Impaired release of interleukin-6 from human osteoblastic cells in the uraemic milieu

Simon J. Steddon1, Christopher W. McIntyre2, Neil J. Schroeder3, Jacky M. Burrin3 and John Cunningham4

1 Department of Renal Medicine and Transplantation, Bart's and the Royal London Hospitals, London, 3 Department of Endocrinology, St Bartholomew's and the Royal London School of Medicine and Dentistry, Queen Mary University of London, 4 Department of Nephrology, University College London Hospitals, The Middlesex Hospital, London and 3 Department of Renal Medicine and Centre for Integrated Systems Biology and Medicine, Derby City General Hospital, Derby, UK

Correspondence and offprint requests to: Professor John Cunningham, Department of Nephrology, University College London Hospitals, The Middlesex Hospital, Mortimer St, London W1T 3AA, UK. Email: john.cunningham{at}uclh.org



   Abstract
 Top
 Abstract
 Introduction
 Subjects and methods
 Results
 Discussion
 References
 
Background. Osteoblast-derived interleukin-6 (IL-6) affects bone metabolism and is linked with a number of pathological states characterized by increased bone resorption, including osteoporosis and renal osteodystrophy. To examine the possibility that uraemia directly influences the release of this cytokine in bone, we have investigated the effect of human uraemic serum on the release of IL-6 from human osteoblast-like cells.

Methods. Individual serum samples collected from healthy male volunteers or male haemodialysis patients prior to and during a dialysis treatment were assayed for IL-6, interleukin-1ß (IL-1ß) and soluble IL-6 receptor (sIL-6R) using specific enzyme-linked immunosorbent assays. MG-63 and SaOS-2 cells were cultured in media containing pooled sera from both groups and alongside matching charcoal-stripped sera. IL-6 concentrations were determined in harvested cell supernatants after 24 h. In further experiments, media containing individual sera obtained from five patients at regular intervals during their haemodialysis treatment were incubated with MG-63 cells to determine the effects of the dialysis process on IL-6 secretion.

Results. Haemodialysis patients had significantly higher (n = 10, P<0.001) circulating concentrations of IL-6 (7.0±1.6 pg/ml) than normal subjects (0.4± 0.1 pg/ml), but there were no significant differences in the concentrations of either IL-1ß or sIL-6R. These serum concentrations did not change significantly during 80 min of dialysis. IL-6 release by MG-63 cells incubated with charcoal-stripped serum from normal or from uraemic subjects was similar. Incubation with untreated sera from normal subjects increased IL-6 release by ~6-fold above the charcoal-stripped control, whereas sera from uraemic subjects increased IL-6 release by only ~2- to 3-fold (normal vs uraemic of 6878±595 and 2579±169 pg/ml, respectively, P<0.001). Similar results were seen with SaOS-2 cells. Haemodialysis did not restore the capacity of uraemic serum to augment IL-6 release to the same degree as normal serum.

Conclusions. These data show that the augmentation of IL-6 release from human osteoblastic cells after incubation with normal serum is greater than after uraemic serum. This may indicate the presence of an inhibitor of IL-6 release in uraemic serum that is involved in the deranged bone turnover of uraemic patients.

Keywords: bone remodelling; haemodialysis; interleukin-6; osteoblast; renal osteodystrophy; uraemic serum



   Introduction
 Top
 Abstract
 Introduction
 Subjects and methods
 Results
 Discussion
 References
 
Renal osteodystrophy describes a spectrum of skeletal abnormalities seen in patients with renal disease and encompasses a series of disorders of bone remodelling [1]. Historically, the predominant group of bony abnormalities have been those attributable to hyperparathyroidism, with associated increases in bone turnover resulting in the histological appearances of osteitis fibrosa. However, low turnover bone lesions in the absence of associated aluminium toxicity have been increasingly recognized [2,3]. Several studies in end-stage renal disease (ESRD) patients have identified normal bone turnover co-existing with moderately elevated levels of parathyroid hormone (PTH) and thus to the recommendation that PTH concentrations are maintained at 2–3 times the upper limit of the normal reference range [4,5]. Recently published K/DOQI guidelines have gone even further, advising a target PTH concentration of 150–300 pg/ml (16.5–33.0 pmol/l) for all those on dialysis or with a glomerular filtration rate of <15 ml/min [i.e. stage five chronic kidney disease] [6]. While the realization that many PTH assays detect potentially antagonistic fragments in addition to intact PTH provides at least a partial explanation of the need for such levels [7], it seems likely that additional factors present in the uraemic milieu may also serve to depress bone cell function.

Since bone remodelling requires coupling of osteoclastic bone resorption and osteoblastic bone formation, disruption of these linked processes may play a role in the aetiology of osteodystrophy. A large number of osteotropic factors such as bone morphogenetic proteins cytokines and growth factors influence the principal mediators of osteoclastic resorption, namely osteoprotegerin and RANK-L [8]. Amongst these, the pluripotent cytokine interleukin-6 (IL-6) is thought to play an important role in the initiation and maintenance of osteoclastogenesis [9] and has been implicated in the pathogenesis of a wide variety of skeletal pathologies characterized by accelerated bone remodelling and resorption. There is evidence for the involvement of IL-6 in the pathogenesis of renal osteodystrophy [10], and we have demonstrated previously that IL-6 release by osteoblast-like cells can be modulated by vitamin D-like compounds [11].

In the present study, the circulating levels of IL-6, interleukin-1ß IL-1ß, and soluble IL-6 receptor (sIL-6R) in patients prior to and during a haemodialysis (HD) session are examined and the effect of both normal and uraemic serum on IL-6 release from human osteoblastic cells explored. In addition, the capacity of both charcoal stripping and therapeutic HD to remove those substances potentially interfering with the serum-stimulated release of osteoblastic IL-6 is evaluated.



   Subjects and methods
 Top
 Abstract
 Introduction
 Subjects and methods
 Results
 Discussion
 References
 
Normal serum samples were obtained from 10 healthy male volunteers with no history of febrile illness in the preceding 4 weeks. Uraemic sera were obtained from 10 male ESRD patients treated by HD (renal diagnoses: small kidneys, uncertain cause, 3; hypertensive nephrosclerosis, 2; renovascular disease, 1; membranous glomerulonephritis, 2; focal and segmental glomerulosclerosis, 1; adult polycystic kidney disease, 1). All patients had been on HD for at least 1 year (mean 26 months, range 13–62) and had a mean age of 39 years (range 28–67). All patients underwent three, 4 h dialysis sessions per week using cellulose acetate dialysers and bicarbonate buffering. Venous access in all subjects was by primary arteriovenous fistulae. Blood was sampled from the arterial access port prior to passing through the dialyser at the start of a HD treatment session. Serum was obtained from blood samples taken into plain glass tubes and allowed to stand upright at room temperature for 20 min before separation by centrifugation at 2000 r.p.m., removal of serum with a sterile pipette and immediate storage at –20°C.

Blood samples were also obtained from five additional patients during an individual dialysis session. The characteristics of these patients are shown in Table 1. Samples were taken prior to dialysis and at 30, 60, 120, 180, 240 and 300 min into their treatment. The first 60 min comprised extra-corporeal circulation alone, with no dialysate flow and no programmed ultrafiltration (hereafter referred to as sham dialysis). After 60 min, dialysate flow was introduced and continued for the remainder of the dialysis treatment.


View this table:
[in this window]
[in a new window]
 
Table 1. Characteristics of the five patients who underwent serum sampling throughout an individual dialysis treatmenta

 
Individual normal and uraemic serum samples were assayed for IL-6, sIL-6R and IL-1ß. IL-1ß is known to increase the release of IL-6 from both MG-63 and SaOS-2 cells [13] and was measured to assess any contribution to the differences in measured IL-6. The sIL-6R is a 50 kDa ligand protein derived from the surface shedding of the gp80 component of the IL-6 receptor. Its ability to bind free IL-6 means it is a potential source of interference with the measured concentration of IL-6. Some aliquots of human sera were ‘inactivated’ by charcoal stripping, a process that removes hydrophobic structures including steroids and growth factors [12]. A 2 g aliquot of activated charcoal was added to 50 ml of serum and mixed at room temperature for 4 h. Serum was recovered by centrifuging for 30 min at 3500 r.p.m. The supernatant was re-centrifuged for a further 20 min at 20 000 r.p.m. at room temperature. Prior to use, the serum was filtered through 0.45 and then 0.2 µm filters (Minisart®, Sartorius AG, Göttingen, Germany). Additional aliquots were heat treated (65°C for 30 min) as an alternative method of growth factor and hormone inhibition.

Ethical approval for these procedures was obtained from the relevant local body (East London and the City Health Authority, reference P/00/241). Patients provided written informed consent.

Cell culture and experimental design
Two human osteoblast-like cell lines were used in these experiments. MG-63 cells produce relatively low levels of alkaline phosphatase and do not form a mineralized matrix in vitro. They are often considered to be of a relatively immature phenotype. SaOS-2 possess some of the characteristics of a more differentiated osteoblastic cell, producing more constitutive alkaline phosphatase and also possessing the ability to form a mineralized matrix in long-term cultures. Cells were cultured and maintained in {alpha}-minimal essential medium ({alpha}-MEM) containing 10% fetal calf serum (FCS) (Gibco Life Technologies Ltd) and antibiotics (benzylpenicillin 100 U/ml, streptomycin 10 µg/ml, amphotericin 2.5 µg/ml) in a humidified 1:19 air/CO2 environment at 37°C. Cells were seeded into 24-well plates at a density of 200 000 cells/well. They were allowed to adhere for 24 h before aspiration of media, washing with sterile phosphate-buffered saline (PBS) and incubation with {alpha}-MEM, antibiotics and 10% charcoal-stripped FCS (CS-FCS) to provide a non-stimulating (growth-arrested) environment for a further 24 h. To examine the effects of human sera on IL-6 secretion, cells were then incubated for 24 h in media supplemented with antibiotics and the human sera of interest at a 10% concentration. Negative controls were provided by refreshing cells with 10% CS-FCS, and positive controls (for IL-6) by using 10% CS-FCS to which IL1-ß (R&D Systems Europe Ltd, Abingdon, Oxon, UK) had been added (at 100 IU/ml). At the end of the 24 h incubation period, media were collected and stored at –20°C for no longer than 1 month before assay. Cells were washed with PBS and subjected to three freeze/thaw cycles between –70°C and room temperature to allow cell lysis for the measurement of total cellular protein. All samples were assayed in at least quadruplicate and individual experiments repeated three or more times.

Cytokine assays
IL-6 was measured in all media and sera by a quantitative sandwich enzyme-linked immunosorbent assay (ELISA) technique (Eurogenetics, Middlesex, UK). IL1-ß and sIL-6R were also assayed by commercially available ELISAs (R&D Systems, Abingdon, UK).

Total protein assay
Cellular protein was assayed using the Bradford dye-binding protein assay (Bio-Rad, Hemel Hempstead, Hertfordshire, UK) in a spectrophotometric microtitre-based format. Absorbance was measured at 595 nm and compared with a standard curve prepared using known concentrations of bovine serum albumin.

Statistical analysis
Comparisons between normal and uraemic serum cytokine concentrations were analysed by unpaired t-test, while other results of more than two groups were analysed by one-way or repeated measures ANOVA with Tukey's post hoc test for inter-group comparisons. P<0.05 was considered significant (NS = not significant) and all results are expressed as the mean±SEM.



   Results
 Top
 Abstract
 Introduction
 Subjects and methods
 Results
 Discussion
 References
 
Serum cytokine concentrations
The serum concentrations of IL-6 were lower in normal subjects than in serum, taken before dialysis, from stable HD patients (0.4±0.1 vs 7.0±1.6 pg/ml, respectively, P = 0.003). However, no significant differences in the concentrations of sIL-6R or IL-1ß were seen in sera taken from normal subjects compared with ESRD patients (sIL-6R, 65±26 vs 96±24 µg/l, NS; IL-1ß, 7.6±1.6 vs 8.7±2.5 pg/ml, NS). In those subjects followed through a dialysis session, no significant changes in the serum levels of IL-6, sIL-6R or IL-1ß were found (Table 1).

Serum effects on osteoblast IL-6 release
Significant differences in IL-6 secretion were seen in MG-63 cells when incubated with the various sera (P<0.0001 by one-way ANOVA, Figure 1). Serum that had been charcoal stripped, from either normal or uraemic subjects, resulted in similar amounts of basal IL-6 release (1134±127 vs 853±78 pg/ml, NS). Untreated normal serum augmented IL-6 release by ~6-fold (to 6878±595 pg/ml) over basal (charcoal-stripped) concentrations. This increase was significantly attenuated with untreated uraemic serum, which increased release by ~2- to 3-fold (to 2579± 169 pg/ml) over basal concentrations. The augmentation of IL-6 release by normal sera was significantly greater than that by uraemic sera (6878±595 vs 2579±169 pg/ml, P<0.001). Heat treatment of uraemic serum had no effect on the release of IL-6 from MG-63 cells (1952±172 heat-treated vs 1872± 165 pg/ml non-heat-treated). IL-1ß was by far the most powerful stimulus to IL-6 production (31 802± 3250 pg/ml), whilst serum-free medium was the weakest (187±33 pg/ml).



View larger version (17K):
[in this window]
[in a new window]
 
Fig. 1. IL-6 release from MG-63 cells after 24 h incubation with media containing normal and uraemic serum. CS = charcoal stripped. Significant differences between treatments were found (P<0.0001 by one-way ANOVA; for each column n = 12). The data were reproduced in two further experiments.

 
Quantitatively similar results were observed with SaOS-2 cells after incubation with human sera. The augmentation of IL-6 release was 6-fold greater with normal serum compared with uraemic serum (374±6 vs 61±4 pg/ml, respectively).

Effect of haemodialysis on osteoblastic IL-6 release in response to uraemic serum
Having found significant differences in IL-6 production, further experiments were designed to explore the capacity, if any, of HD to restore the ability of uraemic serum to promote IL-6 production to the same degree as normal serum (Figure 2). The initial period of sham dialysis was necessary to explore the possible consequences of both heparinization and extracorporeal circulation of blood for subsequent effects in cell culture. Therapeutic dialysis aided the investigation into the potential removal of mediators of the effect. Although for some patients there was variability in osteoblastic IL-6 secretion at different time points, no significant differences or trends were evident.



View larger version (17K):
[in this window]
[in a new window]
 
Fig. 2. IL-6 release from MG-63 cells after 24 h incubation with media containing sera harvested from five individual patients during a single haemodialysis session. The first 60 min represent ‘sham’ dialysis, with no dialysate flow. The abscissas depict the time of blood sampling.

 
Total cellular protein
There were no significant differences in total extracted cellular protein as compared with growth-arrested control cells following incubation with any of the sera under investigation (7.22±0.48 mg/ml for uraemic, 8.1±0.32 mg/ml for charcoal-stripped uraemic, 7.64±0.47 mg/ml for normal serum, 7.97±0.32 mg/ml for charcoal-stripped normal serum and 7.97± 0.52 mg/ml for pooled post-HD serum).



   Discussion
 Top
 Abstract
 Introduction
 Subjects and methods
 Results
 Discussion
 References
 
These studies demonstrate that the release of IL-6 from human osteoblastic cells is substantially greater during exposure to media containing serum from non-uraemic subjects than during incubation with charcoal-stripped sera from the same individuals. The release of IL-6 is substantially attenuated in the presence of media containing serum from uraemic patients established on maintenance HD, though even uraemic serum significantly augmented IL-6 release in comparison with charcoal-stripped serum. Charcoal-stripped serum from normal and uraemic subjects yielded similar results. These findings suggest that uraemic serum itself might be able to influence bone turnover. In agreement with previous experiments [11,13], the constitutive release of IL-6 from SaOS-2 cells was significantly (~10-fold) lower than from MG-63 cells; however, the effect of both normal and uraemic sera on the SaOS-2 cells paralleled that seen in the MG-63 cells. In both these cell lines, IL-6 release was not augmented with the negative control (CS-FCS) but was strongly augmented (>10-fold) with 100 IU of IL-1ß.

Charcoal stripping is a commonly utilized means of removing steroid hormones, including androgen and oestradiol metabolites, cortisol and thyroid hormones from serum [12]. The process is also likely to remove other growth factors and cytokines, although its exact quantitative effect in this regard is ill defined. IL-6 production after exposure to uraemic serum was enhanced with respect to exposure to charcoal-stripped normal or uraemic sera (Figure 1). We were not able to document any consistent effect of HD on the capacity of uraemic serum to augment IL-6 release (Figure 2). Sequential samples taken from five patients during sham dialysis for 60 min, followed by conventional HD for 240 min, evoked a similar and near constant level of augmentation. These results imply that, in contrast to charcoal stripping, the factor(s) in uraemic serum affecting IL-6 release were not removed by haemodialysis. This may be a clue to inherent properties of the molecules involved, as charcoal stripping is able to remove larger, often more complex, molecules than HD across standard membranes.

The difference in IL-6 concentration in media harvested from MG-63 cells cultured with the various sera is not due to pre-existing circulating IL-6. Even though a disparity in the concentration of IL-6 in normal and uraemic sera was present (similar to those differences reported by Herbelin et al. [14] in patients with chronic renal failure±dialysis), these were trivial compared with those in the supernatants at the completion of the experiments. Furthermore, the uraemic and normal serum were diluted 10-fold in media before use. There was no effect of HD itself on circulating IL-6 concentrations (also reported by Herbelin et al. [14]). IL-6, in the presence of sIL-6R, is known to induce its own synthesis in osteoblasts [15] and, if an effect from pre-existing IL-6 in the sera were present, then it might be anticipated that uraemic, rather than normal, serum would increase osteoblast IL-6 secretion.

IL-1ß is known to increase the release of IL-6 from the cell lines studied [13] and, indeed, addition of IL-1ß to the media as a positive control led to a dramatic increase in IL-6 secretion. There were no significant differences in the concentration of IL-1ß in any of the sera used in the incubation experiments and, as HD had no effect on the serum concentrations of IL-1ß (Table 2), it seems unlikely that this cytokine played a significant role. The ability of dialysis membranes to influence IL-6 release has been reported previously. Both circulating IL-1ß and IL-1ß release from harvested peripheral blood monocytes have been shown to be elevated by dialysis involving bioincompatible (cuprophane) membranes [16], though not by cellulose acetate-based membranes akin to those used in this study [17]. Thus, not only is IL-1ß unlikely to have contributed to differences in the release of IL-6, but it is also unlikely to have had any tonic effects; the final concentrations that the MG-63 cells were exposed to were all less than those that we have reported previously as having an effect on IL-6 release from this cell line [11].


View this table:
[in this window]
[in a new window]
 
Table 2. Effect of haemodialysis on serum cytokines in eight subjects followed for 80 min during a single haemodialysis session

 
sIL-6R is a 50 kDa ligand-binding protein derived from surface shedding of the gp80 IL-6 receptor [18] and might be expected to interfere with measured IL-6 levels by binding to free IL-6, possibly through interference with immunoassay detection. Artefacts of this kind seem unlikely as we found no significant differences in the concentrations of sIL-6R in any of the sera from patients or control subjects and dialysis did not influence the circulating concentrations of sIL-6R in the uraemic patients.

There are precedents for components of the uraemic milieu affecting cellular function. In 1976, Wills and Jenkins reported that pre-dialysis serum from uraemic patients was able to inhibit the resorptive effect of parathyroid extract on bone, a phenomenon that was not reproducible by the addition of known uraemic metabolites. Furthermore, this effect could be removed by dialysis, following which there was a net increase in resorption [19]. Uraemic toxins have been shown to affect the normal functioning of calcitriol by inhibiting the binding of calcitriol receptors to vitamin D response elements in a uraemic rat model [20]. We have reported previously that calcitriol does not have an effect on IL-6 transcription or release from MG-63 cells at concentrations < 10–9 M, far higher than those encountered in the cell culture media [11]. Thus, calcitriol is unlikely to be responsible for the effects on IL-6 release in this cell system. Many small proteins in uraemic serum may have profound effects on cellular functions [21] and it might be expected that some of these, for example PTH, may exert influence in the current model. Our previous observation that PTH has no effect on IL-6 production in MG-63 cells at concentrations below 10–9 M [11] coupled with the demonstration that concentrations as high as 10–7 M do not increase IL-6 release in primary cell osteoblast cultures [22] suggests that this is unlikely to be the case. The PTH concentrations in the culture media fell below the lower of these concentrations. Furthermore, heat treatment would have been expected to destroy PTH, and no such effect was observed. Srivastava et al. reported that uraemic serum contains a humoral substance capable of modulating colony-forming unit counts in mice, an effect that could be modified by dialysis [23]. In the current study, an effect of the various sera on cellular proliferation is unlikely to be of relevance as there were no differences in the levels of total cellular protein extracted from the various experiments. This is compatible with, although not proof of, a neutral effect on proliferation. Furthermore, Picton et al. reported that a variety of uraemic toxins exerted no effect on the proliferation of SaOS-2 cells, despite incubation times up to three times longer than those used in this set of experiments [24].

In conclusion, IL-6 release from human osteoblast-like cells is augmented in the presence of normal human serum, but to a much lesser degree by uraemic serum. The removal of certain molecules, including cytokines and growth factors, through charcoal stripping of the serum, removes its ability to augment IL-6 production, though it remains unaffected by either sham or therapeutic dialysis. This suggests there may be a common factor(s) in uraemic serum, apparently not influenced by primary renal diagnosis, causing reduced stimulation of IL-6 release from osteoblasts and therefore potentially involved in the aberrant local signalling in uraemic bone.



   Acknowledgments
 
This work was supported by the National Kidney Research Fund, the Special Trustees of the Royal London Hospital and the Joint Research Board of the Special Trustees of St Bartholomew's Hospital.

Conflict of interest statement. None declared.



   References
 Top
 Abstract
 Introduction
 Subjects and methods
 Results
 Discussion
 References
 

  1. Hruska KA, Teitelbaum SL. Renal osteodystrophy. N Engl J Med 1995; 333: 166–174[Free Full Text]
  2. Sherrard D, Hercz G, Pei Y. The spectrum of bone disease in end stage renal failure—an evolving disorder. Kidney Int 1993; 43: 436–442[ISI][Medline]
  3. Ballantini P, Wedard M, Bonucci E. Frequency of adynamic bone disease and aluminium storage in Italian uraemic patients—retrospective analysis of 1429 iliac crest biopsies. Nephrol Dial Transplant 1996; 11: 663–667[Abstract]
  4. Quarles LD, Lobaugh B, Murphy G. Intact parathyroid hormone overestimates the presence and severity of parathyroid mediated osseus abnormalities in uremia. J Clin Endocrinol Metab 1992; 75: 145–150[Abstract]
  5. Wang M, Hercz G, Sherrard DJ et al. Relationship between intact 1–84 parathyroid hormone and bone histomorphometric parameters in dialysis patients without aluminium toxicity. Am J Kidney Dis 1995; 26: 836–844[ISI][Medline]
  6. Eknoyan G, Levin A, Levin NW. Bone metabolism and disease in chronic kidney disease. Am J Kidney Dis 2003; 42 [4 Suppl 3]: 1–201
  7. Slatopolsky E, Finch J, Clay P et al. A novel mechanism for skeletal resistance in uremia. Kidney Int 2000; 59: 2375–2376
  8. Aubin JE, Bonnelye E. Osteoprotegerin and its ligand: a new paradigm for regulation of osteoclastogenesis and bone resorption. Osteoporos Int 2000; 11: 905–13[CrossRef][ISI][Medline]
  9. Adebanjo OA, Moonga BS, Yamate T et al. Mode of action of interleukin-6 on mature osteoclasts. J Cell Biol 1998; 142: 1347–1356[Abstract/Free Full Text]
  10. Langub MC Jr, Koszweski NJ, Turner HV, Monier-Faugere M-C, Geng Z, Malluche H. Bone resorption and mRNA expression of IL-6 and IL-6 receptor in patients with renal osteodystrophy. Kidney Int 1996; 50: 515–520[ISI][Medline]
  11. McIntyre C, Schroeder NJ, Burrin J, Cunningham J. Effects of new analogues of vitamin D on bone cells: implications for treatment of uraemic bone disease. Kidney Int 1999; 55: 500–511[CrossRef][ISI][Medline]
  12. Darbre P, Yates J, Curtis S, King RJ. Effect of estradiol on human breast cancer cells in culture. Cancer Res 1983; 43: 349–354[Abstract]
  13. Siddiqi A, Burrin JM, Wood DF, Monson JP. Tri-iodothyronine regulates the production of interleukin-6 and interleukin-8 in human bone marrow stromal and osteoblast-like cells. J Endocrinol 1998; 157: 453–461[Abstract/Free Full Text]
  14. Herbelin A, Urena P, Nguyen AT, Zingaff J, Descamps-Latscha B. Elevated circulating levels of interleukin-6 in patients with chronic renal failure. Kidney Int 1991; 39: 954–960[ISI][Medline]
  15. Franchimont N, Rydziel S, Canalis E. Interleukin-6 is autoregulated by trancriptional mechanisms in cultures of rat osteoblastic cells. J Clin Invest 1997; 100: 1797–1803[Abstract/Free Full Text]
  16. Lin YF, Chang DM, Shaio MF et al. Cytokine production during haemodialysis: effects of dialytic membrane and complement activation. Am J Nephrol 1996; 16: 293–299[ISI][Medline]
  17. Qian J, Yu Z, Dai H, Zhang Q, Chen S. Influence of haemodialysis membranes on gene expression and plasma levels of interleukin-1 beta. Artif Organs 1995; 19: 842–846[ISI][Medline]
  18. Mullberg J, Schooltink H, Stoyan T et al. The soluble interleukin-6 receptor is generated by shedding. Eur J Immunol 1993; 23: 473–480[ISI][Medline]
  19. Wills MR, Jenkins MV. The effect of uraemic metabolites on parathyroid extract induced bone resorption in vitro. Clin Chim Acta 1976; 73: 121–125[CrossRef][ISI][Medline]
  20. Patel S, Ke H-Q, Vanholder R, Koenig RJ, Hsu CH. Inhibition of calcitriol receptor binding to vitamin D response elements by uremic toxins. J Clin Invest 1995; 96: 50–59[ISI][Medline]
  21. Haag-Weber M, Mai B, Cohen G, Hörl WH. GIP and DIP: a new view of uraemic toxicity. Nephrol Dial Transplant 1994; 9: 346–347[ISI][Medline]
  22. Littlewood AJ, Russell J, Harvey GR, Hughes DE, Russell RG, Gowen M. The modulation of the expression of IL-6 and its receptor in human osteoblasts in vitro. Endocrinology 1991; 129: 1513–1520[Abstract]
  23. Srivastava A, Agarwal S, Malhotra KK, Sood SK. The effect of serum from patients of chronic renal failure on colony forming units (CFU-S) in mice. Indian J Pathol Microbiol 1990; 33: 166–70[Medline]
  24. Picton ML, Hutchinson AJ, Gokal R, Hoylan JA. Effect of uraemic toxins on the proliferation of human osteoblast like cells in vitro [abstract]. Kidney Int 1999; 55: 2097–2098A
Received for publication: 11.12.03
Accepted in revised form: 10. 8.04





This Article
Abstract
FREE Full Text (PDF)
All Versions of this Article:
19/12/3078    most recent
gfh491v1
Alert me when this article is cited
Alert me if a correction is posted
Services
Email this article to a friend
Similar articles in this journal
Similar articles in ISI Web of Science
Similar articles in PubMed
Alert me to new issues of the journal
Add to My Personal Archive
Download to citation manager
Search for citing articles in:
ISI Web of Science (1)
Disclaimer
Request Permissions
Google Scholar
Articles by Steddon, S. J.
Articles by Cunningham, J.
PubMed
PubMed Citation
Articles by Steddon, S. J.
Articles by Cunningham, J.