IL-1 RI Research Articles

Interleukin-1 receptor antagonist inhibits metastatic potential by down-regulating CXCL12/CXCR4 signaling axis in colorectal cancer

BackgroundThe aim of this study was to investigate the co-operative role of CXCR4/CXCL12 axis and IL-1Ra in metastatic processes mechanism by interactions between colorectal cancer cells and stromal cells in their microenvironment.MethodsExpression of IL-1α, interleukin-1 receptor type I (IL-1 RI), CXCL12 and CXCR4 mRNA and proteins were determined by RT-PCR and Western blot. The effect of secreted level of CXCL12 by IL-1Ra on fibroblasts was measured by ELISA. CXCL12 regulate metastatic potential of colorectal cancer was evaluated by proliferation, invasion and angiogenesis assays, respectively, in which invasion and angiogenesis assays used an in vitro system consisting of co-cultured colorectal cells and stromal cells.ResultsIL-1α was expressed in high liver metastatic colorectal cancer cell lines (HT-29 and WiDr). The colorectal cancer cell-derived IL-1α and rIL-1α significantly promoted CXCL12 expression by fibroblasts, and this enhancing effect can be significantly inhibited by IL-1Ra (P < 0.01). CXCL12 not only enhanced the migration and proliferation of human umbilical vein endothelial cells, but also significantly enhanced angiogenesis (P < 0.01). Furthermore, the high liver-metastatic colorectal cancer cell line (HT-29), which secretes IL-1α, significantly enhanced angiogenesis compared to the low liver-metastatic cell line (CaCo-2), which does not produce IL-1α (P < 0.01). On the contrary, IL-1Ra can significantly inhibit migration, proliferation and angiogenesis (P < 0.01).ConclusionAutocrine IL-1α and paracrine CXCL12 co-enhances the metastatic potential of colorectal cancer cells; IL-1Ra can inhibit the metastatic potential of colorectal cancer cells via decrease IL-1α/CXCR4/CXCL12 signaling pathways.Graphical 7Qx8An2hePDujdT1wSZV-LVideo

Pro-Inflammatory Cytokine IL-1b on Megakaryopoiesis: A Possible Explanation of Thrombocytosis in Inflammation

Pro-inflammatory cytokine interleukin-1b (IL-1b) is multifunctional and exerts its effects on a wide variety of cell types. Among many functions, IL-1b enhances the hematopoietic system by inducing T and B cell proliferation and works in combination with other cytokines on early stem and progenitor cell expansion and differentiation. Megakaryopoiesis and platelet production is maily controlled by thrombopoietin (TPO, Mpl-ligand), which is a growth factor for the megakaryocyte/ platelet lineage. Other early acting cytokines including IL-1, IL-3, IL-6 and IL-11 are also involved in the regulation of this lineage at different degree. Thrombocytosis is observed in various inflammation diseases. IL-1b, as a highly pro-inflammatory cytokine, induces thrombocytosis in mice. However, its mechanism on megakaryopoiesis is still not well understood. Our hypothesis is that IL-1b possesses a direct stimulatory effect on megakaryocytes via its receptors and the transcription factor NF-E2. The expression of IL-1 type I (IL-1 RI) and type II receptors was studied in megakaryotic cell lines: Meg-01, Dami, CHRF-288-11 and M-07e by RT-PCR, southern blot hybridization, flow cytometry or immuno-fluorescent staining. The effect of IL-1b on the formation of megakaryocytic colonies from human and murine stem and progenitor cells was investigated by the plasma clot culture method. The expression of the transcription factor p45 NF-E2, in response to IL-1b, was analyzed by Western blot. Our results show that IL-1 RI and IL-1 RII were expressed in the four human megakaryocytic cells. IL-1b alone or in combination with TPO induced megakaryocyte colony forming units (CFU-MK) in the culture system. The mitogenic effect of IL-1b on the enhancement of TPO-induced megakaryopoiesis was stronger than that of IL-3, IL-6, Flt-3 ligand and stem cell factor. The expression of NF-E2 was demonstrated in all four cell lines. In Meg-01 cells, the expression of NF-E2 was increased at both transcriptional and translational level after treatment with IL-1b for 4 hours. These observations strengthen the evidence of the direct effect of IL-1b on megakaryopoiesis. Our findings also provide a possible explanation of thrombocytosis during inflammation conditions. DisclosuresNo relevant conflicts of interest to declare.

The Effect of Tanshinone-IIa on IL-1β Induced-Thrombocytosis in an Immune Vasculitis Model

Inflammation cytokines may be involved in the pathogenesis of thrombocytosis with vasculitis. Our previous study showed that inflammation cytokine IL-1β plays an important role on in-vitro megakaryopoiesis (Yang M et al, Br J Haematol 2000). The role of IL-1β and Tanshinone IIA (TIIA) (Isolated from Danshen, Radix Salviae Miltiorrhiza Bge) on platelets and megakaryocytes (MKs) in immune vasculitis model was investigated in this study. Rabbit model with immune vasculitis was established by injection (iv) of BSA. After treatment with BSA for 7 days, the platelet count, platelet aggregation and the expression of AnnexinⅤ were significantly increased in this vasculitis model group compared with normal control group (n=7). IL-1β levels was also significantly higher in vasculitis model. There were positive correlations between platelet count and IL-1β levels (R=0.65), platelet aggregation and IL-1β levels (R=0.60). Treatment with TIIA (5 mg/kg/day, iv) and aspirin significantly decreased all these parameters. MKs and CFU-MK number were also significantly increased in vasculitis group as compared to normal group. Treatment with TIIA and aspirin significantly reduced the number of MKs and CFU-MK in this model. Study further demonstrated that IL-1β alone or in combination with TPO induced in-vitro CFU-MK formation. The mRNA of of IL-1 type I and type II receptors (IL-1 RI and RII) were detected in cultured MK (CD61+ CD41+) cells. The expression of IL-1 RI and RII was also confirmed by immunofluorescence staining in bone marrow MKs. Moreover, the IL-1R bloker can reduced IL-1β induced megakaryopoiesis. TIIA on in-vitro megakaryopoiesis was also investigated. TIIA at 10-30 ug/ml significantly inhibited CFU-MK formation. TIIA also induced the apoptosis of MKs in a dose dependent manner by Anexin V assay. Caspase 3 assay showed the activation of Caspase 3 increased from 5% to 16%. Using JC-1 assay found that the depolarized cells increased from 9% to 17% suggesting the involvement of intrinsic apoptotic pathway. IL-1β may play an important role in the thrombocytosis of immune vasculitis by inducing megakaryopoiesis. TIIA has anti-platelet effect in this model which may be mediated via inhibiting IL-1β induced-megakaryopoiesis. DisclosuresNo relevant conflicts of interest to declare.

IL-1β Plays An Important Role On Thrombocytosis In An Immune Vasculitis Model Via Promoting Megakaryopoiesis

Thrombocytosis and inflammation cytokines may be involved in the pathogenesis of vasculitis. Our previous study have showed that major inflammation cytokine IL-1β play an important role on in-vitro megakaryopoiesis (Yang M et al, Br J Haematol 2000). In this study, we investigated the changes of IL-1β and megakaryopoiesis and the effect of aspirin in an immune vasculitis model. Rabbit immune vasculitis model was established by intravenous injection of bovine serum albumin. In this model, platelet number and function of periphery blood, megakaryocyte number and the CFU-MK formation of the bone marrow, and serum levels of inflammatory cytokines were investigated. After treatment with BSA for 7 days, the platelet count, platelet aggregation and the expression of AnnexinⅤ were significantly increased in this vasculitis model group compared with normal control group (n=6). The serum levels of inflammatory cytokine IL-1β was also significantly higher in vasculitis model. There were positive correlations between platelet count and IL-1β levels (R=0.55), platelet aggregation and IL-1β levels (R=0.603). Treatment with aspirin (100 mg/kg/d) significantly decreased all these parameters, showing aspirin had anti-platelets and anti-inflammation effects. Our results also demonstrated that megakaryocyte number and the formation of CFU-MK were significantly increased in vasculitis group as compared to those in normal group. Treatment with aspirin significantly reduced the number of megakaryocytes and the formations of CFU-MK in bone marrow in this immune vasculitis model. Our study further demonstrated that IL-1β alone or in combination with TPO induced in-vitro CFU-MK formation. Using RT-PCR techniques, the mRNA of of IL-1 type I and type II receptors (IL-1 RI and RII) were detected in cultured CD61+ CD41+ cells and four megakaryocytic cell lines. The expression of IL-1 RI and RII was also confirmed by flow cytometry and immunofluorescence staining in bone marrow megakaryocytes. Moreover, the IL-1R bloker can reduced IL-1β induced megakaryopoiesis. This sudy showed that IL-1β may play an important role in the pathogenesis of immune vasculitis. Aspirin has anti-inflammation effects in this model which may be mediated via inhibiting megakaryopoiesis and platelet formation. Disclosures:No relevant conflicts of interest to declare.

Interleukin-1 β up-regulates tumor necrosis factor receptors in the mouse airways

Cytokines like interleukin-1 β (IL-1 β) and tumor necrosis factor α (TNF α), released during the inflammatory process, play important roles in the development of airway hyperresponsiveness. The effects of these cytokines are mediated by cell surface receptors, specific for each cytokine. The expression of cytokine receptors is a dynamic process, where receptors can be up- or down-regulated in response to changes in the environment. One such environmental factor is the presence of cytokines per se. The present study was designed to evaluate the effects of IL-1 β on the expression of its corresponding receptor IL-1 RI, as well as on the closely related TNF α receptors TNF RI and TNF RII in airways using a mouse organ culture assay and intranasal inoculation model. Immunohistochemical staining was used to quantify expressional differences between fresh and cultured tracheal segments. In the fresh, uncultured, segments, IL-1 RI and TNF RI were seen in the epithelial layer and TNF RI in the smooth muscle layer. After 4 days of culture, the expression of TNF RI decreased in the epithelial layer, whereas the corresponding expression of IL-1 RI and TNF RI in the smooth muscle remained unchanged. When culture was performed in the presence of IL-1 β, the expression of IL-1 RI and TNF RI in the epithelial cells and TNF RI in the smooth muscle cells increased. TNF RII was not detected in either fresh or cultured trachea, but after treatment with IL-1 β an expression was found in both the epithelial layer and in the smooth muscle cells. The IL-1 β-induced increased expression, on TNF RI and TNF RII in the smooth muscle ex vivo and in the lung parenchyma after intranasal challenge in vivo, was verified at the mRNA level using real-time RT PCR. To summarize, presence of IL-1 β increases the expression of IL-1 R1 and TNF RI and induces expression of TNF RII in the airway wall. It is not inconceivable that these alterations of the IL-1 and TNF receptors may have important functional implications for the development of hyperresponsiveness in inflammatory airway diseases like asthma.

Biochemical entities that influence membrane-associated TNF RII (80-kDa) and IL-1 RI (80-kDa) complex expression and receptor fragment production in adherent vascular endothelium

The research aim of the present investigation was to identify leukocyte enzyme-proteases that have the capacity to biochemically recruit the passive participation of vascular endothelium in cytokine receptor ‘shedding’ phenomenon involving membrane-associated TNF RII (80-kDa) and IL-1 RI (80-kDa) complexes. Achieving this research objective involved the design of a laboratory approach that delineated to what extent enzyme-proteases released by activated macrophages directly interact with, and liberate soluble fragments of membrane-associated cytokine receptor complexes. Results from this segment of the investigation revealed that cathepsin-D, a leukocyte carboxyl/aspartate protease, altered the integrity and generated soluble fragments of TNF RII (80-kDa) and IL-1 RI (80-kDa) receptor complexes expressed by vascular endothelium. Furthermore, laboratory findings also suggested that cathepsin-D possessed the ability to variably deplete biologically functional membrane-associated TNF RII (80-kDa) and IL-1 RI (80-kDa) complexes. Complementary investigations isolated a carboxyl/aspartate protease from activated macrophages utilizing pepstatin-A affinity chromatography. Exposure of vascular endothelium to pepstatin-A binding proteins resulted in a detectable depletion of membrane-associated TNF RII (80-kDa) and IL-1 RI (80-kDa) in addition to the generation of soluble receptor fragments. Analysis of macrophage pepstatin-A binding proteins by SDS-PAGE identified a primary fraction with a molecular mass of 47–52-kDa that closely correlated with the known molecular mass of leukocyte cathepsin-D. Evaluation of macrophage pepstatin-A binding-protein fractions by non-denaturing Hb-PAGE detected a lucent proteolytic band at 47–52-kDa compatible with the known molecular mass of leukocyte cathepsin-D. Macrophage pepstatin-A binding proteins also hydrolyzed a synthetic enzyme-specific substrate that selectively recognizes cathepsin-D biochemical activity. In conclusion, the leukocyte carboxyl/aspartate protease, cathepsin-D can biochemically alter the integrity and generate soluble fragments of membrane-associated TNF RII (80-kDa) and IL-1 RI (80-kDa) receptor complexes expressed by vascular endothelium. The relevance of this concept is in part based on investigations that have discovered that genetic ‘knock-out’ mice incapable of expressing IL-1 RI (80-kDa) or TNF RI (55-kDa) receptor complexes are highly resistant to developing the pathophysiological alterations classically associated with conditions of endotoxic-shock.

Expression of interleukin (IL) 1 type I and type II receptors in megakaryocytic cells and enhancing effects of IL-1beta on megakaryocytopoiesis and NF-E2 expression.

Megakaryocytopoiesis is regulated by thrombopoietin (TPO) and cytokines such as interleukin 3 (IL-3), IL-6 and IL-11. This study investigated the in vitro effects of IL-1beta on megakaryocytopoiesis and the expression of IL-1 type I and type II receptors (IL-1 RI and RII) on mega-karyocytic cell lines and primary cells. Our results demonstrated that IL-1beta alone or in combination with TPO induced megakaryocyte colony forming units (CFU-MK) from murine and human haematopoietic cells. Using reverse-transcription polymerase chain reaction (RT-PCR) and Southern hybridization techniques, the mRNA of IL-1beta, IL-1 RI, IL-1 RII and the transcription factor NF-E2 were detected in CD61+ CD41+ cells cultured from cord blood and four megakaryocytic cell lines, Meg-01, DAMI, CHRF-288-11 and M-07e. The expression of IL-1 RI and RII proteins was confirmed by flow cytometry and immunofluorescence staining. In Meg-01 cells, the expression of NF-E2 was increased at both mRNA and protein levels after treatment with IL-1beta for 4 h. This study demonstrated for the first time the presence of IL-1 receptors on megakaryocytic cells and the induction of NF-E2 by IL-1beta. The mitogenic effect of IL-1beta on this lineage could be mediated through IL-1 receptors and the activation of NF-E2.

Induction of an angiogenic phenotype in endometriotic stromal cell cultures by interleukin-1beta.

Activated peritoneal macrophages are associated with endometriosis and may play a central role in its aetiology by releasing interleukin-1beta (IL-1beta) in response to refluxed endometrium. Pari passu with the establishment of endometriotic implants is the development of a vascular supply. In this study we investigated the angiogenic properties of two endometrial proteins, vascular endothelial growth factor (VEGF) and interleukin-6 (IL-6), and assessed their production in response to IL-1beta stimulation in human stromal cells isolated from normal endometrium (NE) and endometriotic lesions (EI). Proliferation of bovine brain capillary endothelial cells (BBCE) with a [(3)H]-thymidine incorporation assay was observed when VEGF (2.1 +/- 0.2-fold; P < 0.05) or VEGF and IL-6 (1.8 +/- 0.1-fold; P < 0.05) were added in vitro, relative to saline-treated control cultures. Northern blot analysis showed induction of VEGF mRNA (2.6-fold; P < 0.05) and IL-6 mRNA (6.3-fold; P < 0.05) transcripts in EI cells, but not NE cells, exposed to IL-1beta. A similar induction was seen with VEGF and IL-6 protein secretion in the responsive EI cells. Reverse transcription-polymerase chain reaction (RT-PCR) for the IL-1 receptor type I (IL-1 RI) indicated that the differential effects of IL-1beta on NE and EI cells was associated with 2.4 +/- 0.1-fold more receptor mRNA in EI versus NE cells. We propose that the ability of IL-1beta to activate an angiogenic phenotype in EI stromal cells but not in NE cells, is mediated by the IL-1 RI.

Mechanisms of behavioral and neuroendocrine effects of interleukin-1 in mice.

Interleukin-1 beta is a key molecule in brain-immune interactions that, apart from its immune effects, stimulates the hypothalamo-pituitary-adrenal (HPA) axis and induces behavioral alterations. However, its physiological role during stress responses remain to be elucidated. The possible mechanisms involved in IL-1-mediated stimulation of the HPA axis during stress were assessed by using different approaches. They were first studied in mice deficient for the IL-1 beta-converting enzyme (ICE) gene. Mature IL-1 beta derives from a precursor, the pro-IL-1 beta, devoid of any conventional signal sequence that is mainly processed by ICE. After immune or stress stimulation, ICE-deficient mice were shown to have a hyperactive HPA axis and to able to produce immunoreactive IL-1 beta. This indicates that the greater reactivity of the HPA axis could result from a higher sensitivity to non-ICE-matured IL-1 beta, as suggested by a higher basal transcription of hypothalamic IL-1 receptor type I (IL-1 RI) in ICE-deficient mice. The biological effects of IL-1 beta are mediated by IL-1 RI associated with the IL-1 receptor accessory protein (IL-1RAcP). IL-1RAcP is an essential component for IL-1 action at the periphery, but its role in the brain is not well known. Therefore, the effects of i.c.v. IL-1 beta were studied in IL-1RAcP-deficient mice. In normal mice, i.c.v. IL-1 beta depresses peripheral immune responses, induces the production of plasma IL-6, and stimulates the HPA axis. None of these effects were observed in IL-1RAcP-deficient mice, indicating that IL-1RAcP is necessary for the induction of the main neuroendocrine and immune effects of central IL-1 beta. In normal mice, the role of IL-1 beta was assessed by pretreating the animal with the IL-1 receptor antagonist (IL-1Ra). IL-1Ra did modify the activation of the HPA axis observed during stress, except when the animals were previously sympathectomized. This suggests that the sympathetic nervous system can downregulate the IL-1 beta-induced stimulation of the HPA axis. Finally, the modulation of the production and physiological activities of IL-1 were studied in normal mice, taking advantage of interindividual differences in brain-immune interactions linked to cerebral lateralization. Behavioral/brain lateralization was shown to be related to behavioral response to peripheral administration of IL-1, and to the production of IL-1 and IL-6 in response to LPS. This suggests that cytokines, and especially IL-1 beta, may represent one of the factors responsible for interindividual differences in brain-immune interactions.

Residual bodies and IL-1 α stimulate expression of mRNA for IL-1 α and IL-1 receptor type I in cultured rat Sertoli cells

The cytokine interleukin (IL)-1 α may be produced both by Sertoli cells and immature male germ cells from rat and is thought to play a role in autocrine and/or paracrine regulation of the spermatogenesis. The localization of IL-1 receptors in seminiferous tubules is unknown. In this study we found a constitutive expression of IL-1 receptor type I (IL-1 RI) mRNA in cultured Sertoli cells and peritubular cells from rat, whereas no such transcripts were observed in immature germ cells (pachytene spermatocytes and round spermatids). An autostimulation of IL-1 α mRNA synthesis has previously been described in other cell types. Stimulation of Sertoli cells with recombinant IL-1 α for 0–7 h resulted in a rapid increase in both IL-1 α and IL-1 RI mRNA. When Sertoli cells were cultured with residual bodies for 0–48 h, mRNA levels for both IL-1 α and IL-1 RI were increased in a biphasic manner. We suggest that phagocytosis of residual bodies triggers an autocrine IL-1 α loop in Sertoli cells where both IL-1 α and one of its receptors are stimulated.

Podocytes are the major source of IL-1α and IL-1β in human glomerulonephritides

To address the question of in situ production of IL-1 alpha and IL-1 beta in proliferative and non-proliferative forms of human glomerulonephritis (GN), we performed immunocytochemical and in situ hybridization studies on renal biopsies from patients with mesangial IgA-GN (N = 38), idiopathic membranous GN (MGN; N = 12), minimal change disease (MCD; N = 9), focal segmental glomerulosclerosis (FSGS; N = 5) and acute endocapillary GN (AGN; N = 3). Normal kidneys (N = 10) served as controls. Concomitantly, the expression of IL-1 receptor type I (IL-1 RI), IL-1 receptor type II (IL-1 RII) and of IL-1 receptor antagonist (IL-1 RA) was analyzed. Antibodies against antigens expressed on podocytes (PP-44), endothelial cells (CD31) and monocytes/macrophages (CD11b, CD14, CD68) were applied to attribute the expression of IL-1/IL-1 related peptides to intrinsic glomerular and/or blood-derived infiltrating cells. Our results demonstrate that IL-1 RII is constitutively expressed on endothelial cells, and its expression can be induced in proximal tubular cells and in the interstitium. In diseased glomeruli podocytes are capable of producing IL-1 alpha/beta. In MGN and MCD/FSGS, the expression of both IL-1 forms is particularly noted in early stages of the disease and is not only accompanied by a marked reactivity for IL-1 RI, but also for IL-1 RA. In segmental sclerosing lesions in FSGS and in IgA-GN with marked glomerular proliferation and/or sclerosis, a reduced expression of the PP-44 antigen and a diminished ability of podocytes to produce IL-1/IL-1 related peptides are noted. These results suggest that intrinsic glomerular production of IL-1 may be of relevance for the protection of glomeruli from continuing injury.

Autocrine interleukin-1 receptor antagonist can support malignant growth of glioblastoma by blocking growth-inhibiting autocrine loop of interleukin-1.

In situ hybridization (ISH) of human glioblastoma tissue sections revealed expression of interleukin-1 (IL-1)alpha and/or beta and IL-1 receptor types I and II (IL-1R I and II) in the majority of cases evaluable. To understand the function of IL-1-family members in human glioblastomas, we have studied 6 glioblastoma cell lines. RT-PCR, ISH, ELISA and 125I-IL-1-binding assays revealed expression of IL-1 and high-affinity receptors for human (h)IL-1 in all but 1 cell line. Using a colony growth assay in semi-solid media for testing serial plating efficacy (PE, number of colonies per number of cells seeded in %), only the IL-1R-negative cell line was not influenced by recombinant human (rh)IL-1alpha or -beta, whereas IL-1 down-regulated the self-renewal of clonogenic cells of the other glioblastomas. Tritiated thymidine uptake was down-regulated by rhIL-1 in all cell lines studied. Cell viability remained unchanged by rhIL-1. Wherever growth modulation by rhIL-1 was detected, it could be reversed by either soluble IL-1R I or II or by rhIL-1 receptor antagonist (ra). IL-1ra not only was able to reverse rhIL-1-induced growth modulation but alone could modulate glioblastoma growth in comparison with control in cell lines producing IL-1. Our results show the presence of public autocrine loops for IL-1 leading to growth inhibition in some glioblastomas. To understand these loops, we have studied expression and function of IL-1ra in glioblastomas. ISH of human glioblastoma tissue sections revealed expression of hIL-1ra in all 8 cases evaluable. In 4 of 6 cell lines, IL-1ra was found in the supernatant under constitutive conditions, the IL-1R-negative line being among the 2 non-producers. The other non-producing cell line, HTB 17, showed expression of hIL-1R II. Most interestingly, a neutralizing antibody against IL-1ra down-regulated growth of IL-1- and IL-1ra-producing glioblastoma cells to approx. 30% of the controls. Thus, public autocrine loops for IL-1 in human glioblastomas exist and result in growth inhibition. An autocrine production of IL-1-antagonizing molecules such as IL-1ra by these tumors can counteract this IL-1 function and represent a basic escape mechanism supporting malignant growth in some glioblastomas.

THREE DISTINCT PROMOTERS DIRECT TRANSCRIPTION OF DIFFERENT 5′ UNTRANSLATED REGIONS OF THE HUMAN INTERLEUKIN 1 TYPE I RECEPTOR: A POSSIBLE MECHANISM FOR CONTROL OF TRANSLATION

The 5′ untranslated regions (UTR) of the human interleukin 1 (IL-1) type I receptor (IL—1RI) are encoded by one common exon (exon 2) but one of three distinct exons 1 (termed exon 1A, 1B, and 1C). These exons span approximately 50 Kb of genomic DNA. Exons 1A and 1B have multiple transcriptional initiation sites, whereas the promoter for exon 1C uses a single start site. There are no ‘TATA’ or ‘CAAT’ boxes, indicating that these promoters belong to the family of housekeeping gene promoters. Computer sequence analysis of exons 1A, 1B, and 1C predicts the potential to form stable secondary structures (ΔG° 1A= −72.2 Kcal/mol, ΔG° 1B= −125.8 Kcal/mol, ΔG° 1C= −255.4 Kcal/mol). Exon 1C appears to be the most stable whereas exon 1A would yield a mRNA species more likely to be translated than those derived from exon 1B or 1C. The 5′ UTR of exon 1C is also rich (75%) in GC which might inhibit expression. Therefore, we studied the effect of exon 1C on chloramphenicol acetyltransferase (CAT) activity. Deletion of 183 or 296 base pairs from this GC rich region was shown to increase CAT activity. In addition, insertion of a GC-rich fragment of exon 1C inhibited CAT activity driven by SV40 promoter. These results suggest that the 5′ UTR exon 1C of the human IL—1RI may exert a suppressive effect on the translation of IL-1RI transcripts.

Modulation of interleukin-1 receptor expression by transforming growth factor-β in cultured rabbit articular chondrocytes: analysis by reverse transcription-polymerase chain reaction

Interleukin-1 receptor type I (IL-1RI) expression in cultured rabbit articular chondrocytes (RAC) was studied by reverse transcription-polymerase chain reaction (RT-PCR). A cDNA probe specific for the rabbit IL-1RI gene was constructed using primers derived from the sequence data of the human, murine and chick receptors. Transforming growth factor-beta 1 (TGF beta-1) was shown to transiently increase the level of expected 900-bp PCR product at 1 h of incubation and decrease the expression at 48 and 72 h with no effect at 24 h. In receptor binding assays using [125I]-IL-1 alpha, TGF beta decreased IL-1R bioactivity at all time points. These results suggest that TGF beta-induced down-regulation of IL-1 RI could be responsible for its ability to antagonize the effect of IL-1 and that TGF beta may have a role in the repair of articular cartilage.

Expression of type I interleukin-1 receptor mRNA and its regulation in cultured astrocytes

The expression of type I interleukin-1 receptor (IL-1 RI) mRNA in enriched astrocytes and microglia was investigated by Northern blot analysis. 1L-1 R1 mRNA was expressed in untreated astrocytes, but not in microglia. This expression was markedly suppressed by a treatment with isoproterenol, a β-adrenoceptor agonist (1 × 10 −8 to 1 × 10 −6 M). This suppression was clearly reversed by propranolol, a β-adrenoceptor antagonist. Dibutyryl cAMP (dbcAMP) also suppressed the expression of IL- 1RI MRNA. The suppression by isoproterenol or dbcAMP was blocked by H8 or H89, inhibitors of cAMP-dependent protein kinase. The present results suggest that IL-1RI mRNA expression in cultured astrocytes may be regulated by intracellular cAMP, elevated upon the activation of β-adrenoceptors as an example, and cAMP-dependent protein kinase.

Linkage analysis of bovine interleukin 1 receptor types I and II (IL-1R I, II)

Linkage analysis of bovine interleukin 1 receptor types I and II (IL-1R I, II)