Urinary hydroxyproline and serum alkaline phosphatase in sickle cell disease.

Serum alkaline phosphatase, alkaline phosphatase isoenzymes, and urinary hydroxyproline excretion were studied in 20 young adult sickle cell patients and 58 matching normal controls. Total alkaline phosphatase was significantly higher in the sickle cell patients than in controls. Heat inactivation test and isoenzyme electrophoresis indicated that bone is the predominant isoenzyme in patients. Hydroxyproline excretion was significantly higher in the sickle cell patients than in controls. Serum total alkaline phosphatase correlated well with urinary hydroxyproline excretion in sickle cell patients (r = 0.73). Both alkaline phosphatase and hydroxyproline increased with age in the sickle cell patients. This study suggests that delayed growth and/or bone destruction may contribute to the elevated levels of alkaline phosphatase and urinary hydroxyproline.     

Age and sex dependency of the biochemical indices of bone remodelling

The values for the bone isoenzyme of serum alkaline phosphatase peak in the first two years of age, between 6 and 7 years of age, before the end of puberty and in the postmenopause. A population between the ages of 29 and 45 provides a reference population to which all other age groupings can be compared. A significant positive correlation was found between bone isoenzyme of serum alkaline phosphatase and urinary hydroxyproline excretion in children as well as after puberty. However, in the children the urinary hydroxyproline excretion was significantly higher when compared with the bone isoenzyme of alkaline phosphatase. A significant positive correlation was found between the bone isoenzyme of alkaline phosphatase and plasma tartrate-resistant acid phosphatase, irrespective of age and sex. The biochemical indices of bone remodelling correlated significantly with the growth rate in children and adolescents. The results are in good agreement with the concept of the coupling of bone formation to bone resorption.     

Relationship of the activity of the bone isoenzyme of serum alkaline phosphataseto urinary hydroxyproline excretion in metabolic and neoplastic bone diseases

A significant correlation between the activity of the bone isoenzyme or serum alkaline phosphatase and the urinary hydroxyproline excretion in osteomalacia, osteoporosis, primary hyperparathyroidism with osteodystrophy, Paget's disease, secondary bone tumours, and in a control group was found (P less than 0.001). This close correlation was not observed between these variables in patients with active acromegaly. Diagnosis determined from these indices of formation and turnover of bone matrix agreed with that established by histological and histochemical examination of bone, by X-ray investigation of the skeleton, and by the radionuclear 85Sr test. The relationship between the activity of bone isoenzyme and urinary hydroxyproline excretion differed in metabolic bone diseases with a high bone turnover, in patients with osteoporosis and in patients with early osteoclastic bone metastases.     

Biochemical measurements in Paget''s disease of bone

In a group of eighty-three patients with untreated Paget's disease of bone, plasma alkaline phosphatase activity (AP), plasma non-protein-bound hydroxyproline concentration (PHP) and urinary excretion rate of total hydroxyproline (THP) were closely correlated with each other but not with fasting plasma concentrations of calcium or inorganic phosphate. Probit plots of AP and THP showed log-normal distributions overlapping the normal ranges.     

Measurements of whole body retention of diphosphonate and other indices of bone metabolism in 125 normals: dependency on age, sex and glomerular filtration

Measurements of 24-h whole body retention of 99m-Tc-MDP (WBR) has been performed in 125 normal volunteers, together with determinations of serum alkaline phosphatase, urinary hydroxyproline excretion and creatinine clearance. WBR decreased slightly from the 3rd to the 4th decade, after which it increased gradually in the older age-groups. Serum alkaline phosphatase followed an identical pattern, while the urinary hydroxyproline excretion demonstrated a marked but temporary rise in the post-menopausal age-groups. Finally, the creatinine clearance decreased gradually in the older age groups. Analysis of variance demonstrated that WBR varied independently with serum alkaline phosphatase and creatinine clearance, while no relationship between WBR and the hydroxyproline excretion was found. It seems likely that the increasing retention of diphosphonate in elderly persons reflects rising osteoblastic activity as well as decreasing glomerular filtration.     

Clinical and biological effects of low doses of (3 amino-1 hydroxypropylidene)-1,1-bisphosphonate (APD) in Paget''s disease of bone

Abstract. At doses varying between 500 and 1500 mg/day, the disodium salt of (3 amino-1 hydroxypro-pylidene)-1,1-bisphosphonate (APD) has been shown to induce rapid and complete biochemical responses in Paget's disease of bone, but a short-lived and self-limited fever has been observed in 30–40% of patients. The present study is an open and unrandomized trial, performed to explore the use of lower doses of APD (250 and 50 mg/day) given for 6 months to fourteen patients suffering from Paget's disease of bone. Subjective clinical improvement and complete biochemical remission (as assessed by normalization of urinary hydroxyproline excretion and plasma alkaline phosphatase concentration) was observed in ten out of eleven patients given 250 mg/day, but fever did not occur. Thus, a significant decrease in urinary hydroxyproline excretion was noted as early as 15 days after the beginning of the treatment (from 4.5 to 1.3 μmol/l GF; P > 0.05) whereas plasma alkaline phosphatase concentration fell significantly only 1 month later (from 330 to 195 IU/ml; P > 0.05). Both mean (± SEM) urinary hydroxyproline excretion (0.98 ± 0.08 μmol/l GF) and plasma alkaline phosphatase concentration (70 ± 8 IU/ml) were in the normal range after 6 months of treatment. Plasma calcium and phosphorus concentration, urinary calcium excretion and TmP/GFR decreased whereas plasma immunoreactive parathyroid hormone concentration increased. These changes were statistically significant (P < 0.05) only transiently and disappeared at the time when bone formation (assessed by plasma alkaline phosphatase) returned to normal. An increase in total body retention of ⁴⁷Ca during the second month of treatment was documented in five patients (P < 0.05), compatible with a positive calcium balance. Plasma immunoreactive calcitonin did not change (P > 0.05). After 6 months of treatment at 50 mg/day, APD also induced significant decreases in urinary hydroxyproline excretion (from 4.23 to 1.73 μmol/l GF; P < 0.05) and plasma alkaline phosphatase concentration (from 360 to 162 IU/ml; P < 0.05) but biochemical remissions were not complete and these two variables were significantly higher than the corresponding values in the group of patients receiving 250 mg APD/day. Less marked changes were also noted in all the other variables measured. Biological and clinical tolerance was good in all the patients. It is concluded that 250 mg/day of APD is an effective dose schedule, capable of reducing to normal levels the indirect indices of bone turn-over, but in contrast with higher doses, without inducing fever.     

Bone isoenzyme of serum alkaline phosphatase in acromegaly.

In 37 patients with active acromegaly and in 15 patients with inactive acromegaly, activity of bone isoenzyme of serum alkaline phosphatase correlated (P less than 0.001) with serum concentration of immunoreactive growth hormone. By using stepwise regression analysis, the predication of serum growth hormone values based on serum levels of bone isoenzyme of serum alkaline phosphatase, gamma-glutamyl transferase and calcium in these patients with acromegaly was within 1 S.D. range in 37 patients and in only 2 patients was it out of 2 S.D. range. By using discriminant analysis, based on bone and liver isoenzymes of serum alaline phosphatase and urinary hydroxyproline excretion, 87%, 60% and 97% of the classification of patients with active and inactive acromegaly and healthy adults, respectively, was correct. The multivariate approach offers a quantitative appraisal of the biochemical parameters of peripheral growth hormone action used as an indicator of growth hormone concentration in patients with acromegaly.     

Effects of 1-Sar-8-Ile-angiotensin II on urinary prostaglandin excretion in patients with essential hypertension

To investigate the interaction between the renin angiotensin aldosterone system and the renal prostaglandin (PG), urinary excretion of PGE, urinary excretion of main urinary metabolite (MUM) of PGF2a, urinary excretion of aldosterone, and plasma renin activity were measured before and after infusion of 1-Sar-8-Ile-Angiotensin II, a specific competitive inhibitor of angiotensin II, in 18 patients with essential hypertension under normal and low sodium diets. The values of urinary sodium excretion in these patients before the infusion of the peptide were 160.8 +/- 13.3 and 27.0 +/- 2.7 mEq per day on normal and low sodium diet, respectively. On normal sodium diet, urinary excretion of PGE was found to correlate with the level of plasma renin activity before the infusion (r = 0.6977, p less than 0.01), and it was decreased slightly from 0.37 +/- 0.05 ng/min to 0.26 +/- 0.04 ng/min after the infusion of the antagonist. On low sodium diet, urinary excretion of PGE was not significantly changed by the infusion of the peptide and showed no correlation with the level of plasma renin activity before the infusion, while urinary excretion of PGE showed a significant correlation with the excretion of urinary aldosterone (r = 0.6719, p less than 0.02). Excretion of PGF2aMUM decreased after the infusion of this peptide on both sodium diets, but the changes were not statistically significant. The present data suggest that angiotensin II influences the synthesis or release of renal PG in patients with essential hypertension on normal sodium diet, but not when they are on low sodium diet.     

Multiple forms of alkaline phosphatase in plasma of hemodialysis patients.

We used quantitative assays to measure the activity of the bone, liver, and intestinal forms of alkaline phosphatase in plasma in 75 patients with endstage chronic renal failure undergoing hemodialysis. The results were correlated with radiological and other biochemical indices of bone disease and with biochemical indices of liver disease. The total activity of alkaline phosphatase in plasma increased in 28 patients. In 10 of these patients, nine of whom had increased activity of gamma-glutamyltransferase in plasma, the increase in total activity of alkaline phosphatase was from the liver isoenzyme alone (nine patients) or from the liver and bone isoenzymes together (one patient). Intestinal alkaline phosphatase in plasma, although greater than 23 U/L in eight patients, was solely responsible for the increase in total alkaline phosphatase in one patient (who had normal gamma-glutamyltransferase). Bone alkaline phosphatase in plasma was increased in 25 patients, seven of whom had normal total alkaline phosphatase, and was closely correlated (r = 0.78) with osteocalcin concentration in plasma, which was increased in a much greater proportion of patients (99%). Both total and bone alkaline phosphatase were correlated with parathyrin in plasma (r = 0.46 and 0.50, respectively) and with osteocalcin (r = 0.60 and 0.78, respectively). Osteocalcin and bone alkaline phosphatase, but not parathyrin, decreased with age, implying that the skeletal response to parathyrin may be age dependent. In patients with increased total alkaline phosphatase undergoing hemodialysis, the concurrent measurement of gamma-glutamyltransferase may help identify whether the enzyme increase originates from the liver or bone, but this approach wrongly identified the source of the increase in three of 28 patients. Therefore, we recommend a separate measurement of the bone isoenzyme of alkaline phosphatase.     

Metabolic and calcium kinetic studies in idiopathic hypercalciuria

Calcium balances and calcium kinetic studies using ⁴⁷Ca were performed in nine male patients with idiopathic hypercalciuria and in three normal male subjects. A sharp reduction in calcium intake in eight patients with idiopathic hypercalciuria caused a decrease in urinary calcium excretion, the latter remaining elevated above that reported for normal subjects on a low calcium diet. The hypercalciuric patients had an enlarged miscible calcium pool size, an increased calcium turnover rate, increased bone formation and bone resorption rates, and an elevated true intestinal calcium absorption rate, the increase of the latter three parameters being proportional to the increase of the turnover rate. The fraction of the calcium turnover rate excreted in the urine was elevated whereas that constituted by the endogenous fecal calcium excretion was decreased. Arguments are presented for the concept that the primary abnormality in idiopathic hypercalciuria is neither renal calcium hyperexcretion nor intestinal calcium hyperreabsorption, but a more fundamental disturbance in calcium metabolism of as yet unknown cause, leading to a high calcium turnover.     

Electrolyte and collagen content of rat heart in chronic Mg-deficiency and stress (author's transl)

In chronic Mg-deficiency, there is a decrease in the content of Mg and K+ in the heart muscle. The contents of Ca, Na+ and hydroxyproline increase. There is also an increased urinary excretion of adrenaline and especially noradrenaline. With simultaneous noise stress, these changes are even greater. The magnitude of the changes increases with the degree of Mg-deficiency. The changes in the contents of Na+, K+, Ca, Mg and hydroxyproline correlate with the excretion of noradrenaline. The increase in the collagen content can be verified by electron microscopy, and it is due to a stimulation of the fibrocytes.     

EHDP (ethylidene-1-hydroxy-1,1-diphosphonate) in the treatment of polyosseous Paget disease

13 patients with polyosseous Paget's disease were treated for a mean period of 7 months with disodium etidronate (EHDP, ethylidene-1-hydroxy-1,1-diphosphonate); the average daily dosage was 5 mg/kg body weight. Subjectively, all patients reported a considerable improvement, in particular with regard to pain. Objectively, a significant decrease in plasma alkaline phosphatase activity and in urinary hydroxyproline excretion was observed. Bone scintigraphy showed a decreased activity of bone lesions after therapy, but no clear-cut regression was found radiologically. No serious side-effects were observed during treatment with EHDP and oral administration of the drug proved to be advantageous.     

Effect of oophorectomy and calcium deprivation on bone mass in the rat

1. The effects of a low calcium diet and of oophorectomy, separately and together, on cortical and trabecular bone mass, have been examined in mature female rats. 2. Calcium deprivation caused a significant decrease of weight, cortical cross-sectional area and ratio of cortical to total area in the femur, it significantly reduced the volume of trabecular bone and increased the percentage of osteoid surface in the tail vertebrae, and in addition increased the urinary excretion of phosphate and, initially, of hydroxyproline. 3. Oophorectomy caused similar though smaller changes in trabecular bone and urine, whereas the effects of oophorectomy on cortical bone were greater on a low calcium intake than on a normal intake. 4. The ash weight of the femora, expressed as a percentage of the total dry weight, was unaffected by calcium deprivation or oophorectomy alone but was significantly reduced when the two occurred together. 5. The percentage of resorption surfaces in the vertebrae tended to increase on the low calcium diet and after oophorectomy on the normal diet but decreased after oophorectomy on a low calcium diet. 6. It is concluded that oophorectomy and calcium deficiency each reduce bone mass in the adult rat but the greatest effect is seen when they are combined.     

Clinical significance of free plasma hydroxyproline measurement in metabolic bone disease

Free hydroxyproline was measured in plasma of 67 normal subjects and in 70 patients with bone disease including primary hyperparathyroidism (n = 19), osteoporosis (n = 18), Paget's disease (n = 14), cancer involving bone (n = 8), chronic renal failure (n = 6), and osteomalacia (n = 6), and osteomalacia (n = 5). A good correlation was found between plasma and urinary values of the amino acid in normal subjects (r = 0.66; p less than 0.001). In patients with skeletal disorders a highly significant direct correlation was observed between free plasma hydroxyproline on the one hand and urinary hydroxyproline (r = 0.92; p less than 0.001) and serum alkaline phosphatase activity (r = 0.86; p less than 0.001) on the other, even though there were a few examples of dissociations among these parameters. Free plasma hydroxyproline decreased in the patients with Paget's disease following chronic administration of salmon calcitonin. Following successful parathyroidectomy, free plasma levels of hydroxyproline decreased in all the cases studied. Measurement of free plasma hydroxyproline thus appears to provide a specific index of bone metabolism that may be usefully employed as an alternative to the assay of other markers of bone turnover.     

The Hypercalciurias CAUSES, PARATHYROID FUNCTIONS, AND DIAGNOSTIC CRITERIA

The causes for the hypercalciuria and diagnostic criteria for the various forms of hypercalciuria were sought in 56 patients with hypercalcemia or nephrolithiasis (Ca stones), by a careful assessment of parathyroid function and calcium metabolism. A study protocol for the evaluation of hypercalciuria, based on a constant liquid synthetic diet, was developed. In 26 cases of primary hyperparathyroidism, characteristic features were: hypercalcemia, high urinary cyclic AMP (cAMP, 8.58+/-3.63 SD mumol/g creatinine; normal, 4.02+/-0.70 mumol/g creatinine), high immunoreactive serum parathyroid hormone (PTH), hypercalciuria, the urinary Ca exceeding absorbed Ca from intestinal tract (Ca(A)), high fasting urinary Ca (0.2 mg/mg creatinine or greater), and low bone density by (125)I photon absorption. The results suggest that hypercalciuria is partly secondary to an excessive skeletal resorption (resorptive hypercalciuria). The 22 cases with renal stones had normocalcemia, hypercalciuria, intestinal hyperabsorption of calcium, normal or low serum PTH and urinary cAMP, normal fasting urinary Ca, and normal bone density. Since their Ca(A) exceeded urinary Ca, the hypercalciuria probably resulted from an intestinal hyperabsorption of Ca (absorptive hypercalciuria). The primacy of intestinal Ca hyperabsorption was confirmed by responses to Ca load and deprivation under a metabolic dietary regimen. During a Ca load of 1,700 mg/day, there was an exaggerated increase in the renal excretion of Ca and a suppression of cAMP excretion. The urinary Ca of 453+/-154 SD mg/day was significantly higher than the control group's 211+/-42 mg/day. The urinary cAMP of 2.26+/-0.56 mumol/g creatinine was significantly lower than in the control group. In contrast, when the intestinal absorption of calcium was limited by cellulose phosphate, the hypercalciuria was corrected and the suppressed renal excretion of cAMP returned towards normal. Two cases with renal stones had normocalcemia, hypercalciuria, and high urinary cAMP or serum PTH. Since Ca(A) was less than urinary Ca, the hypercalciuria may have been secondary to an impaired renal tubular reabsorption of Ca (renal hypercalciuria). Six cases with renal stones had normal values of serum Ca, urinary Ca, urinary cAMP, and serum PTH (normocalciuric nephrolithiasis). Their Ca(A) exceeded urinary Ca, and fasting urinary Ca and bone density were normal. The results support the proposed mechanisms for the hypercalciuria and provide reliable diagnostic criteria for the various forms of hypercalciuria.     

Thyrocalcitonin, EGTA, and Urinary Electrolyte Excretion

The infusion of thyrocalcitonin (TCT) into thyroparathyroidectomized rats, given either no exogenous parathyroid hormone or a constant infusion of this hormone, leads to a transient phosphaturia and a decreased excretion of urinary magnesium, calcium, and hydroxyproline without a change in glomerular filtration rate. The changes in phosphate excretion may be due to a direct effect of the hormone upon renal tubular function or they may be a consequence of the fall in plasma calcium brought about by the action of TCT upon bone. In support of this latter alternative is the fact that the infusion of sodium ethylenebis-oxyethylenenitrilotetra?cetic acid (EGTA, a specific chelator of calcium) also leads to phosphaturia presumably as a consequence of hypocalcemia. However, EGTA infusion leads to enhanced urinary hydroxyproline excretion and sustained phosphaturia. These latter observations are interpreted to mean that alterations in the local ionic environment of osteolytic cells lead to changes in their activity and constitute a local regulatory system whose activity is modulated by the hormones, thyrocalcitonin and parathyroid hormone.     

Urinary excretion of nitrogenous and non-nitrogenous compounds in the chronic ethanol-fed rat.

1. The metabolic consequences of chronic ethanol feeding was investigated by assay of urinary metabolites. Male Wistar rats were fed a liquid diet containing 35% of total energy as ethanol or isovolumetric, isocaloric and isonitrogenous amounts of the same diet in which ethanol was substituted by isocaloric glucose (controls). 2. At 6 weeks the entire skeletal muscle mass was reduced by approximately 20%. The urinary excretion of nitrogen, urea and uric acid increased by between 23 and 128%. Urinary creatinine excretion was not significantly altered. 3. Urinary excretion of magnesium was significantly increased by 43%. Urinary excretion of sodium, potassium, calcium and phosphate was increased slightly (i.e. 5-22%), but this change was not statistically significant. 4. Proton n.m.r. spectroscopic analysis showed that ethanol feeding reduced the urinary excretion of citrate and 2-oxoglutarate (by approximately 50%), suggesting decreased citric acid cycle activity. There was an increased excretion of alanine (44%), but excretion of succinate and acetate was not significantly altered. Ethanol in the urine of ethanol-fed rats comprised approximately 2% of total ethanol intake and less than 1% of total energy intake. 5. Lactose was detectable in urine of ethanol-fed rats, but not in control rats, reflecting the reported decreased intestinal lactase activity and increased gut permeability in alcoholics. Urinary galactose excretion decreased by 41%, but relatively large increases in lactate excretion (50%) did not achieve statistical significance. 6. It was concluded that chronic ethanol feeding causes disturbances in whole-body nitrogen homoeostasis and alterations in intermediary metabolism.     

Alkaline phosphatase isoenzymes in the plasma of preterm and term infants: serial measurements and clinical correlations

Serial measurements of the bone and fetal intestinal isoenzymes of alkaline phosphatase (EC 3.1.3.1) in the plasma of 43 term and 43 preterm infants, from birth to six weeks later, indicate that the bone isoenzyme gradually increases over this period in both preterm and term infants fed with unsupplemented commercial formulas. Preterm babies given formula supplemented with calcium (with or without additional phosphate) had significantly lower bone isoenzyme activities for most of the study period. The concentrations of fetal intestinal isoenzyme increased, under the stimulation of milk feeding, from generally undetectable at birth to a peak during the first two weeks postpartum, and then declined. This increase was highly significantly negatively correlated with gestational age, the preterm infants having a much higher and more prolonged increase in this isoenzyme than did term infants. Unlike the adult isoenzyme, fetal intestinal alkaline phosphatase in plasma showed no relationship with blood group status.     

Plasma aldosterone, cortisol and electrolyte concentrations in physical exercise after magnesium supplementation

Plasma aldosterone, cortisol, sodium (Na), potassium (K), calcium (Ca), magnesium (Mg) as well as urine and sweat Na, K, Ca and Mg concentrations were measured in nine male healthy persons during an one hour ergometer exercise before and after a fourteen day magnesium aspartate (Mg) supplementation. The usual aldosterone and cortisol increase during exercise was not observed and cortisol concentration was significantly lower after Mg supplementation. Na and K in plasma increased during the exercise; these changes were not affected by Mg. The Mg concentration was elevated in plasma and erythrocytes after Mg supplementation. During the ergometer course plasma Mg was unchanged but decreased significantly in the red blood cells. Mg and K concentration in sweat decreased during the exercise. No influence of Mg on urinary electrolyte excretion was observed.     

Two new methods for separating and quantifying bone and liver alkaline phosphatase isoenzymes in plasma

We describe two new methods for the separation and quantification of the bone and liver isoenzymes of alkaline phosphatase (EC 3.1.3.1) in plasma. In the first, we use wheat-germ lectin to precipitate the bone isoenzyme. About 80% of this, but minimal liver isoenzyme, is precipitated. The activity of the bone isoenzyme is calculated from measuring the alkaline phosphatase activity in the precipitate, that of liver alkaline phosphatase by subtracting the activity of the bone isoenzyme from total alkaline phosphatase activity. The liver fraction will also contain biliary, intestinal, and placental alkaline phosphatase if these are present in the original plasma, but correction for such activity is readily made. In the second method, samples are separated on cellulose acetate membranes that, before electrophoresis, have been soaked in buffer containing wheat-germ lectin. The bone isoenzyme is retarded and clearly separated from the liver fraction, allowing these isoenzymes to be quantified by densitometry. Both methods are rapid, reproducible, and suitable for use in the diagnostic laboratory.