- Email: [email protected]
P1. Rates of vertebral bone loss measured using two different manufacturers' densitometers
Abstracts from the Spring Meeting, March 1994
132 consistent with a role for mechanical bone formation to bone resorption.
forces in the coupling of
Pl. Rates of vertebral bone loss measured using two different manufacturers’ densitometers NFA Peel, R Eastell Depnrtmerrt of Human Metabolism and Clinical Biochemistry, Ut~iversity of Sheffield, Clinical Sciences Centre, Northern General Hospital, Sheffield
It is unknown whether rates of bone loss measured using different manufacturer’s densitometers are comparablb. Our aim was to determine whether the rate of bone loss from the lumbar spine (LS) was the same measured using a Lunar DPX and a Hologic QDR 1000/W. We measured bone mineral density (BMD) in 17 postmenopausal women at 0 and 2 years. The mean rate of loss of LS-BMD was the same using both densitometers (Lunar, -0.0039 f 0.023 g/cmz/year; Hologic, -0.0039 f 0.012 g/cmz/year). The variance of the rate of loss of LSBMD was greater for Lunar than Hologic (F = 4.1, 95% CI 1.5 to 11.3). We used the approach described by Bland and Altman (Lancet 1986:307) to relate the difference in the rate of loss estimate between the two densitometers to the mean rate of loss (r = 0.64, P = 0.005). This sienificant correlation could be exulained if the precision ‘error of”LSBMD measurements by L&ar DPX was twice that by Hologic QDR 1000/W. This finding raises concerns about combining results for the rate of bone loss measured using different manufacturer’s densitometers in multi-centre studies.
P2. The effect of interleukin 6 (1L6) on osteoclastic resorption in human bone marrow cultures M Stow, AM Flanagan St Mary’s Hospital Medical School, London W2 1 PC
bone
IL6 has been implicated as a cytokine which pLays a major role in the generation of human osteoclast-like cells from human bone marrow in vitro. However bone resorption, a function unique to the osteoclast, has never been quantified in these cultures. Human bone marrow stroma was used as a feeder layer on bone slices and recharged with non-adherent bone marrow cells in the presence and absence of 1,25 dihydroxyvitamin D3 (vit D3) with and without IL6. We found that bone resorption was significantly increased in the presence of vit D3, an effect that could not be reproduced with IL6 IL6 at 100 ng/ml actively inhibited the stimulatory effect of vit D3. Bone resorption was never observed when non-adherent cells were cultured alone in the presence of vit D3 or IL6. These results suggest that it is not the primary role of IL6 to increase bone resorption. But it is not possible to tell from these studies if the inhibitory effect is due to reduced numbers of osteclasts or whether mature forms are inhibited from resorbing bone.
P3. Quantitative studies of cell proliferation in bone formed during leg-lengthening G Li, SJ Gregg-Smith, AHRW Simpson, J Bradley, JT Triffitt’ NufJild Department of Orthopaedic Surgery and lMRC Bone Research Laboratory, Nuffield Orthopaedic Centre NHS Trust, Oxford OX3 7LD To assess the effect of mechanical forces on bone tissue formation, a cell proliferation marker, PC10 antibody, was used to identify proliferating cells. New Zealand white rabbits had mid-tibia1 osteotomies, which were stabilised with external fixators and distracted at 0.33, 0.67, 1.33 and 2.77 mm/day to 20% of the tibia1 length. The distraction zone (DZ) was harvested and histological sections were immunostained with PClO. The DZ
was subdivided by morphology into Primary Mineralization Zone (PMZ), Fibrous Zone (FZ) and Newly Formed Bone Zone (NBZ). Ratios of positive stained nuclei were counted in multiple areas of the three sub-zones in each histological section. Results are mean positive staining index (PSI) + SD. Rate mm/day 0.33 0.67 1.33 2.77
FZ pSI(%) 30.7 f 3.4 31.5 + 3.7 53.3 f 7.8 44.9 * 5.4
PMZ PSI(%) 21.2 f 5.5 36.4 f 6.0 65.3 +.5.3 61.2 f 5.9
NBZ PSI(%) 20.0f 7.8 33.9* 3.0 60.7 + 2.3 47.9 & 4.2
Mean PSI(%) 22.2 f 3.9 34.5 f 1.7 61.7 k 4.3 51.3 f 6.7
We conclude that cell proliferation is dependent on the rate of distraction. Highest PSI were seen in each zone at a distraction rate of 1.33 mm/day and it does not increase further at 2.77 mm/day.
P4. Characteriaation of fracture healing using a chorioallantoic membrane model RS Bale, JG Andrew University Department of Orthopaedic Surgery, Hope Hospital, Salford The use of the chorioallantoic membrane (CAM) model of fracture healing has been described by several workers and has been proposed as a relevant model of bone repair. The advantages of this model are that it is cheat. simule to use. the fracturevsite is readily accessible and can bi &&pulated and it avoids some of the ethical issues of using 1arEer animal models. The initial objective of our research was 70 &aracterise in detail the nature of the fracture healing seen when using this model. Fertiiised chick eggs were incubated for 12 and 18 days to provide the donor femora and 9 days to provide the recipient CAMS. Bones were also obtained from neonatal chicks. Healing fractures were harvested on consecutive days after fracturing Using this technique we have not observed a callus response to fracture despite examining bones from the different ages of chick There is a minimal inflammatory response to fracture and granulation tissue is not seen. The bone heals by periosteal repair with subperiosteal bone formation to bridge the fracture gap. Cells at the fracture site do not appear to participate directly in the repair process. The existence of callus after fracture in this model is controversial. It is known from chimeric studies that the osteoblasts of the graft originate from the donor bone but that the osteoclasts and the marrow components stem from the recipient egg. We are currently studying the reasons for the lack of callus response in this model. The CAM model, whilst having advantages and research potential, cannot be said to represent the repair mechanisms encountered in normal adult fracture healing.
PS. Serum tartrate resistant acid phosphatase as a biochemical marker of bone resorption PA Kyd, HM Snook, A Faimey Bone Research Group, Unit of Metabolic Medicine, St. Mary’s Hospital Medical School, London W2 IPG Serum tartrate resistant acid phosphatase (TRAP), the type 5b isoenzyme produced by osteoclasts has been evaluated as a marker of bone resorption. This spectrophotometric TRAP assay is simple and cheap, and within-batch and between batch precision is 15% and ~10% respectively. However serum TRAP activity decreases when stored frozen at -20°C; long term storage of samples (more than 3 months) must be at -70”. The reference range derived from normal volunteers (n = 40, aged 18 - 65 yrs) was 7.0 - 13.Ou/L
132 consistent with a role for mechanical bone formation to bone resorption.
forces in the coupling of
Pl. Rates of vertebral bone loss measured using two different manufacturers’ densitometers NFA Peel, R Eastell Depnrtmerrt of Human Metabolism and Clinical Biochemistry, Ut~iversity of Sheffield, Clinical Sciences Centre, Northern General Hospital, Sheffield
It is unknown whether rates of bone loss measured using different manufacturer’s densitometers are comparablb. Our aim was to determine whether the rate of bone loss from the lumbar spine (LS) was the same measured using a Lunar DPX and a Hologic QDR 1000/W. We measured bone mineral density (BMD) in 17 postmenopausal women at 0 and 2 years. The mean rate of loss of LS-BMD was the same using both densitometers (Lunar, -0.0039 f 0.023 g/cmz/year; Hologic, -0.0039 f 0.012 g/cmz/year). The variance of the rate of loss of LSBMD was greater for Lunar than Hologic (F = 4.1, 95% CI 1.5 to 11.3). We used the approach described by Bland and Altman (Lancet 1986:307) to relate the difference in the rate of loss estimate between the two densitometers to the mean rate of loss (r = 0.64, P = 0.005). This sienificant correlation could be exulained if the precision ‘error of”LSBMD measurements by L&ar DPX was twice that by Hologic QDR 1000/W. This finding raises concerns about combining results for the rate of bone loss measured using different manufacturer’s densitometers in multi-centre studies.
P2. The effect of interleukin 6 (1L6) on osteoclastic resorption in human bone marrow cultures M Stow, AM Flanagan St Mary’s Hospital Medical School, London W2 1 PC
bone
IL6 has been implicated as a cytokine which pLays a major role in the generation of human osteoclast-like cells from human bone marrow in vitro. However bone resorption, a function unique to the osteoclast, has never been quantified in these cultures. Human bone marrow stroma was used as a feeder layer on bone slices and recharged with non-adherent bone marrow cells in the presence and absence of 1,25 dihydroxyvitamin D3 (vit D3) with and without IL6. We found that bone resorption was significantly increased in the presence of vit D3, an effect that could not be reproduced with IL6 IL6 at 100 ng/ml actively inhibited the stimulatory effect of vit D3. Bone resorption was never observed when non-adherent cells were cultured alone in the presence of vit D3 or IL6. These results suggest that it is not the primary role of IL6 to increase bone resorption. But it is not possible to tell from these studies if the inhibitory effect is due to reduced numbers of osteclasts or whether mature forms are inhibited from resorbing bone.
P3. Quantitative studies of cell proliferation in bone formed during leg-lengthening G Li, SJ Gregg-Smith, AHRW Simpson, J Bradley, JT Triffitt’ NufJild Department of Orthopaedic Surgery and lMRC Bone Research Laboratory, Nuffield Orthopaedic Centre NHS Trust, Oxford OX3 7LD To assess the effect of mechanical forces on bone tissue formation, a cell proliferation marker, PC10 antibody, was used to identify proliferating cells. New Zealand white rabbits had mid-tibia1 osteotomies, which were stabilised with external fixators and distracted at 0.33, 0.67, 1.33 and 2.77 mm/day to 20% of the tibia1 length. The distraction zone (DZ) was harvested and histological sections were immunostained with PClO. The DZ
was subdivided by morphology into Primary Mineralization Zone (PMZ), Fibrous Zone (FZ) and Newly Formed Bone Zone (NBZ). Ratios of positive stained nuclei were counted in multiple areas of the three sub-zones in each histological section. Results are mean positive staining index (PSI) + SD. Rate mm/day 0.33 0.67 1.33 2.77
FZ pSI(%) 30.7 f 3.4 31.5 + 3.7 53.3 f 7.8 44.9 * 5.4
PMZ PSI(%) 21.2 f 5.5 36.4 f 6.0 65.3 +.5.3 61.2 f 5.9
NBZ PSI(%) 20.0f 7.8 33.9* 3.0 60.7 + 2.3 47.9 & 4.2
Mean PSI(%) 22.2 f 3.9 34.5 f 1.7 61.7 k 4.3 51.3 f 6.7
We conclude that cell proliferation is dependent on the rate of distraction. Highest PSI were seen in each zone at a distraction rate of 1.33 mm/day and it does not increase further at 2.77 mm/day.
P4. Characteriaation of fracture healing using a chorioallantoic membrane model RS Bale, JG Andrew University Department of Orthopaedic Surgery, Hope Hospital, Salford The use of the chorioallantoic membrane (CAM) model of fracture healing has been described by several workers and has been proposed as a relevant model of bone repair. The advantages of this model are that it is cheat. simule to use. the fracturevsite is readily accessible and can bi &&pulated and it avoids some of the ethical issues of using 1arEer animal models. The initial objective of our research was 70 &aracterise in detail the nature of the fracture healing seen when using this model. Fertiiised chick eggs were incubated for 12 and 18 days to provide the donor femora and 9 days to provide the recipient CAMS. Bones were also obtained from neonatal chicks. Healing fractures were harvested on consecutive days after fracturing Using this technique we have not observed a callus response to fracture despite examining bones from the different ages of chick There is a minimal inflammatory response to fracture and granulation tissue is not seen. The bone heals by periosteal repair with subperiosteal bone formation to bridge the fracture gap. Cells at the fracture site do not appear to participate directly in the repair process. The existence of callus after fracture in this model is controversial. It is known from chimeric studies that the osteoblasts of the graft originate from the donor bone but that the osteoclasts and the marrow components stem from the recipient egg. We are currently studying the reasons for the lack of callus response in this model. The CAM model, whilst having advantages and research potential, cannot be said to represent the repair mechanisms encountered in normal adult fracture healing.
PS. Serum tartrate resistant acid phosphatase as a biochemical marker of bone resorption PA Kyd, HM Snook, A Faimey Bone Research Group, Unit of Metabolic Medicine, St. Mary’s Hospital Medical School, London W2 IPG Serum tartrate resistant acid phosphatase (TRAP), the type 5b isoenzyme produced by osteoclasts has been evaluated as a marker of bone resorption. This spectrophotometric TRAP assay is simple and cheap, and within-batch and between batch precision is 15% and ~10% respectively. However serum TRAP activity decreases when stored frozen at -20°C; long term storage of samples (more than 3 months) must be at -70”. The reference range derived from normal volunteers (n = 40, aged 18 - 65 yrs) was 7.0 - 13.Ou/L