Ellen Hills

Effect of aluminum on normal and uremic rats: tissue distribution, vitamin D metabolites, and quantitative bone histology.

SummaryThe effects of intraperitoneal aluminum chloride (1.5 mg aluminum/kg/day for 9 weeks) were studied in normal and uremic rats. Parameters measured included tissue aluminum, serum vitamin D metabolites, and quantitative bone histology.Aluminum administration increased tissue concentrations of this metal in uremic and nonuremic animals. Bone aluminum concentrations were higher in uremic rats (121 ± 27 mg/kg compared to 47 ± 4), whereas liver values were higher in the nonuremic group (175 ± 47 mg/kg compared to 100 ± 36). Serum concentrations of 25-hydroxyvitamin D and 24,25-dihydroxyvitamin D were reduced in uremia, but aluminum was without apparent effect on any vitamin D metabolite.Aluminum, in the doses administered, caused no skeletal changes in nonuremic animals. Some uremic, non-aluminum-treated rats developed osteomalacia and marrow fibrosis. However, osteomalacia was more severe and the osteoclast count was higher in the uremic, aluminum-treated rats. In this group of animals the mineral apposition rate was reduced at the metaphyseal endosteum but increased at the periosteum, indicating different control mechanisms at the two sites.

The effect of age on bone composition and viability in the femoral head.

We studied the effect of age on bone composition and osteocyte viability in femoral heads from fifty-one subjects. The assessment included determination of: bone volume, ash weight, calcium, and phosphorus content; osteocyte viability in fresh sections stained for lactate dehydrogenase activity; microfractures in fresh sections after removal of marrow elements; bone area, the presence of metabolic bone disease, and the histology of microfractures in embedded calcified sections; and the extent of trabecular microfractures. Bone area and numbers of microfractures were also assessed in eight elderly hip-fracture patients. Bone volume decreased with age, but there was considerable variation in each age group, and no significant difference between men and women. Ash weight and the bone content of calcium and phosphorus also decreased with age, but were constant if corrected for bone volume. Almost all osteocytes were viable in subjects who were younger than twenty-five years, and thereafter viability progressively decreased to a mean of 74 per cent in the eighth decade of life. There was a significant negative correlation between osteocyte viability and age. There was no evidence of metabolic bone disease in any patient. The numbers of microfractures increased with age and correlated negatively with bone viability (r = -0.31, p less than 0.05); in simple linear correlation a relationship between bone area and microfractures could not be demonstrated but in multiple linear correlation, after the inclusion of bone viability, there was an additional negative correlation between numbers of microfractures and bone area (p less than 0.005). Bone area and numbers of microfractures in hip-fracture patients were similar to those in age-matched controls.

Bone death in hip fracture in the elderly

SummaryWe examined femoral head bone from 50 cadavers and from 21 patients who had suffered pathologic fracture of the femoral neck. We used a histochemical technique for lactate dehydrogenase (LDH) activity to demonstrate osteocyte viability. The femoral heads were removed within 36 hours of death or fracture, as LDH activity persists in the cytoplasm of viable cells for this time at 37° after interruption of the blood supply. In the controls, there was an age-related reduction in mean osteocyte viability, from 88±7% (mean±SD) at age 10–29 years to 58±12% at age 70–89 years. In the hip fracture patients, mean osteocyte viability was 58±21% but there was much variability in both osteocyte viability and bone mass. In 5 fracture patients, there was extensive osteocyte death, suggesting that most of the femoral head bone was nonviable; these patients had little microfracture callus. Others had predominantly viable bone which was usually osteoporotic, and their bone frequently showed microfracture callus. Osteomalacia was not seen in any patient. It is suggested that bone death, in addition to osteoporosis, may sometimes contribute to hip fracture in the elderly.

Effect of aluminum and parathyroid hormone on osteoblasts and bone mineralization in chronic renal failure

SummaryBone aluminum, quantitative bone histology, and plasma parathyroid hormone (PTH) were compared in 29 patients undergoing chronic hemodialysis. Histologic techniques included double tetracycline labeling and histochemical identification of osteoclasts and osteoblasts. Bone aluminum was measured chemically by flameless atomic absorption spectrophotometry, and histochemically. When measured chemically, the bone aluminum was 67±46 (SD) mg/kg dry weight (normal 2.4±1.2 mg/kg); histochemically, aluminum was present at 2.9±4.4% of trabecular surface. The biochemical and histochemical results agreed well (r=0.80,P<0.001). No double tetracycline labels were seen at the mineralization front where aluminum was deposited, indicating cessation of mineralization at these sites. The osteoblast surface correlated positively with plasma PTH (r=0.67,P<0.001) and negatively with bone aluminum level (r=−0.42,P<0.05). Multiple linear regression showed a correlation of aluminum with osteoblasts additional to that of PTH, consistent with a direct effect of aluminum in depressing osteoblast numbers. Though a relationship between PTH and chemically determined bone aluminum level could not be demonstrated, there was a negative correlation between osteoclast count and aluminum, and the nine patients with severe hyperparathyroid bone disease had lower chemically determined aluminum levels than the other patients. These results suggest that aluminum (a) directly inhibits mineralization, (b) is associated with decreased PTH activity and hence osteoblast numbers, and (c) directly reduces osteoblast numbers. In addition to inducing severe, resistant osteomalacia, aluminum appears to contribute to the mild osteomalacia commonly seen in renal failure, characterized by extensive thin osteoid and low tetracycline and osteoblast surfaces.

The Determination of Bone Viability: A Histochemical Method for Identification of Lactate Dehydrogenase Activity in Osteocytes in Fresh Calcified and Decalcified Sections of Human Bone

Summary We have developed a technique for assessing bone viability by the histochemical demonstration of lactate dehydro‐genase (LDH) activity in osteocytes. Fresh sawn and ground (75 mU) sections were prepared from femoral heads removed at operation from patients with osteoarthritis of the hip. the sections were decalcified overnight in cold 10% EDTA, pH 7.0. LDH activity was shown by the tetrazolium‐formazan reaction with nitroblue tetrazolium as indicator and lithium lactate as substrate. Osteocytes were regarded as viable if their cytoplasm stained dark blue, indicating LDH activity; lacunae containing non‐viable osteocytes could be identified by interference contrast illumination. Nearly all osteocytes were viable in the samples studied. Small trabecular fragments, such as could be obtained by needle biopsy, were also suitable for staining after grinding to approximately 50 p. the method should have application both in research and in diagnosis of ischemic bone disease.

The pathogenesis of osteoarthritis of the hip. Evidence for primary osteocyte death.

Osteocyte viability was investigated in femoral head bone removed from 38 patients with chronic hip disease, with the use of a histochemical stain to demonstrate lactate dehydrogenase (LDH) activity in osteocytes. Where the osteocyte cytoplasm did not show LDH activity, the cell was considered dead; when several adjacent osteocytes were dead, the bone in that area was regarded as nonviable. The preoperative diagnoses were idiopathic osteoarthritis in 25, chondrocalcinosis in six, rheumatoid arthritis in two, Pagets disease in two, avascular necrosis in two, and congenital dislocation of the hip in one patient. In 16 of the patients with idiopathic osteoarthritis and the two with avascular necrosis, nonviable osteocytes were present in the central regions of many trabeculae, these areas usually being separated by cement lines from viable bone. The pattern suggested previous necrosis of part of the femoral head, with later new bone formation. The pattern was not observed in either control subjects, or patients with known articular disease, such as chondrocalcinosis. Bone collapse of variable severity was apparent radiographically in nine patients with histologic bone death, but not in other patients. Bone death is commonly present in idiopathic osteoarthritis and could be a cause rather than a result of the arthritis.

Growth retardation and renal osteodystrophy in children with chronic renal failure

Height, expressed as standard deviation scores for chronological age and for bone age, was studied in relation to glomerular filtration rate, bone age delay, and bone histology in 47 children with chronic renal disease and GFR less than 80 ml/min/1.73 m2. In multiple regression in all 47 patients, only GFR and bone age delay significantly affected height; 40% of children were short (height standard deviation score less than -2) for chronological age, and 9% were short for bone age. Renal osteodystrophy, which only occurred at GFR less than 30 ml/min/1.73 m2, significantly affected height only in children with congenital renal disease and GFR less than 20 ml/min/1.73 m2. Although radiological and biochemical changes of renal osteodystrophy were seen more often in short children, histological bone disease occurred just as frequently in tall children as in short children. Thus much of the observed height retardation in chronic renal failure is associated with delayed skeletal maturation. In addition, although severe renal osteodystrophy may contribute to growth retardation in advanced renal failure, our data suggest that milder degrees of bone disease evident only on histological study cannot be implicated in the etiology of growth failure in chronic renal impairment.

The Effect of Long-Term Low-Dose Diphosphonate Treatment on Rat Bone

Weaning, male rats were given ethane-1-hydroxy 1,1-diphosphonate (EHDP) or dichloromethylene diphosphonate (Cl2MDP) 0.5 mgP/kg/day for 140 days. Samples prepared after sacrifice were: (1) thin ground (30 micrometers) transverse tibial sections, (2) methacrylate-embedded 5 micrometers sections of the proximal tibial metaphysis, and (3) ultrathin calcified metaphyseal sections for electron microscopy. Cl2MDP reduced the diaphyseal medullary cavity, and EHDP increased the osteoid width. Both drugs increased endosteal bone apposition (possible secondarily to the impaired resorption) and, perhaps as a result, periosteal apposition was increased. Consequently, diaphyseal bone area was unchanged. Metaphyseal bone area was increased and osteoclast numbers reduced, in contrast to the increased osteoclast numbers reported previously in animal experiments with diphosphonates. Acid phosphatase activity within osteoclasts was markedly reduced with EDHP and unchanged with Cl2MDP. The ultrastructural changes in osteoclasts, i.e., a deficiency of ruffled borders, reduced numbers of cytoplasmic vacuoles and the presence of crystals between bone and the osteoclastic plasmalemma, were more marked with EHDP than Cl2MDP.

Bone histology in young adult osteoporosis.

Bone histology was quantitated in 10 osteoporotic patients aged between 17 and 51 years and in six healthy subjects aged between 23 and 43 years. The osteoporosis was of varying aetiology and was clinically stable. All patients were given tetracycline before biopsy and double tetracycline labelling was used in seven patients. Bone forming and resorbing surfaces were defined by the presence of osteoblasts and osteoclasts, respectively, which were identified by histochemical techniques. The associations between bone forming and resorbing surfaces were similar in patients and controls, though the range of values was wider in the patients than in the controls. Mineral apposition rate was normal in the osteoporotic patients, but there was a reduction in mineralising (tetracycline) surface, whether related to osteoid surface or to osteoblast surface. This did not indicate osteomalacia as the directly and indirectly measured mineralisation lag times were normal. The osteoid seams were thinner in osteoporotic patients than in controls. The data suggest that osteoclast and osteoblast numbers were normal in this group of osteoporotic patients but that the metabolic activity of osteoblasts was impaired.

Bone metabolism in idiopathic juvenile osteoporosis: A case report

SummaryA previously healthy 12-year-old boy developed pain on walking and x-rays showed osteoporosis. Over the next 2 years deterioration occurred, the condition became extremely severe, and he was confined to a wheelchair. After 5 years, marked kyphoscoliosis and pigeon chest deformity were present and little increase in height occurred. A wheelchair accident at the age of 17 resulted in several major long bone fractures. Iliac crest biopsies were taken at ages 15 and 17, and subjected to quantitative histology. A histochemical technique for osteoclast recognition by acid phosphatase activity showed resorption parameters to be normal. Double tetracycline labeling and histochemical identification of osteoblasts showed no abnormality of endosteal bone formation. Because of “coupling” of endosteal formation and resorption, these measurements might primarily reflect bone turnover. Failure of periosteal bone formation as shown by failure of radial growth of long bones and of epiphyseal growth was clearly evident. It is likely that osteoporosis developed in this patient due to a reduction in bone formation of unknown etiology rather than by increased bone resorption.