phospho-p42/44 at 5–60 min, phospho-p38 at 15–30 min, phospho-IκB and phospho-Stat1 at 30–60 min and phospho-ELK1, c-Fos, phospho-Stat3 at 60 min of ConA treatment was observed. Pharmacological inhibitors were also used to check the cascade of activation of tyrosine kinase, PKC, PI3 kinase, p42/44, p38, JAK kinase and release of Ca ++ from intracellular storage to sort out the signaling pathways involved in the release of IL-1β by macrophages on treatment with ConA in vitro.

RETRACTED: Quantitative role of p42/44 and p38 in the production and regulation of cytokines TNF-α, IL-1β and IL-12 by murine peritoneal macrophages in vitro by Concanavalin A

In the present study the quantitative role of p42/44 and p38 in the production of TNF-alpha, IL-1beta and IL-12 by murine peritoneal macrophages, in vitro, on treatment with Concanavalin A (ConA) has been investigated. Maximum expression/production of cytokines TNF-alpha, IL-1beta and IL-12 was observed after 16 h by RT-PCR and 24 h by ELISA, on in vitro treatment with ConA. To investigate the role of MAP kinases in the production of cytokines, pharmacological inhibitors of MAP kinases--PD98059, SB202190 and SP600125, were used. The expression of TNF-alpha, IL-1beta and IL-12 was down regulated in the presence of PD98059 and SB202190 in a dose dependent manner, suggesting the involvement of p42/44 and p38 in ConA induced production of TNF-alpha, IL-1beta and IL-12 by macrophages. It was observed that SP600125 did not have any effect on the expression of TNF-alpha, IL-1beta and IL-12. Using different combinations of MAPK inhibitors, it was found that 45% signal is conveyed via p42/44 and 25% via p38 in the production of these cytokines, by ConA treated macrophages while 30% signal passes through unidentified pathways.

Opposite Regulation of IL-1β and Secreted IL-1 Receptor Antagonist Production by Phosphatidylinositide-3 Kinases in Human Monocytes Activated by Lipopolysaccharides or Contact with T Cells

The unbalanced production of IL-1beta and its natural, specific inhibitor, the secreted IL-1R antagonist (sIL-1Ra), plays an important role in chronic/sterile inflammation. Relevant to this condition is direct cellular contact with stimulated T cells which is a potent inducer of cytokine production in human monocytes/macrophages. We previously demonstrated that activation of PI3Ks is a prerequisite of the transcription of the sIL-1Ra gene in human monocytes activated by IFN-beta. In this study, we addressed the question of PI3K involvement in the production of IL-1beta and sIL-1Ra in monocytes activated by cellular contact with stimulated T cells (mimicked by CHAPS-solubilized membranes of stimulated T cells (CE(sHUT))), and a crude preparation of LPS, to compare stimuli relevant to chronic/sterile and acute/infectious inflammation, respectively. In monocytes activated by either CE(sHUT) or LPS, the inhibition of PI3Ks abrogated sIL-1Ra transcript expression and sIL-1Ra production, demonstrating that PI3Ks control the induction of sIL-1Ra gene transcription. In contrast, PI3K inhibition increased the production of IL-1beta protein in both CE(sHUT)- and LPS-activated monocytes, the enhancement being drastically higher in the former. This was not due to changes in IL-1beta mRNA steady-state levels or transcript stability, but to the involvement of PI3Ks in the repression of IL-1beta secretion. The downstream PI3K effector, Akt, was implicated in this process. The present results demonstrate that PI3Ks are involved in the inhibition of IL-1beta secretion and in the induction of sIL-1Ra production in human blood monocytes by controlling different mechanisms in conditions mimicking chronic/sterile (CE(sHUT)) and acute/infectious (LPS) inflammation.

Regulation of the Endothelin/Endothelin Receptor System by Interleukin-1β in Human Myometrial Cells

Proinflammatory cytokines produced at the fetomaternal interface, such as IL-1beta, have been implicated in preterm and term labor. The present study was performed to evaluate the influence of IL-1beta on the endothelin (ET)/ET receptor system in human myometrial cells. We report that myometrial cells under basal conditions not only respond to but also secrete ET-1, one of the main regulators of uterine contractions. Prolonged exposure of the cells to IL-1beta led to a decrease in prepro-ET-1 and ET-3 mRNA correlated with a decrease in immunoreactive ET-1 and ET-3 levels in the culture medium. Whereas ETA receptor expression at both protein and mRNA levels was not affected by IL-1beta treatment, we demonstrated an unexpected predominance of the ETB receptor subtype under this inflammatory condition. Whereas the physiological function of ETB remains unclear, we confirmed that only ETA receptors mediate ET-1-induced myometrial cell contractions under basal conditions. By contrast, prolonged exposure of the cells to IL-1beta abolished the contractile effect induced by ET-1. Such a regulation of IL-1beta on the ET release and the balance of ETA to ETB receptors leading to a loss of ET-1-induced myometrial cell contractions suggest that complex regulatory mechanisms take place to constraint the onset of infection-induced premature contractions.

Inhibition of MAP kinases by crude extract and pure compound isolated from Commiphora mukul leads to down regulation of TNF-α, IL-1β and IL-2

The anti-inflammatory effect of the medicinal plant, Commiphora mukul gum was studied in peripheral blood mononuclear cells (PBMC). Bioassay-guided fractionation using conventional solvent extraction procedures, subsequent column fractionation, followed by monitoring specific activity in PBMC led to the isolation of a lead compound. Both crude ethyl acetate extract and the lead compound, thus isolated, showed inhibitory effect on proliferative response of PBMC in mitogenic lymphocyte proliferation and MLR assays. Further studies on inflammatory mediators such as IFN-γ, IL-12, TNF-α, IL-1β and NO showed down regulation, whereas no inhibition was observed in the case of anti-inflammatory cytokine IL-10. Immunoblot analysis revealed the inhibitory effect of crude ethyl acetate extract on phosphorylation of all the three mitogen activated protein kinases (MAPK) such as ERK, JNK and p38 MAPK. In contrast treatment with pure compound showed no inhibitory effect on ERK. c-fos and c-jun mRNA levels were also reduced in PMA stimulated cells on treatment with crude extract and pure compound. This reduction in c-fos and c-jun levels, when taken together with inhibition of MAPK activation, provides a possible mechanism by which both crude ethyl acetate extract and purified compound isolated from C. mukul exert its action.

Opiate regulation of IL-1β and TNF-α in cultured human articular chondrocytes

In order to investigate if β-endorphins anti-inflammatory effect in cartilage-damaging states is mediated via tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β), we examined its influence on these two cytokines in vitro. Human articular chondrocytes were obtained from patients undergoing total knee arthroplasty and stimulated with β-endorphin (60–6000 ng/ml). Protein levels of TNF-α and IL-1β were measured by ELISA in supernatants from articular chondrocyte cultures. β-Endorphin significantly increased the levels of IL-1β for all concentrations used after 15 min incubation, and when stimulated with 600 and 6000 ng/ml after 24 h incubation. The opioid-induced increase in IL-1β was blocked by naltrexone in the group tested. TNF-α expression was also significantly stimulated by 60 and 600 ng/ml β-endorphin after 15 min, an effect blocked by naltrexone in the group tested. These findings indicate that the mechanism of β-endorphins anti-inflammatory influence in cartilage-damaging states is not apparently mediated via these two cytokines modulation.

A human oral keratinocyte cell line responds to human heat shock protein 60 through activation of ERK1/2 MAP kinases and up- regulation of IL-1β

Heat shock proteins (HSP) are released by cells in response to stress signals. It is hypothesized that pathogenic bacteria stimulate the cells in the periodontium to up-regulate the expression of HSP60, which would stimulate macrophages, and possibly other cells, to produce proinflammatory cytokines. We sought to determine whether oral keratinocytes responded to recombinant human HSP60 and to identify the signalling pathways involved. In addition, whether oral keratinocytes are a source of endogenous HSP60 was also investigated. RT-PCR revealed that rhHSP60 induced expression of the IL-1beta gene in the Human Oral Keratinocyte (HOK-16B) cell line and it was highest at the lowest concentration used (0.1 microg/ml). These responses were mediated via activation of p44/42 MAP-kinases and to a lesser extend the MAP-kinase SAP/JNK. Similar data was obtained from analysis of intracellular signalling pathways in HOK-16B cells by rhHSP70 and LPS (from both E. coli and the oral pathogen Porphyromonas gingivalis). However, there was little activation of p38 by rhHSP60. Blocking of the p44/42 pathway decreased HSP60-induced IL-1beta gene expression and protein secretion. In addition, we discovered that self-HSP60 proteins were constitutively secreted by HOK-16B cells. Secretion of self-HSP60 was up-regulated in cells treated with LPS from P. gingivalis, but down-regulated with LPS from E. coli. To summarize, oral keratinocytes respond to exogenous HSP60 by triggering expression of the inflammatory cytokine IL-1beta through activation of p44/42 MAP kinase. Oral keratinocytes are also a source for self-HSP60 and the secretion of this protein may be differentially modified by LPS from different bacterial species. These results highlight the importance of oral keratinocytes and HSPs in the development of an immune response against bacterial infection.

Overexpressed nuclear factor κB correlates with enhanced expression of interleukin-1β and inducible nitric oxide synthase in aged murine lungs to endotoxic stress

Background Transcriptional regulation is a major determinant of interleukin-1β (IL-1β) protein synthesis. Nuclear factor κB (NF-κB) plays a central role in the regulation of IL-1β and subsequent IL-1β-dependent inflammatory processes. Previously, we observed in a murine endotoxic stress model a progressive increase with age in the amount of IL-1β mRNA. We test the aging pulmonary response of NF-κB and NF-κB-dependent genes, IL-1β, and inducible nitric oxide synthase (iNOS) in the same model. Methods Young (2-month-old) and senescent (25-month-old) mice were given 0.5 mg/kg lipopolysaccharide (LPS) intraperitoneally. Lung and blood samples were harvested after 4 hours. IL-1β production in blood samples and the expression levels of protein and mRNA of IL-1β and iNOS in lung tissues were measured. NF-κB binding activity in lung tissues was also determined. Results LPS induced higher levels of IL-1β in the sera and lungs of senescent mice over young mice. Northern and Western blot analyses showed that mRNA and protein signals of IL-1β and iNOS were significantly higher in old lungs than in young lungs. Electrophoretic mobility shift assay also showed that NF-κB activation was significantly higher in the older animals. Conclusions Our results suggest that elevated activation of NF-κB, at least in part, contributes to the dysregulated expression of IL-1β and iNOS in the lungs of senescent animals. Thus increased expression of proinflammatory cytokines and inflammatory responsive genes in the lung may play a role in the increased susceptibility in aging animals to endotoxic stress.

NEGATIVE REGULATION OF IL-1β PRODUCTION AT THE LEVEL OF TRANSCRIPTION IN MACROPHAGES STIMULATED WITH LPS

The IL-1β gene is rapidly and transiently expressed in LPS-stimulated macrophages. While several studies have addressed the molecular basis of LPS-induced transcriptional activity, the mechanisms which underlie the subsequent decrease in IL-1β gene expression have not been as extensively examined. In this regard, we found that the characteristic decrease in IL-1β production after LPS stimulation could be abrogated by treatment of macrophages with the protein kinase inhibitor staurosporine. This inhibitor mediated an enhancement of IL-1β production which was first evident 8–12h after LPS stimulation and continued at peak levels for the rest of the incubation period (24h). IL-1β production was correlated with the level of mRNA specific for the cytokine. Staurosporine also mediated an enhancement of LPS-induced IL-1β promoter activity measured in RAW 264.7 cells transiently transfected with an IL-1β reporter plasmid. This increase paralleled the enhancement of IL-1β mRNA by staurosporine both in intensity and time after LPS stimulation, suggesting that the negative regulation of IL-1β is exerted primarily at the level of transcription. This regulation may be at least partially due to an observed inhibition of nitric oxide production by staurosporine in LPS-activated macrophages, which was correlated with enhanced IL-1β production. However, the intensity of the observed effects suggested that additional staurosporine-sensitive regulatory mechanisms are in operation at the level of promoter activity.

Differential Regulation of IL-1β and TNF-α RNA Expression by MEK1 Inhibitor after Focal Cerebral Ischemia in Mice

Activation of the extracellular-signal-responsive kinase (ERK 1/2) by MAP kinase/ERK kinase (MEK1/2) following ischemia/reperfusion in the brain has been associated with cell death since inhibition of MEK1/2 provides neuroprotection in cerebral ischemia injury. Since inflammation has been implicated in ischemic brain injury, the present study investigated whether MEK1/2 modifies expression of two key inflammatory cytokines, IL-1β and TNFα, that have been shown to exacerbate ischemic brain injury. A mouse model of transient cerebral ischemia was deployed to test the effect of selective MEK1/2 inhibitor (SL327) on infarct size and cytokine expression. SL327 (100 mg/kg, i.p.) administered 15 min prior to ischemia resulted in 64% reduction in infarct size over controls (n = 8, P < 0.01). Under the same condition, SL327 significantly reduced peak expression of IL-1β mRNA (59% reduction compared to vehicle, P < 0.01, n = 4) but not TNF-α mRNA. A parallel reduction in IL-1β protein (67%, P < 0.05, n = 6) was also observed using ELISA analysis. These data suggest that the neuroprotective effect of MEK1/2 inhibition may be mediated by suppression of IL-1β. The study also demonstrates for the first time that these two cytokines are differentially regulated by kinase mediated signaling pathways.

Interleukin (IL)-1beta regulation of IL-1beta and IL-1 receptor antagonist expression in cultured human endometrial stromal cells.

The interleukin (IL)-1 system is a major regulator of local cellular interactions during embryonic implantation. Because IL-1beta and IL receptor antagonist (IL-1ra) are both expressed in human endometrium, we hypothesized that an appropriate ratio of IL-1beta to IL-1ra might favor the process of embryo implantation. Therefore, we investigated IL-1 regulation of the quantitative ratio of IL-1beta/IL-1ra messenger RNA (mRNA) expression in human endometrial stromal cells using quantitative competitive PCR, as well as intracellular protein expression after stromal cell solubilization. Confluent stromal cell cultures were stimulated with human IL-1beta (0-1000 IU/mL) for 24 h. After 24 h, total RNA was extracted, reverse transcribed, and coamplified by PCR with a defined amount of internal standard. The quantitative ratio was determined by the density of target to the internal standard. After culture with IL-1beta for 24 and 48 h, stromal cells were solubilized, and the intracellular protein levels of IL-1beta and IL-1ra were measured by enzyme-linked immunosorbent assay. The IL-1beta and IL-1ra mRNA were both up-regulated, and IL-1R tI mRNA was down-regulated, by IL-1beta in a dose-dependent manner. The quantitative ratio of IL-1beta to IL-1ra mRNA was constant with the presence of increasing concentrations of IL-1beta (1-1000 IU/mL). IL-1beta and IL-1ra protein was not detected in conditioned media of cultures before addition of IL-1beta. IL-1beta and IL-1ra protein levels increased with increasing amounts of IL-1beta after solubilization of stromal cells. The IL-1beta was detectable after 12 h of culture, in comparison with IL-1ra, which was detectable after 24 h of IL-1beta stimulation. These results suggest that IL-1 may play a crucial role in embryo-maternal interaction by regulating stromal cell expression of IL-1beta and IL-1ra, resulting in an appropriate ratio during the process of embryonic implantation.

Induction of Interleukin-4 (IL-4) byLegionella pneumophilaInfection in BALB/c Mice and Regulation of Tumor Necrosis Factor Alpha, IL-6, and IL-1β

Infection of BALB/c mice with a sublethal concentration of Legionella pneumophila causes an acute disease that is resolved by innate immune responses. The infection also initiates the development of adaptive Th1 responses that protect the mice from challenge infections. To study the early responses, cytokines induced during the first 24 h after infection were examined. In the serum, interleukin-12 (IL-12) was detectable by 3 h and peaked at 10 h, while gamma interferon was discernible by 5 h and peaked at 8 h. Similar patterns were observed in ex vivo cultures of splenocytes. A transient IL-4 response was also detected by 3 h postinfection in ex vivo cultures. BALB/c IL-4-deficient mice were more susceptible to L. pneumophila infection than were wild-type mice. The infection induced higher serum levels of acute-phase cytokines (tumor necrosis factor alpha [TNF-alpha], IL-1beta, and IL-6), and reducing TNF-alpha levels with antibodies protected the mice from death. Moreover, the addition of IL-4 to L. pneumophila-infected macrophage cultures suppressed the production of these cytokines. Thus, the lack of IL-4 in the deficient mice resulted in unchecked TNF-alpha production, which appeared to cause the mortality. Monocyte chemoattractant protein-1 (MCP-1), a chemokine that is induced by IL-4 during Listeria monocytogenes infection, was detected at between 2 and 30 h after infection. However, MCP-1 did not appear to be induced by IL-4 or to be required for the TNF-alpha regulation by IL-4. The data suggest that the early increase in IL-4 serves to regulate the mobilization of acute phase cytokines and thus controls the potential harmful effects of these cytokines.

Extracellular matrix proteins in organ transplantation.

Extracellular matrix proteins in organ transplantation.

Treatment of rheumatoid arthritis with interleukin 1 receptor antagonist

Rheumatoid arthritis (RA) is characterised by chronic synovial inflammation of multiple large and small joints in the upper and lower limbs.1 The pathogenesis of RA is complex and involves the...

Differential Effects of One and Repeated Endotoxin Treatment on Pituitary- Adrenocortical Hormones in the Mouse: Role of Interleukin-1 and Tumor Necrosis Factor-α

The role of endogenous interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF-α) in modulating the hypothalamic-pituitary-adrenal (HPA) axis response was examined in male C57BL/6 mice injected with endotoxin (lipopolysaccharide, LPS) or saline at 24-hour intervals for 4 or 8 consecutive days. The mice were divided into four groups: (1) LPS injections for 4 or 8 days and LPS injection on day 5 or 9, respectively (LPS-LPS); (2) LPS injections for 4 or 8 days and saline injection on day 5 or 9, respectively (LPS-saline); (3) saline injections for 4 or 8 days and LPS injection on day 5 or 9, respectively (saline-LPS), and (4) saline injections for 4 or 8 days and saline injection on day 5 or 9 (saline-saline). The mice were sacrificed by decapitation 2 h after the last injection and plasma levels of hormones and cytokines and tissue levels of IL-1β were measured. Plasma adrenocorticotropin (ACTH) levels were significantly attenuated in the LPS-LPS group compared with the dramatic increases in the saline-LPS group following 4 or 8 days of endotoxin treatment. Plasma corticosterone concentrations were comparable in the LPS-LPS group after 4 days’ treatment, but significantly lower following 8 days of treatment when compared with saline-LPS group. Repeated endotoxin treatment followed by a single saline injection (LPS-saline) did not alter the levels of IL-1β in plasma or any of the tissues examined. IL-1β levels in the hippocampus, hypothalamus, adrenal gland and plasma were elevated to comparable levels in the saline-LPS and LPS-LPS groups after 4 days of treatment. In contrast to the plasma IL-1β response, TNFα levels were dramatically increased in the saline-LPS group but not in the LPS-LPS group following the 4-day treatment regimen. Increases in IL-1β concentrations were seen in all tissues following one endotoxin challenge in the saline-LPS group following the 8-day treatment regimen, while increases were significantly attenuated in the hypothalamus, adrenal gland and plasma in LPS-LPS for 8 days. The sustained increases in tissue levels of IL-1β following 4-day endotoxin treatment appears to have functional consequences since [¹²⁵I]IL-1α binding was significantly decreased in the LPS-saline group compared with the saline-saline group. Furthermore, [¹²⁵I]IL-1α binding was markedly reduced in the LPS-LPS group compared with the saline-LPS group. There was a significant positive correlation between plasma ACTH and IL-1β after a single and repeated LPS treatment for 4 days, while a significant correlation was seen between plasma ACTH and TNFα following one but not repeated LPS treatment. These data demonstrate a differential regulation of IL-1β and TNFα by repeated endotoxin treatment and suggest that while TNFα may be important modulating the attenuated pituitary adrenocortical response following the 4-day endotoxin treatment, IL-1β appears to be the primary regulator of the response following the 8-day endotoxin treatment in the regulation of the HPA axis.

The rat intraovarian interleukin (IL)-1 system: cellular localization, cyclic variation and hormonal regulation of IL-1β and of the type I and type II IL-1 receptors

An increasing body of evidence supports the possibility that intraovarian interleukin (IL)-1 plays an intermediary role in the periovulatory cascade. To gain further insight into the intraovarian IL-1 hypothesis, we studied the cellular localization cyclic variation and hormonal regulation of IL-1β, as well as of the type I and type II IL-1 receptors (IL-1R) in immature rats. In situ hybridization localized IL-1β and type I IL-1R transcripts to the granulosa cell compartment, the innermost layers of the theca interna and to the oocyte of the untreated immature ovary. Molecular probing of whole ovarian material in the course of a simulated estrous cycle revealed a progressive preovulatory increase in IL-1β and type I IL-1R transcripts to an in vivo peak at the time of ovulation (3.0- and 2.5-fold increases over untreated controls; P<0.05). Comparable efforts to localize and probe for type II IL-1R transcripts failed to elicit a detectable signal. The basal in vitro expression pattern of IL-1β and type II IL-1R transcripts by whole ovarian dispersates revealed an early (4 h) spontaneous increase to a peak (2.1- and 5.8-fold increases over time 0; P<0.05) followed by a gradual decline to a 48 h nadir. Treatment of whole ovarian dispersates with the IL-1 receptor antagonist (IL-1RA) or with IL-1β failed to alter the initial (4 h) burst of IL-1β or of type II IL-1R expression thereby suggesting IL-1-independence. Treatment with hCG proved equally ineffective. However, longer-term treatment of whole ovarian dispersates with IL-1β produced a significant secondary increase (5.9-fold over time 0; P<0.05) in IL-1β (but not type II IL-1R) transcripts by 48 h. This IL-1 effect was completely blocked by co-treatment with IL-1RA thereby suggesting mediation via a specific IL-1 receptor. Qualitatively comparable but quantitatively reduced results obtained for isolated granulosa cells. The basal in vitro expression pattern of type I IL-1R transcripts by whole ovarian dispersates revealed a progressive spontaneous increase (3.1-fold increase overall) over the 48 h culture. Treatment with IL-1β produced a significant ( P<0.05) increase (5-fold) in type I IL-1R transcripts by 48 h, an effect which was completely blocked by co-treatment with IL-1RA. Taken together, these observations: (1) localize IL-1β and its type I receptor to granulosa cells, the innermost layers of the theca interna and to the oocyte; (2) confirm their periovulatory in vivo expression pattern; (3) document their expression by untreated cultured whole ovarian dispersates; and (4) demonstrate their in vitro responsiveness to receptor-mediated/IL-1-driven autocrine amplification. The type II IL-1R was undetectable in vivo, its in vitro expression pattern proving IL-1- and hCG-independent. The periovulatory expression pattern of IL-1β and its receptor (type I) is compatible with the notion that the intraovarian IL-1 system may play an intermediary role in the ovulatory process.