Reversible inhibition of osteoclastic activity by bone-bound gallium (III).

Gallium(III) is a new therapeutic agent for hypercalcemia. Ga3+ reduces osteoclast action, but how it inhibits the cell's physiology is unknown. In vivo, 7-12 microM Ga(III) reduces calcium release from bone, but surprisingly, 10-100 microM Ga3+ added to isolated avian osteoclasts did not reduce their degradation of L-(5-3H)-proline bone. 3H-proline labels bone collagen specifically, and collagenolysis is an excellent indicator of bone dissolution because collagen is the least soluble component of bone. Ga(III) greater than 100 microM inhibited osteoclasts in vitro, but also killed the cells. To resolve this apparent conflict, we measured 67Ga distribution between bone, cells, and media. Gallium binds avidly but slowly to bone fragments. One hundred micrograms of bone clears 60% of 1 microM gallium from 500 microliters of tissue culture medium, with steady state at greater than 24 h. Osteoclasts on bone inhibited gallium binding capacity approximately 40%, indicating a difference in available binding area and suggesting that osteoclasts protect their substrate from Ga binding. Less gallium binds to bone in serum-containing medium than in phosphate-buffered saline; 30% reduction of the affinity constant suggests that the serum containing medium competes with bone binding. Consequently, the effect of [Ga] on bone degradation was studied using accurately controlled amounts of Ga(III) pre-bound to the bone. Under these conditions, gallium sensitivity of osteoclasts is striking. At 2 days, 100 micrograms of bone pre-incubated with 1 ml of 1 microM Ga3+, with 10 pmoles Ga3+/micrograms bone, was degraded at 50% the rate of control bone; over 50 pM Ga3+/micrograms bone, resorption was essentially zero. In contrast, pre-treatment of bone with [Ga3+] as high as 15 microM had no significant effect on bone resorption rate beyond 3 days, indicating that gallium below approximately 150 pg/micrograms bone acts for a limited time and does not permanently damage the cells. We conclude that bone-bound Ga(III) from medium concentrations less than 15 microM inhibits osteoclasts reversibly, while irreversible toxicity occurs at solution [Ga3+] greater than 50 microM.     

Punch shear testing of extracted vital and endodontically treated teeth

Plano-parallel specimens of human dentin cut from vital and endodontically treated teeth were tested by the punch shear test. Shear strength values were found to positively correlate with approximate toughness values. Statistically significant differences were found between shear strength and toughness values for vital and endodontically treated teeth, the latter showing lower values. The clinical impression that endodontically treated teeth are weaker and more brittle than vital teeth has therefore been quantitated. Anatomically different teeth or the methods used to store and cut teeth could not be consistently correlated with punch shear and toughness values. When dentin slices were constrained during punching so that bending was prevented, the precision of the results was improved and higher values were recorded.     

Bone marrow-derived mononuclear phagocytes autoregulate mannose receptor expression

This study extends our previous observation that surface mannose receptor expression by pure populations of CSF-1-dependent bone marrow-derived macrophages increases with time (Clohisy, D. R., Bar-Shavit, Z., Chappel, J. C., and Teitelbaum, S. L. (1987) J. Biol. Chem. 262, 15922-15929). We presently find, however, that the progressive enhancement of 125I-mannose-bovine serum albumin (125I-Man-BSA) binding per cell reflects cell number rather than duration of culture. In fact, macrophages plated at high density bind 8-fold more 125I-Man-BSA than do their low density counterparts, with no difference in receptor-ligand affinity. Furthermore, cells cultured at high density are ultimately subjected to lower levels of exogenously provided macrophage growth factor, and fewer are in interphase. By obtaining synchronous populations of quiescent bone marrow macrophages, however, we demonstrate that neither cell cycling nor attendant levels of colony stimulating factor-1 influence mannose receptor expression. Our next series of experiments established that density-related mannose receptor expression reflects removal, by marrow macrophages, of a "down-regulating" factor contained in culture medium. To this end, we treated mononuclear phagocytes with either macrophage- or control-conditioned medium and found that, via a fetal calf serum-residing protein(s), only control medium is capable of noncompetitively reducing 125I-Man-BSA binding in a dose-dependent manner. Moreover, reconstituted 20-40% (NH4)2SO4-precipitable fractions derived from either sham-conditioned medium or fetal calf serum are capable of down-regulating mannose receptor expression. Alternatively, the same fraction obtained from macrophage-conditioned medium contains no such activity. Finally, initial characterization of the down-regulating factor reveals it to be acid-activable and trypsin-sensitive, yet resistant to heating to at least 80 degrees C, ribonuclease A, or freezing and thawing. We conclude that bone marrow macrophages up-regulate expression of their own plasma membrane mannose receptor by inactivating a noncompetitive, serum-residing inhibitory protein(s).     

1,25-dihydroxyvitamin D3 and macrophage colony-stimulating factor-1 synergistically phosphorylate talin

Macrophage colony stimulating factor (CSF-1) and 1,25-dihydroxyvitamin D₃ (1,25(OH)₂D₃) are potent inducers of macrophage differentiation. Both appear to modulate protein phosphorylation, at least in part, through protein kinase C (PKC) raising the question as to whether they concurrently impact on macrophage-like cells. In this regard, we utilized the CSF-1 dependent murine macrophage-like line BAC 1.25F5. CSF-1 treatment of these cells for 30 min leads to particular phosphorylation of a 165 kDa protein, the putative CSF-1 receptor, and a 210 kDa moiety. 1,25(OH)₂D₃ exposure for 24 h prior to addition of CSF-1 enhances phosphorylation of the 165 kDa species and, especially, the 210 kDa protein. Phosphorylation of the latter protein is 1,25(OH)₂D₃ dose- and time-dependent and the molecule is specifically immunoprecipitated with a rabbit polyclonal anti-talin antibody. Experiments with okadaic acid show that the enhanced phosphorylation of talin does not result from serine phosphatase inhibition. CSF-1 and 1,25(OH)₂D₃, alone or in combination, do not increase talin protein expression. The tyrosine kinase inhibitor, genestein, blocks 1,25(OH)₂D₃/CSF-1 induced phosphorylation of the putative CSF-1 receptor but has no effect on talin phosphorylation which occurs exclusively on serine. In contrast to genestein, staurosporin, an inhibitor of PKC, inhibits phosphorylation of talin. Moreover, exposure of 1,25(OH)₂D₃ pretreated cells to phorbol 12-myristate 13-acetate (PMA) in place of CSF-1 also prompts talin phosphorylation. Finally, 1,25(OH)₂D₃ enhances ³[H]PDBu binding, indicating that the steroid increases PMA receptor capacity. Thus, CSF-1 and 1,25(OH)₂D₃ act synergistically via PKC to phosphorylate talin, a cytoskeletal-associated protein.     

Low codon bias and high rates of synonymous substitution in Drosophila hydei and D. melanogaster histone genes

We have evaluated codon usage bias in Drosophila histone genes and have obtained the nucleotide sequence of a 5,161-bp D. hydei histone gene repeat unit. This repeat contains genes for all five histone proteins (H1, H2a, H2b, H3, and H4) and differs from the previously reported one by a second EcoRI site. These D. hydei repeats have been aligned to each other and to the 5.0-kb (i.e., long) and 4.8-kb (i.e., short) histone repeat types from D. melanogaster. In each species, base composition at synonymous sites is similar to the average genomic composition and approaches that in the small intergenic spacers of the histone gene repeats. Accumulation of synonymous changes at synonymous sites after the species diverged is quite high. Both of these features are consistent with the relatively low codon usage bias observed in these genes when compared with other Drosophila genes. Thus, the generalization that abundantly expressed genes in Drosophila have high codon bias and low rates of silent substitution does not hold for the histone genes.      

Chemoembolization in the treatment of hepatic malignancy.

Numerous factors must be taken into account to best provide for the health and well-being of refugee patients in developed countries. One issue that is rarely considered is the awful and not uncommon occurrence of political torture. Large numbers of refugees and other displaced persons are survivors of political torture, and health care professionals must be prepared for this possibility when treating refugee patients. The effects of torture are pervasive, and we provide some practical considerations for health professionals who care for survivors of torture. Specific challenges include problems relating to exile and resettlement, somatic symptoms and pain, and the "medicalization" of torture sequelae.     

Cognitive and behavioural impairment among elderly people in institutions providing different levels of care.

OBJECTIVE: To compare the prevalence and degree of cognitive and behavioural impairment in elderly patients in institutions providing different levels of care. DESIGN: Prevalence study. SETTING: A nursing home, a home for the aged and psychogeriatric wards in a provincial psychiatric hospital. PATIENTS: Only subjects 65 years of age or older were eligible for inclusion. A random sample was selected comprising 25% of the residents in the nursing home and the home for the aged; of the 119 asked to participate 95 agreed (44 in the nursing home and 51 in the home for the aged). All 50 on the psychogeriatric wards agreed to participate. MAIN OUTCOME MEASURES: The Mini-Mental State Examination (MMSE) and the Kingston Dementia Rating Scale (KDRS). RESULTS: An MMSE score of less than 24 (cognitive impairment) was given to 37 (84%) of the residents in the nursing home, 43 (84%) of those in the home for the aged and 48 (96%) of the patients in the psychiatric hospital; the corresponding numbers for a KDRS score of more than 0 (cognitive impairment) were 41 (93%), 48 (94%) and 50 (100%). The seven patients receiving the highest level of care at the home for the aged (special care) had more behavioural problems than those in the psychiatric hospital did (p less than 0.001). CONCLUSIONS: Cognitive and behavioural impairment was widespread in the three institutions regardless of the level of care. When planning services and allocating resources government funding agencies should consider the degree and prevalence of such impairment among elderly people in institutions.     

Effects of diaphragmatic ischemia on the inspiratory motor drive.

To assess the effect of diaphragmatic ischemia on the inspiratory motor drive, we studied the in situ isolated and innervated left diaphragm in anesthetized, vagotomized, and mechanically ventilated dogs. The arterial and venous vessels of the left diaphragm were catheterized and isolated from the systemic circulation. Inspiratory muscle activation was assessed by recording the integrated electromyographic (EMG) activity of the left and right costal diaphragms and parasternal intercostal and alae nasi muscles. Tension generated by the left diaphragm during spontaneous breathing attempts was also measured. In eight animals, left diaphragmatic ischemia was induced by occluding the phrenic artery for 20 min, followed by 10 min of reperfusion. This elicited a progressive increase in EMG activity of the left and right diaphragms and parasternal and alae nasi muscles to 170, 157, 152, and 128% of baseline values, respectively, an increase in the frequency of breathing efforts, and no change in left diaphragmatic spontaneous tension. Thus the ratio of left diaphragmatic EMG to tension rose progressively during ischemia. During reperfusion, only the frequency of breathing efforts and alae nasi EMG recovered completely. In four additional animals, left diaphragmatic ischemia was induced after the left phrenic nerve was sectioned. Neither EMG activity of inspiratory muscles nor respiratory timing changed significantly during ischemia. In conclusion, diaphragmatic ischemia increases inspiratory motor drive through activation of phrenic afferents. The changes in alae nasi activity and respiratory timing indicate that this influence is achieved through supraspinal pathways.     

Genetic control of susceptibility to experimental allergic encephalomyelitis in mice

The genetic control of susceptibility to experimental allergic encephalomyelitis (EAE) was studied in mice. The results indicate that sensitivity to disease is not inherited in a simple Mendelian dominant way. Susceptibility to EAE is governed by genes located outside of the major histocompatibility complex and not byH-2-linkedIr genes. No correlation was observed between susceptibility to disease and the cellular immune response toward the small encephalitogenic protein.     

A comparison of diversity estimators applied to a database of host–parasite associations

Understanding the drivers of biodiversity is important for forecasting changes in the distribution of life on earth. However, most studies of biodiversity are limited by uneven sampling effort, with some regions or taxa better sampled than others. Numerous methods have been developed to account for differences in sampling effort, but most methods were developed for systematic surveys in which all study units are sampled using the same design and assemblages are sampled randomly. Databases compiled from multiple sources, such as from the literature, often violate these assumptions because they are composed of studies that vary widely in their goals and methods. Here, we compared the performance of several popular methods for estimating parasite diversity based on a large and widely used parasite database, the Global Mammal Parasite Database (GMPD). We created artificial datasets of host–parasite interactions based on the structure of the GMPD, then used these datasets to evaluate which methods best control for differential sampling effort. We evaluated the precision and bias of seven methods, including species accumulation and nonparametric diversity estimators, compared to analyzing the raw data without controlling for sampling variation. We find that nonparametric estimators, and particularly the Chao2 and second-order jackknife estimators, perform better than other methods. However, these estimators still perform poorly relative to systematic sampling, and effect sizes should be interpreted with caution because they tend to be lower than actual effect sizes. Overall, these estimators are more effective in comparative studies than for producing true estimates of diversity. We make recommendations for future sampling strategies and statistical methods that would improve estimates of global parasite diversity.