Biomedical subjects

M Silbermann

Publications and source records attributed to M Silbermann.

At least 91 records · Page 5Linked to original sources

PubMed1988

Picrotoxin, a gamma-aminobutyric acid-receptor antagonist, retards craniofacial development in the weaning rat: I. Effect on mandibular bone growth.

The in vivo effects of elevated doses of picrotoxin, a gamma-aminobutyric acid (GABA) A-receptor antagonist, were studied in the skulls of weaning rats. Twenty-one-day-old male rats were treated daily with 2 mg/kg of pictroxin for a period of 3 weeks. This study revealed that chronic administration of the agent caused a reduction in bone formation in various growth sites in the skull along with a significant decrease in the calcium content and alkaline phosphatase activity in the mandible. Serum levels of calcium were unchanged, but the activity of alkaline phosphatase decreased. The decrease in bone alkaline phosphatase was accompanied by structural changes in the developing mandible. The latter was manifested by qualitative changes in the structure of ossification sites, in the appearance of the osteoblasts, and in the pattern of bone mineralization. These findings indicate that picrotoxin affects the normal growth of the craniofacial skeleton in an intact growing animal, probably because of central changes in GABA-ergic control on motor function along with possible alteration in corticosteroid secretion.

Alkaline Phosphatase↗

PubMed1988

Picrotoxin, a gamma-aminobutyric acid-receptor antagonist, retards craniofacial development in the weaning rat: II. Effect on mandibular condylar cartilage.

The in vivo effects of picrotoxin, a gamma-aminobutyric acid (GABA)-receptor antagonist, were studied in the mandibular condyles of weaning rats. Male rats 21 days old were treated daily with 2 mg/kg of picrotoxin for a period of 3 weeks. This study revealed that chronic administration of the agent caused a reduction in bone formation in various sites in the mandible, along with significant changes in the structure of the condylar cartilage and its ossification front. The length of the chondroblastic zone increased, yet the length of the hypertrophic zone was reduced. The latter phenomenon was manifested by qualitative changes in the overall structure of various cellular zones, in the appearance of the osteoblasts, and in the pattern of cartilage mineralization. The changes in the condylar cartilage cannot be attributed to a direct effect of picrotoxin; in vitro studies indicated no significant change in the incorporation of 3H-thymidine and 35S-sulfate in picrotoxin-treated cultures. These findings indicate that picrotoxin affects the normal growth of the mandible in an intact, growing animal, probably through an indirect route involving neurons in the central nervous system.

Animals↗

PubMed1988

Glucocorticoid hormone adversely affects the growth and regeneration of cartilage in vitro.

Apical portion of condylar cartilage of neonatal mice served as an experimental model to study the in vitro effects of elevated concentration of triamcinolone, a fluorinated synthetic analogue of cortisol on cartilage cell growth and development. Triamcinolone acetonide (10(-6) M) led to a significant decrease in the proliferative rate of chondroprogenitor cells along with an inhibition of the chondrogenic differentiation pathway. By 5 days in culture, control explants exhibited a pronounced increase in size and succeeded to reconstitute their native form. In hormone-treated cultures, the explants also elongated yet the increase was due to an accelerated cellular hypertrophy rather than increase in cell number. The findings of the present study which were based upon quantitative 3H-thymidine autoradiography as well as upon determination of 3H-thymidine incorporation suggest that in vitro corticosteroid hormone depresses pre-chondroblasts proliferation along with an interference in the normal differentiative pathway of the above cells.

Animals↗

PubMed1987

In vitro induction of osteosarcomalike lesion by transformation of differentiating skeletal precursor cells with FBR murine osteosarcoma virus.

This study analyzed the transforming potential of murine viruses in organ cultures of mouse fetal condylar cartilage: Finkel-Biskis-Jinkins-Murine sarcoma virus (FBJ-MuSV) and Finkel-Biskis-Reilly-Murine sarcoma virus (FBR-MuSV). It was only the FBR-MuSV isolated from a radiation-induced osteosarcoma, that induced morphological changes as early as 24 hours following the infection. The latter manifested itself by a marked enlargement of the number of progenitor cells concomitant with an accumulation of spindle-like cells, giant cells, and pleomorphic cells along with large bone spicules and heavy mineralization of the remaining cartilage. The newly formed tissue synthesized type I collagen and revealed profound invasive characteristics. By day 7, FBR-MuSV-infected cultures acquired the appearance of an osteosarcomatouslike lesion. Electron microscopy examinations revealed that both the matrix and the osteogenic cells in the induced tumors differed markedly from that encountered in normal mammalian osseous tissue. To determine which cells within the condylar tissue served as the target for the FBR-MuSV, we used antibodies against viral P30 protein for an indirect immunoperoxidase reaction. The chondroprogenitor cells were the only ones that reacted positively for the virus-specific protein. Further, the in vitro-induced tumor was tumorigenic in syngeneic mice, hence, brought about the development of osteo-fibrosarcoma subcutaneously. By contrast to FBR-MuSV, the FBJ-MuSV did not elicit similar transformative effects in vitro. Since both viruses possess the fos oncogene, it has been suggested that the unique tumorigenic potential of the FBR-MuSV may be linked to structural alterations in the fos oncogene product.

Animals↗

PubMed1987

Effects of increased doses of 1,25 dihydroxyvitamin D3 on matrix and DNA synthesis in condylar cartilage of suckling mice.

The in vivo effects of high doses of 1,25(OH)2D3 were studied in condylar cartilage of suckling mice. Seven-day-old animals were treated with 20 ng of the hormone for 7 consecutive days. Biochemical assays on collagen content and synthesis were complemented by structural studies using light and electron microscopy. Indirect immunofluorescent methods were used for the localization of type I and II collagens and for fibronectin. This study revealed that the protein content of the condyle decreased substantially following the administration of the hormone. Protein synthesis increased in hormone-treated animals during the first 4 days but was significantly inhibited thereafter. Collagen synthesis, however, was inhibited instantaneously, followed by a decrease in the percentage of cold hydroxyproline of the total protein. Hormone-treated condyles showed a marked decrease in the distribution of type I collagen, no apparent change in the distribution of type II collagen, but an enhanced reactivity for fibronectin especially around hypertrophic chondrocytes. SDS-gel electrophoresis of collagen chains suggested that the hormone did not induce a significant change in the ratios of type I and II collagen chains, yet additional peaks became evident in 1,25(OH)2D3-treated specimens. The decrease in collagen synthesis was accompanied by ultrastructural changes in the appearance of the extracellular collagen bundles. They later appeared as a dense meshwork of collagen fibrils, a feature that was lacking in control tissues. The changes in collagen fibrillogenesis could be explained by our in vitro studies indicating a marked depression of 35S-sulfate incorporation secondary to treatment with 1,25(OH)2D3. The hormone was also found to suppress the incorporation of 3H-thymidine, hence it may be concluded that 1,25(OH)2D3, when used in high concentrations, possesses an inhibitory effect upon both the proliferative activity of the cartilage progenitor cells as well as upon the metabolic activity of the condylar cells as related to collagen and glycosaminoglycans synthesis.

Animals↗

PubMed1987

The in vitro behavior of fetal condylar cartilage in serum-free hormone-supplemented medium.

Mandibular condyles of fetal mice 19-20 days in utero were kept in a serum-free organ culture system for up to 14 days and were investigated for their capacity to develop osteoid and to mineralize in vitro. After 3 days in culture, the cartilage of the mandibular condyle appeared to have maintained all its inherent structural characteristics, including its various cell layers: chondroprogenitor, chondroblastic, and hypertrophic. After 7-9 days in culture, no chondroblasts could be seen; instead, most of the cartilage consisted of hypertrophic chondrocytes. In addition, various areas throughout the explant revealed the appearance of osteoidlike material. The process of matrix mineralization progressed with time, and by the 14th day new bonelike material was found to occupy a larger portion of the explant. The newly formed matrix reacted positively with antibodies against type I and type III collagens, as well as against bone alkaline phosphatase. Electron microscopic examination showed that the mineralization appeared to be associated with collagen fibers as well as the matrix vesicles. In composition, the in vitro-formed mineral deposits resembled hydroxyapatite crystals. Biochemical assays indicated that the newly formed tissue reacted strongly for alkaline phosphatase and incorporated 45Ca. The findings of the present study imply that fetal condylar cartilage possesses the potential to develop in vitro osseouslike tissue even in a system that is serum-free. Due to the fact that the newly formed extracellular matrix mineralized and reacted positively to bone markers as well as to cartilage macromolecules, it would seem justifiable to define the new tissue as chondroid bone.

Alkaline Phosphatase↗

PubMed1987

Acceleration of cartilage and bone differentiation on collagenous substrata.

Chondroprogenitor cells of newborn murine mandibular condyles were cultured on top of collagen sponges for up to 18 days. After 24 h in culture, new chondroblasts developed which subsequently matured showing signs of hypertrophy, while the extracellular matrix revealed positive reactivity for type II collagen, cartilage proteoglycans and mineralization. Light and electron microscopy examinations showed signs of new osteoid formation, a feature that was preceded by positive immunohistochemical reaction for type I collagen, fibronectin and bone specific sialoprotein. A close temporal and spatial association was noted between the development of mature, mineralized cartilage and new osteoid. The differentiation of new cartilage and bone cells was linked to an increased activity of DNA synthesis and cellular proliferation. The de novo bone formation was accompanied by increasing rates of alkaline phosphatase activity and uptake of [45Ca] features that were found to be tightly correlated to each other. The collagen substrata appeared also to facilitate the migration of cells, their replication and their subsequent differentiation to their respective cellular lineage. Hence, collagen sponges in vitro appear to serve as a promising substrata for culture systems involved with the growth and differentiation of mineralizing tissues such as cartilage and bone.

Alkaline Phosphatase↗

PubMed1987

Further characterization of the chondroprogenitor zone in mandibular condyles of suckling mice. An ultrastructural and cytochemical study.

In the neonatal mouse the mandibular condyle serves as an important growth center for the developing mandible. The youngest cells in this organ are the chondroprogenitor cells that are the source for new differentiating chondroblasts. The present study provides new data concerning the fine structure and cytochemical characteristics of the cartilage precursor cells as seen in suckling mice. The condylar chondroprogenitor cells normally reveal a mesenchyme-like appearance with multiple, elongated cell processes that enable close contact between neighboring cells. These cells also exhibit a variety of pinocytotic vesicles, coated pits and appear to be actively involved in the internalization of a fluid-phase marker horseradish peroxidase. Further, the progenitor cells were found to contain trimetaphosphatase reaction products within lysosomal bodies and alkaline phosphatase reactivity along their plasma membrane. Precipitates of calcium complexes in the form of calcium pyroantimonate could be demonstrated in association with the plasmalemma of the cells as well as with the extracellular collagen fibrils. Matrix granules, representing cartilage proteoglycans, became a distinct feature following staining with ruthenium red and were found to be in close contact with the extracellular collagen fibrils and with the plasmalemma. Hence, in addition to their active role in cell proliferation, the progenitor cells are also involved in the synthesis and secretion of major extracellular macromolecules such as collagen and proteoglycans.

Acid Anhydride Hydrolases↗

PubMed1987

Effects of long-term running stress on the ultrastructure of the aging mouse heart.

Apical regions of the left ventricular myocardium of female C57BL/6J mice, aged between 6 and 27 months, that had been subjected to endurance running for 2, 10 or 15 months, were examined by light and electron microscopy. Focal degenerative changes were found in myocytes of these experimental mice, including loss of myofibrils, erosion of myocytes and changes in nuclei, lipofuscin content and mitochondria. The interstitial connective tissue showed an increase in collagen in aged myocardia. Endurance-trained runners frequently showed signs of platelet aggregation or microthrombi in small capillaries associated with myocytes. These were most common in endurance-trained middle-aged and very elderly mice, but were not found in sedentary age-matched controls. Our experiments indicate that running stress may be damaging to cardiac tissue in aged animals. Our findings may be of significance in helping the understanding of the development and the etiology of stress-induced cardiovascular damage in the elderly.

Aging↗

PubMed1987

Giant mitochondria in the myocardium of aging and endurance-trained mice.

During the course of an ultrastructural study on the apex region of the left ventricular myocardium of experimental female C57BL/6J mice, aged between 6 and 27 months, and that had been subjected to long-term daily enforced endurance running schedules, we encountered some instances of unusual giant mitochondria. These mitochondria showed degenerative changes including the disruption, disorganization and loss of cristae and the development of a very electron-lucent matrix. In many instances, these giant mitochondria showed dense inclusions within the matrix. The giant mitochondria appear to have developed as a result of fusion between adjacent hypertrophic mitochondria followed by a sequence of progressive degenerative changes. These giant mitochondria were most common in the myocardium of mice that had followed endurance training schedules for 10 or 15 months and were not encountered in age-matched nonrunning control mice.

Aging↗

PubMed1987

Effect of short-term physical stress on DNA and collagen synthesis in the femur of young and old mice.

Aging of long bone in the hindleg of the mouse is accompanied by a progressive cavitation of the cortical bone along with clear atrophic structural changes characteristic of senile osteopenia. Quantitative analysis indicates that there is a nonsignificant change in the DNA content, but an increase in the protein percentage of the total bone wet weight. Bones of old animals incorporate significantly less (3H)-thymidine and markedly less (3H)-proline. This study revealed that endurance training in old animals induced a significant increase in DNA and collagen synthesis, yet the overall wet weight of the corresponding bone decreased. It thus became apparent that the femur of old animals possesses the capacity to respond to enforced training, yet the latter contains elements characteristic of a negative type of adaptation.

Aging↗

PubMed1987

Dexamethasone impairs growth and collagen synthesis in condylar cartilage in vitro.

The in vitro effects of dexamethasone on condylar cartilage from normal newborn mice were tested by measuring protein and DNA content, collagen synthesis, prolyl hydroxylase activity, collagen chains and by immunofluorescence the localization of type I and II collagen and fibronectin. The biochemical assays were complemented by structural studies of hormone-treated and control cultured specimens. It became apparent that both the protein and DNA content of the tissue decreased immediately following the addition of dexamethasone of the incubation system. Protein synthesis was significantly decreased by the hormone by 24 h. The degree of collagen hydroxylation was decreased by 24 h. Dexamethasone-treated condyles did not reveal a significant increase in the percentage of cold hydroxyproline of the total protein. Using the indirect immunofluorescence method, hormone-treated condyles revealed an enhancement of positive reactivity for type I collagen and fibronectin. SDS-gel electrophoresis of 3H-labeled collagen chains isolated by CM-cellulose chromatography indicated that dexamethasone did not significantly affect the ratios of the collagen chains. For further characterization, each chain was subjected to cyanogen bromide cleavage showing that the peptide maps of alpha 1(I) and alpha 1(II) chains were not different in dexamethasone-treated tissues in comparison to controls.

Animals↗

PubMed1987

Age-related changes in the role of matrix vesicles in the mandibular condylar cartilage.

A combined approach of light microscopy, immunofluorescence, transmission electron microscopy and electron energy loss spectroscopy (EELS) was used to study age-related changes in the condylar cartilage in mice. Chondrocalcin, a cartilage matrix calcium-binding protein, was demonstrated by indirect immunofluorescence microscopy using monospecific antibodies. In one week old animals the most intense staining was observed in the matrix around the hypertrophic cells in the mineralising zone, to a lesser degree around the cells in the zone of chondroblasts, while no staining was noted in the zone of chondroprogenitor cells and in the matrix around the early hypertrophic cells. In the mineralisation zone the distribution of chondrocalcin correlated with that of mineral deposits as revealed by the von Kossa stain. The matrix between the early hypertrophic cells as shown by transmission electron microscopy revealed the presence of matrix vesicles and demonstrated a gradual accumulation of hydroxyapatite in the mineralising zone. In one month old animals chondrocalcin localisation was mainly confined to the lower hypertrophic zone which also demonstrated positive von Kossa staining was seen along the articular surface. In older animals multiple electron-dense structure that resembled matrix vesicles were observed in the non-mineralising portions of the condylar cartilage. Use of the EELS method confirmed the almost complete lack of calcium ions in these structures. In contrast, with the use of the same method, detectable amounts of calcium were recorded in vesicles in the mineralising zones of all age groups. Hence what appear ultrastructurally as structures similar to matrix vesicles represent atypical vesicles that might characterise an ageing and degenerative articular cartilage and are not necessarily associated with the mineralisation process.

Aging↗

PubMed1987

Further characterisation of the extracellular matrix in the mandibular condyle in neonatal mice.

This study provides newer information concerning the extracellular matrix of neonatal condylar cartilage--a genuine representative of a secondary type of cartilage. In addition, the data presented hereby indicate that the condylar cartilage contains a population of progenitor cells that synthesise Type I collagen rather than Type II. Under normal conditions in vivo local biomechanical factors influence the progenitor cells to differentiate into cartilage cells and thereby to shift their synthetic pathway from Type I collagen to Type II collagen--the typical collagen of cartilage extracellular matrix. In the absence of such biomechanical effects the condylar progenitor cells seem to proceed with their inherent differentiation pathway and express an osteogenic phenotype (Fig. 21).

Animals↗

PubMed1987

Age-related trend for osteopenia in femurs of female C57BL/6 mice.

Age-related changes in femoral cortical bone were quantified in female C57BL/6 mice. Variables included in this study were cortical thickness; number of osteocytes and periosteal preosteoblasts; bone DNA, protein, calcium, and phosphorus content; and uptake of 3H-thymidine, 3H-proline, and 45CaCl2. Also, the activity of bone alkaline and acid phosphatase was measured as well as the hydroxyproline content of the tissue. Femurs of old mice 28 months old, compared to those of young mice 7 months old, showed significant decreases in cortical thickness, number of bone cells, bone mineral content, and enzyme activity. The latter were accompanied by a marked reduction in ability to incorporate radiolabeled precursors of DNA and collagen synthesis as well as of the mineralization process. Hence, aging female C57BL/6 mice reveal structural and biochemical features indicative of developing osteopenia.

Aging↗

PubMed1987

Chondroid bone arises from mesenchymal stem cells in organ culture of mandibular condyles.

Mandibular condyles of fetal mice 19 to 20 days in utero comprising clean cartilage and its perichondrium were cultured for up to 14 days, and their capacity to develop osteoid and to mineralize in vitro was examined. After 3 days in culture the cartilage of the mandibular condyle appeared to have lost its inherent structural characteristics, including its various cell layers: chondroprogenitor, chondroblastic, and hypertrophic cells. At that time interval no chondroblasts could be seen; instead, most of the cartilage consisted of hypertrophic chondrocytes. By that time, the surrounding perichondrium, which contains pluripotential mesenchymal stem cells, revealed the first signs of extracellular matrix enclosing type I collagen, bone alkaline phosphatase, osteonection, fibronectin, and bone sialoprotein as demonstrated by immunofluorescent techniques. Electron microscopic examinations of the newly formed matrix revealed foci of mineralization within and along collagen fibers as is normally observed during bone development. The composition of the latter mineral deposits resembled calcium pyrophosphate crystals. Following 14 days in culture larger portions of the condyle revealed signs of osseous matrix, yet the tissue reacted positively for type II collagen. Hence, the condylar cartilage, a genuine representative of secondary-type cartilage, elaborated in vitro a unique type of bone that would be most appropriately defined as chondroid bone. Biochemical assays indicated that the de novo formation of chondroid bone was correlated with changes in alkaline phosphatase activity and 45Ca incorporation. The findings of the present study imply that mesenchymal stem cells that ordinarily differentiate into cartilage possess the capacity to differentiate into osteogenic cells and form chondroid bone.

Animals↗

PubMed1986

Supraphysiological concentrations of dexamethasone induce elevation of calcium uptake and depression of [3H]-thymidine incorporation into DNA in cartilage in vitro.

When condylar cartilages of neonatal mice are treated in vitro with dexamethasone they respond with a significant depression of DNA synthesis. Concomitant with the retardative effect upon the incorporation of [3H]-thymidine, dexamethasone enhances the uptake of 45Ca by the condylar tissue. The hormone's effect upon calcium uptake is relatively rapid and takes place within 2 hours whereas the effect upon the incorporation of [3H]-thymidine takes place after a lag period of 24 hours. The present findings tend to indicate the casual relationship between the above effects of dexamethasone in fast growing cartilage. Further, there is evidence that both of the above responses are specific and are protein and mRNA synthesis dependent. Control studies using both a calcium ionophore (A23187) and a calcium blocker (D-600) provided data that further substantiate the proposed association among dexamethasone, calcium metabolism, and DNA synthesis in neonatal growth cartilage.

Animals↗

PubMed1986

Parathyroid hormone stimulates proliferation of chondroprogenitor cells in vitro.

Mandibular condylar explants of newborn ICR mice were maintained as serum-free organ culture systems and were used to study the effects of 0.1-10.0U/ml parathyroid hormone (PTH) on the morphology of the organ and the ultrastructure of the chondroprogenitor cells. Parameters of proliferation such as 3H-thymidine autoradiography and incorporation into the explants were also studied. The chondroprogenitoric zone gradually increased with increasing dosages of the hormone up to a maximum of 5-fold of the control with 5.0 U/ml PTH. Autoradiographic studies showed a 3-fold increase in the number of 3H-thymidine-labeled cells in the chondroprogenitoric zone of PTH-treated explants. This was matched by a dose-dependent stimulation of 3H-thymidine incorporation, reaching maximal values at 5.0 U/ml PHT. At this concentration, the stimulated incorporation of 3H-thymidine was found to be dependent on the Ca2+ concentration of the medium. Chondroprogenitor cells located adjacent to the chondroblastic zone tended to pile up and aggregate in "syncytium"-like clusters, establishing intercellular gap junctions. All PTH-treated chondroprogenitor cells demonstrated large deposits of glycogen and highly elaborated stacks of their Golgi systems; the latter were associated with large numbers of vesicular elements. On the other hand, the chondroblastic zone was significantly reduced in size. Hence, it seems that PTH possesses a rather intense mitogenic effect upon chondroprogenitor cells and might possibly interfere with their normal pattern of differentiation into mature cartilage cells.

Animals↗