Interleukin-6 and CAAT/Enhancer Binding Protein β-Deficient Mice Act as Tools to Dissect the IL-6 Signalling Pathway and IL-6 Regulation

doi:10.1016/S0171-2985(97)80035-6

Immunobiology

Volume 198, Issues 1–3, December 1997, Pages 144-156

https://doi.org/10.1016/S0171-2985(97)80035-6 Get rights and content

Abstract

Interleukin-6 (IL-6) is a pleiotropic cytokine playing important roles in immunity, hemopoiesis and inflammation. IL-6 signalling is known to involve the activation of two independent transcription factors: Stat3 (through phosphorylation by Jak kinases) and C/EBPβ (through activation of the ras pathway). In addition, C/EBPβ is believed to act as a transcriptional activator of the IL-6 gene itself. Making use of IL-6-deficient mice, we have recently demonstrated that IL-6 is essential for the induction of acute phase mRNAs in the liver upon localized tissue damage, but not upon systemically induced inflammation. Here we show that the defective mRNA induction is paralleled by a defective activation of Stat3, thus establishing a direct relationship between IL-6 function, Stat3 activation and acute phase genes induction. On the other hand, making use of C/EBPβ-deficient mice, we show that the induction of IL-6 by a variety of stimuli does not require C/EBPβ activity. In contrast to the predicted activating role of C/EBPβ, IL-6 levels are increased in the C/EBPβ-deficient mice, suggesting that C/EBPβ may act as a down-modulator of the IL-6 gene. Through the generation of C/EBPβ, IL-6 double mutant mice we show that IL-6 hyperproduction is responsible for the development of the Castleman's like lymphoproliferative disease described in the C/EBPβ-deficient mice, since the disorder is completely blocked by inactivating the IL-6 gene.

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Abnormal expression of CEBPB promotes the progression of renal cell carcinoma through regulating the generation of IL-6
2023, Heliyon
The CCAAT/enhancer-binding protein beta (CEBPB), a transcription factor regulating immune and inflammatory responses, has been implicated in the pathogenesis of various malignancies. However, its specific regulatory mechanism in renal cell carcinoma (RCC) remains poorly understood.
The expression of CEBPB was detected in RCC cells and tissues using qRT-PCR, western blotting and immunohistochemistry. ELISA assay was used to detect the immune factors regulated by CEBPB in supernatants. Additionally, western blotting was employed to measure the phosphorylation level of STAT3 and the expression levels of its downstream target genes.
CEBPB was found to be overexpressed in both RCC tissues and cell lines, and its higher expression was associated with a lower survival rate. In RCC cells, CEBPB enhances the expression of IL6, consequently promoting the phosphorylation of STAT3 and the expression of its downstream target genes. This mechanism ultimately facilitates tumor progression.
The dysregulated expression of CEBPB facilitates RCC progression through the IL6/STAT3 pathway. CEBPB is a potential diagnostic markers and a novel effective therapeutic target for RCC patients.
Novel insights and therapeutic approaches in idiopathic multicentric Castleman disease
2018, Blood
Citation Excerpt :
Janus kinase (JAK)/signal transducer activator of transcription 3 (STAT3), mitogen-activated protein kinase (MAPK), and phosphatidylinositol-3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) are the 3 primary pathways activated by IL-6. Knocking out CCAAT/enhancer binding protein b (C/EBPb) causes an iMCD-like phenotype in mice because of unopposed STAT3.37 Pierson et al found that MAPK and PI3K/Akt/mTOR signaling are both enriched among the most up- and downregulated plasma proteins during iMCD flare, and the greatest number of compounds (6 of the top 20) that downregulate expression (in vitro) of the most upregulated proteins in iMCD-TAFRO patients, including VEGF, CXCL13, and other chemokines, target PI3K and/or mTOR.29
Castleman disease (CD) describes a heterogeneous group of hematologic disorders that share characteristic lymph node histopathology. Patients of all ages present with either a solitary enlarged lymph node (unicentric CD) or multicentric lymphadenopathy (MCD) with systemic inflammation, cytopenias, and life-threatening multiple organ dysfunction resulting from a cytokine storm often driven by interleukin 6 (IL-6). Uncontrolled human herpesvirus-8 (HHV-8) infection causes approximately 50% of MCD cases, whereas the etiology is unknown in the remaining HHV-8-negative/idiopathic MCD cases (iMCD). The limited understanding of etiology, cell types, and signaling pathways involved in iMCD has slowed development of treatments and contributed to historically poor patient outcomes. Here, recent progress for diagnosing iMCD, characterizing etio-pathogenesis, and advancing treatments are reviewed. Several clinicopathological analyses provided the evidence base for the first-ever diagnostic criteria and revealed distinct clinical subtypes: thrombocytopenia, anasarca, fever, reticulin fibrosis/renal dysfunction, organomegaly (iMCD-TAFRO) or iMCD-not otherwise specified (iMCD-NOS), which are both observed all over the world. In 2014, the anti-IL-6 therapy siltuximab became the first iMCD treatment approved by the US Food and Drug Administration, on the basis of a 34% durable response rate; consensus guidelines recommend it as front-line therapy. Recent cytokine and proteomic profiling has revealed normal IL-6 levels in many patients with iMCD and potential alternative driver cytokines. Candidate novel genomic alterations, dysregulated cell types, and signaling pathways have also been identified as candidate therapeutic targets. RNA sequencing for viral transcripts did not reveal novel viruses, HHV-8, or other viruses pathologically associated with iMCD. Despite progress, iMCD remains poorly understood. Further efforts to elucidate etiology, pathogenesis, and treatment approaches, particularly for siltuximab-refractory patients, are needed.
Castleman Disease Pathogenesis
2018, Hematology/Oncology Clinics of North America
HHV-8-negative, idiopathic multicentric Castleman disease: Novel insights into biology, pathogenesis, and therapy
2014, Blood
Citation Excerpt :
Also, knocking out C/EBPβ in mice causes a lymphoproliferative disease nearly identical to iMCD, and symptoms can be reversed with IL-6 blockade.70 The elevated IL-6 secretion in these mice, which was not anticipated, was hypothesized to occur because of unopposed STAT3 signaling or preferential expansion of type 2 helper (TH2) pools.70 A candidate aberrantly activated kinase is the anaplastic lymphoma kinase (ALK) gene, which drives the growth of inflammatory myofibroblastic tumors and causes a Castleman-like syndrome following a translocation event.49
Multicentric Castleman's disease (MCD) describes a heterogeneous group of disorders involving proliferation of morphologically benign lymphocytes due to excessive proinflammatory hypercytokinemia, most notably of interleukin-6. Patients demonstrate intense episodes of systemic inflammatory symptoms, polyclonal lymphocyte and plasma cell proliferation, autoimmune manifestations, and organ system impairment. Human herpes virus-8 (HHV-8) drives the hypercytokinemia in all HIV-positive patients and some HIV-negative patients. There is also a group of HIV-negative and HHV-8-negative patients with unknown etiology and pathophysiology, which we propose referring to as idiopathic MCD (iMCD). Here, we synthesize what is known about iMCD pathogenesis, present a new subclassification system, and propose a model of iMCD pathogenesis. MCD should be subdivided into HHV-8-associated MCD and HHV-8-negative MCD or iMCD. The lymphocyte proliferation, histopathology, and systemic features in iMCD are secondary to hypercytokinemia, which can occur with several other diseases. We propose that 1 or more of the following 3 candidate processes may drive iMCD hypercytokinemia: systemic inflammatory disease mechanisms via autoantibodies or inflammatory gene mutations, paraneoplastic syndrome mechanisms via ectopic cytokine secretion, and/or a non-HHV-8 virus. Urgent priorities include elucidating the process driving iMCD hypercytokinemia, identifying the hypercytokine-secreting cell, developing consensus criteria for diagnosis, and building a patient registry to track cases.
Stat3 activation links a C/EBPδ to myostatin pathway to stimulate loss of muscle mass
2013, Cell Metabolism
Citation Excerpt :
The C/EBP transcription factors (C/EBPα, -β, -γ, -δ, -ω, and -ζ) are expressed in several tissues and act to regulate inflammatory and metabolic processes (Ramji and Foka, 2002). C/EBPβ or -δ can stimulate intracellular signaling in hepatocytes or inflammatory cells (Poli, 1998; Akira et al., 1990; Alonzi et al., 1997) and in muscles of mice responding to an excess of glucocorticoids, the expression and binding activity of C/EBPβ and -δ are increased (Penner et al., 2002; Yang et al., 2005). A potential mechanism that includes C/EBPδ involves increased myostatin expression because the myostatin promoter contains recognition sites for members of the C/EBP family of transcription factors (Ma et al., 2001; Allen et al., 2010).
Catabolic conditions like chronic kidney disease (CKD) cause loss of muscle mass by unclear mechanisms. In muscle biopsies from CKD patients, we found activated Stat3 (p-Stat3) and hypothesized that p-Stat3 initiates muscle wasting. We created mice with muscle-specific knockout (KO) that prevents activation of Stat3. In these mice, losses of body and muscle weights were suppressed in models with CKD or acute diabetes. A small-molecule that inhibits Stat3 activation produced similar responses, suggesting a potential for translation strategies. Using CCAAT/enhancer-binding protein δ (C/EBPδ) KO mice and C2C12 myotubes with knockdown of C/EBPδ or myostatin, we determined that p-Stat3 initiates muscle wasting via C/EBPδ, stimulating myostatin, a negative muscle growth regulator. C/EBPδ KO also improved survival of CKD mice. We verified that p-Stat3, C/EBPδ, and myostatin were increased in muscles of CKD patients. The pathway from p-Stat3 to C/EBPδ to myostatin and muscle wasting could identify therapeutic targets that prevent muscle wasting.
C/EBP-δ induction by gp130 signaling: Role in transition to myelin gene expressing phenotype in a melanoma cell line model
2004, Journal of Biological Chemistry
Expression of genes encoding structural myelin proteins marks the inception of the myelinating Schwann cell (SC) phenotype. Earlier embryonic SC as well as adult non-myelinating SC produce the intermediate filament glial fibrillary acid protein (GFAP), which disappears from the myelinating SC. We previously observed that triggering of the gp130 receptor system by the IL6RIL6 ligand, comprising interleukin-6 (IL-6) fused to the soluble IL-6 receptor, induces myelin gene expression in rat embryonic dorsal root ganglia (DRG) cultures as well as in the murine melanoma cell line B16/F10.9. Study of target genes regulated by IL6RIL6 indicates a strong and selective induction of the transcriptional regulator C/EBP-δ in DRG cultures and in the F10.9 cell line. As shown here, silencing of C/EBP-δ mRNA and protein expression by introduction of small interference RNA-producing plasmids in the F10.9 cells prevented the induction of myelin protein zero (P0) and myelin basic protein (MBP) mRNAs by IL6RIL6. Doxycycline-regulated overexpression of C/EBP-δ was sufficient to induce accumulation of P0 and MBP mRNAs, the effect being selective, because C/EBP-δ did not affect several other genes strongly regulated by IL6RIL6. Interestingly, GFAP was inhibited by C/EBP-δ overexpression, leading to a modulation of the ratio between myelin gene products versus GFAP and suggesting that C/EBP-δ plays a role in the switch to a myelinating phenotype. The down-regulation of Pax3, also typical of the transition to myelinating cells, was observed after C/EBP-δ expression in correlation to P0 induction and to decrease of melanogenesis and cell growth. In cultures of dissociated cells of embryonic rat DRG, where we knocked-down the C/EBP-δ mRNA, we found an inhibition of P0 mRNA induction by IL6RIL6, showing that the role of C/EBP-δ on this myelin gene is not unique to the melanoma system.

View all citing articles on Scopus

¹: Dr. Valeria Poli, Department of Biochemistry, Medical Sciences Institute, University of Dundee, Dundee DD1 4HN, UK.

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Interleukin-6 and CAAT/Enhancer Binding Protein β-Deficient Mice Act as Tools to Dissect the IL-6 Signalling Pathway and IL-6 Regulation

Abstract

Immunol. Today

Blood

Blood

Cell

Cell

J. Biol. Chem.

Cell

Cell

J. Biol. Chem.

Blood

Transcriptional regulation of acute phase genes by IL-6 and related cytokines

A nuclear factor for IL-6 expression (NF-IL6) is a member of a C/EBP family

EMBO J

C/EBPβ is required for the late phases of acute phase genes induction in the liver and for tumor necrosis factor-α, but not interleukin-6, regulation

Cell Death & Differentiation

Transcriptional responses to polypeptide ligands: the Jak-Stat pathway

Annu. Rev. Biochem.