Effect of glutamine synthetase inhibition on astrocyte swelling and altered astroglial protein expression during hyperammonemia in rats.

Inhibition of glutamine synthesis reduces astrocyte swelling and associated physiological abnormalities during acute ammonium acetate infusion in anesthetized rats. We tested the hypothesis that inhibition of glutamine accumulation during more prolonged ammonium acetate infusion in unanesthetized rats reduces cortical astrocyte swelling and immunohistochemical changes in astrocytic proteins. Rats received a continuous i.v. infusion of either sodium acetate or ammonium acetate for 24 h to increase plasma ammonia from about 30-400 mumol/l. Cohorts were pretreated with vehicle or l-methionine-S-sulfoximine (MSO; 0.83 mmol/kg). MSO reduced glutamine synthetase activity by 57% and glutamine synthetase immunopositive cell number by 69%, and attenuated cortical glutamine accumulation by 71%. Hyperammonemia increased the number of swollen astrocytes in cortex and MSO reduced this increase to control values. The number of glial fibrillary acidic protein immunopositive cells in cortex was greater in hyperammonemic rats and the increase in superficial cortical layers was attenuated by MSO. Immunoreactivity for the gap junction protein connexin-43 in the neuropil, assessed by optical density, was greater in the hyperammonemic group compared with controls, but this increase was not attenuated by MSO. No changes in the optical density of GLT1 glutamate transporter immunoreactivity in cortex were detected in any group. We conclude that glutamine synthetase inhibition reduces astrocyte swelling and ameliorates some of the reactive astroglial cytoskeletal alterations seen at 24 h of hyperammonemia, but that gap junction changes in astrocytes occur independently of glutamine accumulation and swelling.

[Cognitive disorders appearing before the age of 65 in patients of the Alzheimer Centre of the VU Medical Centre: diagnoses and clinical characteristics]. / Cognitieve stoornissen optredend vóór de leeftijd van 65 jaar bij patiënten van het Alzheimercentrum van het VU Medisch Centrum: diagnosen en klinische kenmerken.

OBJECTIVE: To obtain a profile of the causes and clinical characteristics of cognitive disorders in patients referred to a memory clinic before the age of 65 years. DESIGN: Retrospective case-note study. METHOD: Data were collected from 127 subjects with objective cognitive disorders who visited the Alzheimer Centre of the VU Medical Centre in Amsterdam, the Netherlands, in the period from 1 January 2001 to 31 December 2003 with an onset of complaints before the age of 65. Besides the diagnoses, we investigated the clinical presentations, the occurrence of cardiovascular risk factors, the family history, and the presence of noncognitive neurological signs. RESULTS: The most common causes of cognitive decline under the age of 65 were Alzheimer's disease (46%) and frontotemporal dementia (23%). Vascular dementia was seen in 5% and dementia with Lewy bodies in 2%; 9% had mild cognitive impairment but no dementia. Hypertension and a positive family history for dementia were each present in 40% of the patients. Non-cognitive neurological abnormalities were found only in cases of non-Alzheimer dementia. During the period under investigation, the number of patients with objective cognitive disorders increased more than did the number without a cognitive disorder. CONCLUSION: Within the population of a memory clinic, Alzheimer's disease was the most frequent cause of cognitive decline under the age of 65, followed by frontotemporal dementia. The distribution differed from causes of dementia at an older age, where vascular dementia had the second place.

Concentration of bone elements in osteoporosis.

In aging and in osteoporosis, decreased bone density is associated with decreased bone mass. However, changes in the bone mineral phase remain a matter for investigation. In particular, it is unknown whether bone mineral loss is directly related to reduction in bone mass or associated with changes in the concentration of mineral elements in mineralized bone tissue. In this study, the cortical bone concentration of elements was determined in biopsies of the ilium from 33 subjects (12 controls and 21 individuals with untreated severe osteoporosis). Calcium and phosphorus concentrations were evaluated in cortical and trabecular bone using energy-dispersive x-ray (EDX) microanalysis and inductively coupled plasma optical emission spectrometry (ICPOES). Bone concentrations of Na, K, Mg, Cu, Zn, Fe, Sr, Al, B, and Si were also determined in cortical bone using ICPOES. Additionally, the concentration of F in cortical bone was measured with a specific ion electrode and the concentration of Pb was determined by atomic absorption spectrometry. In mineralized bone tissue there was no significant age-dependent variation in the concentration of Ca, P, or other elements either in controls or in osteoporotic subjects. Moreover, the concentration of elements in bone tissue did not differ in the two groups. These results suggest that the decrease in bone density in osteoporosis is directly related to evolution of the bone mass, without detectable changes in the concentration of elements in bone.

Altered trabecular architecture induced by corticosteroids: a bone histomorphometric study.

Prolonged corticosteroid (CS) therapy induces osteoporosis and fractures. Osteoporosis is characterized at the histomorphometric level by reduced bone volume (BV/TV) and disruption of the three-dimensional (3D) trabecular architecture. Several stereological methods have been proposed to characterize these alterations: measurements of trabecular thickness and trabecular number, star volumes, interconnectivity index (ICI) of the bone marrow spaces, and trabecular bone pattern factor (TBP(f)). These methods were computerized with a single program running on an image analyzer to evaluate the bone changes in a series of iliac biopsies performed on 31 male patients. All of them were asthmatic and had received CS for a long period of time. BV/TV was reduced when compared with age-matched controls. In the CS-treated population, exponential relationships were obtained between bone volume and the different connectivity parameters. The various methods used to measure connectivity were well correlated. When the population was divided into two groups (BV/TV greater or less than an 11% threshold), the architectural disturbances were found to imply two mechanisms. A progressive decline in trabecular thickness was noted in both groups versus controls. Trabecular perforations were not established in the group with BV/TV> 11% with the star volume or ICI, although some alterations were detected by trabecular bone pattern factor measurement. However, perforations were revealed in the group with BV/TV < 11% by all the different methods. Perforations seemed to occur when the trabecular thickness was below 70 mu m. This strongly suggests that bone histomorphometry should take into consideration bone volume in combination with detailed 3D descriptors of the trabecular architecture. Several histological methods need to be used in combination to appreciate the 3D architecture of trabecular bone.

Effects of tiludronate on bone loss in paraplegic patients.

Immobilization secondary to spinal cord injury is associated with a marked and rapid atrophy of trabecular bone (disuse osteoporosis). This is due to an early increase of osteoclastic bone resorption associated with a pronounced decreased osteoblastic bone formation. Bisphosphonates are antiosteoclastic compounds and they have been effective in preventing disuse osteoporosis. However, some of them also depress osteoblastic activity and may impair the mineralization process. Tiludronate was shown effective in reducing bone resorption in several metabolic bone diseases without inducing mineralization defects. Twenty paraplegic patients (6 females and 14 males) were randomly assigned to three groups: 6 patients entered the placebo group; 7 patients received tiludronate 200 mg/day; and 7 received 400 mg/day. Histomorphometric analysis was performed on transiliac bone biopsies before and after 3 months treatment. An insignificant decrease of bone volume was observed in the placebo group and the 200 mg group. In patients receiving 400 mg/day, a slight increase was noted. Osteoid parameters changed nonsignificantly in three groups although the 400 mg group exhibited a slight tendency to decrease osteoid volume and thickness. Eroded surfaces increased in all groups. The number of osteoclasts (identified histochemically by TRAP staining) increased in the placebo group but decreased in groups receiving tiludronate. Tiludronate appears effective in reducing bone resorption without impairing bone formation in a manner that preserved bone mass and bone cell coupling.

Isolation of osteoclasts from Pagetic bone tissue: morphometry and cytochemistry on isolated cells.

Giant osteoclasts and other cells were isolated from Pagetic bone tissue using 0.5 mM ethylene diamine tetraacetic acid on bone samples from 8 patients with Paget's disease. The cell suspension contained osteoclasts and osteoblasts as well as some mononuclear cells such as monocytes. The number of nuclei in isolated osteoclasts (33.85 +/- 20.92 nuclei/osteoclast) correlates fairly well (p less than 0.02) with the number of nuclei counted on histologic sections (15.88 +/- 11.80 nuclei/osteoclast) for samples from each patient. Enzyme histochemistry demonstrated acid phosphatase activity in isolated osteoclasts and in mononucleated cells, such as monocytes. Alkaline phosphatase was detected only in osteoblasts while succinate dehydrogenase was observed in osteoclasts, osteoblasts and monocytes. Esterases, such as nonspecific aliesterase and specific naphthol AS-D acetate esterase, were identified in osteoclasts and in macrophages. Inhibition of specific naphthol AS-D acetate esterase in osteoclasts by addition of sodium fluoride suggests that the enzyme could be of monocytic origin.

Fat in bone xenografts: importance of the purification procedures on cleanliness, wettability and biocompatibility.

Xenografting is a promising alternative to allografts and autografts. The remaining lipids in bone are known to influence the biocompatibility. A comparative study of wettability was done on standardized blocks of two biomaterials. A highly purified and defatted bovine bone graft (T650) was found to retain more water (2.06 g/block) than a less defatted biomaterial (T360, 0.3 g/block). Wettability, observed in the laboratory, may reflect an important in vivo property: the rapidity for extracellular fluids and blood cells to invade the graft and carry bone forming cells. When implanted in rabbit cancellous bone, T650 appears to be osteoconductive in a manner that allows trabecular architecture to be restored within 6 months.

Sarcomatous degeneration in Paget's bone disease.

The authors report 12 cases (8 men and 4 women) of sarcomatous degeneration in Paget's bone disease, with an average age of 72.3 years. Sarcomatous degeneration occurred often in polyostotic Paget's disease, and osteitis deformans was seen in 4 cases. Femur and pelvis were the most affected bones. Pain was a constant feature, whereas tumefaction and fracture were less common. Osteolytic lesions were more frequent than condensed or mixed lesions and radiological signs of malignancy were usually found. Seven cases were histologically classified as osteogenic sarcoma and 3 cases as fibrosarcoma. Electron microscopy was performed on 2 osteogenic sarcomas and in 1 case revealed microcylindrical inclusions in Pagetic osteoclasts and in multinucleated giant tumor cells, but none in mononucleated tumor cells. The average survival time for the patients in this study was only 4.5 months.

Somatostatin and adrenocorticotrophic hormone like immunoreactivity in small cell carcinoma of the lung.

The immunocytological detection of adrenocorticotrophic hormone (ACTH) and somatotropin release inhibitor factor (SRIF) like immunoreactivity was carried out on tumour cells from bronchial brush smears in 39 cases of lung tumours. Results obtained were compared with the cytological and histological diagnosis and confirmed the high incidence of ACTH synthesis by malignant bronchial carcinoma cells: the same phenomenon also seems to occur for somatostatin. The concomitant detection of ACTH and SRIF like immunoreactivity seems to be highly suggestive of small cell carcinoma and indicates that the immunocytological detection of hormones carried out at the same time as cytological examination can improve the accuracy of the diagnosis.

Osteoclast cytomorphometry in patients with femoral neck fracture.

In patients with femoral neck fracture, nutritional deficiencies have been shown to be common. A low calcium diet and/or a reduced vitamin D intake have been suspected to cause secondary hyperparathyroidism responsible for increased bone turn over and bone loss. Parathyroid hormone (PTH) levels are increased in these patients, data which are in accordance with the pronounced changes observed on bone biopsies reflecting a true hyperparathyroidism. We have used a cytomorphometrical approach to characterize PTH-induced changes on the osteoclastic population. Osteoclasts were detected histochemically (by tartrate resistant acid phosphatase staining) on bone biopsies from 10 control subjects, 8 patients with primary hyperparathyroidism and 10 patients with a femoral neck fracture of osteoporotic origin. The maximum Feret's diameter of each osteoclast (Oc.Le) was determined with a semiautomatic image analyzer. In all groups, the frequency distribution of Oc.Le appeared positively skewed. In both hip fractured patients and primary hyperparathyroid patients, the mode of the distribution was higher (25-30 microns) than in controls (20-25 microns). When graphically converted on a probability graph, the osteoclastic populations appeared homogeneous and well described by a lognormal distribution in the three groups. However, osteoclasts appeared similarly enlarged in the groups of patients with primary hyperparathyroidism and with femoral neck fracture. PTH has been shown to increase both the recruitment of mononucleated precursors and their fusion into larger osteoclasts than controls. In the present study, a cytomorphometric method appeared able to identify the border line hyperparathyroidism in the hip fractured patients.

Transposed ciliary microtubules in Kartagener's syndrome. A case report with electron microscopy of bronchial and nasal brushings.

Report is made of the case of a 44-year-old white woman with Kartagener's syndrome marked by respiratory disorders and repeated serous otitis since infancy. The technique of cell sampling through bronchial and nasal brushings facilitated observation of ciliary structures in electron microscopy. The results revealed a specific anomaly in the organization of the ciliary microtubules. The doublet transposition observed may be associated with ciliary dyskinesia.

[Cellular aspects of reorganization of pagetic bone tissue]. / Aspects cellulaires des remaniements du tissu osseux pagétique.

The light microscopy features of the cellular reorganisation of tone tissue in Paget's disease are well known. They result in the constitution of abnormal bone tissue. Ultrastructural studies have enabled a more detailed study of osteoblasts, osteocytes and osteoclasts. Osteoclasts are the only cells which present abnormalities. In particular, we observe intranuclear and intracytoplasmic inclusions composed of microcylinders. The morphological analysis is able to relate the presence of these formations with a viral infection of the paramyxovirus group. The results obtained by immunocytochemical techniques support this hypothesis. The osteoclast lesion could be a determining factor in the abnormal reorganization of bone tissue in Paget's disease, which may be considered to be a disease due to a slow acting virus.

[Current aspects of Paget's bone disease]. / Aspects actuels de la maladie osseuse de Paget.

The clinical and laboratory signs, as well as the imaging and course of Paget's disease of bone, are now well known. This chronic and usually benign disease is characterized by excessive remodelling of bone tissue, associated with an increase, sometimes considerable, of osteoclast resorption and osteoblast formation activities. Studies conducted during the last two decades were aimed at determining more precisely some aspects of the disease, notably: its epidemiological aspect and in particular its geographical distribution; its aetiological aspect using data obtained from electron microscopy, immunocytology and hydridization in situ, which has led to the hypothesis of a viral origin, and finally its therapeutic aspect with the recent introduction of truly effective treatments such as calcitonin and biphosphonates.

[Paget's bone disease and virus (author's transl)]. / Maldie osseuse de Paget et virus.

In spite of the large number of theories advanced to clarify the etiology of Paget's disease, its cause is still being discussed, and no satisfactory conclusion has been reached. The possibility of a viral origin was raised by the discovery of inclusion bodies, detectable by electron microscopy, in the nuclei and cytoplasm of the osteoclasts in the affected bone tissue, in 1974. In fact, the microcylindrical structures described by various authors, and visible only in osteoclasts and only in Paget's disease, if one excepts certain giant-cell bone tumors, have a close analogy, morphologically, with the nucleocapsids of paramyxovirus of the measles group, described in experimental infections or human diseases (subacute sclerosing panencephalitis). Various morphological arguments, drawn from studies of inclusions in richly nucleated giant osteoclasts found in Paget's disease, are in favor of the viral nature of these formations. Immunocytological methods have constituted another approach to the problem raised by the discovery of inclusions. They have demonstrated the existence of an antigenic material in the osteoclasts found in Paget's disease which reacts positively with antiserums containing anti-measles antibodies or with produce a crossed reaction with them. Controlled tests have confirmed these findings. Biological arguments are presently sufficient, therefore, for the possibility of a viral etiology of Paget's disease to be validly accepted from among the pathogenic hypotheses proposed for a disease that was first described a century ago.

[Sterno-costo-clavicular hyperostosis (author's transl)]. / Hyperostose sterno-costo-claviculaire. A propos de deux observations dont une forme partielle.

The authors report two cases of sterno costo clavicular hyperostosis. Six cases have been described in previous reports by Köhler (five cases) and CAMUS (one case). This disease begins with sterno costo clavicular pain that develops with exacerbations over several years. Biologically, there is a no specific inflammatory syndrome. Radiologically, clavicles sternum and first ribs are enlarged and increased in density. There is also an ossification of the sterno clavicular and sterno costal junctions. Phlebography sometimes show subclavian veins occlusion. Histological finding is hyperostosis without osteoclast inclusions. The main differential diagnosis is the Paget's disease.

[Viral origin of Paget's disease of bone?]. / L'origine virale de la maladie osseuse de Paget: quels arguments?

The viral aetiology of Paget's bone disease was suspected by the finding, in 1974, of microcylindric paramyxovirus nucleocapsid-like inclusions in nuclei and cytoplasm of pagetic osteoclast. Paramyxovirus antigens were detected, using monoclonal antibodies, in the osteoclasts of pagetic lesions. Paramyxovirus RNA sequences were identified in Paget's bone tissue, predominantly in osteoclasts, using specific hybridization. However, these last results are conflicting because some reports failed to reproduce hybridization with Paramyxovirus. Nevertheless, Paramyxovirus could be responsible for the cytopathologic aspect of multinucleated osteoclasts for the stimulation of hyperexpression, in Paget's disease, of interleukin 6 and of c-fos oncogene known to enhance osteoclastic resorption activity.

[Paget's bone disease. Epidemiology and etiology]. / Maladie osseuse de Paget. Epidémiologie et étiologie.

Paget's bone disease is a chronic and usually benign bone disorder mainly found in people over fifty. The condition is characterized by excessive bone remodelling related to bone cell overactivity leading to abnormal bone structure. The viral origin of the disease, first proposed on the basis of ultrastructural observations, is now supported by immunocytological and molecular hybridization studies. Genetic and environmental elements may be involved, favouring and modulating the expressivity of this slow virus infection.