Differential response of neuronal cells to a fusion protein of ciliary neurotrophic factor/soluble CNTF-receptor and leukemia inhibitory factor

Ciliary neurotrophic factor (CNTF) displays neurotrophic activities on motor neurons and neural cell populations both in vivo and in vitro. On target cells lacking intrinsic expression of specific receptor alpha subunits cytokines of the IL-6 family only act in the presence of their specific agonistic soluble receptors. Here, we report the construction and expression of a CNTF/soluble CNTF-receptor (sCNTF-R) fusion protein (Hyper-CNTF) with enhanced biological activity on cells expressing gp130 and leukemia inhibitory factor receptor (LIF-R), but not membrane-bound CNTF-R. At the cDNA level, the C-terminus of the extracellular domain of human CNTF-R (amino acids 1-346) was linked via a sing…

AN IL-6/IL-6 SOLUBLE RECEPTOR (IL-6R) HYBRID PROTEIN (H-IL-6) INDUCES EPO-INDEPENDENT ERYTHROID DIFFERENTIATION IN HUMAN CD34+CELLS

H-IL-6 is a hybrid protein constructed to contain IL-6 and its soluble receptor linked by a flexible peptide chain. Here we show that H-IL-6 strongly enhances proliferation of human CD34(+)cells in serum-free liquid culture, and that the majority of the cells generated belong to the erythroid lineage, being positive for the marker Glycophorin A. Conversely, H-IL-6 does not increase the number of myeloid, CD13-positive cells. Comparable effects are observed on progenitors from cord blood and adult peripheral blood. Therefore, H-IL-6 triggers an erythroid-inducing signal in haematopoietic progenitor cells, independently from erythropoietin (EPO).

Extramedullary Expansion of Hematopoietic Progenitor Cells in Interleukin (IL)-6–sIL-6R Double Transgenic Mice

Soluble cytokine receptors modulate the activity of their cognate ligands. Interleukin (IL)-6 in association with the soluble IL-6 receptor (sIL-6R) can activate cells expressing the gp130 signal transducer lacking the specific IL-6R. To investigate the function of the IL-6–sIL-6R complex in vivo and to discriminate the function of the IL-6–sIL-6R complex from the function of IL-6 alone, we have established a transgenic mouse model. Double-transgenic mice coexpressing IL-6 and sIL-6R were generated and compared with IL-6 and sIL-6R single-transgenic mice. The main phenotype found in IL-6–sIL-6R mice was a dramatic increase of extramedullary hematopoietic progenitor cells in liver and spleen…

Model Based Targeting of IL-6-Induced Inflammatory Responses in Cultured Primary Hepatocytes to Improve Application of the JAK Inhibitor Ruxolitinib

IL-6 is a central mediator of the immediate induction of hepatic acute phase proteins (APP) in the liver during infection and after injury, but increased IL-6 activity has been associated with multiple pathological conditions. In hepatocytes, IL-6 activates JAK1-STAT3 signaling that induces the negative feedback regulator SOCS3 and expression of APPs. While different inhibitors of IL-6-induced JAK1-STAT3-signaling have been developed, understanding their precise impact on signaling dynamics requires a systems biology approach. Here we present a mathematical model of IL-6-induced JAK1-STAT3 signaling that quantitatively links physiological IL-6 concentrations to the dynamics of IL-6-induced …

Liver regeneration induced by a designer human IL‐6/ sIL‐6R fusion protein reverses severe hepatocellular injury

The cytokine IL-6 plays a significant role in liver regeneration in conjunction with additional growth factors (HGF, TNF-α, and TGF-α). Many IL-6 effects depend on a naturally occurring soluble IL-6 receptor (sIL-6R). Here, the chimeric protein hyper-IL-6, constructed from the human IL-6 protein fused to a truncated form of its receptor, was found to have superagonistic IL-6 properties, and as such, enhanced liver cell regeneration. Hyper-IL-6 reversed the state of hepatotoxicity and enhanced the survival rates of rats suffering from fulminant hepatic failure after D-galactosamine administration. The hyper-IL-6 protein has a significant potential for use in the treatment of severe human liv…

The designer cytokine hyper-IL-6 mediates growth inhibition and GM-CSF-dependent rejection of B16 melanoma cells.

The low immunogenic B16 melanoma cell line was transfected with a mammalian expression vector containing the complementary DNA for a sIL-6R/IL-6 fusion protein, termed Hyper-IL-6 (H-IL-6), which was shown to have biological activities at 100-1000-fold lower concentrations than IL-6 in combination with sIL-6R. The secreted p84 glycoprotein was detected in the supernatant of transfected cells and was fully active on BAF3/gp130 cells, which respond to IL-6/sIL-6R but not to IL-6 alone. Administration of recombinant H-IL-6 to C57BL/6 mice resulted in a prolonged acute phase protein gene expression indicating long systemic persistence of the fusion protein. Transfected B16 cells (B16/H-IL6 cells…

Activation of gp 130 by IL-6/soluble IL-6 receptor induces neuronal differentiation

Interleukin-6 (IL-6) on target cells binds to the specific IL-6 receptor (IL-6R) and subsequently induces homodimerization of the signal-transducing protein gp130. Cells which express gp130 but no IL-6R and which therefore do not respond to IL-6 can be stimulated by the complex of IL-6 and soluble IL-6R (slL-6R). Here we show that on rat pheochromocytoma cells (PC12), the combination of IL-6 and slL-6R but not IL-6 alone induces expression of c-fos, GAP-43 and neuron-specific enolase followed by neuron-specific differentiation and formation of a neuronal network. The differentiation was dose-and time-dependent and followed the same kinetics as nerve-growth factor (NGF)-induced differentiati…

Hyper-IL-6 (H-IL-6), a Fusion Protein of Soluble IL-6 Receptor (Sil-6R), and Interleukin-6 (IL-6), Acts Synergistic with Thrombopoietin (TPO) and Stem Cell Factor (SCF) in Expanding Megakaryocyte Progenitors from Human Cd34++/Cd90+ Cell

It has been shown that signalings from c-kit and gpl30, the signal-transducing receptor component of the IL6 receptor, act synergistic for the ex-vivo expansion of multipotential hematopoietic progenitors. A similar synergistic effect has been demonstrated for signalings from c-kit, c-mpl, and flt3. While c-kit is activated by stem cell factor (SCF), gpl30 can be activated by the complex of soluble IL-6 receptor (sIL-6R) and interleukin-6 (IL-6). Recently, a bioactive designer cytokine, H-IL-6, a fusion protein consisting of SIL-6R and IL-6 linked by a flexible peptide chain has been shown to expand human hematopoietic colony-forming cells. We tested the activity of H-IL-6 alone and in comb…

The function of the soluble IL-6 receptor in vivo.

Interleukin-6 (IL-6) is considered an important mediator of acute inflammatory responses. Moreover, IL-6 functions as a differentiation and growth factor of hematopoietic precursor cells, B-cells, T-cells, keratinocytes, neuronal cells, osteoclasts and endothelial cells. IL-6 exhibits its action via a receptor complex consisting of a specific IL-6 receptor (IL-6R) and a signal-transducing subunit (gp130). Soluble forms of both receptor components are generated by shedding and are found in patients with various diseases such as AIDS, rheumatoid arthritis and others. The function of the soluble IL-6R in vivo is unknown. To discriminate between the biologic function of hIL-6 alone and that of …

Soluble interleukin-6 receptor (sIL-6R) makes IL-6R negative T cell line respond to IL-6: it inhibits TNF production

The receptor for interleukin-6 (IL-6) consists of two subunits: a ligand specific IL-6Ralpha and gp130 that is responsible for signal-transduction. A soluble form of the ligand specific chain was described that when complexed to IL-6 is capable of binding to the membrane-bound gp130 subunit and thus can elicit signal-transduction. This soluble receptor can act on cells that express only the gp130 but not the ligand-specific subunit of the IL-6R. This phenomenon, called trans-signaling, introduced a novel aspect of cytokine action. In this study we examined the response of Jurkat cells, that are known not to express IL-6Ralpha, to IL-6, the soluble IL-6 receptor (sIL-6R) and a covalent compl…

Recombinant Human Single Chain Fv Antibodies Recognizing Human Interleukin-6

A human antibody library was displayed on the surface of filamentous bacteriophage and screened for binding to human interleukin-6 (IL-6). Two antibody-bearing phages were selected that bound IL-6. The complementary-determining region 3 loops of the variable heavy chains of these two antibodies differed in length and sequence and recognized two distinct epitopes. One of the single chain Fv fragments isolated (H1) was found to bind human (but not murine) IL-6 with an affinity comparable to that of the human IL-6 receptor. H1 also recognized newly synthesized human IL-6 intracellularly, as shown by indirect immunofluorescence. H1 did not neutralize human IL-6, and the H1 epitope was mapped to…

The membrane distal half of gp130 is responsible for the formation of a ternary complex with IL-6 and the IL-6 receptor

AbstractGp130 is the signal transducing subunit of the interleukin-6 receptor. Signaling is initiated by the complex formation of gp130 with IL-6 bound to the IL-6 receptor (IL-6R). We have subdivided the extracellular domain of gp130 in two parts and expressed the mutant proteins as soluble IgG fusion proteins in COS-7 cells. By studying the formation of the ternary complex we show that the membrane distal half of gp130 which contains a cytokine receptor domain is responsible for the interaction with the IL-6/IL-6R complex. Interestingly this is the same region which is believed to be involved in specific recognition of the related cytokines LIF, OM, and probably also of CNTF and IL-11.

IL-6 trans-Signaling

The molecular consequence of the fever response has been illuminated by a recent study showing that a temperature shift to 40 degrees C resulted in increased leukocyte adhesion to tissue sections, which was mediated by L-selectin activation in lymphocytes. This L-selectin activation during heat responses was dependent on IL-6 trans-signaling via the soluble IL-6R.

Tissue inhibitor of metalloproteinases-2 (TIMP-2) in rat liver cells is increased by lipopolysaccharide and prostaglandin E2

AbstractTo explore the functional role of TIMP-2 in liver, we determined TIMP-2 mRNA levels in primary rat hepatocytes and in total rat liver. Rat hepatocytes constitutively express TIMP-2 mRNA at a low level. Incubation with dexamethasone, prostaglandin E2 and a combination of inflammatory cytokines leads to an up-regulation of TIMP-2 mRNA. In rats in vivo we found a dramatic increase of TIMP-2 expression after intraperitoneal injection of lipopolysaccharide. Compared to our previous findings on TIMP-1 we conclude that TIMP-2 mRNA expression is regulated in a distinct and partially opposite manner. Over-production of TIMP-2 could inhibit the activity of metalloproteinases and thus lead to …

Generation of tumor-reactive CTL against the tumor-associated antigen HER2 using retrovirally transduced dendritic cells derived from CD34+ hemopoietic progenitor cells.

Abstract Ag-specific CD8+ CTL are crucial for effective tumor rejection. Attempts to treat human malignancies by adoptive transfer of tumor-reactive CTL have been limited due to the difficulty of generating and expanding autologous CTL with defined Ag specificity. The current study examined whether human CTL can be generated against the tumor-associated Ag HER2 using autologous dendritic cells (DC) that had been genetically engineered to express HER2. DC progenitors were expanded by culturing CD34+ hemopoietic progenitor cells in the presence of the designer cytokine HyperIL-6. Proliferating precursor cells were infected by a retroviral vector encoding the HER2 Ag and further differentiated…

Cytokines and Neurotrophins Interact in Normal and Diseased States

Neurotrophins (NTs) such as nerve growth factor (NGF) as well as cytokines, for example, interleukin-6 (IL-6), are communicators between the nervous and immune systems. There is evidence for mutual interactions between NTs and cytokines. Strategies are being developed to elucidate the molecular mechanism/s of interactions and to understand how cytokines are involved in health and disease. Analysis of underlying signaling pathways in glial cells indicates that different transcription factors, such as NF-kappa B, cAMP-responsive-element binding protein (CREB), and activator protein 1 (AP-1), are involved in NT induction. IL-6 and NTs of the NGF family are coexpressed at sites of nerve injury.…

Optimization of retroviral-mediated gene transfer to human NOD/SCID mouse repopulating cord blood cells through a systematic analysis of protocol variables

Abstract Retroviral transduction of human hematopoietic stem cells is still limited by lack of information about conditions that will maximize stem cell self-renewal divisions in vitro. To address this, we first compared the kinetics of entry into division of single human CD34 + CD38 − cord blood (CB) cells exposed in vitro to three different flt3-ligand (FL)-containing cytokine combinations. Of the three combinations tested, FL + hyperinterleukin 6 (HIL-6) yielded the least clones and these developed at a slow rate. With either FL + Steel factor (SF) + HIL-6 + thrombopoietin (TPO) or FL + SF + interleukin 3 (IL-3) + IL-6 + granulocyte-colony-stimulating factor (G-CSF), >90% of the cells th…

Soluble gp130 is the natural inhibitor of soluble interleukin-6 receptor transsignaling responses

Signal transduction in response to interleukin-6 (IL-6) requires binding of the cytokine to its receptor (IL-6R) and subsequent homodimerization of the signal transducer gp130. The complex of IL-6 and soluble IL-6R (sIL-6R) triggers dimerization of gp130 and induces responses on cells that do not express membrane bound IL-6R. Naturally occurring soluble gp130 (sgp130) can be found in a ternary complex with IL-6 and sIL-6R. We created recombinant sgp130 proteins that showed binding to IL-6 in complex with sIL-6R and inhibited IL-6/sIL-6R induced proliferation of BAF/3 cells expressing gp130. Surprisingly, sgp130 proteins did not affect IL-6 stimulated proliferation of BAF/3 cells expressing …

Yeast expression of the cytokine receptor domain of the soluble interleukin-6 receptor

Abstract The complex of the soluble interleukin-6 receptor (sIL-6R) and IL-6 (IL-6) is a potent agonist on cells expressing the signal transducing protein gp130. In contrast, IL-6 alone only stimulates cells which express a membrane bound form of the IL-6R and gp130. The natural occurring sIL-6R is generated by shedding of the membrane receptor and to a lesser extend by alternative splicing. We have inserted the coding sequence of the 323 amino acid residues of the human sIL-6R into an expression/secretion vector suitable for the methylotrophic yeast Pichia pastoris . We obtained, however, no detectable expression and secretion of the recombinant protein. When we used only the coding sequen…

The substrate degradome of meprin metalloproteases reveals an unexpected proteolytic link between meprin β and ADAM10

The in vivo roles of meprin metalloproteases in pathophysiological conditions remain elusive. Substrates define protease roles. Therefore, to identify natural substrates for human meprin α and β we employed TAILS (terminal amine isotopic labeling of substrates), a proteomics approach that enriches for N-terminal peptides of proteins and cleavage fragments. Of the 151 new extracellular substrates we identified, it was notable that ADAM10 (a disintegrin and metalloprotease domain-containing protein 10)—the constitutive α-secretase—is activated by meprin β through cleavage of the propeptide. To validate this cleavage event, we expressed recombinant proADAM10 and after preincubation with meprin…

The transcription factor NFATc2 controls IL-6–dependent T cell activation in experimental colitis

The nuclear factor of activated T cells (NFAT) family of transcription factors controls calcium signaling in T lymphocytes. In this study, we have identified a crucial regulatory role of the transcription factor NFATc2 in T cell–dependent experimental colitis. Similar to ulcerative colitis in humans, the expression of NFATc2 was up-regulated in oxazolone-induced chronic intestinal inflammation. Furthermore, NFATc2 deficiency suppressed colitis induced by oxazolone administration. This finding was associated with enhanced T cell apoptosis in the lamina propria and strikingly reduced production of IL-6, -13, and -17 by mucosal T lymphocytes. Further studies using knockout mice showed that IL-…

Cutting Edge: Trans-Signaling via the Soluble IL-6R Abrogates the Induction of FoxP3 in Naive CD4+CD25− T Cells

Abstract Chronic inflammatory diseases may develop when regulatory T cells (Tregs) fail to control the balance between tolerance and immunity. Alternatively, activated immune cells might prevent the induction or activation of Tregs in such diseases. In this study, we demonstrate that trans-signaling into T cells via the soluble IL-6 receptor completely abrogates the de novo induction of adaptive Tregs. Mechanistically, IL-6 trans-signaling augmented the expression of the TGF-β signaling inhibitor SMAD7. Consequently, SMAD7 overexpression in T cells using newly created transgenic mice rendered CD4+CD25− T cells resistant to the induction of FoxP3. Finally, IL-6 trans-signaling inhibited Treg…

Human herpes virus 8 interleukin-6 homologue triggers gp130 on neuronal and hematopoietic cells

Human herpes virus-8 (HHV8) encodes a cytokine named viral interleukin-6 (vIL-6) that shares 25% amino-acid identity with its human homologue. Human IL-6 is known to be a growth and differentiation factor of lymphatic cells and plays a potential role in the pathophysiology of various lymphoproliferative diseases. vIL-6 is expressed in HHV8-associated-diseases including Kaposi's sarcoma, Body-cavity-based-lymphoma and Castleman's disease, suggesting a pathogenetic involvement in the malignant growth of B-cell associated diseases and other malignant tumours. We expressed vIL-6 in Escherichia coli as a fusion protein with recombinant periplasmic maltose binding protein. After cleavage from the…

Astrocytic alterations in interleukin-6/Soluble interleukin-6 receptor alpha double-transgenic mice.

Interleukin-6 (IL-6), a major cytokine with diverse effects on cells mainly of the immune and hematopoietic systems, has been linked to several neurological disorders such as acquired immune deficiency syndrome dementia, multiple sclerosis, and Alzheimer's disease. Central nervous system (CNS)-specific expression of IL-6 caused neurodegeneration, massive gliosis, and vascular proliferation in transgenic mice. However, the effects of systemically circulating IL-6 and its receptor IL-6Ralpha on the CNS are unknown. IL-6Ralpha is the specific component of the IL-6 receptor system and hence an important co-factor of IL-6. IL-6Ralpha is bioactive in a membrane-bound and in a soluble (s) form. We…

Efficient retrovirus-mediated gene transfer to transplantable human bone marrow cells in the absence of fibronectin

Abstract The low frequency of transplantable hematopoietic stem cells in adult human bone marrow (BM) and other differences from cord blood stem cells have impeded studies to optimize the retroviral transduction of stem cells from adult sources. To address this problem, first a cytokine combination was defined that would both maximize the kinetics of adult BM CD34+CD38− cell mitogenesis and minimize the period of prestimulation required for the transduction of these cells by a MSCV-GFP/neor virus in tissue culture dishes in the absence of fibronectin. Three days of stimulation with flt3-ligand, Steel factor, interleukin (IL)-3, and hyper-IL-6 proved both necessary and sufficient to obtain 8…

Possible role of human interleukin-6 and soluble interleukin-6 receptor in hepatitis B virus infection

Human interleukin-6 has been shown to promote hepatitis B virus (HBV) infection. However, it is not clear whether this influence is the result of a direct interaction between interleukin-6 (IL-6) and the HBV envelope proteins or of a rather indirect mechanism. A direct interaction of IL-6 and the preS region of the large envelope protein (L-protein) of HBV has been reported. In this study we assessed the binding of IL-6 and of the IL-6 receptor subunits to the preS region of the L-protein of HBV. Binding of IL-6 and IL-6 receptor subunits sIL-6R and gp130 to preS was assessed by immunoprecipitation with recombinant preS proteins. In patient sera IL-6 and sIL-6R concentrations were analysed …

The function of the soluble interleukin 6 (IL-6) receptor in vivo: sensitization of human soluble IL-6 receptor transgenic mice towards IL-6 and prolongation of the plasma half-life of IL-6.

Interleukin 6 (IL-6) is considered an important mediator of acute inflammatory responses. Moreover, IL-6 functions as a differentiation and growth factor of hematopoietic precursor cells, B cells, T cells, keratinocytes, neuronal cells, osteoclasts, and endothelial cells. IL-6 exhibits its action via a receptor complex consisting of a specific IL-6 receptor (IL-6R) and a signal transducing subunit (gp130). Soluble forms of both receptor components are generated by shedding and are found in patients with various diseases such as acquired immune deficiency syndrome, rheumatoid arthritis, and others. The function of the soluble (s)IL-6R in vivo is unknown. Since human (h)IL-6 acts on human and…

Signals regulating neurotrophin expression in glial cells

IgG Fc sialylation is regulated during the germinal center reaction following immunization with different adjuvants

Background: Effector functions of IgG Abs are regulated by their Fc N-glycosylation pattern. IgG Fc glycans that lack galactose and terminal sialic acid residues correlate with the severity of inflammatory (auto)immune disorders and have also been linked to protection against viral infection and discussed in the context of vaccine-induced protection. In contrast, sialylated IgG Abs have shown immunosuppressive effects.Objective: We sought to investigate IgG glycosylation programming during the germinal center (GC) reaction following immunization of mice with a foreign protein antigen and different adjuvants.Methods: Mice were analyzed for GC T-cell, B-cell, and plasma cell responses, as wel…

Gp130-signaling synergizes with FL and TPO for the long-term expansion of cord blood progenitors

We investigated the effect of a new fusion protein of IL-6 and the soluble IL-6R, H-IL-6, on the long-term ex vivo expansion of hematopoietic progenitors derived from AC133+cord blood cells. H-IL-6, which acts on both IL-6Ralpha-positive and IL-6Ralpha-negative cells, effectively synergized with FL and TPO with or without SCF for the propagation of primitive progenitors. However, IL-6 showed a greater synergistic effect with FL and TPO than H-IL-6 for long-term progenitor propagation. During the first 6 weeks of culture under stroma-free serum-containing conditions, IL-6 induced a 1.96 +/- 0.64-fold higher expansion of nucleated cells, a 2.28 +/- 0.33-fold higher expansion of CD34+ cells an…

Generation and function of the soluble interleukin-6 receptor

Regulation of endotoxin-induced IL-6 production in liver sinusoidal endothelial cells and Kupffer cells by IL-10

SUMMARY Sinusoidal endothelial cells and Kupffer cells are the first cell populations in the liver that come into contact with gut-derived endotoxin in portal blood. Although endotoxin concentrations as high as 1 ng/ml are physiologically present in portal blood, no local inflammation is seen. We show that the proinflammatory cytokine IL-6, which is central to the development of inflammatory reactions in the liver, is produced by sinusoidal endothelial cells and Kupffer cells in response to low concentrations of endotoxin (100 pg/ml to 1 ng/ml). The anti-inflammatory cytokine IL-10 down-regulated endotoxin-induced IL-6 release in endothelial and Kupffer cells. Importantly, Kupffer cells sec…

IL-6 Signaling Promotes Tumor Growth in Colorectal Cancer

Recent investigations support an important role for TGF-beta in the development of colorectal cancer. However, the molecular consequences of TGF-beta signaling in the colon remains incompletely understood. In a recent study in Immunity, we analyzed the role of TGF-beta in a murine model of colon cancer. Using transgenic mice overexpressing TGF-beta or a dominant negative TGF-beta receptor II under control of the CD2 minigene, we show that TGF-beta signaling in tumor infiltrating T lymphocytes regulates the growth of dysplastic colon epithelial cells, as determined by histology and a novel system for high resolution chromoendoscopy in vivo. At the molecular level, TGF-beta signaling in T cel…

Interleukin-6 (IL-6) and its soluble receptor support survival of sensory neurons

The cytokine interleukin-6 (IL-6) has multiple functions in the immune and hematopoietic systems. IL-6 is related to ciliary neurotrophic factor (CNTF), a trophic factor for motoneurons, sensory dorsal root ganglion (DRG) neurons, and other neuronal subpopulations. Both act via related receptor complexes, consisting of one ligand-specific α-receptor subunit (IL-6R and CNTFR, respectively) and two signal-transducing receptor components. Even though IL-6 is expressed by neurons and glia, the functions of IL-6 in the nervous system are poorly understood. Here, we report that exogenous human IL-6 promotes the survival of dissociated newborn rat DRG neurons in vitro if supplemented with soluble …

Blockade of interleukin 6 trans signaling suppresses T-cell resistance against apoptosis in chronic intestinal inflammation: Evidence in Crohn disease and experimental colitis in vivo

The pro-inflammatory cytokine interleukin (IL)-6 (refs. 1-5) can bind to cells lacking the IL-6 receptor (IL-6R) when it forms a complex with the soluble IL-6R (sIL-6R) (trans signaling). Here, we have assessed the contribution of this system to the increased resistance of mucosal T cells against apoptosis in Crohn disease (CD), a chronic inflammatory disease of the gastrointestinal tract. A neutralizing antibody against IL-6R suppressed established experimental colitis in various animal models of CD mediated by type 1 T-helper cells, by inducing apoptosis of lamina propria T cells. Similarly, specific neutralization of sIL-6R in vivo by a newly designed gp130-Fc fusion protein caused suppr…

Local blockade of IL-6R signaling induces lung CD4+ T cell apoptosis in a murine model of asthma via regulatory T cells.

We previously reported high levels of the soluble form of the IL-6R (sIL-6R) in the airways of asthmatic subjects. Here, we analyzed the IL-6R effects on Th2 cell survival in the lung by locally antagonizing sIL-6R-mediated trans-signaling with a designer fusion protein (gp130-Fc) as well as IL-6R signaling with an antibody against the gp80 unit of the IL-6R (alphaIL-6R) in a murine model of asthma after ovalbumin peptide (OVA) sensitization and challenge. Blockade of the sIL-6R led to a significant decrease in inflammatory cells by an apoptosis-independent mechanism. In contrast, local treatment with alphaIL-6R antibodies that also block signaling via the membrane-bound IL-6R (mIL-6R) led …

The Soluble Interleukin-6 Receptora

Gene Therapy of Human Melanoma — from Animal Experiments to the Clinical Trial

Low immunogenecity of tumour cells is one of the reason that specific immune response is insufficient to destroy malignant cells. In an attempt to augment weakly immunogenic B78H1 mouse melanoma cell line, these cells were transfected with genomic DNA from a line of human melanoma cells expressing a 96kD melanoma associated antigen (MAA) that is intercellular adhesion molecule 1 (ICAM 1). The transfected cells expressed fivefold higher quantities of the melanoma associated antigen from which the DNA was obtained. Human ICAM 1 expressed by mouse melanoma cells appeared to be highly immunogenic leading to the rejection of the modified mouse melanoma cells. The transfected cells appeared to be…

Pore-forming toxins trigger shedding of receptors for interleukin 6 and lipopolysaccharide.

Cleavage of membrane-associated proteins with the release of biologically active macromolecules is an emerging theme in biology. However, little is known about the nature and regulation of the involved proteases or about the physiological inducers of the shedding process. We here report that rapid and massive shedding of the interleukin 6 receptor (IL-6R) and the lipopolysaccharide receptor (CD14) occurs from primary and transfected cells attacked by two prototypes of pore-forming bacterial toxins, streptolysin O and Escherichia coli hemolysin. Shedding is not induced by an streptolysin O toxin mutant which retains cell binding capacity but lacks pore-forming activity. The toxin-dependent c…

Introduction to stem cell biology in vitro. Threshold to the future.

Abstract: Transplantable hematopoietic cells with multilineage reconstituting ability can be quantitated in suspensions of human or murine cells using similar assay procedures. The incorporation into these assays of stringently defined functional endpoints ensures a high degree of specificity for the cells detected. Application of these assays to stem cell-containing suspensions after they have been stimulated for several days with defined cytokines in vitro, or by a mixture of defined and/or undefined factors in vivo, has shown that net amplifications in these populations can be obtained under both circumstances. Such studies have allowed cytokine conditions that support stem cell self-ren…

Gene Therapy of Human Melanoma. Immunization of Patients with Autologous Tumor Cells Admixed with Allogeneic Melanoma Cells Secreting Interleukin 6 and Soluble Interleukin 6 Receptor. University School of Medical Sciences at GreatPoland Cancer Center, Poznań, Poland

SHEDDING OF THE INTERLEUKIN-6 RECEPTOR: MECHANISMS AND PHYSIOLOGICAL CONSEQUENCES

Reduced virus specific T helper cell induction by autologous dendritic cells in patients with chronic hepatitis B-restoration by exogenous interleukin-12

SUMMARYInsufficient stimulatory capacities of autologous dendritic cells (DC) may contribute in part to impaired T cell stimulation and therefore viral persistence in patients with chronic hepatitis B virus (HBV) infection. In order to characterize the antigen presenting functions of DC from chronic HBV carriers and controls antigen specific T cell responses were analysed. CD34+ peripheral blood progenitor cells were differentiated to immature DC in the presence of GM-CSF, IL-6/IL-6R fusion protein and stem cell factor. Proliferative CD4+ T cell responses and specific cytokine release were analysed in co-cultures of DC pulsed with HBV surface and core antigens or tetanus toxoid and autologo…

More about genetically modified tumour vaccines

Rapid Differentiation of a Rare Subset of Adult Human Lin−CD34−CD38− Cells Stimulated by Multiple Growth Factors In Vitro

Recently, several reports of lineage-negative (lin(-)) CD34(-) cells with in vivo hematopoietic activity have focused interest on the properties and growth factor response characteristics of these cells. We have now identified a combination of 5 growth factors that are necessary and sufficient to stimulate a marked mitogenic and differentiation response by a subset of human lin(-)CD34(-)CD38(-) cells present in normal adult human marrow and granulocyte colony-stimulating factor (G-CSF)-mobilized blood. Less than 0.1% of the cells in highly purified (including doubly sorted) lin(-)CD34(-)CD38(-) cells from these 2 sources formed colonies directly in semisolid medium or generated such cells a…