PubMed Health⌕ Search

Biomedical subjects

S Rose-John

Publications and source records attributed to S Rose-John.

At least 19 recordsLinked to original sources
PubMed2006

STAT3 activation via interleukin 6 trans-signalling contributes to ileitis in SAMP1/Yit mice.

BACKGROUND AND AIMS: SAMP1/Yit mice spontaneously develops intestinal inflammation. Previously, we demonstrated that the signal transducer and activator of transcription (STAT)-3/suppressor of cytokine signalling (SOCS)-3 pathway is pivotal in human inflammatory bowel disease. In our studies in SAMP1/Yit mice, the aim was to investigate whether STAT3 activation contributes to ileitis and to examine the therapeutic effects of this signal blockade. METHODS: Intestinal expression of phospho-STAT3 in SAMP1/Yit mice and control AKR/J mice was examined by western blotting and immunohistochemistry. SOCS3 and interleukin 6 (IL-6) mRNA were determined by northern blotting and reverse transcription-polymerase chain reaction, respectively. We also examined the effects of intravenously injected hyper-IL-6, an IL-6/soluble IL-6 receptor fusion protein, and of soluble gp130-Fc, a specific inhibitor of soluble IL-6 receptor signalling, on STAT3 phosphorylation and disease severity in SAMP1/Yit mice. RESULTS: Phospho-STAT3 was expressed strongly during the disease course in SAMP1/Yit mice but only transiently in AKR/J mice. Phospho-STAT3 was localised to epithelial and mononuclear cells in the diseased intestine of SAMP1/Yit mice. SOCS3 as well as IL-6 mRNAs were expressed in affected intestine. Administration of hyper-IL-6 caused disease exacerbation and enhancement of STAT3 phosphorylation. In contrast, soluble gp130-Fc administration ameliorated the disease and suppressed STAT3 phosphorylation. CONCLUSION: STAT3 signalling is critical in the development of intestinal inflammation in SAMP1/Yit mice. Blockade of this signalling pathway by soluble gp130-Fc may have therapeutic effects in inflammatory bowel disease.

Animals↗
PubMed2006

Interleukin-6 trans-signalling in chronic inflammation and cancer.

Interleukin-6 (IL-6) is a cytokine, which plays an important role in many chronic inflammatory diseases. IL-6 belongs to a family of 10 cytokines, which all act via receptor complexes containing the cytokine receptor subunit gp130. On cells, IL-6 first binds to a specific membrane-bound IL-6R and the complex of IL-6 and IL-6R interacts with gp130 leading to signal initiation. Whereas gp130 is widely expressed throughout the body, the IL-6R is only found on some cells including hepatocytes and some leucocytes. A soluble form of the IL-6R is an agonist capable of transmitting signals through interaction with the gp130 protein. In vivo, the IL-6/soluble IL-6R complex stimulates several types of target cells, which are unresponsive to IL-6 alone, as they do not express the membrane-bound IL-6R. We have named this process trans-signalling. We provided evidence that a soluble form of the IL-6 family signalling receptor subunit gp130 is the natural inhibitor of IL-6 trans-signalling responses. We showed that in chronic inflammatory diseases such as inflammatory bowel disease, peritonitis, rheumatoid arthritis, asthma as well as in colon cancer, IL-6 trans-signalling is critically involved in the maintenance of the disease state. Moreover, in all these animal models, the progression of the disease can be interrupted by specifically interfering with IL-6 trans-signalling using recombinant-soluble gp130Fc protein. The pathophysiologic mechanisms by which the IL-6/soluble IL-6R complex perpetuates the inflammatory state are discussed.

Animals↗
PubMed2006

Designer cytokines for human haematopoietic progenitor cell expansion: impact for tissue regeneration.

Haematopoietic stem cells after bone marrow transplantation can expand 100-fold and thereby completely reconstitute the haematopoietic system. Such an expansion has so far not been achieved in vitro using cytokines and growth factors. An interesting parallel exists between haematopoietic stem cells and mouse embryonic stem cells. In both cell types, massive gp130 stimulation results in an inhibition of cellular differentiation. For both cell types, several intrinsic transcription factors are known which, when overexpressed, lead to massive self-renewal of the cells. It is currently unknown how in haematopoietic stem cells these intrinsic transcription factors, which include HOXB4 and Bmi-1, can be stimulated by extrinsic signals. Once such signals have been identified, they will be combined with the hyperstimulation of gp130 using designer cytokines. Additional strategies include a cell-permeable version of the HOXB4 protein and members of the WNT family ofligands. These strategies might eventually lead to an expansion of haematopoietic stem cells to a similar extent as that observed after bone marrow transplantation. Should haematopoietic stem cell expansion prove feasible and safe, there is great potential for the treatment of leukaemic disorders but also for the treatment of less severe diseases and non-malignant genetic disorders.

Animals↗
PubMed2005

Fast modulation of heat-activated ionic current by proinflammatory interleukin 6 in rat sensory neurons.

The pro-inflammatory cytokine interleukin-6 (IL-6) together with its soluble receptor (sIL-6R) induces and maintains thermal hyperalgesia. It facilitates the heat-induced release of calcitonin gene-related peptide from rat cutaneous nociceptors in vivo and in vitro. Here we report that exposure of nociceptive neurons to the IL-6-sIL-6R complex or the gp130-stimulating designer IL-6-sIL-6R fusion protein Hyper-IL-6 (HIL-6) resulted in a potentiation of heat-activated inward currents (I(heat)) and a shift of activation thresholds towards lower temperatures without affecting intracellular calcium levels. The Janus tyrosine kinase inhibitor AG490, the selective protein kinase C (PKC) inhibitor, bisindolylmaleimide 1 (BIM1), as well as rottlerin, a selective blocker of the PKCdelta isoform, but not the cyclooxygenase inhibitor indomethacin, effectively reduced the effect. Reverse transcription-polymerase chain reaction (RT-PCR) and in situ hybridization revealed expression of mRNA for the signal-transducing beta subunit of the receptor gp130 in neuronal somata, rather than satellite cells in rat dorsal root ganglia. Together, the results suggest that IL-6-sIL-6R acts directly on sensory neurons. It increases their susceptibility to noxious heat via the gp130/Jak/PKCdelta signalling pathway.

Acetophenones↗
PubMed2005

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 cells regulated STAT-3 activation in tumor cells via IL-6. IL-6 signaling required tumor cell derived soluble IL-6R rather than membrane bound IL-6R and suppression of such TGF-beta-dependent IL-6 trans-signaling prevented tumor progression in vivo. Similar to these observations in mice, here we show that human colon cancer tissue expressed only low amounts of membrane bound IL-6R. In contrast, expression and activity of the matrix metalloproteinase TACE were increased. In summary, our data provide novel insights into the role of TGF-beta signaling in colorectal cancer and suggest novel therapeutic approaches for colorectal cancer based on an inhibition of TGF-beta-dependent IL-6 trans-signaling.

ADAM Proteins↗
PubMed2002

Murine stromal cells producing hyper-interleukin-6 and Flt3 ligand support expansion of early human hematopoietic progenitor cells without need of exogenous growth factors.

Expansion of primitive hematopoietic progenitor cells (HPC) is a major challenge in stem cell biology. Stimulation by growth factors (GF) is essential for proliferation of HPC, while the role of stromal cell coculture for maintenance of progenitor/stem cell potential is unclear. We evaluated the potential of a murine stromal cell layer providing hematopoietic GF to support expansion of human CD34(+) cells. Murine MS-5 cells were transfected with the cDNA encoding huFlt3 ligand and the interleukin6/sinterleukin-6R fusion protein hyper-IL-6. Expansion of CFC and week6 CAFC was at least as efficient in transfected clones compared to control cocultures supported with exogenous GF. Cell numbers reached 17.5- to 62.3- (day 14) and 17.4- to 92.4-fold (day 21) of input cells. Expansion of CFU-GM/Mix was 4.0- to 12.8-fold (day 14) and 4.9- to 11.7-fold (day 21). Primitive week6 CAFC were expanded up to 6.5-fold (day 14) and 6.2-fold (day 21) without exogenous GF. When direct contact of HPC and stromal cells was inhibited, a loss of CFC and much more of CAFC potential was observed with unaffected overall cell proliferation. Here, we show the generation of GF producing murine stromal cells which efficiently support early hematopoiesis without exogenous GF. Direct stromal cell-HPC contact is advantageous for maintenance of differentiation potential.

Animals↗
PubMed2002

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

Insufficient 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 autologous CD4+ T cells. Cultured under identical conditions DC from chronic HBV carriers, individuals with acute resolved hepatitis B and healthy controls expressed similar phenotypical markers but chronic HBV carriers showed less frequent and weaker HBV antigen specific proliferative T helper cell responses and secreted less interferon-gamma while responses to the tetanus toxoid control antigen was not affected. Preincubation with recombinant IL-12 enhanced the HBV specific immune reactivities in chronic HBV patients and controls. In conclusion, the weak antiviral immune responses observed in chronic hepatitis B may result in part from insufficient T cell stimulating capacities of DC. Immunostimulation by IL-12 restored the HBV antigen specific T cell responses and could have some therapeutical benefit to overcome viral persistence.

Antigen Presentation↗
PubMed2001

In vitro reconstitution of recognition and activation complexes between interleukin-6 and gp130.

Gp130 is a shared signal-transducing receptor for a family of four-helix cytokines, of which interleukin-6 is a prototypic member. IL-6-type cytokines activate gp130 to elicit downstream intracellular JAK/STAT signaling cascades through formation of hetero-oligomeric receptor complexes. Interleukin-6 must first complex with its specific alpha-receptor (Ralpha) in order to bind and activate gp130. We have dissected the extracellular activation pathway of human gp130 by human IL-6 through reconstitution of soluble complexes representing intermediate and final states in the hierarchical assembly of the IL-6/IL-6Ralpha/gp130 signaling complex. To isolate these hetero-complexes, we have applied a protein engineering strategy of covalently linking IL-6 to its Ralpha, which results in a "hyperactive" single-chain complex (hyper-IL-6) which we express in both Escherichia coli and insect cells. We have determined that IL-6/IL-Ralpha and the cytokine-binding homology region (CHR) of gp130 (D2D3) form a stable trimolecular "recognition" complex (trimer) consisting of 1IL-6,1 IL-6Ralpha, and 1 gp130-CHR. Addition of the N-terminal (D1) Ig-like domain (IGD) of gp130 to the CHR results in a transition to a hexameric "activation" complex containing 2 IL-6, 2IL-6Ralpha, and 2 gp130. These results clearly demonstrate that the recognition and activation complexes are disparate hetero-oligomeric molecular species linked by the recruitment of the gp130 IGD by the unique site III epitope present on all gp130-class cytokines. The results of these studies are relevant to other members of the IL-6 family of gp130-cytokines and address a longstanding question concerning the respective roles of the gp130 CHR and IGD in assembly of the active signaling oligomer.

Antigens, CD↗
PubMed2001

Structure of an extracellular gp130 cytokine receptor signaling complex.

The activation of gp130, a shared signal-transducing receptor for a family of cytokines, is initiated by recognition of ligand followed by oligomerization into a higher order signaling complex. Kaposi's sarcoma-associated herpesvirus encodes a functional homolog of human interleukin-6 (IL-6) that activates human gp130. In the 2.4 angstrom crystal structure of the extracellular signaling assembly between viral IL-6 and human gp130, two complexes are cross-linked into a tetramer through direct interactions between the immunoglobulin domain of gp130 and site III of viral IL-6, which is necessary for receptor activation. Unlike human IL-6 (which uses many hydrophilic residues), the viral cytokine largely uses hydrophobic amino acids to contact gp130, which enhances the complementarity of the viral IL-6-gp130 binding interfaces. The cross-reactivity of gp130 is apparently due to a chemical plasticity evident in the amphipathic gp130 cytokine-binding sites.

Antigens, CD↗
PubMed2001

Up- and down-regulation of granulocyte/macrophage-colony stimulating factor activity in murine skin increase susceptibility to skin carcinogenesis by independent mechanisms.

The role of granulocyte-macrophage colony-stimulating factor (GM-CSF) in tumorigenesis is complex. On the one hand, GM-CSF can promote tumor cell growth, survival, and even metastasis. On the other hand, it can stimulate tumor cell rejection. In skin, it is early expressed after topic application of tumor-promoting agents and therefore may be responsible for changes that correlate with skin tumor promotion (e.g., epidermal hyperproliferation and inflammation). To analyze GM-CSF function in skin tumorigenesis, we generated transgenic mice epidermally overexpressing either GM-CSF or a GM-CSF antagonist. Both types of transgenic mice exhibited significantly increased numbers of benign tumors in a two-step skin carcinogenesis experiment using 7',12'-dimethylbenz[a]anthracene (DMBA) as initiator and 12-O-tetradecanoylphorbol-CSF displayed a significantly elevated carcinoma burden following a single-step carcinogenesis protocol consisting of tumor initiation only. Therefore, endogenous promotion is responsible for elevated tumor development in GM-CSF-overexpressing mice. In antagonist transgenic animals, an increased tumorigenicity of modified B16 tumor cells after cutaneous transplantation as compared with nontransgenic or GM-CSF transgenic mice was observed. Thus, the antitumor activity leading to the repression of tumor cell growth in control mice is GM-CSF dependent and is compromised in mice expressing the antagonist. We suggest that both, up-regulation and down-regulation of GM-CSF activity in skin, increase the incidence and growth of tumors via two independent mechanisms: endogenous tumor promotion in the case of increased GM-CSF activity and compromised tumor cell rejection in the case of decreased GM-CSF activity.

9,10-Dimethyl-1,2-benzanthracene↗
PubMed2001

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) showed morphological alterations in combination with a dramatic growth inhibition in vitro. Subcutaneous injection in C57BL/6 mice resulted in an almost complete rejection of B16/H-IL6 cells. This effect was partially abolished in FVB/BL/6 mice transgenic for a GM-CSF receptor antagonist, indicating a GM-CSF-dependent rejection of H-IL-6 transfected B16 cells. These results demonstrate that the anti-tumor effect of cytokines like IL-6 which are secreted by transfected melanoma cells at least in part depends on GM-CSF activity.

Animals↗
PubMed2001

Distinct role of gp130 activation in promoting self-renewal divisions by mitogenically stimulated murine hematopoietic stem cells.

Previous studies have demonstrated hematopoietic stem cell amplification in vitro after the activation of three cell-surface receptors: flt3/flk2, c-kit, and gp130. We now show flt3-ligand and Steel factor alone will stimulate >85% of c-kit(+)Sca-1(+)lin(-) adult mouse bone marrow cells to proliferate in single-cell serum-free cultures, but concomitant retention of their stem cell activity requires additional exposure to a ligand that will activate gp130. Moreover, this response is restricted to a narrow range of gp130-activating ligand concentrations, above and below which hematopoietic stem cell activity is lost. These findings indicate a unique contribution of gp130 signaling to the maintenance of hematopoietic stem cell function when these cells are stimulated to divide with additional differential effects dictated by the intensity of gp130 activation.

Animals↗
PubMed2001

Recognition sequences and structural elements contribute to shedding susceptibility of membrane proteins.

Although regulated ectodomain shedding affects a large panel of structurally and functionally unrelated proteins, little is known about the mechanisms controlling this process. Despite a lack of sequence similarities around cleavage sites, most proteins are shed in response to the stimulation of protein kinase C by phorbol esters. The signal-transducing receptor subunit gp130 is not a substrate of the regulated shedding machinery. We generated several chimaeric proteins of gp130 and the proteins tumour necrosis factor alpha (TNF-alpha), transforming growth factor alpha (TGF-alpha) and interleukin 6 receptor (IL-6R), which are known to be subject to shedding. By exchanging small peptide sequences of gp130 for cleavage-site peptides of TNF-alpha, TGF-alpha and IL-6R we showed that these short sequences conferred susceptibility to spontaneous and phorbol-ester-induced shedding of gp130. Importantly, these chimaeric gp130 proteins were functional, as shown by the phosphorylation of gp130 and the activation of signal transduction and activators of transcription 3 ('STAT3') on stimulation with cytokine. To investigate minimal requirements for shedding, truncated cleavage-site peptides of IL-6R were inserted into gp130. The resulting chimaeras were susceptible to shedding and showed the same cleavage pattern as observed in the chimaeras containing the complete IL-6R cleavage site. Surprisingly, we could also generate cleavable chimaeras by exchanging the juxtamembrane sequence of gp130 for the corresponding region of leukaemia inhibitory factor ('LIF') receptor, a protein that like gp130 is not subject to regulated or spontaneous shedding. Thus it seems that there is no minimal consensus shedding sequence. We speculate that structural changes allow the access of the protease to a membrane-proximal region, leading to shedding of the protein.

Amino Acid Sequence↗
PubMed2001

Induction of strong hepatitis B virus (HBV) specific T helper cell and cytotoxic T lymphocyte responses by therapeutic vaccination in the trimera mouse model of chronic HBV infection.

Humanized BALB/c mice (termed trimera mice) conditioned by lethal total body irradiation and bone marrow transplantation from SCID mice have been described to support rapid engraftment of human peripheral blood mononuclear cells (PBMC) and the induction of strong B and T cell responses after immunization in vivo. Moreover, these mice can be infected with the hepatitis B and C viruses (HBV, HCV). The current study employed this model to study therapeutic vaccination approaches against the HBV. Thus, strong primary Th cell responses against the HBV core (HBc) and the Borrelia burgdorferi control antigen were induced by transfer of antigen-loaded dendritic cells together with autologous PBMC from HBV-naive donors as well as by vaccination with high doses of antigen or a DNA plasmid encoding for HBcAg. Moreover, primary peptide-specific CTL responses against the immunodominant epitope HBc(18 - 27) were induced by HBc particle or DNA vaccination of chimera engrafted with HBV-naive PBMC. Finally, strong HBc-specific Th cell and antibody responses were induced by HBc or DNA vaccination of mice reconstituted with PBMC from a chronic HBV patient. Thus, since HBc represents the immunodominant antigen in self-limited HBV infection, HBc particles or DNA vectors are good candidates for therapeutic vaccination, that will be further studied in our model and clinical studies.

Animals↗
PubMed2001

A novel and rapid prediction assay for the effectiveness of IL-6 receptor specific antisense oligonucleotides by proliferation inhibition of an interleukin-6 dependent cell line.

Interleukin-6 (IL-6) belongs to a family of cytokines that use receptors consisting of a common signal-transducing chain (gp130). Baf/3 cells transfected with the human IL-6 receptor (IL-6R) and gp130 (Baf/3-gp130/IL-6R) can only grow in medium containing IL-6. We attempted to interrupt the signal transducing pathway of IL-6 with the help of antisense oligonucleotides (ASOs) designed against the IL-6R. We used 18 different kinds of antisense oligonucleotides of overlapping sequences around the translational start codon of the human IL-6R. Sense ASOs were used as a control. The proliferation of cells was analysed by H-thymidine incorporation. Cell surface expression of the IL-6R was assessed by FACS analysis. We identified three ASOs which strongly inhibited the proliferation of IL-6 dependent transfected Baf/3 cells. Flow cytometric studies on the suppression of surface expression of IL-6R by ASOs showed a similar pattern. These results should help to clarify the structural requirements of functionally effective ASOs in the inhibition of IL-6R.

Base Sequence↗
PubMed2001

Hyper-IL-6 gene therapy reverses fulminant hepatic failure.

Fulminant hepatic failure is a catastrophic condition caused by massive hepatocellular apoptosis and necrosis. Inhibition of hepatocyte apoptosis and the enhancement of the endogenous potential for liver regeneration could potentially form an effective basis for treatment of this condition. In response to injury in the liver, IL-6 mediates the acute-phase response and induces both cytoprotective and mitogenic functions. Hyper-IL-6 is a superagonistic designer cytokine consisting of human IL-6 linked by a flexible peptide chain to the secreted form of the IL-6 receptor. In a mouse model of acute liver failure induced by d-galactosamine administration, a single low dose of a hyper-IL-6-encoding adenoviral vector, in contrast to an adeno-IL-6 vector, maintained liver function, prevented the progression of liver necrosis, and induced liver regeneration, leading to dramatically enhanced survival. Thus, hyper-IL-6 gene therapy may be useful for the treatment of fulminant hepatic failure, which is often fatal even following treatment by transplantation.

Adenoviridae↗