SUMMARYEfforts to identify the signal transduction pathways used by interleukin-10 (IL-10) have resulted in limited success. The anti-inflammatory effects elicited by IL-10, and the mechanisms by which these are mediated, are still relatively unknown. Understanding the signalling mechanisms behind the suppression of cytokine expression by IL-10 could be of potential therapeutic interest. Although the consensus is that the Janus kinase, Jak1, as well as the signal transducer and activator of transcription STAT3 are central, much controversy exists about the participation and roles of many other signalling pathways targeted by IL-10. The mechanisms of cytokine suppression proposed by various groups have included transcriptional, post-transcriptional and post-translational regulation of IL-10 target genes; nevertheless no unifying model has emerged thus far. Here we would like to highlight novel findings and discuss their implications in the context of current understanding of IL-10 signalling.

There is general agreement that signal transducer and activation of transcription 3 (STAT3) is required to mediate the antiinflammatory activities of interleukin (IL)-10. However, STAT3 is activated by multiple factors that do not share the anti-inflammatory activity of IL-10. The question remains whether STAT3 is sufficient for the anti-inflammatory effects or whether there are other signals required, as had been suggested previously. We set out to map the human IL-10 receptor and to identify the key elements involved in transducing the cytokine-suppressive effects of IL-10. We were able to show an absolute requirement for both of the tyrosine residues found within the YXXQ-STAT3-docking site within the IL-10 receptor 1 and that no other signals appeared to be required. We used a constitutively active STAT3 to determine whether expression of this factor could suppress lipopolysaccharide-induced tumor necrosis factor and IL-6 production. Our data show that STAT3 activity can suppress both IL-6 and tumor necrosis factor production in lipopolysaccharide-stimulated macrophages. However, in synovial fibroblasts, STAT3 did not suppress IL-6 production, suggesting that the cellular environment plays an important role in dictating whether STAT3 drives a pro-or anti-inflammatory response.

IL-10-deficient mice exhibit spontaneous enterocolitis and other symptoms akin to Crohn’s disease, indicating that IL-10 might regulate normal physiology in the gut. However, clinical trials with IL-10 in Crohn’s disease were disappointing, although some patients showed healing of intestinal mucosa. This study searched for genetic polymorphisms within the IL-10 pathway. We decided to screen for mutations of the IL-10R1 cDNA in healthy volunteers and Crohn’s disease patients and identified two novel variants: a serine 138-to-glycine (S138G) and a glycine 330-to-arginine (G330R) substitution. The allelic frequency in a European cohort was relatively high (16% for the S138G and 33% for the G330R), and S138G was in strong linkage disequilibrium with G330R. A similar allele frequency was found in a group of Crohn’s patients. In IL-10R1 G330R-expressing monocytes, the inhibitory effect of IL-10 on TNF-α production was diminished, indicating that this variant may be a loss-of-function allele. No such difference was observed between haplotypes 4 (G330R only) and 7 (S138G and G330R). In addition, these IL-10R1 variants had no influence on the IL-10R1 expression density. Structural analysis of the S138G variant revealed that the substitution of S138G may interfere with binding of IL-10 to IL-10R1.

Aims-To establish whether the multinucleate cells in lesions of patients with cherubism are also osteoclasts and if this is the case whether they were responsive to calcitonin; to carry out cytogenetic studies on two members of the same family aVected by cherubism in an attempt to identify any major chromosomal defects; and to perform an in-depth modern biochemical study of four children in the same family. Subjects and methods-Four related children with cherubism were studied. Tissue taken from one of the children at elective decompression of an optic nerve was submitted to in vitro bone resorption studies. Cytogenetic studies were done on two of the children and biochemical studies on all four. Results-The multinucleate cells in the cherubic lesions were shown to be osteoclasts since they synthesised tartrate resistant acid phosphatase, expressed the vitronectin receptor, and resorbed bone. Bone resorption by the cultured multinucleate cells was significantly inhibited by calcitonin. High resolution cytogenetic studies failed to detect any chromosomal abnormalities in two children with cherubism. The biochemistry profile of all four children with cherubism showed that serum calcium, parathyroid hormone, parathyroid related hormone, calcitonin, and alkaline phosphatase were within normal levels. Urine analysis of pyridinium and deoxypyridinium cross links, hydroxyproline, and calcium in relation to urine creatinine were measured to assess bone resorption in these children, and the values were at the upper end of the normal range in all four. Conclusions-Further studies are required to determine whether calcitonin treatment will control this grossly deforming disease until the time when the physiological changes that occur at puberty rectify the pathology. It is not recommended that biochemical markers of bone resorption are used in isolation to monitor the activity of cherubism in individuals because the results are based on a small number of children and because of reports of marked interindividual variation in the levels of these markers, particularly in children. (J Clin Pathol 1998;51:831-837)

Macrophage colony-stimulating factor (M-CSF) is essential for murine osteoclast formation and its role in human hematopoiesis in vitro is not fully defined. Therefore, we have investigated the effect of M-CSF on the formation of human osteoclasts in vitro. M-CSF was found to induce substantial bone resorption and osteoclast formation in a dose-responsive and time-dependent manner above that induced by 1,25 dihydroxyvitamin D3 (1,25 vitamin D3) in cultures of human bone marrow (BM) stromal cells sedimented onto devitalized bone. By day 14 there was a mean of approximately 50% of the surfaces of the bone slices resorbed compared with only 6% in cultures treated with 1,25 vitamin D3 alone. Osteoclasts were identified as 23c6+ cells (an antibody that recognizes the vitronectin receptor), 87.5% of which coexpressed the calcitonin receptor. The number of 23c6+ cells correlated strongly with bone resorption spatially, and in a dose-responsive and time-dependent manner; the correlation coefficient in cultures treated with 1,25 vitamin D3 alone was 0.856 and those treated with both M-CSF and 1,25 vitamin D3 was 0.880. Granulocyte-macrophage colony-stimulating factor, IL-1 beta, IL-3, IL-6, tumor necrosis factor-alpha, transforming growth factor-beta, leukemia inhibitory factor, and IL-11 did not increase bone resorption above that in 1,25 vitamin D3-treated cultures. We also found that 1,25 vitamin D3 increased, to a minor but significant degree, both bone resorption and the concentration of M-CSF in the culture supernatants above that in vehicle-treated cultures, indicating that M-CSF is present in our BM cultures, but that there is insufficient to induce substantial osteoclast formation. These results define a critical role for M-CSF in the formation of human osteoclasts.