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The effects of fluoride (F) on ectopic bone formation induced in rats by implants of demineralized cortical bone tissue were studied. Test rats received sodium fluoride (NaF), 6 mmol/liter in drinking water, and controls received fluoride-free water. Implant accumulations of tracer hydroxyproline ([3H](OH)P), 45Ca, and stable Ca were determined 24 h after injections of tritiated proline ([3H]P) and 45Ca, to estimate rates of collagen synthesis, mineralization, and net mineral mass, respectively. Conventional histology on undemineralized implant sections was done. Mineralized bone was first observed by implant histology, 2 weeks after implantation and continued to increase up to 8 weeks. A few chondrocytes were observed. Prior to bone formation, dense fibrous tissue was observed within the marrow space of the original implant. The rate of collagen synthesis peaked at 1 week, again at 3 weeks, and then continued at a slower rate up to 8 weeks. The rates of mineralization paralleled the rates of collagen synthesis between 2 and 8 weeks, indicating bone mineralization over this period. During the first 2 weeks after implantation no mineral deposition was observed. The initial peak of collagen synthesis without mineralization (0-2 weeks) indicates fibrous tissue formation and is in agreement with the histological analysis. Fluoride treatment increased rates of collagen synthesis during both the initial period of fibrous tissue formation and later bone formation. The ratio of mineralization rate to collagen synthesis rate (45Ca/[3H](OH)P) was decreased by fluoride throughout the 2-8 week period, but net mineral mass was comparable to control rats by 8 weeks, indicating that fluoride delays, but does not prevent, bone mineralization. Fluoride stimulation of both fibrous tissue and bone suggests effects on precursor cells, possibly cells of the immune system.

We have developed a localized noninvasive nuclear magnetic resonance (NMR) method for determining the accumulated bone fluoride content in human index fingers. Using a 27 MHz "split-ring" resonator probe, we measure the total fluoride (F) content within a 6.5 cm length of the distal end of the index finger from a calibrated determination of the intensity of the observed 19F free induction decay (FID) signal. Since fluoride impurities accumulate almost exclusively in bone mineral, the 19F resonance signal is broadened by rigid lattice magnetic dipole-dipole interactions, causing the FID signal to have a relatively short lifetime of approximately 75 microseconds. This short signal lifetime, plus the weakness of the signal strength prevents conventional magnetic resonance imaging equipment from observing the 19F in bone mineral. Nevertheless, we have achieved an in vivo sensitivity of +/- 0.5 mg fluoride in an observation period of 30 min. Assuming an index finger has between 0.25 and 0.5 g of bone calcium, this sensitivity is just sufficient to detect fluorides in the index finger of most adults whose bone fluoride concentration is greater than 2.0 mg fluoride/g calcium (0.8 mg/g ash weight). We are particularly interested in using this new NMR technique to monitor the rate of bone fluoride accumulation in osteoporotic patients receiving therapeutic levels of fluoride in their diets.

The value of a program for the rehabilitation of osteoporotic patients (PRO) was assessed from a 4-year follow up of 139 patients referred to the program over its initial 2 years of operation. The program consisted of educational seminars, social activities and regular exercise supervision. Patients had annual clinical assessments, and bone mass measurements by neutron activation analysis (reported as CaBI). Fitness was assessed by performance on a treadmill (reported in terms of VO2max, ml/kg/min). Seventy-eight of the 139 patients remained in the program over the 4-year follow-up. This unusually high level of commitment to the program is indicative of the psychological value that patients have derived from it. The effect of the program on the osteoporosis process was inconclusive. Group 2, the 37 patients who obtained the greatest improvement in fitness (VO2max > 6 ml/kg/min), had a significantly greater reduction in back pain than did group 1, the 36 with less significant improvement (VO2max < or = 6 ml/kg/min). The bone mass and incidence of vertebral fractures on entry into the program were not significantly different between the two groups. Group 2 had on average a greater increase in bone mass over the 4 years; mean increases in CaBI, (+/- SD) were 0.09 +/- 0.09 and 0.05 +/- 0.10 for groups 2 and 1 respectively. Group 2 had on average fewer new vertebral fractures (0.08 +/- 0.36 and 0.28 +/- 0.75 for groups 2 and 1 respectively).(ABSTRACT TRUNCATED AT 250 WORDS)

A non-destructive method based on instrumental neutron activation analysis (INAA) for the assay of aluminum in bone samples is described. The28Al signal obtained upon neutron irradiation includes contributions from both the reaction27Al(n,γ)28Al and31P(n,α)28Al. The first reaction is with the thermal neutrons and the second one is with the fast neutrons. The contribution from the31P reaction is calculated from the fact that Ca/P ratio in bone mineral is constant and the amount of calcium can be measured from the thermal reaction48Ca(n, γ)49Ca. The aluminum values obtained by the INAA procedure agreed within 10% of those obtained by atomic absorption spectrophotometry. With this assay the levels of aluminum in normal bones (

Bioelectric impedance analysis (BIA) has been widely used for assessment of body composition in healthy subjects but has not been validated in malnourished patients. This study compared calculation of fat-free mass (FFM) by five methods to determine whether the currently used equations for total body water (TBW) as assessed by BIA were applicable to 19 malnourished patients with Crohn's disease. When compared with TBW assessed by H2(18)O dilution, BIA was higher by 5.9 +/- 1.1% (P < 0.005). A stepwise-multiple-regression equation was derived to validate BIA: TBW (kg) = 0.25 (ht2/resistance) + 0.29 (wt) + 3.63 (r = 0.97, SEE = 0.28). Comparison of the hydration of FFM (TBW by 18O dilution/FFM) between methods showed that total body potassium (TBK) gave a significantly higher value when compared with the reference method of body protein, mineral, and 18O analysis. Dual-energy x-ray absorptiometry (DXA) provided a value for TBW/FFM in close agreement with the reference method. We conclude that BIA overestimates TBW, and TBK underestimates FFM in malnourished patients. DXA provides an accurate measurement of body fat in malnutrition.

Anorexia nervosa is an eating disorder which occurs most frequently in adolescents and young women. It is characterized by fear of obesity and consequent self-starvation in the pursuit of thinness resulting in marked weight loss and amenorrhea1,2. It is associated with many complications, involving almost all the organs of the body. Several reports indicate that some patients have asymptomatic osteopenia and a few of them develop fractures3,4,5,6,7. We investigated bone mass and the relationship between bone mass and soft tissue compartment measurements in adolescent patients with anorexia nervosa.

Osteoporosis is a common problem after menopause. There are a number of methods to assess bone mass. Among them dual-energy X-ray absorpticmetry (DXA) is one with very low radiation and high precision1,2 but its discriminatory ability (Sensitivity and Specificity) in the diagnosis of osteoporosis is only fair.3,4,5 In the present study, an attempt was made to improve this sensitivity and specificity.

Dual energy X-ray absorptiometry (DXA) is widely used for bone mineral measurements and is recently being used to measure soft tissue composition. In-vivo measurements are reproducible with excellent precision (1-2%) for bones and somewhat poorer precision for soft tissue composition. We have measured the bone mineral content (BMC), bone area and density (BMD, g/cm2) using aluminum plates of known weight and area in place of bone. In order to simulate tissue, cellulose (paper in the form of books) was used as the surrounding medium. Further experiments were done to determine whether the presence of thin bones would affect the results of soft tissue analysis.

Dual energy x-ray absorptiometry (DXA) is used widely for measurement of bone mass. With DXA, reproducibility is good (1–2%), radiation exposure is negligible (

Vertebral morphometry on thoracic and lumbar spine radiographs and bone mass measurements were carried out on 215 patients investigated for postmenopausal osteoporosis. Bone mineral mass was measured on the central third of the skeleton by neutron activation analysis and the result, normalized for body size, expressed as a calcium bone index (CaBI). The normal CaBI value for females (20-40 years) is 0.97 (0.11) with a lower limit for these young, normal women, of 0.75. Vertebral compression deformity was defined as a mean height more than 15% lower than adjacent normal vertebrae. Thoracic and lumbar anterior wedge deformities and central compression were defined as anterior/posterior (A/P) or mid/posterior (M/P) height ratios of less than 0.75. For the 129 patients without vertebral deformities, the mean CaBI was 0.80 (0.12) (1 SD) and 32% of these patients had CaBI values below the normal young adult range (CaBI less than 0.75). In 20 patients, vertebral deformities were limited to 1 or 2 mid-thoracic vertebrae, and the mean CaBI values for these 20 patients was 0.81 (0.15), equal to that for patients without any vertebral deformity. For the remaining 67 patients, (i.e., patients with one or more vertebral deformities involving at least one distal thoracic or one lumbar vertebra) the mean CaBI value was 0.66 (0.10), 17% below the value for patients without vertebral deformities. Low CaBI values (CaBI less than 0.75) were observed in 87% of these patients, consistent with the diagnosis of osteoporotic fractures. Based on our CaBI results, however, mid-thoracic deformity was not associated with significant osteopenia and is not, therefore, diagnostic of osteoporotic fracture.

Flameless Atomic Absorption Spectrophotometry was found to be a sensitive (2·10−12 g detection limit), accurate but destructive method for cadmium assay in bone biopsy samples (about 30 mg dry weight). The inductively coupled plasma emission technique was poorer in sensitivity (1.2·10−9 g) and is also a destructive method. Activation Analysis is still less sensitive (2·10−8 g detection limit) but a nondestructive one. Cadmium was found to accumulate in bone of rats fed, for 5 weeks, 0, 50, and 100 mg Cd/l in drinking water and the bone concentrations were 0.16, 1.09, and 2.6 mg Cd/kg bone (dry wt). Histological examination of the bones showed that cadmium induced increased osteoid surface in the bone with no evidence of accompanying kidney damage. This suggests a primary effect of cadmium on bone rather than secondary effect due to kidney damage.

Partial-body calcium measurements by in vivo neutron activation analysis (IVNAA) were correlated with radiographic measurements of bone mineral mass and density, external diameter, and combined cortical thickness of the proximal radius in three groups of subjects: normals, osteoporotics, and chronic renal failure patients. The radial bone mineral mass (in g/cm) correlated best with IVNAA measurements in normal subjects (r=0.94), reasonably well in osteoporotics (r=0.83), and less well in chronic renal failure patients (r=0.65). Simple combined cortical thickness measurements correlated moderately well both in normal subjects (r=0.82) and in osteoporotic subjects (r=0.74).

Measurement of nitrogen in the human body gives a measure of total protein in the body. Nitrogen (N) may be measured in vivo by a nuclear technique in which the characteristic nitrogen gamma rays emitted after capture of neutrons are detected and counted. The accuracy (as determined by comparison of the measurement of nitrogen in a pig by nuclear and chemical methods) is better than 10%. The reproducibility is less than 6%. The radiation dose is about 5 millirads. This method had been used with 43 patients and 23 normal volunteers and the results are reported. The data are presented in the from of total N content, percentage N by weight, and as a comparison of N and potassium (K) in the body. It is found that the mean percentage of N in the normal males is 2.5%, giving a value of 1.75 kg in the 70-kg standard man. The mean percentage values are not greatly different between normal volunteers and malnourished patients. Taking all subjects, N/K ratios ranged from 7.5 to 20 g/g. Thus K cannot be used as a predictor of N. Nitrogen content of normals correlates well (r = 0.68) with height squared; for malnourished patients there is again a good correlation (r = 0.81) but the regression line for patients is approximately 34% below that for the volunteers consistent with their depleted state.

This work describes an in vivo neutron activation analysis facility for small samples, such as rats or human hand, using two 100 μg252Cf neutron sources. The irradiation area is a cylindrical space, of 12 cm diameter and about 15 cm length, with fairly uniform neutron flux distribution. Experimental data on the reproducibility, effects of volume and other conditions for in vivo measurements are given. Comparative atomic absorption data on calcium measurements on rats are reported. The facility is now used for animal experiments as well as human hand irradiations in clinical investigations involving calcium metabolism and bone diseases.

Total body nitrogen (TBN) was measured in 121 volunteers. TBN depends on body size, with the best simple relationship being TBN α (HAS) 2.6 for both men and women, where HAS is a mean of height and arm span. This relationship was used as a predictor of normal TBN. Weight and total body potassium (TBK) were considered for improving normalization, but rejected. Five patients with anorexia nervosa had only 71% of predicted nitrogen. Twelve patients with malabsorption syndrome initially had 68% of normal, but after a year of total parenteral nutrition this increased to 95%. On the other hand, 14 patients with renal failure initially had near normal nitrogen but after a year on continuous ambulatory peritoneal dialysis, nitrogen fell by 20%. TBN, when normalized for body size, provides a reliable index of protein status in clinical investigation and cannot be accurately predicted by weight or TBK.

verely malnourished patient, the exact rec-ognition of early and subtle malnutrition isnot as simple. Furthermore, the results ofsuch marginal deficiency are not clearlyknown. Since the recognition and knowledgeof the effects of early malnutrition are bothimportant in defining the need for nutritionalsupport, several investigators have attemptedto study this area. They have related a varietyof parameters of anthropometry and bio-chemistry

In rats, phosphorus deficiency (P−) has been shown previously to stimulate the linear bone apposition rate (BAR) and this P− effect is dependent on adequate intake of vitamin D 7 . To investigate further the relative importance of the vitamin D 3 metabolites, 1,25(OH) 2 D 3 , 24,25(OH) 2 D 3 , and 25(OH)D 3 , in BAR stimulation, we studied, in P− rats, the relationships between BAR and plasma levels of these three vitamin D 3 metabolites following vitamin D 3 deprivation. Three groups of rats were placed on diets differing only in phosphorus (P) and vitamin D 3 (D 3 ) content, with one group diet deficient in both P and D 3 , one diet, P−, D 3 replete, and one diet both P and D 3 replete. Plasma levels of the three vitamin D 3 metabolites, plasma Ca and P, isotopic Ca absorption and BAR measurements were carried out at 1, 3, and 5 weeks after onset of the test diets. In P−, D 3 replete rats, both plasma levels of 1,25(OH) 2 D 3 and BAR were increased throughout the 1 to 5 week study period, while 25(OH)D 3 and 24,25(OH) 2 D 3 levels were not significantly different from P and D 3 replete controls. In P−, D 3 restricted rats, BAR was decreased by one week, prior to any reduction in plasma levels of 25(OH)D 3 and 24,25(OH) 2 D 3 and while plasma 1,25(OH) 2 D 3 levels were still well above control values. In this P− rat model, the vitamin D dependent BAR stimulation does not appear to be directly related to alterations in the plasma levels of 1,25(OH) 2 D 3 , 24,25(OH) 2 D 3 , or 25(OH)D 3 .

The association between elevated brain aluminum levels and Alzheimer's disease (AD) is examined and critically reviewed. We found elevated aluminum levels in the brains of patients with AD (greater than 4 micrograms/g dry wt.) compared with normal subjects (approximately 1.5 micrograms/g dry wt.). Nine laboratories from different geographical regions have confirmed this finding. Two laboratories did not find any differences between AD and control brains. This discrepancy is traced to differences in sample sizes used for the aluminum assay and the sample selection criteria. It is found that it is essential to use small sizes (approximately 10 mg dry wt.) and to ensure that control brains do not contain neurofibrillary tangles (NFT) and that AD brains do. The exact pathogenic role of aluminum in AD is, as yet, unclear. It is the only element (other than calcium, which non-specifically accumulates at all degenerating tissue sites) that is found in elevated concentrations in NFTs. It is found elevated at four loci in the brain, i.e. the DNA-containing structures of the nucleus, the protein moities of NFTs, the amyloid cores of senile plaques and cerebral ferritin. The evidence thus far indicates that aluminum is toxic to the brain and it is probable that it has a pathogenic role in Alzheimer's disease.

The effect of peritoneovenous shunting on body composition has been studied in 7 cirrhotic patients undergoing a successful shunt and 3 patients in whom the shunt was unsuccessful. In the 7 patients with functioning shunts, their weight had decreased by a mean of 9 kg (p less than 0.001) by 6 wk after initial diuresis, natriuresis, and kaliuresis, and was associated with a decrease in total body potassium (TBK) but not total body nitrogen (TBN). This resulted in a significant decrease in the TBK/TBN ratio from 2.12 +/- 0.74 to 1.66 +/- 0.20 (p less than 0.01). By a mean of 14 mo, in these 7 patients there was a significant increase in mean TBN (from 1.54 +/- 0.10 to 1.84 +/- 0.10, p less than 0.005) associated with an improvement in the mean nitrogen index (from 0.74 +/- 0.04 to 0.88 +/- 0.04, p less than 0.005). These changes were associated with a significant increase in nonalcoholic calories, a nonsignificant increase in protein consumption, and a positive nitrogen balance. After the initial kaliuresis, mean potassium balance remained constantly positive (+22.7 +/- 3.4 mmol/day), serum aldosterone levels normalized, and TBK increased. In contrast, 3 patients with failed peritoneovenous shunting continued to lose weight significantly despite the presence of ascites; TBN and nitrogen index also decreased. In conclusion, body composition studies appear to have confirmed the clinical observation that cirrhotic patients with massive ascites have depleted body protein which is gradually repleted only after successful shunting. In this situation TBK, long used as a measure of lean body mass, is less satisfactory than TBN and nitrogen index. This improvement in body protein appears to be explained by an increased dietary intake associated with improved nitrogen balance, but these changes are not found in patients in whom the shunt failed.

A procedure has been developed to measure fluoride concentration in bone biopsies by neutron activation analysis /NAA/. The NAA procedure is non-destructive so that the bone biopsies can be used subsequently for histological evaluation. The fluoride content is expressed as F/Ca ratio in the bone samples. The fluoride and calcium are measured using the reactions:19F/n, γ/20F /t=11.2 s/ and48Ca/n,γ/49Ca/t=8.8 m/, respectively. The F/Ca ratio normalizes the fluoride to bone mineral avoiding the use of bone weight which is unreliable with fresh biopsy samples. This ratio also corrects for variations in neutron flux and gamma counting efficiencies. Results by this procedure were compared to biochemical determinations using an ion-selective electrode for fluoride and atomic absorption for calcium. The two methods gave results which agreed within ±5% which is the precision of the NNA procedure. The NAA method provides a simple and non-destructive procedure for fluoride measurement in bone biopsies for clinical studies. The method is now routinely used in our clinical studies for the fluoride measurements on biopsies from osteoporotic patients treated with fluoride therapy for nearly four years.

Measurements were made of duodenal calcium-binding protein (CaBP) on rats during development of rickets and, subsequently, following vitamin-D2 treatment.Results showed a poor inverse correlation between duodenal CaBP and rickets. In rats fed a phosphate-deficient rachitogenic diet, duodenal CaBP concentration finally fell below detectable limits, but CaBP was still readily measurable 2 weeks after rickets was clearly established. Following a massive dose of vitamin D2 (50000 I.U.) to rachitic animals, CaBP was formed. However, a small dose of vitamin D2 (500 I.U. daily for 4 days) was insufficient to demonstrate CaBP synthesis even though it was sufficient to demonstrate calcification of cartilage. Administration of normal rat chow to rachitic rats stimulated greater CaBP synthesis than vitamin-D treatment alone. The rachitogenic diet supplemented with phosphate, which caused osteoporosis but not rickets, inhibited CaBP synthesis. The results suggest that nutritional deficiencies from the rachitogenic diet, in addition to vitamin-D deficiency, inhibited CaBP synthesis.

Thirty-two patients with postmenopausal osteoporosis have been followed for periods of 3 yr or more to assess the effects of treatment on bone mineral mass based on sequential measurements of bone mineral in the central third of the skeleton. Neutron activation analysis (NAA) was used to measure calcium in the trunk and proximal femurs. The NAA measurement is expressed as an index, CaBI, which relates the subject's Ca value to the estimated mean value for normal subjects of the same size based on height and arm span. Bone and serum fluoride concentrations were measured, and bone biopsies were carried out in many cases. The treatments consisted of various combinations of NaF (25 mg, twice daily), calcium (1 g/day), vitamin D2 (50,000 IU twice weekly), and estrogen (0.625 mg/day). NaF was always given in combination with Ca and vitamin D2. During the 3-yr treatment period, the 32 patients showed changes in bone mineral mass from losses of −10.5% to increases of 21%. The coefficient of variation of 9% for ch...

The rate of bone apposition was studied by labeling bone with four or five sequential doses of oxytetracycline given at equal intervals of 2 or 4 days in rats and rabbits. The rate was estimated in bone formation sites bearing all the doses. This method has the advantage of assessing the rate of new bone mineralization during the active bone-forming phase only. This index of bone formation was found to be independent of the location of bone samples in the skeleton, and therefore might be under the influence of general body mechanisms only. Preliminary study in the rats indeed yielded results indicating that this bone apposition rate is accelerated by dietary deficiency in calcium.

Measurements of body composition were carried out on 56 healthy female volunteers 20–80 years of age. Anthropometric measurements were made to estimate lean body mass (ANLBM). Total body potassium (K) was measured from the naturally occuring 40K; body nitrogen (N) and calcium (Ca) were measured by neutron activation techniques. Body K, N and Ca reflect intracellular water, protein and bone mineral mass respectively. The 23 older women (mean (±SD) age=62.3±8.5) showed no significant differences from the 33 young adults (mean age 35.2±3.8) with respect to height, total body mass, ANLBM and intracellular water (K) but the older group had significantly less protein (by 17%) and less bone mineral (16%) than younger adults. When normalized for body size, based on height, ANLBM was significantly correlated to intracellular water (r=0.52) and to bone mineral (r=0.39) but correlated to either ANLBM (r=0.05) or to intracellular water (r=0.10). Protein was correlated to bone mineral (r=0.39). Simple anthropometric measurements may provide reasonable estimates of the fat and lean fractions of total body mass. However the composition of lean tissue varies with aging. To determine the essential components of lean tissue, direct measurements of protein and bone mass are required.

Using the technique of short interval sequential tetracycline labeling, it was documented that the apposition of mineralized bone matrix in adult male Sprague-Dawley rats was inhibited by hydrocortisone. The inhibition occurred as early as six days after the onset of the treatment and was dose dependent over a dose range of 0.62 to 20 mg per kg body weight per day. Vitamin D2 supplements by injection protected bone from this hydrocortisone action. 64 I. U. of vitamin D2 injected daily was able to prevent the inhibition of bone apposition by 20 mg per kg body weight per day of hydrocortisone. The results imply that vitamin D or its metabolites may compete with hydrocortisone in some cellular mechanisms and support the usefulness of vitamin D supplements in the treatment and the prevention of steroid-induced osteoporosis.

A radioimmunoassay for porcine intestinal calcium-binding protein (CaBP) has been developed. 125 I-labeled CaBP was prepared using chloramine-T. Antisera to porcine CaBP were raised in guinea pigs and rabbits. Antibody-bound CaBP was separated from free CaBP using talc. The immunoassay was highly sensitive, measuring concentrations of CaBP in duodenal extracts as low as 1 ng/ml with good reproducibility. This is sufficiently sensitive to measure CaBP in small amounts of duodenal tissue obtained by peroral capsule biopsy. The method was specific for CaBP, showing no reactivity toward other duodenal proteins or several peptide hormones. Measurements of CaBP by immunoassay showed a good correlation with measurements of CaBP-specific calcium-binding. The immunodilution curves obtained when assaying pig renal cortical extracts against pure duodenal CaBP were parallel when tested with six different antisera, demonstrating a high degree of immunochemical similarity between kidney and duodenal CaBP. Clearly, the radioimmunoassay possesses distinct advantages for the study of the biologic role of CaBP.

A 252 Cf neutron-irradiation facility designed specifically for clinical in vivo measurement of calcium in the hand is described. Results of preliminary measurements are presented. Hand phantoms were exposed to the neutron beam for 10 min and the induced 49 Ca activities were counted for 10 min after an elapsed time of 2 min. The results indicate that with a 2 × 100 μg 252 Cf neutron source and two 20.3 × 12.7 cm NaI crystals, the counts per gram of bone mineral mass changes by about 4% for each 100 cm 3 change in the overall volume (soft tissue plus bone) of the hand. For hands of equal volumes the counts per gram are expected to be almost indepdendent of the bone volume. With an absorbed dose equivalent of 150 mSv (15 rem), the sensitivity is about 200 counts per 10 min per gram Ca. The statistical reproducibility of the results is better than 3% for the average value of 11 g Ca in the normal hand.

Nitrogen in the human body is measured in vivo using prompt neutron capture gamma-rays. The quantity of nitrogen can then be used as measured of protein. Data are presented on three groups of subjects; volunteers of different ages, those with liver ailments, and those on peritoneal dialysis. The data show that the nitrogen measurements given information (in accord with clinical findings) which is not given by indirect methods of estimating lean body mass.

The effects of nutritional support on body weight, nitrogen and potassium were investigated and were compared with changes in serum albumin and delayed cutaneous hypersensitivity (DCH). Fifteen malnourished patients (maintained by total parenteral nutrition (TPN) were studied for periods of 5 to 33 months. The TPN consisted of infusions of amino acids with a dual substrate of glucose and lipids. Body nitrogen was measured by prompt gamma ray analysis and potassium was determined by measurement of the naturally occurring radio-active 40 K in a whole body counter. Results of sequential measurements show on average a significant increase in body nitrogen of 30% (P Thus, changes in body protein are not necessarily reflected by measurements of the body parameters of nutritional status. For a reliable estimate of body protein, the direct measurement of body nitrogen is required.

Rapid weight gain has been observed following kidney transplantation. To determine the accompanying changes in body composition, we studied 8 patients (7 females, 1 male) aged 3-17.5 years who underwent renal transplantation. Body composition measurements included weight, height, triceps skinfold thickness, total body potassium, total body water, and extracellular water. Excessive weight gain was observed in most of the patients. Weight as a percentage of ideal weight for height increased from 96.0 +/- 13.8 to 116.3 +/- 13.0% (p less than 0.01). This was accompanied by a gain in fat mass in the first 3 months and a subsequent increase in lean body mass in the next 3 months. Extracellular water was increased before transplantation (32.9 +/- 6.5% of body weight) and returned to normal (27.3 +/- 8.6%) 3 months after transplantation (p less than 0.01). The weight gain following kidney transplantation in children resulted mainly from increases in adipose tissue and lean body mass, and was not related to water retention.

The effect of treatment on calcium kinetic and balance studies was investigated on six patients with osteoporosis (group A). The results were compared with data obtained from similar studies on four patients with calcium abnormalities secondary to steatorrhea and on one infant with vitamin D-dependent rickets (group B). The studies were carried out before treatment and subsequently at various periods up to 2 yr after the onset of treatment. The effect of treatment on bone calcium kinetics differed significantly between the two groups. In osteoporosis (group A), accretion rates of calcium to bone were lower as compared to pretreatment values. In contrast, in four of the five subjects of group B, accretion rates increased from pretreatment values. The results show that in osteoporosis changes in bone calcium metabolism were observed for as long as 2 yr on treatment, despite the fact that the balance data were normal at this time.

In a group of 12 patients with steatorrhea secondary to various etiologies, defective calcium absorption and negative calcium balance occurred in all patients with biochemical osteomalacia. Losses of body calcium resulted from obligatory endogenous fecal losses. High endogenous fecal calcium values appeared to be due to increased digestive juice secretions as well as defective reabsorption. Bone calcium turnover in osteomalacia varied from below to well above the normal range in agreement with reported data on bone biopsy studies. Markedly elevated bone calcium accretion rates were found in idiopathic steatorrhea alone and were associated with advanced bone disease.

Toronto General Hospital records show that 26 persons received Thorotrast myelography in the years 1940 and 1941. Of these, 15 have been located and the activity in nine has been measured for residual thorium activity using a whole-body counter and thoron exhalation techniques. For comparison, measurements of thorium burdens of nine subjects who had Thorotrast arteriograms are presented. Of the 15 myelography cases located, 12 have had serious progressive cauda equina deficit. Even on the assumption that the 11 unlocated are all well, the incidence of myelopathy is still nearly 50 per cent. On certain assumptions the thorium burdens of the nine measured are ascertained to be between 0·03 and 1·0 g, with a mean value of 0·45g. It is pointed out that these levels are all comparable to, and in some cases less than, the maximum permissible body burdens (ICRP) and yet severe effects have occurred.

A calcium binding protein has been demonstrated in human duodenal mucosa which we believe to be human vitamin D dependent CaBP. Sephadex gel filtration demonstrated that the duodenal mucosa of both the human and pig contained a calcium binding protein with a similar molecular weight to the reported vitamin D dependent rat CaBP (M.W. 12 000–13 000) but dissimilar to chick CaBP (M.W. 24 000–28 000). In addition the rat, pig, and human mucosa contained a high molecular weight calcium binding protein which is probably not vitamin D dependent.A relatively simple procedure, utilizing the tagging of CaBP by 47Ca, has been developed to partially purify normal pig CaBP by Sephadex gel filtration. Further fractionation of the 12 000–13 000 M.W. area, using disc electrophoretic procedures, separated two calcium binding proteins which had similar electrophoretic mobilities and calcium binding formation constants (3.5 and 5.5 × 106, respectively), indicating that both are forms of CaBP but that either one or both have been altered during the procedures. The electrophoretic mobilities of both of these proteins are relatively low in the presence of calcium ions but much greater when calcium ions are removed by chelation with EDTA. This finding should facilitate both the identification and purification of mammalian CaBP.

A new procedure for the determination of urinary calcium concentrations as low as 1 mg./L. is presented. Calcium is separated from interfering urinary constituents by oxalate precipitation with strontium carrier and then determined by emission flame photometry. Small calcium losses in the separation procedure are estimated by 47Ca-tracer technics. Wet-ashing ensures complete exchange of tracer and sample calcium. Experimental evidence demonstrated the accuracy of the procedure and a precision of ± 3% of the observed results. The use of tracer may be omitted if systematic errors up to 5% are acceptable. Wet-ashing has also been omitted with equally good results but there is an additional risk of error due to possible interference effects by constituents of some pathological samples.

A calcium-binding protein from porcine intestinal mucosa has been characterized by a variety of physical and chemical techniques. Chromatography on Sephadex G-75 suggests a molecular weight around 12,000, while electrophoresis in sodium dodecyl sulfate-urea-polyacrylamide gel indicates a molecular weight in the region of 7000. Sedimentation equilibrium analyses give evidence for aggregation, with the monomeric form having a molecular weight between 7000 and 9000. The amino acid composition is as follows: Lys11–12, Arg1, Asp7, Thr1, Ser6–7, Glu17, Gly4, Ala5, Val3, Ile3, Leu10, Tyr1, Phe5. There is no histidine, half-cystine, methionine, or tryptophan. The composition requires a minimal molecular weight of 9000. No free terminal amino group was detectable, but digestion with penicillocarboxypeptidases S1 and S2 indicated the following COOH-terminal sequence: .... (Gly, Thr, Asp)Ala-Ile-Val(Phe, Ser)Leu-Lys-GlnOH. The circular dichroism spectrum of the binding protein in the near ultraviolet shows a positive peak at 276 nm with a shoulder at 282 nm characteristic of tyrosine, two positive peaks at 264 nm and 257 nm, and two negative peaks at 268 and 261 nm characteristic of phenylalanine. In the far ultraviolet range two negative bands at 222 nm and 207 nm are observed. Removal of calcium ions causes a marked decrease in the tyrosine band at 276 nm to less than 25% of its initial value, but no difference is observed in the far ultraviolet range. Ultraviolet difference spectra between the calcium-containing and the calcium-free protein show peaks characteristic of tyrosine. No difference in sedimentation rates was observed. It appears that the binding of calcium causes a local perturbance of the environment of the single tyrosine residue, but no gross conformational changes.

Lean body mass by anthropometry (AMLBM), total body potassium (TBK), and total body nitrogen (TBN) by prompt gamma analysis, are reported in 38 malnourished patients during total parenteral nutrition. Over long periods (greater than 2 months) TBN increased 32% while AMLBM and TBK rose only by 9.2 and 9.5%, respectively. Changes in AMLBM and in K were significantly correlated, but changes in nitrogen were not correlated with either. From short-term studies, the same conclusions can be obtained; although there are changes in lean body mass and in potassium, there is no correlation with such changes in nitrogen. With repletion, changes in body protein are not necessarily related to changes in AMLBM or to TBK: the latter two reflecting total and intracellular water, respectively, but not protein content.

It has been widely believed that phosphate deficiency causes osteomalacia. Based on this belief, the rickets of familial hypophosphatemia has been attributed to phosphate deficiency associated with the hypophosphatemia. The present studies on rats have, however, demonstrated significant differences between the effects of phosphate deficiency on bone metabolism and the characteristic features of rickets. Weanling rats, maintained on a mildly phosphate deficient diet, had hypercalcemia and hypophosphatemia, and impairment of body growth, bone growth, and bone mineralization. The maximum effect was observed at 5 wk; between 5 and 20 wk the rats improved despite persistent hypophosphatemia. Histologically, at 5 wk the bone showed thick unmineralized osteoid seams covering most bone surfaces, but the epiphyseal cartilage was normal. In addition, the excess osteoid readily incorporated tetracycline indicating normal mineralization and, based on a new sequential pulse labeling technique, the linear bone apposition rate (LBA) was significantly (p < 0.001) increased above control values. This increase was observed within the initial 4 days of phosphate (P) deficiency and persisted up to 15 wk. This effect of P deficiency on LBA was dependent on vitamin D activity. At 4 wk, the mean LBA was 0.106 +/- 0.003 (1 SE) in control rats, 0.149 +/- 0.008 microns/hr in P deficient rats, 0.083 +/- 0.004 microns/hr in vitamin D deficient rats and 0.086 +/- 0.006 microns/hr in rats deficient in both P and vitamin D. We have reported a similar increase in LBA with parathyroid hormone activity. With vitamin D deficiency, phosphate deficient rats showed all the characteristic features of rickets; disorganization of epiphyseal cartilage, excessive unmineralized osteoid, and reduced mineralization based on the incorporation of tetracycline. We conclude that the effects of phosphate deficiency on bone metabolism more closely resembles the effects of PTH activity than the characteristic effects of osteomalacia and rickets.

Studies were done to investigate nephrocalcinosis produced in weanling female Wistar rats fed pelleted, semisynthetic diets. The rats were fed diets varying in concentrations of Ca and P supplied as inorganic salts for periods of 4–6 weeks and results compared with control rats fed laboratory rodent chow for the same period of time. Measurement of renal Ca and P concentrations showed that nephrocalcinosis was produced by semisynthetic diets with inorganic phosphate concentrations as low as 0.5% on a weight basis; in contrast, rats fed regular laboratory chow (P = 0.72%) showed no evidence of nephrocalcinosis. The severity of the lesion was proportional to dietary phosphate concentrations from 0.5 to 1.0% but other dietary factors modified the severity of the lesion. With the lower dietary phosphate of 0.5%, increasing dietary Ca from 0.5 to 1.0% decreased the severity of the renal calcification. Decreasing protein concentrations from 25 to 15% casein increased the severity of the renal lesions. Other dietary factors also appear to modify the phosphate-induced nephrocalcinosis since no lesions occurred in rats on laboratory chow. It is suggested that the availability of dietary phosphate may be a factor. The phosphate in the semisynthetic diets was totally inorganic while the natural foods of laboratory chow contain, at least in part, organic phosphate.

In the procedure of in vivo neutron activation analysis for calcium and other elements, the thickness of the body affects both neutron flux distribution in the body and self-absorption of gamma rays emitted by the body. Measurements have been carried out to find the effects of these factors individually and also when combined. With the PuBe neutron source and NaI detector arrangements used at Toronto for activation analysis of the trunk, the combined changes in calcium counts are about 5 per cent for each 1 cm change in the thickness of the subject.

The effect of vitamin D activity on the major renal Ca-binding protein has been compared with that on the intestinal Ca-binding protein. Using a method based on Ca-binding capacity, these proteins were measured in vitamin D deficient rats after vitamin D treatment for varying periods up to 5 days. Since P deficiency has been shown to stimulate synthesis of the active metabolite 1,25-dihydroxycholecalciferol, a similar experiment was done on rats fed a P-deficient diet for periods up to 21 days. The renal Ca-binding protein was unchanged by vitamin D treatment to vitamin D deficient rats and was only slightly increased (50%) by phosphate deficiency. By comparison, the intestinal protein was increased twofold by vitamin D treatment and fivefold by phosphate deficiency. Results indicate that vitamin D activity has no direct effect on the major renal Ca-binding protein.

The effect of dietary Ca in response to fluoride (F) treatment was investigated in rats. Rats were maintained on either adequate (0.5%) or high (2.0%) dietary Ca and given for 5 weeks, NaF in drinking water. The minimum NaF levels that inhibited body growth and reduced survival were 300 mg/L with 0.5% diet Ca and 550 mg/L with 2.0% diet Ca. With these toxic F doses, bone histology showed increased formation surfaces and thickened osteoid seams (osteoid index 6–7%). Fluoride doses 30% below toxic levels (200 and 350 mg/L for 0.5 and 2.0% diet Ca, respectively) had no demonstrable effect on bone. Additional diet Ca reduced F absorption from 76 ± 3 to 47 ± 3% for 0.5 and 2.0% diet Ca, respectively. Comparable absorbed doses of F produced comparable effects on bone and body growth but, with additional dietary Ca, these effects were observed with 50% lower serum and bone F levels. Variable response to NaF therapy can be produced in rats by alterations in dietary Ca alone. Results indicate that for clinical treatment the NaF dose needs to be adjusted on an individual basis but neither serum nor bone F levels can be used reliably to establish optimal doses.

Patients receiving long term parenteral nutrition may develop metabolic bone disease. In all 11 patients studied, histologic studies of bone showed excessive unmineralized bone tissue despite normal plasma 25-hydroxyvitamin D levels. Three patients also had bone pain and fractures and severe urinary loss of calcium and phosphate. Withdrawal of vitamin D from parenteral nutrition solutions was associated with improved histologic findings of bone in all patients, shown by a decrease in osteoid tissue and an increase in tetracycline uptake. In the three patients with symptoms, bone pain subsided, fractures healed, and urinary loss of calcium and phosphate decreased. Thus, vitamin D may be a factor in the genesis of parenteral nutrition-induced metabolic bone disease.

Osteoporosis is a common crippling bone disease affecting post-menopausal and elderly women. It results in fractures, spinal deformity and chronic pain leading to decreased mobility and functional disability. A preventive program in Toronto, described in this article, aims to prevent bone loss and maximise the functional capacity of osteoporotic patients through a program with educational, social and exercise components. Evaluation of the program, also reported here, showed that 80 per cent of patients complied with the requirements of the exercise program and that those who exercised reported improvement in general well being, stamina, mobility and pain tolerance. The exercise group also showed a significant improvement in VO 2 max (p less than 0.001) and bone mass (p less than 0.02) after one year of exercise. None of the patients developed fracture as a direct result of the exercise. The exercise prescribed for osteoporosis is safe and beneficial. The program is a social success.