New methodology for evaluating osteoclastic activity induced by orthodontic load

Orthodontic tooth movement (OTM) is a dynamic process of bone modeling involving osteoclast-driven resorption on the compression side. Consequently, to estimate the influence of various situations on tooth movement, experimental studies need to analyze this cell.

Objectives

The aim of this study was to test and validate a new method for evaluating osteoclastic activity stimulated by mechanical loading based on the fractal analysis of the periodontal ligament (PDL)-bone interface.

Material and Methods

The mandibular right first molars of 14 rabbits were tipped mesially by a coil spring exerting a constant force of 85 cN. To evaluate the actual influence of osteoclasts on fractal dimension of bone surface, alendronate (3 mg/Kg) was injected weekly in seven of those rabbits. After 21 days, the animals were killed and their jaws were processed for histological evaluation. Osteoclast counts and fractal analysis (by the box counting method) of the PDL-bone interface were performed in histological sections of the right and left sides of the mandible.

Results

An increase in the number of osteoclasts and in fractal dimension after OTM only happened when alendronate was not administered. Strong correlation was found between the number of osteoclasts and fractal dimension.

Conclusions

Our results suggest that osteoclastic activity leads to an increase in bone surface irregularity, which can be quantified by its fractal dimension. This makes fractal analysis by the box counting method a potential tool for the assessment of osteoclastic activity on bone surfaces in microscopic examination.     

Fragmentation of the Golgi apparatus of motor neurons in amyotrophic lateral sclerosis (ALS). Clinical studies in ALS of Guam and experimental studies in deafferented neurons and in beta,beta'-iminodipropionitrile axonopathy.

Previous morphological immunoenzymatic studies with organelle-specific antibodies have disclosed an apparent fragmentation of the Golgi apparatus in large numbers of motor neurons in 12 cases of sporadic, non-Guamanian amyotrophic lateral sclerosis (ALS) in three cases of other types of motor neuron disease and in one case of a mitochondrial myopathy with cytochrome c oxidase deficiency. Motor neurons with fragmented Golgi apparatus were moderately atrophic; in these cells, discrete immunostained elements of the organelle were twice as many as in normal neurons, and the size of each Golgi element and the percentage of the cytoplasmic area occupied by the Golgi apparatus were reduced (Am J Pathol 1992, 140: 731-737). In this report we have confirmed the fragmentation of the organelle of motor neurons in the spinal cord in six sporadic cases of Guamanian ALS. In four of the six cases the clinical course was 1 to 2 years. The percentages of motor neurons with fragmented Golgi apparatus varied from 38 to 92. Motor neurons from three additional cases of Guamanian ALS of clinical duration from 5 to 7 years did not show fragmentation of the Golgi apparatus. In two cases of Guamanian ALS and in one non-Guamanian ALS, all neurons with ubiquitin-positive skein-like or granular inclusions believed to be pathognomonic for ALS had fragmented Golgi apparatus. To examine whether the fragmentation of the Golgi apparatus results from reactions to either neuronal deafferentation or to lesions of proximal axons, we conducted two experimental studies. In the first study, we examined in cats the Golgi apparatus of deafferented neurons of the dorsal lateral geniculate nucleus. In the second study, we examined the Golgi apparatus of motor neurons in the spinal cord of rats with proximal axonopathy induced by beta,beta'-iminodipropionitrile. In these two experiments, the neuronal Golgi apparatus studied by immunoenzymatic techniques and morphometry, was not fragmented. Taken together, the results of these studies strongly suggest that the fragmentation of the Golgi apparatus of motor neurons in ALS represents an important and perhaps early change of the organelle that may be involved in the pathogenesis of ALS. The fragmentation of the Golgi apparatus of motor neurons is a fairly specific and easily recognizable marker of ALS and may be used together with other criteria for comparisons between the human disease and proposed animal models of the disorder.     

Immunocytochemical study of the glial fibrillary acidic protein in human neoplasms of the central nervous system

The distribution of the glial fibrillary acidic protein (GFAP) was investigated in sections of 131 paraffin-embedded brain neoplasms obtained at surgery or at autopsy. The unlabeled antibody immunoperoxidase (peroxidase-antiperoxidase, PAP) method was used. Equally good results were obtained from 17-year-old material and from recent material derived at surgery or autopsy and fixed with Bouin fluid or phosphate-buffered formalin. The perikaryons and processes of reactive astrocytes showed the most intense stain for GFAP. Positive reaction to antibody against GFAP of varying intensity was demonstrated in astrocytomas of various grades of malignancy (32 of 32), glioblastoma multiforme (10 of 10), subependymal giant cell astrocytoma (1 of 1), ependymoma (2 of 10), subependymoma (4 of 4), and astrocytes in mixed neoplasms (8 of 8). In two neoplasms diagnosed as malignant astrocytomas and in four neoplasms diagnosed as glioblastoma multiforme, GFAP stain was limited to a few neoplastic cells. Usually the stain was more intense over processes than in perikaryons, with the exception of gemistocytic astrocytomas and the giant cells in glioblastoma multiforme, which showed an equally intense stain over perikaryons and processes. The periphery of Rosenthal fibers was intensely positive for GFAP. In astrocytic neoplasms the number of GFAP-positive cells and the intensity of the stain were inversely proportional to the degree of malignancy. In the following neoplasms the reaction for GFAP was negative: oligodendroglioma (3), oligodendroblastoma (1), medulloblastoma (3), medulloepithelioma (1), neuroblastoma (1), pineocytoma (1), typical teratoma of the pineal (1), fibrosarcoma (1), pituitary adenoma (2), craniopharyngioma (1), chordoma (1), chemodectoma of globus jugulare (1), metastatic carcinoma (17), and lymphoma (8). In one of 18 meningiomas, endogenous peroxidase activity was seen in mast cells. All meningiomas studied were negative for GFAP. In one of six neurinomas a positive reaction for GFAP was detected over processes. The authors concluded that the immunostain for GFAP is useful in the diagnoses of astrocytic neoplasms and of mixed gliomas.     

Stem and Progenitor Cell Expansionin Co-culture of Mobilized CD34~ Cells and Osteopetrotic Mouse Stroma

1 IntroductionCulture systems capable of expanding and/or maintaining hematopoietic stem cells will not only facilitate our understanding of stem cell biology, but also broaden clinical applications. Among various in vitro hematopoietic culture systems, co-cultures of marrow or CD34~ cells with an adherent stromal layer that can produce cytokines and extracellular matrix components most effectively supports long-term hematopoiesis (LTC), mimicking the bone marrow micro-environment.The OP-9 stromal cells ar...     

Nerve growth factor induced changes in the Golgi apparatus of PC-12 rat pheochromocytoma cells as studied by ligand endocytosis, cytochemical and morphometric methods

Summary Cells of the PC-12 rat pheochromocytoma cell line respond to nerve growth factor (NGF) by sprouting neurites and biochemically differentiating into sympathetic ganglion-like cells. NGF-stimulated (differentiated) and unstimulated (undifferentiated) cells were studied by cytochemical techniques for the localization of the enzymes acid phosphatase (ACPase) and thiamine pyrophosphatase (TPPase), and by a morphometric analysis of the distribution of endocytosed wheat-germ agglutinin labelled with horseradish peroxidase (WGA-HRP). Both cytochemical stains showed the enzymes to be distributed in lysosomes and certain cisternae of the Golgi apparatus in both NGF stimulated and unstimulated cells. ACPase was not confined to GERL (Golgi-endoplasmic reticulum-lysosome) as in certain other cells. The morphometric studies demonstrated that the reaction product of the internalized WGA-HRP occupied 4.7% of the cytoplasmic area in unstimulated cells and 4.5% in NGF-stimulated ones. Despite this similarity, the distribution of the WGA-HRP among the studied intracellular compartments in these two cell groups varied. In the NGF-stimulated cells 3.3% of the WGA-HRP reaction product was found in the innermost Golgi cisterna(e) while in unstimulated cells only 0.3% was seen in this compartment. Similarly, 4.3% of the WGA-HRP stain was found in small vesicles at the trans aspect of the Golgi apparatus in stimulated cells, when only 0.3% of the stain occupied this compartment in undifferentiated cells. The morphometric analysis also revealed that when the PC-12 cells were stimulated with NGF, the Golgi apparatus increased in area by approximately 70%. These findings are consistent with the hypothesis that NGF induced differentiation of PC-12 cells is coupled with enhanced endocytosis of WGA and probably of its receptor to the innermost Golgi cisterna(e) and the closely associated vesicles.     

Effect of Whitening Agents on Dentin Bonding

Background: Several studies have shown a reduction in enamel bond strengths when the bonding procedure is carried out immediately after vital bleaching with peroxides. This reduction in bond strengths has become a concern in cosmetic dentistry with the introduction of new “in‐office” and “waiting‐room” bleaching techniques. The aim of this in vitro study was to evaluate the effect of three bleaching regimens: 35% hydrogen peroxide (HP), 35% carbamide peroxide (CP), and 10% CP, on dentin bond strengths. Materials and Methods: One hundred and twenty fresh bovine incisors were used in this study. The labial surface of each tooth was ground flat to expose dentin and was subsequently polished with 600‐grit wet silicon carbide paper. The remaining dentin thickness was monitored and kept at an average of 2 mm. The teeth were randomly assigned to four bleaching regimens (n = 30): (A) control, no bleaching treatment; (B) 35% HP for 30 minutes; (C) 35% CP for 30 minutes; and (D) 10% CP for 6 hours. For each group, half of the specimens (n = 15) were bonded with Single Bond/Z100 immediately after the bleaching treatment, whereas the other half was bonded after the specimens were stored for 1 week in artificial saliva at 37°C. The specimens were fractured in shear using an Instron machine. Results: For the groups bonded immediately after bleaching, one‐way analysis of variance (ANOVA) followed by the Duncan's post hoc test revealed a statistically significant reduction in bond strengths in a range from 71% to 76%. For the groups bonded at 1 week, one‐way ANOVA showed that group B (35% HP for 30 min) resulted in the highest bond strengths, whereas 10% CP resulted in the lowest bond strengths. Student's t‐test showed that delayed bonding resulted in a significant increase in bond strengths for groups B (35% HP) and C (35% CP); whereas the group bleached with 10% CP (group D) remained in the same range obtained for immediate bonding. Storage in artificial saliva also affected the control group, reducing its bond strengths to 53% of the original.