- Email: [email protected]
P1. The expression of thyroid hormone receptors in human bone
688
Abstracts
Bone Vol. 19, No. 6 December 1996:685-701
who were taking part in a double-blind, placebo-controlled study of the efficacy of MTX in PBC. 12 women received treatment with MTX (7.5 mg weekly), and none received drugs known to affect bone metabolism. They were compared to 16 age-matched women from a population-based group (mean age 61.8 years). The patients did not differ from the controls with respect to height, weight, or years postmenopause. Bone mineral density (BMD) of the lumbar spine (LS), femoral neck (FN), total body (TB) and right hand were measured by DXA (Hologic QDR 1000/W) at baseline, and after 1 year in the PBC group and 2 years in the controls. The table shows the annual % rates of change in BMD at each site.
of positive cells and the intensity of staining were greatest for the 131 isoform, followed by al and 02 respectively. Our results demonstrate, for the first time, thyroid hormone receptor isoforms in human bone in situ and suggest that the skeletal actions of thyroid hormones are mediated via these receptors.
PBC No MTX P Total (n=15) (MTXv (n=27) no MTX) -1.45 0.8 -1.30 -3.45 0.8 -3.60 -2.06 0.7 -2.16 -1.70 0.6 -1.95
Department of Medicine and Therapeutics, University of Aberdeen Medical School, Foresterhill. Aberdeen AB9 2ZD
MTX (n-12) LS FN TB Hand
-1.12 -3.76 -2.28 -2.23
Controls (n=16) -0.45 -0.44 -0.52 -1.08
P (controls v PBC) 0.3 0.001 0.0002 0.2
We conclude that postmenopausal women with PBC have generalised acceleration of bone loss, and that MTX therapy in PBC does not affect the rate of bone loss. O14. B o n e loss o b s e r v e d in i n f l a m m a t o r y b o w e l disease is associated with suppression o f b o n e formation CL Lin, C Moniz, TJ Chambers, JWM Chow
King's College School of Medicine and Dentistry, lzmdon and St. George's Hospital Medical School, London SW170RE Although it is well established that idiopathic inflammatory bowel disease (IBD) is associated with osteoporosis, the pathogenesis of the bone loss is unclear. Therefore, we decided to investigate the bone loss and underlying pathogenetic mechanisms using a rat model of inflammatory bowel disease. Colitis was induced by intrarectal administration of 2,4,6-trinitrobenzenesulphonic acid (TNBS). Animals inspected 1 week after TNBS administration exhibited a mild colitis. After 3 weeks a florid transmural colitis was observed, followed by recovery of the bowel to near normal 6 weeks after induction of IBD. We found no significant loss of bone, or alteration in bone turnover, in animals killed 1 week after TNBS administration. There was a 33% loss of cancellous bone volume in animals killed 3 weeks after TNBS administration compared with control animals• This was associated with a marked suppression of the cancellous bone formation rate to less than 10% that of control pair-fed artimals. The increase in static parameters of bone resorption in the colitic animals was not statistically significant. The bone volume of animals sacrificed 6 weeks after induction of IBD remained low, but the bone formation rate was increased to 7-fold that of control animals, indicating recovery of the bone in parallel with the colon. Our data suggests that bone loss may occur rapidly in IBD, and that the bone loss is associated with suppression of bone formation.
PI. The expression o f thyroid h o r m o n e receptors in h u m a n bone EO Abu, A Homer, S Bord, VKK Chatter~ee, JE Compston
Department of Medicine, University of Cambridge Clinical School, Addenbrooke's Hospital, Cambridge CB2 2QQ Thyroid hormones have well-documented effects on bone turnover and studies with radiolabelled T3 have demonstrated specific and saturable binding sites in bone. Recently, isoform specific thyroid hormone receptors have been reported in rat osteosarcoma cell lines but their presence in human bone has not been reported. We have used immunolocalisation to study the expression of thyroid hormone receptors in human bone in situ. Ten , m cryostat sections of heterotopic or osteophytic bone were applied to APES coated slides. Sections for polyclonal antibodies (anti-ctl and anti-cz2), were fixed in neutral-buffered formalin and those for the monoclonal antibody (anti-131) were fixed in acetone. Immunostaining was performed using the standard ABC staining procedures with appropriate controls. TRAP staining was used to identify osteoclasts. The thyroid hormone receptor isoforrns al, cz2 and 131 were widely expressed by osteocytes, osteoblasts and chondrocytes. The number
P2. Expression of cytokines, growth factors and proto-oncogenes in renal b o n e disease: s t u d i e s u s i n g r e v e r s e - t r a n s c r i p t i o n polymerase chain reaction (RT/PCR) A Kumar, S H Ralston
Background: Renal osteodystrophy is characterised by marked abnormalities of bone turnover as the result of secondary hyperparathyroidism. Whilst PTH acts directly on bone, it may also act indirectly by stimulating production of local mediators in the bone micro-environment. In order to investigate this possibility, we studied expression of cytokines, growth factors and protooncogenes by RT/PCR in renal bone disease. Method: Patients with chronic renal failure (n=28) and normal subjects (n=17) were studied. RNA was extracted from transiliac bone biopsies in each group and cDNA prepared as template for PCR. Results: These are shown in the figure below. We found surprisingly little difference _ _ between the groups in number of patients ~.Fo~i showing expression of c.~yc i individual cytokines, TGr~~ growth factors, or protoIGF4 I cop I oncogenes. O f interest was TaAPr ~,~b the high frequency of c-Fos cTn rr and c-Myc expression TNF a ~ IIRer~l ,~-~r (-80%) in both groups, IL-1 b [ ~ ; however. The calcitonin ..... ¢=:::= receptor (CTR) was 0 20 40 60 80 100 expressed significantly •**,,~,,,~°., more often innormals (28% vs 3%; p<0.02) whereas tartrate resistant acid phosphatase (TRAP) mRNA was expressed more often in renal patients, especially those with raised serum PTH values (62% vs 17%; p<0.008). Conclusion: Renal osteodystrophy is associated with increased expression for TRAP and decreased expression of CTR, but no difference in expression of other factors• Renal osteodystrophy may largely result from direct effects of PTH on bone, rather than local induction of cytokines or growth factors. P3. Clinical evaluation of a radioimmunoassay for the m e a s u r e m e n t of d e o x y p y r i d i n o l i n e in urine and serum KE Naylor, A Blumsohn, RA Hannon, R Eastell
Department of Human Metabolism and Clinical Biochemistry, University of Sheffield, Clinical Sciences Centre, Northern General Hospital, Sheffield 55 7AU The aim of this study was to evaluate a new radioimmunoassay for free immunoreactive deoxypyridinoline (iFDpd-Nichols) in serum and urine, in comparison with another urine Dpd ELISA, (iFDpdMetra, Pyrilinks-D), and total Dpd by HPLC (Dpd-HPLC). These markers were compared in 4 clinical models: 1) patients with Paget's disease, n=14, treated with oral etidronate; 2) healthy pubertal girls, in comparison with premenopausal adult controls n=81; 3) early postmenopausal women, n=7, following HRT; 4) circadian changes in healthy premenopausal women, n=13. Study 1: Etidronate therapy x 6 months resulted in a 10.5% (+ 10.24 SEM) decrease from baseline for zl~)pd-Nichols, 40.3% (± 5.3 SEM) for iFDpd-Metra and 58.7% (+ 7.6 SEM) for Dpd-HPLC. Serum zl:Dpd-Nichols increased slightly in response to etidronate therapy, (12% increase, P--0.5). Study 2: In mid puberty (Tanner stages II and III), the level of iFDpd-Nichols was 2.3 times higher than adult controls, iFDpd-Metra 4.4 times higher and Dpd-HPLC 9.5 times higher. Study 3: HRT in postmenopausal women resulted in a 13.4% (+ 3.0 SEM) decrease at 6 months for iFDpd-Nichols, 30.6% (±4.2 SEM) for iFDpd-Metra and 45.0% (± 6.7 SEM) for Dpd-HPLC. Study 4: Circadian changes: iFDpd-Nichols did not show any significant circadian rhythm (amplitude <10%). The timing of the circadian
Abstracts
Bone Vol. 19, No. 6 December 1996:685-701
who were taking part in a double-blind, placebo-controlled study of the efficacy of MTX in PBC. 12 women received treatment with MTX (7.5 mg weekly), and none received drugs known to affect bone metabolism. They were compared to 16 age-matched women from a population-based group (mean age 61.8 years). The patients did not differ from the controls with respect to height, weight, or years postmenopause. Bone mineral density (BMD) of the lumbar spine (LS), femoral neck (FN), total body (TB) and right hand were measured by DXA (Hologic QDR 1000/W) at baseline, and after 1 year in the PBC group and 2 years in the controls. The table shows the annual % rates of change in BMD at each site.
of positive cells and the intensity of staining were greatest for the 131 isoform, followed by al and 02 respectively. Our results demonstrate, for the first time, thyroid hormone receptor isoforms in human bone in situ and suggest that the skeletal actions of thyroid hormones are mediated via these receptors.
PBC No MTX P Total (n=15) (MTXv (n=27) no MTX) -1.45 0.8 -1.30 -3.45 0.8 -3.60 -2.06 0.7 -2.16 -1.70 0.6 -1.95
Department of Medicine and Therapeutics, University of Aberdeen Medical School, Foresterhill. Aberdeen AB9 2ZD
MTX (n-12) LS FN TB Hand
-1.12 -3.76 -2.28 -2.23
Controls (n=16) -0.45 -0.44 -0.52 -1.08
P (controls v PBC) 0.3 0.001 0.0002 0.2
We conclude that postmenopausal women with PBC have generalised acceleration of bone loss, and that MTX therapy in PBC does not affect the rate of bone loss. O14. B o n e loss o b s e r v e d in i n f l a m m a t o r y b o w e l disease is associated with suppression o f b o n e formation CL Lin, C Moniz, TJ Chambers, JWM Chow
King's College School of Medicine and Dentistry, lzmdon and St. George's Hospital Medical School, London SW170RE Although it is well established that idiopathic inflammatory bowel disease (IBD) is associated with osteoporosis, the pathogenesis of the bone loss is unclear. Therefore, we decided to investigate the bone loss and underlying pathogenetic mechanisms using a rat model of inflammatory bowel disease. Colitis was induced by intrarectal administration of 2,4,6-trinitrobenzenesulphonic acid (TNBS). Animals inspected 1 week after TNBS administration exhibited a mild colitis. After 3 weeks a florid transmural colitis was observed, followed by recovery of the bowel to near normal 6 weeks after induction of IBD. We found no significant loss of bone, or alteration in bone turnover, in animals killed 1 week after TNBS administration. There was a 33% loss of cancellous bone volume in animals killed 3 weeks after TNBS administration compared with control animals• This was associated with a marked suppression of the cancellous bone formation rate to less than 10% that of control pair-fed artimals. The increase in static parameters of bone resorption in the colitic animals was not statistically significant. The bone volume of animals sacrificed 6 weeks after induction of IBD remained low, but the bone formation rate was increased to 7-fold that of control animals, indicating recovery of the bone in parallel with the colon. Our data suggests that bone loss may occur rapidly in IBD, and that the bone loss is associated with suppression of bone formation.
PI. The expression o f thyroid h o r m o n e receptors in h u m a n bone EO Abu, A Homer, S Bord, VKK Chatter~ee, JE Compston
Department of Medicine, University of Cambridge Clinical School, Addenbrooke's Hospital, Cambridge CB2 2QQ Thyroid hormones have well-documented effects on bone turnover and studies with radiolabelled T3 have demonstrated specific and saturable binding sites in bone. Recently, isoform specific thyroid hormone receptors have been reported in rat osteosarcoma cell lines but their presence in human bone has not been reported. We have used immunolocalisation to study the expression of thyroid hormone receptors in human bone in situ. Ten , m cryostat sections of heterotopic or osteophytic bone were applied to APES coated slides. Sections for polyclonal antibodies (anti-ctl and anti-cz2), were fixed in neutral-buffered formalin and those for the monoclonal antibody (anti-131) were fixed in acetone. Immunostaining was performed using the standard ABC staining procedures with appropriate controls. TRAP staining was used to identify osteoclasts. The thyroid hormone receptor isoforrns al, cz2 and 131 were widely expressed by osteocytes, osteoblasts and chondrocytes. The number
P2. Expression of cytokines, growth factors and proto-oncogenes in renal b o n e disease: s t u d i e s u s i n g r e v e r s e - t r a n s c r i p t i o n polymerase chain reaction (RT/PCR) A Kumar, S H Ralston
Background: Renal osteodystrophy is characterised by marked abnormalities of bone turnover as the result of secondary hyperparathyroidism. Whilst PTH acts directly on bone, it may also act indirectly by stimulating production of local mediators in the bone micro-environment. In order to investigate this possibility, we studied expression of cytokines, growth factors and protooncogenes by RT/PCR in renal bone disease. Method: Patients with chronic renal failure (n=28) and normal subjects (n=17) were studied. RNA was extracted from transiliac bone biopsies in each group and cDNA prepared as template for PCR. Results: These are shown in the figure below. We found surprisingly little difference _ _ between the groups in number of patients ~.Fo~i showing expression of c.~yc i individual cytokines, TGr~~ growth factors, or protoIGF4 I cop I oncogenes. O f interest was TaAPr ~,~b the high frequency of c-Fos cTn rr and c-Myc expression TNF a ~ IIRer~l ,~-~r (-80%) in both groups, IL-1 b [ ~ ; however. The calcitonin ..... ¢=:::= receptor (CTR) was 0 20 40 60 80 100 expressed significantly •**,,~,,,~°., more often innormals (28% vs 3%; p<0.02) whereas tartrate resistant acid phosphatase (TRAP) mRNA was expressed more often in renal patients, especially those with raised serum PTH values (62% vs 17%; p<0.008). Conclusion: Renal osteodystrophy is associated with increased expression for TRAP and decreased expression of CTR, but no difference in expression of other factors• Renal osteodystrophy may largely result from direct effects of PTH on bone, rather than local induction of cytokines or growth factors. P3. Clinical evaluation of a radioimmunoassay for the m e a s u r e m e n t of d e o x y p y r i d i n o l i n e in urine and serum KE Naylor, A Blumsohn, RA Hannon, R Eastell
Department of Human Metabolism and Clinical Biochemistry, University of Sheffield, Clinical Sciences Centre, Northern General Hospital, Sheffield 55 7AU The aim of this study was to evaluate a new radioimmunoassay for free immunoreactive deoxypyridinoline (iFDpd-Nichols) in serum and urine, in comparison with another urine Dpd ELISA, (iFDpdMetra, Pyrilinks-D), and total Dpd by HPLC (Dpd-HPLC). These markers were compared in 4 clinical models: 1) patients with Paget's disease, n=14, treated with oral etidronate; 2) healthy pubertal girls, in comparison with premenopausal adult controls n=81; 3) early postmenopausal women, n=7, following HRT; 4) circadian changes in healthy premenopausal women, n=13. Study 1: Etidronate therapy x 6 months resulted in a 10.5% (+ 10.24 SEM) decrease from baseline for zl~)pd-Nichols, 40.3% (± 5.3 SEM) for iFDpd-Metra and 58.7% (+ 7.6 SEM) for Dpd-HPLC. Serum zl:Dpd-Nichols increased slightly in response to etidronate therapy, (12% increase, P--0.5). Study 2: In mid puberty (Tanner stages II and III), the level of iFDpd-Nichols was 2.3 times higher than adult controls, iFDpd-Metra 4.4 times higher and Dpd-HPLC 9.5 times higher. Study 3: HRT in postmenopausal women resulted in a 13.4% (+ 3.0 SEM) decrease at 6 months for iFDpd-Nichols, 30.6% (±4.2 SEM) for iFDpd-Metra and 45.0% (± 6.7 SEM) for Dpd-HPLC. Study 4: Circadian changes: iFDpd-Nichols did not show any significant circadian rhythm (amplitude <10%). The timing of the circadian