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Biomedical subjects

F R Singer

Publications and source records attributed to F R Singer.

At least 55 records · Page 3Linked to original sources
PubMed1990

Purification and N-terminal amino acid sequence of the tartrate-resistant acid phosphatase from human osteoclastoma: evidence for a single structure.

Tartrate-resistant acid phosphatase type-5 was purified to apparent homogeneity from human osteoclastomas by sequential chromatography on CM-Sepharose, Phenyl-Sepharose, concanavalin A-Sepharose, FPLC Superose-12, and FPLC Mono-S. The purification over the original tissue extract was 1167-fold, with a yield of 16%. An identity in the N-terminal amino acid sequence and Mr was found between this enzyme and two type-5 tartrate-resistant acid phosphatases isolated from hairy cell leukemia spleen. However, they appeared to be different as assessed by amino acid composition. In contrast to a previous report, no evidence was found for two subunits of the tartrate-resistant acid phosphatase.

Acid Phosphatase↗
PubMed1990

Multiple osteocalcin fragments in human urine and serum as detected by a midmolecule osteocalcin radioimmunoassay.

Reliable markers of bone formation are essential to the investigation of metabolic bone disorders. In this regard, evidence indicates that circulating levels of human osteocalcin (OC) correlate with the skeletal isoenzyme of alkaline phosphatase and can be used as an index of bone formation. A disadvantage of using serum OC as a marker of formation is its diurnal variation. To address this problem we carried out our studies to determine the usefulness of urine in the assessment of bone turnover. Using a midmolecule specific human OC RIA, we were able to detect OC in urine of normal adults (42 mugeq/g creatinine), normal children (849 mu/geq/g creatinine), and Paget's disease patients (613 mugeq/g creatinine). Immunoreactive fragments of OC in human urine and human serum were separated by high pressure liquid chromatography. Multiple fragments were found in normal adult urine that were not detected in normal adult serum. Uremic and Paget's disease sera contain several immunoreactive forms of OC, other than the intact molecule, not found in normal adult serum. Additionally, both Paget's disease sera and urine contained a specific peak of immunoreactive material, eluting at 25% acetonitrile, that was not found in any other serum or urine tested. Urinary OC (uOC) correlated with both skeletal alkaline phosphatase (r = 0.91) and serum OC (r = 0.83), indices of skeletal formation. While uOC has a diurnal variation similar to that of serum OC, determinations of 24-h uOC give integrated values of daily bone turnover rates. Z-Score analysis indicates that uOC (z = 14.04) is better able to distinguish between normal children with high bone turnover and normal adults than either skeletal alkaline phosphatase (z = 8.87) or serum OC (z = 9.01).

Adult↗
PubMed1990

Development of an immunoassay for human serum osteoclastic tartrate-resistant acid phosphatase.

A tartrate-resistant acid phosphatase (TrACP), which has been suggested to be very similar to the osteoclastic TrACP, was partially purified from the spleen of a patient with hairy cell leukemia. The purification procedure consisted of carboxymethyl-Sepharose, phosphocellulose, Sephacryl S-200, and phenyl-Sepharose chromatographies. Polyclonal antibodies were generated in guinea pigs with a titer of at least 1:6000. Immunohistochemical staining of fetal rat tibia with the antisera revealed that only the lysosomes of osteoclasts, but not osteoblasts, were stained. An enzyme-linked immunosorbent assay (ELISA) was developed with the antisera. There was no cross-reactivity with 1) partially purified acid phosphatases (ACPs) from normal human and beef spleens, 2) ACPs in extracts of human osteoblastic cells, 3) purified bovine bone matrix TrACP, or 4) commercial prostatic ACP. However, extracts of giant cell bone tumors, containing large amounts of bona fide osteoclasts, showed large amounts of cross-reactive material, which diluted in parallel with the partially purified hairy cell leukemic TrACP in the ELISA. Commercial serum band 5b TrACP also displaced in parallel with the partially purified hairy cell leukemic TrACP. Immunoblotting studies revealed that the antiserum, but not nonimmune guinea pig serum, reacted with the homogeneous hairy cell leukemia splenic band 5 TrACPs, which were recently purified by our laboratory. Preliminary application of the ELISA to sera of patients with metabolic bone diseases revealed that normal healthy individuals had measurable amounts of the immunoreactive material, and patients with Paget's disease or hyperparathyroidism, who should have high bone turnover, had elevated levels of this immunoreactive material in their sera. In contrast, the level of serum osteoclastic TrACP in a patient with an acute lymphatic leukemia was normal. In summary, 1) we have shown that hairy cell leukemia splenic TrACP shares significant immunological similarity with the osteoclastic TrACP and with the serum band 5b TrACP, and 2) the ELISA holds promise for a sensitive and specific assay for bone resorption.

Acid Phosphatase↗
PubMed1990

Role of the bisphosphonate etidronate in the therapy of cancer-related hypercalcemia.

Bisphosphonates (formerly diphosphonates), analogues of endogenous pyrophosphates, are potent inhibitors of osteoclastic bone resorption. While the exact mechanism of action remains poorly understood, etidronate, the only bisphosphonate currently available in the United States for the treatment of hypercalcemia, decreases the elevated serum calcium levels and alleviates the symptoms associated with hypercalcemia of malignancy. Both open and controlled clinical studies have demonstrated that this agent, administered at a dose of 7.5 mg/kg/d in 2-hour intravenous infusions over 3 to 7 days, promotes normocalcemia. In a randomized study, etidronate was shown to be superior to maximally approved doses of calcitonin. Moreover, when administered to patients without overt renal failure (serum creatinine less than 2.5 mg/dL), the drug is remarkably free of significant acute toxicity and side effects. Therefore, an important therapeutic aspect of this agent is that it permits concurrent antibiotic and chemotherapy administration. Judicious use of appropriate and specific pharmacologic intervention strategies in the patient with cancer-related hypercalcemia is expected to enhance quality of life, improve systemic cancer therapy tolerance, and reduce the morbidity of this serious neoplastic complication.

Animals↗
PubMed1989

Measurement of parathyroid hormone.

Interpretation of serum immunoreactive PTH measurements requires an understanding of the secretion, metabolism, and heterogeneity of circulating immunoreactive PTH. Both intact hormone and biologically inactive carboxyl fragments containing the middle and C-terminal regions are secreted by the parathyroid glands. Inactive fragments also are produced peripherally by metabolism of intact hormone by liver and kidney. Inactive fragments represent 75 to 95% of the total immunoreactivity in serum, a consequence of their long half-life in vivo as compared with intact hormone. Immunoassays for PTH can be divided into those measuring intact hormone (N-terminal, intact) and those measuring both inactive fragments and intact hormone (mid-region, C-terminal, polyvalent). The latter principally measures inactive fragments because of their greater concentration as compared with intact hormone in peripheral serum. The clinical utility of PTH assays varies considerably because of differences in their specificity and sensitivity. Serum PTH levels have been more often observed to be elevated in individuals with primary hyperparathyroidism with the use of research quality radioimmunoassays that recognize both inactive fragments and intact hormone than with conventional N-terminal or intact assays. Homologous mid-region assays have provided exceptional clinical sensitivity in confirming primary hyperparathyroidism. Comparison of a sensitive mid-region radioimmunoassay with a recently developed two-site, noncompetitive chemiluminescent immunoassay for intact PTH indicated that both methods were highly useful in the differential diagnosis of hypercalcemia. The mid-region assay provided the best diagnostic sensitivity in primary hyperparathyroidism with more elevated levels of PTH. The sensitivity of the intact assay was good, a significant improvement over conventional N-terminal and intact assays. The specificity of the intact assay was clearly superior, with measured PTH levels found to be suppressed to below normal in most subjects with hypercalcemia associated with malignancy. In contrast, measured levels were primarily normal with the mid-region assay. The higher levels of immunoreactive PTH observed in nonparathyroid hypercalcemia with the mid-region assay are in agreement with the measurement of biologically inactive carboxyl fragments, which continue to be secreted in hypercalcemia.

Amino Acid Sequence↗
PubMed1989

Vitamin D metabolite-mediated hypercalcemia and hypercalciuria patients with AIDS- and non-AIDS-associated lymphoma.

Fifteen patients with lymphoma and hypercalcemia (greater than or equal to 11.0 mg/dL) were identified by screening the serum chemistry profile obtained from patients upon admission to the Los Angeles County/USC Medical Center. Seven of the 15 (47%) possessed a frankly elevated serum concentration of 1,25-dihydroxyvitamin D [1,25-(OH)2-D]. An additional patient with severe hypercalcemia (16.2 mg/dL) had a serum 1,25-(OH)2-D concentration in the midnormal range, not a suppressed value. To examine the potential existence of hypercalciuria in absence of overt hypercalcemia, prospective screening of 23 normocalcemic patients with lymphoma was undertaken. Four of the 23 patients (17%) had increased fractional urinary calcium excretion rates (0.35 +/- 0.3 mg calcium/100 mL glomerular filtrate [GF], mean +/- SE; normal, less than 0.16 mg/100 mL GF); two of the hypercalciuric patients had a frankly elevated serum 1,25-(OH)2-D concentration. Of the 19 hypercalcemic/hypercalciuric lymphoma patients identified, none had an elevated serum immunoreactive parathyroid hormone concentration. Fourteen of the 19 hypercalcemic/hypercalciuric patients (74%) suffered from B-cell neoplasms, three had Hodgkin's lymphoma, and two had adult T-cell leukemia/lymphoma. All hypercalcemic/hypercalciuric patients had widespread disease (stage III or IV). Six patients, four with hypercalcemia and two with hypercalciuria, had acquired immunodeficiency syndrome (AIDS). These data suggest that the deregulated synthesis of a 1,25-(OH)2-D-like metabolite is a common cause of hypercalcemia and hypercalciuria in patients with lymphoma including patients with AIDS-associated tumors.

Adult↗
PubMed1988

Cobalamin and osteoblast-specific proteins.

Cobalamin deficiency has well-known hematologic and neurologic effects, but little is known about its other effects. We therefore studied the effect of cobalamin on osteoblast-related proteins. We found that mean (+/- 1 SD) levels of skeletal alkaline phosphatase in the blood were lower in 12 cobalamin-deficient patients (3.89 +/- 2.19 units per liter) than in 5 nondeficient and 5 iron-deficient control subjects (7.55 +/- 3.99 units per liter). The degree of the megaloblastic anemia correlated with the reduction in skeletal alkaline phosphatase levels (r = 0.67, P less than 0.01). With cobalamin therapy, levels of skeletal alkaline phosphatase rose in 11 of the 12 cobalamin-deficient subjects but not in the controls. The cobalamin-deficient patients also had significantly lower osteocalcin levels than the control subjects (1.11 +/- 0.77 vs. 1.84 +/- 0.49 nmol per liter). During cobalamin therapy, these levels rose in the cobalamin-deficient patients but not in the controls. In contrast to the levels of osteoblast-related proteins, hepatic alkaline phosphatase levels were similar in the patients and controls and were usually unaffected by cobalamin therapy. In vitro studies of calvarial cells from chicken embryos showed that their alkaline phosphatase content was cobalamin-dependent, thus supporting our in vivo observations in humans. Our findings suggest that osteoblast activity depends on cobalamin and that bone metabolism is affected by cobalamin deficiency, but we do not yet know whether cobalamin deficiency produces clinically important bone disease.

Adult↗
PubMed1988

Osteoclasts in human osteopetrosis contain viral-nucleocapsid-like nuclear inclusions.

We report the discovery of nuclear inclusions in the osteoclasts of three unrelated patients with benign osteopetrosis that resemble the osteoclast inclusions characteristic of Paget's disease of bone. These inclusions are morphologically and dimensionally identical to the nucleocapsids of a virus of the Paramyxoviridae family. Supporting a possible viral association with benign osteopetrosis in the observation of the presence of antigens of respiratory syncytial virus, measles virus, and/or mumps virus in the cells of all five patients whose paraffin-embedded bone specimens were tested. These included two patients whose osteoclasts contained nuclear inclusions. No patients with the malignant form of the disease have been studied. There is as yet no proof that a virus is causally related to human osteopetrosis even though a virus can produce an avian form of the disease.

Adult↗
PubMed1988

Hypercalcemia and abnormal 1,25-dihydroxyvitamin D concentrations in leprosy.

Two patients with lepromatous leprosy and hypercalcemia are presented. Serum immunoreactive parathyroid hormone and urinary cyclic adenosine monophosphate concentrations were suppressed. Serum 1,25-dihydroxyvitamin D [1,25-(OH)2D] concentrations were elevated in one patient and normal in the other. Urinary hydroxyproline excretion was slightly high in both patients. Hypercalcemia resolved excretion was slightly high in both patients. Hypercalcemia resolved with prednisone therapy. Abnormal 1,25-(OH)2D production and/or metabolism may play a role in the pathogenesis of hypercalcemia in some patients with leprosy.

Adult↗
PubMed1988

Gallium-67 citrate localization in osteoclast nuclei of Paget's disease of bone.

Gallium-67 citrate scintigraphy has been used to indicate the extent of bone involvement in patients with Paget's disease of bone and is an excellent marker in monitoring the effects of specific therapy. Since gallium uptake is dependent on cellular function, autoradiographic techniques can be applied to cells of Paget's lesions to understand better the mechanism of [67Ga]citrate uptake. Bone biopsies were obtained from sites of increased uptake using [67Ga]citrate scintigraphy in two patients with Paget's disease. In both patients electron microscopic autoradiographs demonstrated a high concentration of silver grains over the nuclei of osteoclasts. The cellular mechanism is unknown but may be related to the known inhibitory effect of calcitonin on osteoclast activity. The association of [67Ga]citrate with the nucleus of the osteoclasts is unique and different from tumor cells in which there is a high association of [67Ga]citrate with the lysosome fraction within the cytoplasm.

Autoradiography↗
PubMed1987

Therapy of hypercalcemia of malignancy.

An appreciation of the pathogenesis of the hypercalcemia of malignancy is essential to its management. At the outset, since most patients with symptomatic hypercalcemia of malignancy are dehydrated, infusion of 2 to 3 liters of saline per day will at least partially reduce serum calcium levels. Induction of calciuresis by infusing larger volumes of saline simultaneously with parenteral administration of furosemide may reduce the serum calcium concentration to normal in the short term. Of major importance in long-term therapy, however, are drugs that inhibit bone resorption, a major cause of hypercalcemia. These include calcitonin, plicamycin, glucocorticoids, prostaglandin synthetase inhibitors, and the diphosphonates. These agents may provide long-term control of hypercalcemia in many patients. Reduction of intestinal calcium absorption by dietary means or by glucocorticoid therapy is often effective in the rare subset of patients with increased serum levels of 1,25-dihydroxyvitamin D. Oral and intravenous phosphorus therapy may be effective via unknown mechanisms in some patients. The diphosphonates, in particular, should greatly facilitate management of both acute and chronic hypercalcemia of malignancy. Daily intravenous infusion of etidronate disodium (etidronate) with saline over a period of three to six days, for example, appears to be a safe and effective means of restoring serum calcium concentrations to the normal range. Study results have shown that more than 90 percent of patients have a response to etidronate. Oral administration of the drug has been demonstrated to maintain normal serum calcium concentrations.

Bone Resorption↗
PubMed1987

The spectrum of pathology in osteomalacia.

A classification and discussion of the histology of osteomalacia and the current histological techniques used in its identification are presented. Three categories of histologic features of osteomalacic bone are proposed: (1) increased osteoid, elevated osteoblast and osteoclast numbers and marrow fibrosis; (2) increased osteoid and osteoblast numbers, and (3) increased osteoid associated with few osteoblasts. Clinical conditions for which quantitative bone histomorphometric data are available are presented and related to the three categories. The consistency of clinical pathophysiology and histologic bone features is emphasized.

Bone and Bones↗
PubMed1987

Critical evaluation of viral antigen data in Paget's disease of bone.

This study evaluates previous viral antigen data obtained from fixed tissue sections and cells grown in culture from bone affected by Paget's disease. Finding antigens to both respiratory syncytial virus (RSV) and measles virus (MV) in the same osteoclasts of ten patients could not be explained on the basis of any previously known cross-reactivity. Therefore, possible causes for these observations were sought. Monoclonal antibodies to viral proteins of RSV and MV were used to label proteins. Polyclonal antibodies that were monospecific and were produced exclusively in nonhuman species were used to rule out nonspecific reaction with human proteins. Antivimentin antibody was used to test the possibility of cross-reactivity with a cytoskeletal protein, as a second antibody F(ab')2 conjugated to fluorescein was used to rule out nonspecific reactivity with Fc receptors. Electron microscopy was used to evaluate bone cell cultures derived from Paget's bone in comparison with Paget's osteoclasts. Results showed that the pattern of monoclonal viral antibody labeling followed different patterns in different patients. Nonspecific reactivity was ruled out by significant negative and positive controls. Cross-reactivity with vimentin could not account for the positive immunofluorescent results because of an entirely different pattern of fluorescence in the same samples of live and fixed cells after colchemid treatment. It was concluded that specific viral antigens are present in osteoclasts and in cells grown from Paget's bone and that the present data are compatible with the possibility that Paget's disease of bone is a slow virus infection.

Antibodies, Monoclonal↗
PubMed1987

Characterization and assay of tartrate-resistant acid phosphatase activity in serum: potential use to assess bone resorption.

We improved the spectrophotometric assay of tartrate-resistant acid phosphatase (TrACP; EC 3.1.3.2) activity in serum. During development of the assay we found that human serum contains a dialyzable, mixed-type noncompetitive inhibitor(s) of TrACP activity, the effects of which on the assay were substantially lessened by diluting the serum sample with water before assay and increasing the substrate concentration. Hemolysis releases into serum a significant amount of TrACP activity from erythrocytes, which can be inactivated by incubating the serum at 37 degrees C for 1 h before assay. Our improved assay was reproducible (CV = 5%), and measured within 10% of the amount of added bovine skeletal TrACP activity. Preliminary application of the assay revealed that the amount of serum TrACP activity in patients with skeletal diseases differed from normal values and changed in the same direction as the expected change in bone turnover, suggesting that TrACP activity in serum could be useful clinically as a marker of bone metabolism, possibly of bone resorption.

Acid Phosphatase↗
PubMed1986

The effects of metabolic acidosis on bone formation and bone resorption in the rat.

Metabolic acidosis (MA) has been implicated in the pathogenesis of both osteomalacia and osteopenia. Alterations in the secretion of parathyroid hormone and in the metabolism of vitamin D may contribute to such skeletal changes. To minimize the influence of these factors, quantitative bone histology and measurements of bone formation using double tetracycline labeling were done in thyroparathyroidectomized (TPTX) rats with MA induced by ammonium chloride (TPTX-A), and in both non-acidotic TPTX (TPTX-C) and intact (C) controls. To evaluate the response of both cortical and trabecular bone to MA, histologic studies were done at three separate sites in the tibia, cortical bone from the mid-shaft, and trabecular bone from the epiphysis and from the metaphysis. Plasma pH was lower in TPTX-A, 7.24 +/- 0.10, than in either TPTX-C, 7.39 +/- 0.03, or C, 7.43 +/- 0.04, P less than 0.01, and urinary hydroxyproline excretion increased from 89.8 +/- 8.7 in TPTX-C to 150.2 +/- 25.9 micrograms/mg/creatinine in TPTX-A, P less than 0.01. Resorption surface at the epiphysis increased from 1.8 +/- 0.6% in TPTX-C to 4.0 +/- 1.6% in TPTX-A, P less than 0.05, values not different from those in C, 3.1 +/- 1.1%. Resorption surface was unchanged at other skeletal sites, but total bone volume at the metaphysis fell from 15.5 +/- 5.6% in TPTX-C to 9.0 +/- 4.3% in TPTX-A, P less than 0.05. Bone formation was reduced at each skeletal site in TPTX-A vs. TPTX-C, P less than 0.05 for all values, but histologic evidence of osteomalacia was not observed.(ABSTRACT TRUNCATED AT 250 WORDS)

Acidosis↗
PubMed1986

Relationship between urinary calcium and calcium intake during calcitriol administration.

The hypercalciuria that occurs when 1,25 (OH)2D3 (calcitriol) is given to humans with normal renal function depends on dietary Ca absorption and may also relate, in part, to enhanced bone resorption. To evaluate the relationship between urinary and dietary Ca during treatment with calcitriol, 12 metabolic balance studies were performed in normal volunteers ingesting a diet containing 350 mg/day of Ca, to which Ca gluconate was added. After 10 days on either 350 mg/day or 1550 mg/day of Ca, calcitriol, 0.5 microgram every 12 hr, was given. Then diet Ca was changed in successive 5-day treatment periods from 350 to 650, 950 and 1550 mg/day (group A) or from 1550 to 950, 650 and 350 mg/day (group B). On the lowest diet Ca, urinary Ca was less than Ca intake during calcitriol treatment (group A, 220 +/- 50 mg/day; group B, 247 +/- 40). As diet Ca was changed during calcitriol treatment, urinary Ca correlated with diet Ca (r = 0.60) until diet Ca reached 950 mg/day. With calcitriol, serum iPTH fell by 18 to 25% (P less than 0.01) and urinary hydroxyproline fell by 11 to 19% (P less than 0.05 to 0.01). Baseline serum levels of 1,25(OH)2D were 47 +/- 8 and 34 +/- 5 pg/ml in group A and B, respectively, and the values increased to 51 +/- 12 and 45 +/- 7.4 pg/ml during treatment with calcitriol. Serum Ca from fasted subjects was not affected by calcitriol, but the mean postabsorptive serum Ca (moon) was increased by 0.35 mg/dl.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗