04307nam a2200397 a 450000100080000000500110000800800410001910000230006024500720008326000420015530000100019750000270020752033170023465000200355165000100357165000160358165000150359765000100361265000160362265000150363865000090365365000100366265000110367265300240368365300170370765300220372465300140374665300230376065300240378365300190380765300220382665300170384865300100386565300170387565300170389213197342014-01-15       1997    bl uuuu  m   00u1 u    #d1 aNICODEMO, M. L. F.  aMeasurement and function of turnover markers in sheep and pig bone.  a[S.l.] : University of Aberdeenc1997  a204p.  aTese Doutorado. CNPGC.  aPlasma osteocalcin and the urinary pyridinium crosslinks, pyridinoline and deoxypyridinoline are widely used as markers of bone tumover though many aspects of their function and metabolism remain obscure. In this thesis several likely physiological determinants of the levels of these markers in blood, bone and urine were examined in sheep and in pigs. Measurement of osteocalcin proved a major problem, immunoassay being unreliable as a method for measuring the levels of this protein in bone. This necessitated the development of an RP-HPLC assay (Chapter 3) in which the protein was separated from bone extracts using a linear 4-60% acetonitrile gradient containing 0. I % TFA run over 45 mins at a flow rate of lml/min. The standard curve was linear over a range of 0.5-15.0 pg osteocalcin injected and with inter- and intra-assay coefficients of variation of 6.9 and 8.8 §lo respectively and with recovery of osteocalcin added to bone extracts averaging 102.7+ 6,16 %. In sheep, plasma osteocalcin levels, determined by ELISA, showed large between animal as well as variability over 24 hours but no evidence of consistent circadian rhythm and decreased with age. In bone, changes in osteocalcin level tended to parallel those for calcium, whereas levels of the pyridinium crosslinks tended to increase with age (Chapter 4). Neither group of markers were sufficiently sensitive to detect differences in bone tumover between lambs of similar age but growing at different rates (Chapter 5). At slaughter the faster growing lambs had heavier though less well mineralised bones but showed no change in the ratio of calcium/osteocalcin. The ratio of calcium/osteocalcin in bone was also unchanged in lambs fed diets deficient in phosphorus (Chapter 6) despite an overall decline in mineral and in plasma and bone osteocalcin contents, which appeared to support the view that plasma and bone osteocalcin contents, which appeared to support the view thatosteocalcin may play a functional role in the mineralization process. However, against this, pigs fed diets deficient in both calcium and phosphorus showed no change in either plasma or bone osteocalcin levels despite marked reductions in bone calcium content (Chapter 7). This suggests that the amount of osteocalcin deposited in bone is not simply related to the amount of mineral present and so calls into question whether it has a direct role in the mineralization process. Chapter 8 describes an experiment in which adult sheep were treated with one of the new bisphosphonates (Ibandronate) and its effects on the metabolism of both pyridinoline and deoxypyridinoline were monitored. Given at rates which have been shown to be effective in reducing bone resorption in humans, this compound had little effect on the overall rates of excretion of these  crosslinks but did alter the proportions excreted in free or peptide bound form. Another feature of this study was the high variability seen in the escretion of these markers in urine both between animals and between collection periods with coeficients of variation of 48 and 36 % for total pyridinoline and deoxypyridinoline and 62 and 50 % for their free fractions. Similar high variability has been reported by other workers and has clear implications in regard to the accuracy of the these markers.  abone metabolism  abones  aosteocalcin  aphysiology  aswine  aDiagnostico  aFisiologia  aOsso  aOvino  aSuíno  aBiochemical markers  aBone markers  aDeoxypiridinoline  aDiagnosis  aDioxidipirionolina  aMarcador bioquimico  aMarcador osseo  aMetabolismo osseo  aOsteocalcina  aOvine  aPiridinolina  aPyridinoline