The regulation of gp130 surface expression on monocytes : implications in chronic inflammation

The glycoprotein 130 (gp130) is the signal transducing element common to all interleukin (IL)-6 type cytokines. Signals mediated by gp130 are involved in several biological processes such as hematopoiesis, cell differentiation and immune reactions. To initiate downstream signaling, a complex of gp130 and the ligand-specific alpha-receptor chain needs to be formed. Pathways activated by gp130-mediated signaling include the Janus kinase/signal transducer and activator of transcription (JAK/STAT) and the Src homology 2 domain containing tyrosine phosphatase/extracellular signal-regulated kinase (SHP2/ERK) cascade. While gp130 is found on most cells, expression of ligand-binding receptor chains is restricted to particular cell types. It has therefore long been assumed that the expression pattern of alpha receptors determines the sensitivity of cells to IL-6 type cytokines. However, depending on the regulatory state of a cell, expression of gp130 may vary and thus alter responsiveness. The importance of strictly regulated gp130 surface expression is further underlined by the fact that the soluble alpha-receptor of IL-6, sIL-6R, can replace the membrane-bound receptor and induce signaling in cells expressing only gp130. In hepatocytes, a rapid downregulation of gp130 surface expression by pro-inflammatory cytokines has been described. This downregulation was mediated by the stress-induced mitogen-activated protein kinase (MAPK) p38, phosphorylation of serine 782 within the cytoplasmic tail of gp130 and its subsequent internalization and degradation. The reduced availability of the receptor on the cell surface restricted IL-6-induced STAT3 activation. Since IL-6 plays an important role in several immune reactions, we asked whether this regulatory mechanism might be of relevance in human monocytes. Different approaches confirmed that gp130 was internalized in response to the pro-inflammatory stimulus IL-1beta. Internalization of gp130 was dependent on p38 and stimulation of monocytes with IL-1beta led to an increased phosphorylation of serine 782. Furthermore, downstream signaling of IL-6 was affected by IL-1beta, which again depended on p38. To study these regulatory effects in vivo, two transgenic mouse lines were generated. Overexpression of gp130 wildtype (wt) or gp130 mutant (mut) was restricted to hematopoietic cells. In gp130 mut mice serine residues in the cytoplasmic domain were mutated to alanine to create non-phosphorylatable residues, which was assumed to prevent internalization and stabilize gp130 surface expression in response to stress stimuli. Two preliminary findings might support our expectations: 1) IL-1beta had a stronger inhibitory effect on gp130 surface expression on cluster of differentiation (CD)11b+ splenocytes from gp130-wt mice compared to gp130-mut animals. 2) In a model of leukocyte infiltration into the peritoneal cavity more cells were recruited in gp130-mut than in gp130-wt mice. Given the known role of IL-6 in migration, this supports the assumption that the altered stability of gp130 surface expression influences cell behavior. A particular role in chronic inflammation is attributed to IL-6. Raised serum levels of IL-6 and sIL-6R have been associated with different chronic inflammatory disorders such as inflammatory bowel disease (IBD) or juvenile idiopathic arthritis (JIA). In this study gp130 surface expression on peripheral monocytes from pediatric IBD patients was shown to be increased compared to healthy controls. This was reflected in the level of downstream signaling. Specifically, monocytes from IBD patients displayed enhanced STAT3 phosphorylation. The clinical and pathophysiologic significance of this differential regulation remained unclear. However, these findings were reproduced in a murine model of chronic colitis, thus presenting a platform for future analyses. Reduced gp130 surface expression was found on synovial fluid (SF) monocytes compared to peripheral blood (PB) monocytes in JIA patients. By incubating healthy peripheral blood mononuclear cells (PBMCs) with SF biochemical abnormalities of SF monocytes were partially reproduced. This included gp130 downregulation, which depended on p38. In summary, in the framework of this thesis it was shown that gp130 surface expression on human monocytes is regulated by the pro-inflammatory cytokine IL-1beta, which impairs IL-6-mediated downstream signaling. This mechanism might be of relevance in inflammatory conditions exemplified by inflamed joints of JIA patients. The herein generated transgenic mice may be of value for future investigations of the role of crosstalk regulation in chronic inflammatory situations