The soluble interleukin-6 (IL-6) receptor/IL-6 fusion protein enhances in vitro maintenance and proliferation of human CD34(+)CD38(-/low) cells capable of repopulating severe combined immunodeficiency mice.
(17/863)
In vitro maintenance and proliferation of human hematopoietic stem cells is crucial for many clinical applications. Early hematopoietic cells express low levels of FLT-3 and c-kit receptors, as well as the interleukin-6 (IL-6) receptor signal transducing element, gp130, but do not express IL-6 receptor itself. Therefore, we have attempted to maintain human cord blood or bone marrow CD34(+) cells ex vivo in serum-free cultures containing stem cell factor (SCF) and FLT-3 ligand (FL) alone or together with a new recombinant molecule of soluble IL-6 receptor fused to IL-6 (IL6RIL6 chimera). The effect of IL6RIL6 chimera on the proliferation and differentiation of CD34(+) cells was compared with that of each chimera component added separately. The engraftment potential of in vitro-cultured cells was determined using our recently established functional in vivo assay for primitive human severe combined immunodeficiency (SCID)-repopulating cells (SRC). We report here that IL6RIL6 chimera induced significantly higher levels of progenitors and SRC compared with SCF + FL alone or together with IL-6 and soluble IL-6 receptor. IL6RIL6 chimera prolonged in vitro maintenance of SRC for up to 14 days. Stimulation of CD34(+)CD38(-/low) enriched cells with IL6RIL6 chimera maintained the early CD34(+)CD38(-/low) cell subpopulation, which could be detected in vitro for up to 14 days. Moreover, IL6RIL6 chimera preferentially stimulated the growth of early CD34(+)38(-/low) cells, resulting in significantly higher levels of progenitors compared with more mature CD34(+)38(+) cells. Taken together, these findings demonstrate the importance of IL6RIL6 chimera in stimulating the proliferation of early CD34(+). CD38(-)gp130(+)IL-6R(-) cells in vitro and extended maintenance of progenitors and SRC. (+info)
Enhanced production of interleukin-6 (IL-6), oncostatin M and soluble IL-6 receptor by cultured peripheral blood mononuclear cells from patients with systemic sclerosis.
(18/863)
OBJECTIVES: To determine whether the spontaneous production of interleukin 6 (IL-6), oncostatin M (OSM), soluble IL-6 receptor (sIL-6R) and soluble gp130 (sgp130) from peripheral blood mononuclear cells (PBMC) is increased in patients with systemic sclerosis (SSc). METHODS: The culture supernatants of PBMC from patients with SSc (n = 33) and healthy controls (n = 20) were examined by enzyme-linked immunosorbent assay. RESULTS: The production levels of IL-6, OSM and sIL-6R were significantly higher in patients with SSc than in controls. However, sgp130 levels in supernatants from patients with SSc were not significantly elevated when compared with those from controls. Soluble IL-6R levels correlated significantly with the severity of pulmonary fibrosis in patients with SSc. CONCLUSIONS: The enhanced production of IL-6, OSM and sIL-6R from PBMC may cooperatively contribute to the disease process in SSc. In particular, enhanced sIL-6R production from PBMC may be related to the development of pulmonary fibrosis via enhancement of IL-6 signal transduction in SSc, since sIL-6R can act as an agonist of IL-6. (+info)
The influence of age and gender on serum dehydroepiandrosterone sulphate (DHEA-S), IL-6, IL-6 soluble receptor (IL-6 sR) and transforming growth factor beta 1 (TGF-beta1) levels in normal healthy blood donors.
(19/863)
Dysregulation of IL-6 synthesis is thought to play a role in the development of a number of age-related conditions, such as rheumatoid arthritis, osteoporosis, atherosclerosis, Alzheimer's disease and B cell malignancies. Recently it has been suggested that the production of IL-6 is influenced by the adrenal hormone dehydroepiandrosterone (DHEA) and its sulphated derivative DHEA-S. In humans we investigated the relationship between DHEA-S, IL-6, IL-6 sR and TGF-beta1 in the serum of normal healthy male and female blood donors. Using immunoassay techniques we found that the serum levels of DHEA-S significantly (P = 0.0001) decreased with age in both males and females. Furthermore, mean DHEA-S levels in all age groups were significantly (P = 0.0001) higher in males. Such correlations were not apparent for IL-6 using a standard assay, but a high sensitivity assay revealed that serum IL-6 was significantly (P = 0.0018) positively correlated with age in males only. In addition, serum levels of DHEA-S were significantly (P = 0.048) negatively correlated with serum IL-6, again in male subjects only. In contrast, serum IL-6 sR and TGF-beta1 levels were not correlated with age in either males or females and were not significantly different between the sexes. However, a significant (P = 0.024) negative correlation between DHEA-S and IL-6 sR was found in males. These studies clearly highlight the complex nature of the relationship between these molecules in the ageing process in normal healthy blood donors and demonstrate the need to use high sensitivity assays when measuring IL-6 in apparently healthy individuals under the age of 70 years. (+info)
Induction of myelin gene expression in Schwann cell cultures by an interleukin-6 receptor-interleukin-6 chimera.
(20/863)
Expression of myelin basic protein (MBP) and Po gene products is induced during the final postnatal maturation of Schwann cells and reinduced during nerve regeneration. We show that a chimeric protein containing interleukin-6 fused to its soluble receptor (IL6RIL6 chimera) induces MBP and Po RNAs and proteins in cultures of dorsal root ganglia (DRG) from 14 day old mouse embryos. Activation of gp130 signaling by IL6RIL6 appears comparable to cyclic AMP elevating agents to induce the myelin gene products in DRG and in pure Schwann cell cultures. (+info)
Human herpesvirus 8 interleukin-6 (vIL-6) signals through gp130 but has structural and receptor-binding properties distinct from those of human IL-6.
(21/863)
Human herpesvirus 8 (HHV-8) has been associated with classical, endemic (African), and AIDS-related Kaposi's sarcoma (KS), body cavity-based primary effusion lymphomas, and multicentric Castleman's disease (MCD). HHV-8 encodes a functional homologue of interleukin-6 (IL-6), a cytokine that promotes the growth of KS and myeloma cells and is found at elevated levels in MCD lesions and patient sera. We have previously reported that the viral IL-6 (vIL-6) gene product can support the growth of the IL-6-dependent murine hybridoma cell line, B9, and that the gp80 (IL-6 receptor [IL-6R]) component of the IL-6 receptor-signal transducer (gp180) complex plays a role in mediating this activity. However, it has been shown by others that vIL-6 can function in human cells independently of IL-6R. Here we have extended our functional studies of vIL-6 by identifying transcription factors and pathways used in human Hep3B cells, investigating the utilization of gp130 and IL-6R by vIL-6, and undertaking mutational analyses of vIL-6 and gp130. The data presented here establish that vIL-6, in common with its endogenous counterparts, can mediate signal transduction through gp130 and activate multiple transcription factors, map residues within the vIL-6 protein that are and are not important for vIL-6 signalling, and identify a gp130 mutant that is nonfunctional with respect to vIL-6 signalling in the absence of IL-6R but that retains the ability to mediate vIL-6 and human IL-6 (hIL-6) signal transduction when IL-6R is coexpressed. The data presented demonstrate functional and mechanistic similarities between vIL-6 and endogenous IL-6 proteins but also highlight differences in the structural and receptor-binding properties of vIL-6 relative to its human counterpart. (+info)
Novel path to activation of vascular smooth muscle cells: up-regulation of gp130 creates an autocrine activation loop by IL-6 and its soluble receptor.
(22/863)
This study describes a novel path to the activation of smooth muscle cells (SMC) by the IL-6/soluble IL-6 receptor (sIL-6R) system. Human vascular SMC constitutively express only scant amounts of IL-6R and so do not respond to stimulation with this cytokine. We show that SMC also do not constitutively express appreciable levels of gp130, which would render them sensitive to transsignaling by the IL-6/sIL-6R complex. Because gp130 is generally believed not to be subject to regulation, SMC would thus appear not to qualify as targets for the IL-6/sIL-6R system. However, we report that treatment of SMC with IL-6/sIL-6R provokes marked up-regulation of gp130 mRNA and surface protein expression. This is accompanied by secretion of IL-6 by the cells, so that an autocrine stimulation loop is created. In the wake of this self-sustaining system, there is a selective induction and secretion of MCP-1, up-regulation of ICAM-1, and marked cell proliferation. The study identifies SMC as the first example of cells in which gp130 expression is subject to substantive up-regulation, and discovers a novel amplification loop involving IL-6 and its soluble receptor that drives SMC into a proinflammatory state. (+info)
Whole blood production of monocytic cytokines (IL-1beta, IL-6, TNF-alpha, sIL-6R, IL-1Ra) in haemodialysed patients.
(23/863)
BACKGROUND: The production of monocytic cytokines by isolated mononuclear cells after stimulation by phytohaemagglutinin (PHA) and lipopolysaccharide (LPS) is generally increased in haemodialysed (HD) patients. We performed whole blood (WB) cultures to evaluate cytokine production by blood cells inside their complex cellular and humoral network. METHODS: Diluted whole blood from HD patients (collected before dialysis) and controls was cultured alone with PHA (2.5 microg/ml) or LPS (1 and 3 microg/ml). Supernatants were collected after 24 and 48 h of culture, and concentrations of IL-1 beta, IL-6, TNF-alpha, sIL-6R and IL-1Ra were determined by ELISA. RESULTS: The low spontaneous production of IL-1beta, IL-6 and TNF-alpha in both patients and controls was not significantly modified by PHA. The lower dose of LPS (1 microg/ml) induced a significant but lower increase in production of IL-1beta, IL-6 and TNF-alpha in patients than in controls. In contrast, while it did not further increase their production in controls, the higher concentration of LPS (3 microg/ml) still increased their production in patients to the same level than in controls. The plasma concentrations of sIL-6R were higher in patients than in controls. In both groups, the sIL-6R concentration did not vary during the culture period whether the cells were stimulated or not with LPS or PHA. This suggests that the increased plasma levels of sIL-6R were not produced by blood cells. Despite a similar significant LPS and PHA induced production of IL-1Ra, the IL-1Ra/IL-1beta ratio was always higher in patients than in controls. CONCLUSION: Monocytes from HD patients in WB cultures are hyporesponsive to PHA and LPS for their IL-1beta, TNFalpha and IL-6 production in contrast to isolated monocytes that demonstrate signs of activation. If it reflects the in vivo situation it could partly explain the immune defect in uraemic and haemodialysed patients. Higher sIL-6R/IL-6 and IL-1Ra/IL-1beta ratios could also participate to the complex immune disturbances of HD patients by reducing the biological activity of two cytokines playing a major role in the immune and inflammatory network. (+info)
Myeloma cells release soluble interleukin-6Ralpha in relation to disease progression by two distinct mechanisms: alternative splicing and proteolytic cleavage.
(24/863)
Multiple myeloma (MM) is a plasma-cell malignancy characterized by the accumulation of malignant plasma cells within the bone marrow. Interleukin (IL)-6 is an essential survival and growth factor for myeloma cells that exerts its activity through a cell surface receptor composed of an 80-kDa ligand binding molecule (IL-6Ralpha) and a 130-kDa signal-transducing molecule. Of major interest, the soluble form of the IL-6Ralpha (sIL-6Ralpha) is an agonistic molecule able to potentiate IL-6 activity and a strong prognostic factor in MM. In the present study, we demonstrate that purified myeloma cells from all of the patients with MM and human myeloma cell lines release sIL-6Ralpha. The level of sIL-6Ralpha release correlates with disease activity and is clearly up-regulated during tumoral expansion in vivo and immortalization in vitro. Of note, this sIL-6Ralpha release is strongly reduced (50%) by a hydroxamate-based metalloproteinase inhibitor underlying the importance of shedding in the production of sIL-6Ralpha by myeloma cells. Using specific IL-6Ralpha primers flanking the transmembrane domain, we demonstrate by PCR the presence of two IL-6R mRNAs corresponding to the membrane IL-6Ralpha and to the sIL-6Ralpha generated through alternative splicing in myeloma cells. In conclusion, we show that: (a) native myeloma cells and human myeloma cell lines release sIL-6Ralpha by two distinct mechanisms: alternative splicing and proteolytic cleavage of the membrane IL-6Ralpha; and (b) the release of the sIL-6Ralpha, which is an agonist of IL-6, correlates with disease progression, explaining in part its strong prognostic value in vivo. (+info)