Patterning Chronic Active Demyelination in Slowly Expanding/Evolving White Matter MS Lesions.

BACKGROUND AND PURPOSE: Slowly expanding/evolving lesions measured by conventional T1-weighted/T2-weighted brain MR imaging may contribute to progressive disability accumulation in MS. We evaluated the longitudinal change in myelin and axonal tissue integrity in white matter slowly expanding/evolving lesions by means of the magnetization transfer ratio and DTI radial diffusivity. MATERIALS AND METHODS: Slowly expanding/evolving lesions were detected within the Study to Assess the Efficacy, Safety, Tolerability, and Pharmacokinetics of BIIB033 in Participants With Relapsing Forms of Multiple Sclerosis When Used Concurrently With Avonex (SYNERGY) Phase 2 clinical trial dataset (NCT01864148), comprising patients with relapsing-remitting and secondary-progressive MS (n = 299) with T1-weighted/T2-weighted MR imaging at all trial time points (baseline to week 72). RESULTS: Compared with non-slowly expanding/evolving lesions (areas not classified as slowly expanding/evolving lesion) of baseline nonenhancing T2 lesions, slowly expanding/evolving lesions had a lower normalized magnetization transfer ratio and greater DTI radial diffusivity, both in patients with relapsing-remitting MS (n = 242) and secondary-progressive MS (n = 57, P < .001 for all). Although the changes with time in both the normalized magnetization transfer ratio and DTI radial diffusivity between slowly expanding/evolving lesions and non-slowly expanding/evolving lesions were positively correlated (P < .001), a decrease in the normalized magnetization transfer ratio and a greater increase in DTI radial diffusivity were observed in slowly expanding/evolving lesions versus non-slowly expanding/evolving lesions from baseline to week 72 in relapsing-remitting MS and secondary-progressive MS (P < .001 for all). CONCLUSIONS: Patterns of longitudinal change in the normalized magnetization transfer ratio and DTI radial diffusivity in slowly expanding/evolving lesions were consistent with progressive demyelination and tissue loss, as seen in smoldering white matter MS plaques.

Treatment with denosumab reduces the incidence of new vertebral and hip fractures in postmenopausal women at high risk.

CONTEXT: The FREEDOM (Fracture REduction Evaluation of Denosumab in Osteoporosis every 6 Months) trial showed denosumab significantly reduced the risk of fractures in postmenopausal women with osteoporosis. OBJECTIVE: We evaluated the effect of denosumab on the incidence of new vertebral and hip fractures in subgroups of women at higher risk for these fractures. DESIGN: FREEDOM was a 3-yr, randomized, double-blind, placebo-controlled, phase 3 trial. PARTICIPANTS AND SETTING: Postmenopausal women (N = 7808) with osteoporosis were enrolled at 213 study sites worldwide. INTERVENTIONS: Subjects received s.c. denosumab (60 mg) or placebo every 6 months and daily supplements of calcium (≥1000 mg) and vitamin D (≥400 IU). MAIN OUTCOME MEASURES: This post hoc analysis evaluated fracture incidence in women with known risk factors for fractures including multiple and/or moderate or severe prevalent vertebral fractures, aged 75 yr or older, and/or femoral neck bone mineral density T-score of -2.5 or less. RESULTS: Compared with placebo, denosumab significantly reduced the risk of new vertebral fractures in women with multiple and/or severe prevalent vertebral fractures (16.6% placebo vs. 7.5% denosumab; P < 0.001). Similarly, denosumab significantly reduced the risk of hip fractures in subjects aged 75 yr or older (2.3% placebo vs. 0.9% denosumab; P < 0.01) or with a baseline femoral neck bone mineral density T-score of -2.5 or less (2.8% placebo vs. 1.4% denosumab; P = 0.02). These risk reductions in higher-risk individuals were consistent with those seen in patients at lower risk of fracture. CONCLUSIONS: Denosumab reduced the incidence of new vertebral and hip fractures in postmenopausal women with osteoporosis at higher risk for fracture. These results highlight the consistent antifracture efficacy of denosumab in patients with varying degrees of fracture risk.

[Bone quality: from theory to reality]. / La qualité osseuse: de la théorie a lareálité.

Bone quality is an essential element of bone strength. Bone quality refers not only to bone architecture but also to its material properties which are directly dependent on bone remodeling (i.e., turnover). Therefore, maintaining bone architecture but also bone turnover is critical for optimal bone quality and for providing the ability of bone to resist fractures.

Increased expression of receptor activator of NF-kappaB ligand (RANKL), its receptor RANK and its decoy receptor osteoprotegerin in the colon of Crohn's disease patients.

Crohn's disease (CD) is associated with low bone mass due to chronic inflammation and other factors. Receptor activator of NF-kappaB ligand (RANKL), its receptor RANK and its decoy receptor osteoprotegerin (OPG) are potentially involved in this process as they regulate osteoclastogenesis and are influenced by pro-inflammatory cytokines. The aim of this study was to determine the levels of soluble RANKL (sRANKL), RANK and OPG expression both in the serum and in the colon of CD patients. Levels of sRANKL and OPG were assessed in the serum and the supernatants of cultured colonic biopsies in patients with CD and controls by ELISA. RANK expression was explored by immunostaining and immunofluorescence of fixed colonic samples. OPG and sRANKL levels were higher in the serum of CD patients as compared to age- and sex-matched controls. Levels of sRANKL and OPG were significantly enhanced in cultured colonic biopsies from CD, and OPG levels correlated with histological inflammation, and pro- and anti-inflammatory cytokine levels. No significant correlation was found for sRANKL. RANK+ cells were increased in the colon of CD, particularly in inflamed areas. These cells were positive for CD68 or S100 protein. We conclude that serum and local levels of sRANKL and OPG are increased in CD. Moreover, RANK is expressed in the colonic mucosa by subpopulations of activated macrophages or dendritic cells at higher levels in CD compared to normal colon.

Rapid improvement of bone metabolism after infliximab treatment in Crohn's disease.

BACKGROUND: Crohn's disease is associated with low bone mineral density and altered bone metabolism. AIM: To assess the evolution of bone metabolism in Crohn's disease patients treated with infliximab. METHODS: We studied 71 Crohn's disease patients treated for the first time with infliximab for refractory Crohn's disease. Biochemical markers of bone formation (type-I procollagen N-terminal propeptide, bone-specific alkaline phosphatase, osteocalcin) and of bone resorption (C-telopeptide of type-I collagen) were measured in the serum before and 8 weeks after infliximab therapy and compared with values in a matched healthy control group. RESULTS: Eight weeks after treatment with infliximab, a normalization of bone markers was observed with a median increase in formation markers of 14-51% according to marker and a lower but significant decrease in resorption marker (median 11%). A clinically relevant increase in bone formation markers was present in 30-61% of patients according to the marker. A clinically relevant decrease in C-telopeptide of type-I collagen was present in 38% of patients. No association was found with any tested demographic or clinical parameter. CONCLUSION: Infliximab therapy in Crohn's disease may rapidly influence bone metabolism by acting either on bone formation or bone resorption. This improvement seems to be independent of clinical response to infliximab.

[Alteration of bone metabolism in HIV-infected patients treated by HAART]. / Anomalies du métabolisme osseux chez les patients infectés par le HIV et traités par trithérapie.

For several years already, a growing number of studies reports modifications in the bone metabolism among HIV-infected patients. Some of these studies, published even before the use of HAART, involved the infection itself. With the experience already available as concerns HAART, antiretroviral treatments (ART) seem however to be called into question. Data are divergent yet. Some studies tend to invalidate the collected data about the harmful role of HAART and prove the absence of effect or even the beneficial action of ART on bone. Moreover, the three important classes of ART are implied, even if the proteases inhibitors are most commonly charged. Pathogenic mechanism remain hypothetical. While the impact on morbidity seems to be weak for the time being, long-term repercussions are still unknown, in particular when children are concerned. In such conditions, it appears difficult to set up coherent politics of screening, prevention and treatment. Nevertheless beyond the divergences, the multifactorial character of alteration of HIV-infected patient's bone metabolism seems to be undeniable. The identification of the different parameters should in the future clarify the situation and enable the publishing of exact criteria of screening, prevention and treatment.

Increased matrix metalloproteinase-3 serum levels in rheumatic diseases: relationship with synovitis and steroid treatment.

OBJECTIVE: To determine matrix metalloproteinase-3 (MMP-3) serum levels in patients with rheumatic diseases and to study the relation between MMP-3 and C reactive protein (CRP) levels. METHODS: MMP-3 serum levels were determined by enzyme linked immunosorbent assay (ELISA) in (a) patients with active inflammatory rheumatic diseases: rheumatoid arthritis (RA), psoriatic arthritis, polymyalgia rheumatica, acute crystal arthritis, and ankylosing spondylitis; (b) patients with active inflammatory systemic diseases: cutaneo-articular or renal systemic lupus erythematosus (SLE), systemic sclerosis, and vasculitides; (c) patients with non-inflammatory rheumatic diseases: osteoarthritis and fibromyalgia; (d) critically ill patients without rheumatic diseases, representing an acute inflammatory control group; (e) healthy controls. RESULTS: MMP-3 serum levels were significantly increased in patients with active RA, psoriatic arthritis, and polymyalgia rheumatica, whether treated or not by corticosteroids, and in female patients with acute crystal arthritis. MMP-3 serum levels were normal in steroid-free patients with active cutaneo-articular or renal SLE, systemic sclerosis, and vasculitides but were significantly increased in steroid treated patients. MMP-3 levels were normal in fibromyalgia, osteoarthritis, ankylosing spondylitis, and acute inflammatory controls. MMP-3 was significantly correlated with CRP in RA (r=0.5, p=0.0004) but not in any of the other disease groups. CONCLUSIONS: MMP-3 serum levels are increased in inflammatory rheumatic diseases characterised by joint synovitis, such as RA, polymyalgia rheumatica, psoriatic arthritis, and acute crystal arthritis-that is, whether the diseases are acute or chronic, erosive or not. They are normal in SLE, systemic sclerosis, and vasculitides as well as in non-rheumatic inflammatory controls, but are significantly increased by steroids. These data strongly suggest that serum MMP-3 reflects synovial inflammation.

Schistosoma mansoni induces the synthesis of IL-6 in pulmonary microvascular endothelial cells: role of IL-6 in the control of lung eosinophilia during infection.

The nature of the interactions between the intravascular parasite Schistosoma mansoni and the host pulmonary vasculature is critical in determining the outcome of infection. In this report, we show that lung schistosomula selectively induce the synthesis of IL-6 mRNA and protein in cultured human and mouse lung microvascular endothelial cells (EC) and that parasite excretory/secretory lipophilic compounds, particularly prostaglandin E(2), are responsible for this effect. In vivo, a striking increase of IL-6 expression is observed in the pulmonary microvasculature of S. mansoni-infected C57BL/6 mice suggesting that, in vivo, parasites also induce the synthesis of IL-6 in lung EC. In infected mice, IL-6 deficiency results in an accelerated mobilization of eosinophils into the lung tissue and in a dramatic increased number of recruited leukocytes, particularly eosinophils, in the airway. This effect is associated with an enhanced production of eotaxin (CCL11) and IL-5 in the lungs of IL-6 knockout (KO) animals. Finally, compared to wild-type mice, we detect a dramatic increased level of parasite mortality in the lungs of IL-6 KO mice. Taken together, we suggest that parasite larvae activate EC to produce IL-6 to escape the inflammatory reaction that develops in the lungs of infected hosts. Finally, we show that the parasite-induced IL-6 synthesis is mediated by a protein kinase A-dependent pathway that principally targets the cAMP-response element and the nuclear factor-kappaB sites from the -256/+20 region of the IL-6 promoter.

Relationship between bone mass density and tensile strength of the skin in women.

BACKGROUND: Skin and bone may be affected similarly during the climacteric and during long-term corticotherapy. Little is known about the correlation between the respective alterations in bone mass density (BMD) and tensile strength of the skin. MATERIALS AND METHODS: A total of 100 women aged 19-88 years, receiving, or not, hormone replenishment therapy or systemic corticosteroids, were enrolled in the study. Tensile strength of the inner forearm skin was measured using the suction method operated in both the steep and progressive-force application modes. BMD was measured by dual X-ray absorptiometry at the hip, femoral neck and lumbar spine. RESULTS: Being heterogeneous, the population of volunteers yielded a wide range of BMD and cutaneous tensile strength values. However, significant correlations were found between BMD and tensile skin parameters. In particular, a positive correlation was yielded between the biological elasticity of skin and the BMD of the hip and femoral neck. CONCLUSION: An overall correlation is shown between skin elasticity on a relatively sun-protected area and cortico-trabecular BMD.

Increased synovial fluid levels of interleukin-12, sCD25 and sTNF-RII/sTNF-RI ratio delineate a cytokine pattern characteristic of immune arthropathies.

The assessment of cytokines and their soluble receptors in the synovial fluid (SF) of inflammatory arthropathies may be useful in studying pathogenetic and immunoregulatory mechanisms underlying different diseases. The aim of this work was to study the cytokine network occurring in inflammatory arthropathies and to identify a cytokine profile which is characteristic of an immune-mediated synovitis. Levels of IL-12, as well as IL-4, IL-8, IL-10, IFN-gamma, sCD25, TNF-alpha and its soluble receptors were measured in the SF of various arthropathies, i.e. non-inflammatory arthropathies: "control" meniscus pathology (n = 21), osteoarthritis (n = 22) and chronic crystal arthritis (n = 9); a non-immune inflammatory arthropathy: acute crystal arthritis (n = 11); 2 immune inflammatory arthropathies: reactive arthritis (ReA) (n = 23) and rheumatoid arthritis (RA) (n = 44). SF levels of IL-10, TNF-alpha and sTNF-RII were found to be increased in the three inflammatory arthropathies compared to the "control" meniscus group. Within the inflammatory group, acute crystal arthritis was characterized by a significantly higher sTNF-RI/TNF-alpha ratio and ReA by a significantly lower sTNF-RII/TNF-alpha ratio compared to the two other diseases. The two immune arthropathies, RA and ReA, were characterized by increased SF levels of IL-12, sCD25 and of the sTNF-RII/sTNF-RI ratio. ReA differed however from RA by showing lower IL-8 and IL-4 levels, higher IFN-gamma levels and a higher IL-12/IL-10 ratio, suggesting a more prevalent Th1 profile in ReA SF. Our data indicate that the measurement of SF cytokines and soluble receptors may discriminate between each inflammatory arthropathy and might be useful in clinical practice.

Increased synovial fluid levels of soluble CD23 are associated with an erosive status in rheumatoid arthritis (RA).

Synovial fluid (SF) levels of soluble CD23 (sCD23) were determined in 96 patients presenting with an inflammatory knee effusion (73 with RA and 23 with reactive arthritis (ReA) serving as a control inflammatory non-erosive group) and were correlated with the degree of joint destruction, with local immune parameters (IL-1beta, IL-3, IL-4, IL-6, IL-8, IL-10, IL-12 and sCD25) and with serum markers of inflammation, C-reactive protein and erythrocyte sedimentation rate. RA patients, classified as erosive or not according to Larsen's grade, were separated as follows: (i) 13 patients with non-erosive RA; (ii) 16 RA patients with erosions in hands but not in knees, matched for disease duration with the first group; (iii) 44 RA patients with hand and knee erosions, matched with the second group for rheumatoid factor positivity but of longer disease duration. SF sCD23 levels were significantly increased in both erosive RA groups compared with non-erosive diseases, whether RA or ReA (P < 0.05), whose SF levels were not different. SF IL-10 showed a similar profile to that of SF sCD23 and was the only other parameter characteristic of erosive RA, but no direct correlation was found between the two. SF sCD23 was significantly correlated with IL-12 (r = 0.65, P = 0.0001) and sCD25 (r = 0.39, P = 0.0019) exclusively in the two erosive RA populations. In conclusion, these data showing that increased levels of sCD23 are not only found in the SF of erosive joints but also in knee SF of patients with erosive RA but without knee x-ray-diagnosed erosions suggest that this parameter might be of predictive value for joint destruction. Longitudinal studies are however needed to confirm its potential clinical interest.

Transforming growth factor-beta increases interleukin-6 transcripts in osteoblasts.

Bone remodeling is regulated by local factors and cytokines. Among them, interleukin-6 (IL-6) plays a critical role in bone resorption, and its synthesis is stimulated by osteoresorptive factors. Transforming growth factor-beta (TGF-beta) is present in high amounts in the bone matrix and is a local regulator of bone formation. However, its role in bone resorption remains unclear. In this paper, we report that TGF-beta stimulates IL-6 transcripts in a time- and dose-dependent manner in primary rat osteoblasts isolated from 22-day-old calvariae (Ob cells). The TGF-beta effect on IL-6 mRNA levels does not require de novo protein synthesis because cycloheximide, a protein synthesis inhibitor, does not block the induction. The mechanisms of IL-6 stimulation by TGF-beta is at least partially transcriptional because TGF-beta induces IL-6 heterogenous nuclear RNA, and, to a lesser extent, IL-6 transcription rate as determined by a nuclear run-on assay. Transforming growth factor-beta upregulation of IL-6 may be critical in conditions of increased bone resorption, such as myeloma.

Involvement of insulin-like growth factors in early T cell development: a study using fetal thymic organ cultures.

The expression of insulin-like growth factor (IGF) and IGF receptor genes was investigated by RT-PCR during ontogeny of the murine thymus. IGF-1, IGF-1R, M6P/IGF-2R genes are expressed in the thymus both in fetal and postnatal life, whereas IGF-2 messenger RNAs (mRNAs) decline after birth but are still detectable on the seventh week. By in situ hybridization, IGF-2 transcripts were located in the outer cortex and medulla of the postnatal thymus, and on the whole surface ofthe epithelial-like network in the fetal thymus. The effects of anti-IGFs and IGF-receptors neutralizing Abs on the generation of pre-T cell subpopulations were then investigated using fetal thymic organ cultures (FTOC). FTOC treatment with an anti-IGF-2 mAb, an anti-IGF-1R mAb, or an anti-M6P/IGF-2R polyclonal Ab induced a blockade of T cell differentiation at the CD4-CD8- stage, as shown by a significant increase in the percentage of CD4-CD8- cells and a decrease in the percentage of CD4+CD8+ cells. Moreover, anti-IGF-2 Ab treatment induced an increase in CD8+ cells suggesting that thymic IGF-2 might have a role in determining differentiation into the CD4 or CD8 lineage. Anti-IGF-1 Ab treatment decreased the proportion in CD4-CD8- cells and increased the frequency in CD4+CD8+. FTOC treatment with anti-(pro)insulin did not exert any significant effect on T cell development. These data indicate that the intrathymic IGF-mediated signaling plays an active role in the early steps of T cell differentiation during fetal development.

Synovial fluid matrix metalloproteinase-3 levels are increased in inflammatory arthritides whether erosive or not.

OBJECTIVE: To study the levels of matrix metalloproteinase-3 (MMP-3) in the knee synovial fluid (SF) of inflammatory arthropathies (rheumatoid arthritis whether erosive or not, reactive arthritis, acute crystal arthritis) and degenerative arthropathies [chronic crystal disease, osteoarthritis and (control) meniscus pathology] and to correlate them with the degree of joint destruction, local inflammatory and immune parameters and systemic markers of inflammation. METHODS: SF levels of MMP-3 (precursor, active and tissue inhibitor of MMP-bound forms), tumour necrosis factor (TNF) alpha, soluble TNF receptors I and II, interleukin (IL)-6 and soluble IL-6 receptor were measured by ELISA in 107 inflammatory and 53 degenerative arthropathies. RESULTS: MMP-3 levels in SF were (i) significantly higher in inflammatory than in degenerative arthropathies; (ii) not related to the degree of joint destruction; (iii) significantly correlated with the levels of all SF markers tested and with erythrocyte sedimentation rate and serum levels of C-reactive protein and fibrinogen. CONCLUSION: Increased MMP-3 levels in SF are found in inflammatory arthropathies and are not specific for erosive joint diseases. MMP-3 in SF is therefore a potential candidate for the assessment of the inflammatory process in joints. However, the exclusive determination of the active form could indicate the degree of joint destruction.

Characterization of the insulin-like growth factor axis in the human thymus.

The components of the insulin-like growth factor (IGF) axis have been investigated in the normal human thymus. Using ribonuclease protection assays (RPA), IGF-II transcripts were detected in the normal human thymus. By reverse transcriptase polymerase chain reaction (RT-PCR) analyses, promoters P3 and P4 were found to be active in the transcription of IGF2 gene within human thymic epithelial cells (TEC). No IGF-II mRNA could be detected in human lymphoid Jurkat T cells with 30 cycles of RT-PCR. By Northern blot analyses, IGFBP-2 to -6 (but not IGFBP-1) were found to be expressed in TEC with a predominance of IGFBP-4. Interestingly, Jurkat T cells only express IGFBP-2 but at high levels. The type 1 IGF receptor was detected in Jurkat T cells but not in human TEC. The identification of the components of the IGF axis within separate compartments of the human thymus adds further evidence for a role of this axis in the control of T-cell development. The precise influence of thymic IGF axis upon T-cell differentiation and immunological self-tolerance however needs to be further investigated.

Platelet-derived growth factor induces interleukin-6 transcription in osteoblasts through the activator protein-1 complex and activating transcription factor-2.

Platelet-derived growth factor (PDGF) BB, a mitogen that stimulates bone resorption, increases the expression of interleukin-6 (IL-6), a cytokine that induces osteoclast recruitment. The mechanisms involved in IL-6 induction by PDGF BB are poorly understood. We examined the effect of PDGF BB on IL-6 expression in cultures of osteoblasts from fetal rat calvariae (Ob cells). PDGF BB increased IL-6 mRNA and heterogeneous nuclear RNA levels, the rate of transcription, and the activity of base pairs (bp) -2906 to +20 IL-6 promoter fragments transiently transfected into Ob cells. Deletion analysis revealed two responsive regions, one containing an activator protein-1 (AP-1) site located between bp -276 and -257, and a second, less well defined, downstream of -257. Targeted mutations of a cyclic AMP-responsive element (CRE), and nuclear factor-IL-6 and nuclear factor-kappaB binding sites in a bp -257 to +20 IL-6 construct that was transfected into Ob cells, revealed that the CRE also contributed to IL-6 promoter induction by PDGF BB. Electrophoretic mobility shift assay revealed AP-1 and CRE nuclear protein complexes that were enhanced by PDGF BB. Supershift assays revealed binding of Jun and Fos to AP-1 and CRE sequences and binding of activating transcription factor-2 to CRE. In conclusion, PDGF BB induces IL-6 transcription in osteoblasts by regulating nuclear proteins of the AP-1 complex and activating transcription factor-2.

Interleukin-6 with its soluble receptor enhances the expression of insulin-like growth factor-I in osteoblasts.

Interleukin (IL)-6, a cytokine produced by skeletal cells and known to increase bone resorption, has mitogenic effects for bone cells, possibly by regulating the synthesis of other local factors. We tested the effects of IL-6 and its soluble receptor (IL-6sR) on the expression of insulin-like growth factor (IGF)-I and IGF-II in cultured osteoblast-enriched cells from fetal rat calvariae (Ob cells). IL-6 did not modify IGF-I messenger RNA (mRNA) levels, but when tested in the presence of IL-6sR, IL-6 at 1 to 100 ng/ml increased IGF-I transcripts by up to 3.2-fold after 24 h. IL-6sR caused a small increase in IGF-I mRNA levels when tested alone. IL-6 and IL-6sR increased immunoreactive IGF-I levels by 2.4-fold after 24 h and 6.4-fold after 48 h. Cycloheximide prevented, and indomethacin markedly decreased, the effect of IL-6 and IL-6sR on IGF-I mRNA levels, but hydroxyurea did not. IL-6 and IL-6sR did not alter the decay of IGF-I mRNA in transcriptionally arrested Ob cells, and the half-life of the predominant 6.5-kb IGF-I transcript was about 11 h in control and treated cells. In addition, IL-6 and IL-6sR increased the levels of IGF-I heterogeneous nuclear RNA. IL-11 also increased IGF-I mRNA levels, whereas oncostatin M and leukemia-inhibitory factor did not. In contrast to their effects on IGF-I, IL-6 and IL-6sR caused only a modest increase in IGF-II mRNA and polypeptide levels. In conclusion, IL-6, in the presence of IL-6sR, increases IGF-I synthesis in Ob cells; this effect may lead to a secondary increase in bone formation.

Interleukin 6 is autoregulated by transcriptional mechanisms in cultures of rat osteoblastic cells.

Interleukin 6 (IL-6), a cytokine produced by skeletal cells, stimulates osteoclast recruitment. The IL-6 soluble receptor (sIL-6R) increases IL-6 activity, and IL-6 and sIL-6R levels are increased in conditions of increased bone resorption. We examined the production of IL-6 by primary rat osteoblasts (Ob cells) cultured in the presence of IL-6 and sIL-6R. IL-6 alone did not induce IL-6 transcripts, but IL-6 was stimulatory in the presence of sIL-6R. Furthermore, sIL-6R by itself increased IL-6 transcripts. Cycloheximide superinduced IL-6 transcripts and did not prevent the effect of IL-6 and sIL-6R. IL-6 in the presence of sIL-6R stimulated IL-6 rates of transcription and the activity of IL-6 promoter fragments in transiently transfected Ob cells. 5' deletions of the IL-6 promoter and targeted mutations of the multiple response element (MRE)/cAMP responsive element (CRE), the nuclear factor for IL-6 (NF-IL-6), and the nuclear factor-kappaB (NF-kappaB) binding sites indicated that NF-IL-6 and NF-kappaB, in combination with MRE/CRE, binding sites are required for the induction of the IL-6 promoter by IL-6. In conclusion, IL-6 induces its own synthesis in osteoblasts by transcriptional mechanisms. This positive feedback may be important in conditions of increased bone resorption.

Interleukin-6 and its soluble receptor regulate the expression of insulin-like growth factor binding protein-5 in osteoblast cultures.

Interleukin-6 (IL-6), a cytokine produced by bone cells, is known to influence bone resorption by stimulating the development of osteoclasts from precursor cells and to have mitogenic actions on osteoblastic cells. Insulin-like growth factors (IGFs) are important local regulators of bone formation, and IGF binding protein (IGFBP)-5 stimulates bone cell growth and enhances the effects of IGF-I. We tested the effects of IL-6 in the presence and absence of its soluble receptor (sIL-6R) on IGFBP-5 expression in cultures of osteoblast-enriched cells from 22-day-old fetal rat calvariae (Ob cells). When tested individually, IL-6 and sIL-6R had a modest stimulatory effect on IGFBP-5 messenger RNA (mRNA) levels. In contrast, when IL-6 and sIL-6R were tested in combination, they caused a considerable increase in IGFBP-5 mRNA levels, and IL-6 at 100 ng/ml and sIL-6R at 125 ng/ml increased IGFBP-5 transcripts by 5- to 7-fold after 24 h. The effect of IL-6 and sIL-6R on IGFBP-5 transcripts was not blocked by indomethacin, but cycloheximide markedly inhibited IGFBP-5 mRNA levels in control and treated cultures. IL-6 and sIL-6R did not modify the decay of IGFBP-5 mRNA in transcriptionally arrested Ob cells, and stimulated the rate of IGFBP-5 transcription as demonstrated by a nuclear run-on assay. IL-6 and sIL-6R did not increase intact IGFBP-5 levels in the extracellular matrix and increased IGFBP-5 fragments in the culture medium. Conditioned medium from Ob cells induced the proteolytic fragmentation of an IGFBP-5 standard, an effect that was accelerated and enhanced by conditioned medium from IL-6/sIL-6R-treated cultures and prevented by metalloprotease inhibitors. In conclusion, IL-6, in the presence of sIL-6R, stimulates IGFBP-5 mRNA expression in Ob cells by transcriptional mechanisms, and accelerates the fragmentation of the protein.