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Biomedical subjects

D Lewinson

Publications and source records attributed to D Lewinson.

At least 37 records · Page 2Linked to original sources
PubMed1991

In vitro response of neonatal condylar cartilage to simultaneous exposure to the parathyroid hormone fragments 1-34, 28-48, and 53-84 hPTH.

Mandibular condylar explants of neonatal ICR mice were maintained as serum-free organ culture systems and were used to study the effects of three synthetic fragments of human parathyroid hormone (hPTH) on the morphology of the organ and its ability to incorporate [3H]thymidine. Forty-eight-hour incubation with hPTH (1-34), at a concentration of 0.5 microgram/ml caused an increase of 88% in DNA synthesis and a marked increase in the size of the chondroprogenitor zone. The mitogenic effect of hPTH (1-34) was decreased to 34% over control levels when the fragment hPTH (28-48) was added to the system. However, the addition of the latter fragment brought about a marked enhancement in the mineralization of the cartilaginous extracellular matrix along with the formation of an appreciable amount of new bone. The de novo osseous tissue was attached to the mineralized cartilage. When the carboxyl-terminal fragment hPTH (53-84) was added together with the other two fragments, the mitogenic effect of hPTH (1-34) was completely abolished and the respective cultures incorporated [3H]thymidine even less than untreated control cultures. Moreover, the addition of hPTH (53-84) to the culture system led to distinct structural features throughout the mineralized hypertrophic cartilage. The latter contained a mixture of cells within an unorganized extracellular matrix. Untreated control cultures lacked such structures, but contained the various cell zones as normally seen in neonatal condylar cartilage. Therefore, it seems reasonable to suggest that each of the three fragments tested induces a biological effect on neonatal cartilage and might be involved in the normal process of endochondral ossification.

Animals↗
PubMed1990

Microvascular invasion of rabbit growth plate cartilage and the influence of dexamethasone.

The normal morphological features of growth plate angiogenesis were examined in rabbits and compared with changes induced by dexamethasone. Penetration of growth plate cartilage was led by perivascular cells with some contribution by luminal capillary endothelial cells. There was a close relationship between the invasive perivascular cells and the luminal endothelial cells of the capillary tip. Growth plates from rabbits treated with dexamethasone underwent major changes in the pattern of capillary invasion. Most striking was the appearance of numerous narrow and tortuous channels which penetrated the cartilage, in some cases forming complete loops. These channels were filled with debris or red cells but did not contain capillaries. It is suggested that dexamethasone treatment leads to channel formation by disrupting the normal control of capillary invasion.

Animals↗
PubMed1990

Histochemical study of matrical components in the mandibular condyle of the neonatal mouse.

In the mandibular condyle of the newborn mouse the chondroprogenitor (CP) zone is the only layer that incorporates 3H-thymidine thus serving the source for cells of the cartilage lineage. Ultrastructurally these cells have a mesenchymal appearance surrounded by collagen fibrils as well as by additional filaments that become apparent following fixation with ferrocyanide-reduced OsO4. In addition, electron-dense particles indicative of proteoglycans are scattered throughout the matrix in the CP zone as well as in the chondroblastic and hypertrophic zones. Following labeling with 35S-sulfate the CP zone as well as the other compartments revealed a substantial number of grains following processing for autoradiography. The number of grains per cell was highest in the hypertrophic zone. Indirect immunofluorescence indicated the presence of fibronectin in the articular surface, CP zone and in the hypertrophic zone. The immunogold method localized fibronectin intracellularly in CP cells and extracellularly in the hypertrophic zone. Therefore, in the mandibular condyle the CP cells which are capable for DNA synthesis are also involved in the synthesis of macromolecules of which some are specific for the cartilage phenotype, while others are associated with other functions of connective tissue cells.

Animals↗
PubMed1990

Effects of parathyroid hormone fragments on the growth of murine mandibular condylar cartilage in vitro.

The present study showed that the fragment hPTH (1-34) is mitogenic in organ cultures of neonatal mandibular condylar cartilage, and even more so in late fetal condyles. Three fragments of hPTH were used to clarify which part of the molecule possesses a mitogenic effect alike that of the native hormone: hPTH (1-34), (28-48) and (53-84). [3H]thymidine incorporation into trichloracetic acid insoluble material and quantitative autoradiography were employed in a serum-free medium to assess the effects of these fragments while light and electron microscopy studies served for morphological evaluations. It became evident that the fragment hPTH (1-34) enhanced the incorporation of [3H]thymidine, a fact that could be noted only in serum-free medium. The putative target cells for the effects of hPTH (1-34) were the chondroporogenitor cells which also appeared to have experienced a blockage in their differentiation into chondroblasts. Ultrastructurally, the latter cells responded in the formation of adherent profiles of plasma membranes, whereas the differentiated zone of the cartilage reduced its size. Using serum-free medium, hPTH (1-34) also brought about an inhibition in alkaline phosphatase activity, a fact that was not encountered in medium containing serum. By contrast, hPTH (28-48) had no mitogenic effect, although treated specimens revealed morphological changes in the chondroprogenitor cell zone along with an enhancement of cartilage cells hypertrophy. No significant effects on either mitogenecity or morphology could be noted in hPTH (53-84)-treated cultures.

Animals↗
PubMed1989

Application of the ferrocyanide-reduced osmium method for mineralizing cartilage: further evidence for the enhancement of intracellular glycogen and visualization of matrix components.

The ferrocyanide-reduced osmium (FRO) fixation method was applied to neonatal mouse mandibular condylar cartilage for its processing for electron microscopy. The results were compared to those obtained by the conventional glutaraldehyde-osmium tetroxide fixation method. Three different stages in the life cycle of condylar cartilage cells were examined. FRO enabled the visualization of delicate fibrillar mesh in the matrix of all three zones of the cartilage, resulting in a dense appearance of the intercellular matrix. The classical stellate shape of matric granules seen in cartilage fixed with glutaraldehyde-osmium tetroxide was not observed in FRO-processed tissues. Chondrocytes that were FRO-processed almost entirely filled their lacunar space. In their pericellular area, fibrillar material and electron-dense aggregates could be demonstrated by the FRO method. As a conclusion of this study, it is recommended to supplement a conventional protocol with the FRO fixation method for routine and research purposes.

Animals↗
PubMed1989

Effects of different concentrations of serum on cartilage growth in an organ culture system.

The purpose of the present study was to examine the effects of various concentrations of serum on the behavior of neonatal condylar cartilage when cultured in an organ culture system. Mandibular condylar cartilages were obtained from newborn ICR mice, of which the zone of undifferentiated chondroprogenitor cells along with a few layers of young cartilage cells were cultivated at the medium-air interface. The incubation medium included fetal bovine serum at concentrations ranging from 0 to 10%, and the explants were kept in vitro up to 10 d. The serum-free medium maintained the chondrogenic expression, and the overall size of the cartilagenous portion of the explants increased with the decrease of the concentrations of serum in the medium. When explants were labeled with [3H]thymidine and were then processed for autoradiography, the peak of labeling was noticed at 48 h, a feature that recapitulated itself in all cultures (73, 140, 175, 201, and 129 labeled cells per chondroprogenitor zone in explants grown in 0, 1, 2.5, 5, and 10%, respectively). It can be concluded that serum-free medium maintains the chondrogenic phenotype of condylar cartilage in vitro.

Animals↗
PubMed1989

Effect of thyroid hormone and growth hormone on recovery from hypothyroidism of epiphyseal growth plate cartilage and its adjacent bone.

Hypothyroidism was induced in young female Sprague-Dawley rats by the addition of methimazole (0.67 mg/ml) to drinking water for a period of 7 weeks (7-14 weeks of age). The responses of the articular cartilage, epiphyseal growth plate cartilage, epiphyseal trabecular bone, and metaphyseal trabecular bone in the proximal tibia were assessed by structural parameters. In addition, replacement therapies were introduced for the last 2 weeks of the experimental period. These included 0.7 U/kg BW human GH (hGH), 15 micrograms/kg BW L-T4 (T4), and a combination of hGH and T4 at the same doses. In the hypothyroid rats, the width of epiphyseal growth plate cartilage decreased by 27%, that of articular cartilage by 35%, epiphyseal trabecular bone volume by 30%, and metaphyseal trabecular bone volume by 66% relative to those in age-matched control tissues. T4 treatment led to a full restoration of the epiphyseal trabecular bone and surpassed by 40% the control value. The magnitude of the articular cartilage and the epiphyseal trabecular bone volume returned to control values, while that of metaphyseal trabecular bone was 68% of control values. Treatment with hGH did not improve the epiphyseal growth plate cartilage or articular cartilage. It did restore epiphyseal trabecular bone to almost normal values, but metaphyseal trabecular bone improved to only a small though significant level (45% of control value). The combination of T4 and hGH resulted in an additional enlargement in the width of the epiphyseal growth plate cartilage and an increase in metaphyseal trabecular bone volume compared to those in the T4 group. Qualitative examinations indicated that it was only in the T4 and T4 plus hGH groups that the lowest chondrocytes in the epiphyseal growth plate cartilage resumed their normal hypertrophied size. These results suggest that the change in the hypothyroid state do not rely solely on the lack of pituitary GH synthesis and secretion, as replacement by exogenous GH did not restore normal epiphyseal growth plate cartilage morphology or its remodeling into metaphyseal trabecular bone. Treatment with T4 (which restored endogenous pituitary GH to 30% of control levels) results in full recovery of the epiphyseal growth plate cartilage morphology along with its associated metaphyseal trabecular bone. In addition, it can also be concluded that the decrease in epiphyseal trabecular bone volume observed in the hypothyroid animals was due solely to the GH-deficient state that accompanied hypothyroidism.

Animals↗
PubMed1987

Carbonic anhydrase cytochemistry in mitochondria-rich cells of salamander larvae gill epithelium as related to age and H+ and Na+ concentrations.

Carbonic anhydrase (CAH) was localized in the mitochondria-rich cells (MRC) of 1-week-old salamander larvae gill epithelium, in both MRC and pavement cells of 6-week-old larvae, and in regenerated stems of previously amputated gills. CAH activity of the MRC was measured quantitatively using a microscope densitometric technique. Changes in CAH activity per cell and changes in the numbers of CAH-positive MRC were followed under different H+ and Na+ concentrations at the two age groups. CAH activity per cell increased with age, whereas the numbers of CAH-positive MRC dropped. CAH activity per cell in the 1-week-old age group reached maximal values at pH 7.4 and stayed relatively high in the more alkaline media. Moderate increases of Na+ concentrations had small but significant effects on increasing CAH activity of gill MRC. When taking into consideration not only the changes in cellular activity but also the changes in the number of CAH-positive cells under the different acclimation media, an activity index (ICAH) was calculated. Thus, the ICAH in the 1-week-old was found to be dependent on the decline of ambient H+ concentrations (expressed as increasing pH), reaching maximal effect at pH 8.0. On the other hand, raising the Na+ concentrations of the acclimation media to 110 and 220 mOsm/liter caused a maximal inhibition of tissue CAH activity as expressed by ICAH. In conclusion, it is suggested that salamander larvae gill MRC take part in the adaptation of the larvae to changing H+ concentrations of their milieu rather than in their adaptation to changes in its osmolality.

Aging↗
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↗
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↗
PubMed1984

In vitro precocious accumulation of calcium and matrix vesicles formation in young cartilage cells: specific effects of corticosteroids.

The present study examined the effects of various corticosteroid and noncorticosteroid hormones upon the ultrastructure of chondroprogenitor cells and chondroblasts in an organ-culture system of late fetal condylar cartilage. Corticosteroids, (triamcinolone acetonide, dexamethasone, corticosterone) at concentrations of 10(-6) - 10(-8)M stimulated markedly a precocious formation of matrix vesicles by chondroblasts. This stimulation was accompanied by a significant accretion of calcium complexes intra- and extracellularly in both the chondroprogenitor cell population and chondroblasts in vitro, as well as in the newly induced matrix vesicles. Nonglucocorticoid steroids (progesterone, estradiol, testosterone, cortexolone) did not evoke similar effects. Progesterone and testosterone, however, seemed to adversely affect the ultrastructure of the cartilage cells, whereas estradiol appeared to have a favorable effect on the morphology of cultured condylar cartilage.

Adrenal Cortex Hormones↗
PubMed1984

Calmodulin localization in bone and cartilage.

The localization of calmodulin (CaM), a calcium-dependent regulatory protein, was demonstrated in the following rat skeletal tissues by the indirect immunofluorescence method: a) growth plate cartilage of fetal and juvenile long bones, b) fetal epiphyseal cartilage, c) juvenile hyaline costal cartilage, d) neonatal mandibular condylar cartilage, e) neonatal diaphyseal lamellar bone. CaM was not detected in perichondrial and periosteal cells. Mature and mineralizing chondrocytes demonstrated the highest labelling intensities.

Animals↗
PubMed1984

The effects of trifluoperazine on calcifying tissues in the immature rat.

When the antipsychotic drug trifluoperazine is given to immature rats during stages of rapid bone development, severe localized derangement in their growth plate architecture results. In addition, 3 weeks of treatment with doses of 3 mg/kg body wt/day had marked effects on the anatomic distribution of bone mineral, and on the quantitative chemical composition of the bone matrix. Specifically, ash contents and Ca/P ratios were decreased and mineral crystals appeared larger and/or more perfect in the treated animals. The finding of decreased hexosamine content and alterations in bone lipid composition could be interpreted as specific effects of the drug on calmodulin activity.

Animals↗
PubMed1983

In vitro transformation of chondroprogenitor cells into osteoblasts and the formation of new membrane bone.

Mandibular condyles of fetal mice 19 to 20 days in utero were kept in an organ culture system for up to 10 days. After 2 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 5 days in culture no chondroblasts could be seen and, instead, the entire cartilage was occupied by hypertrophic chondrocytes. At the same time, the mesenchymal cells at the chondroprogenitor zone differentiated with osteoblasts which produced osteoid. Light microscopic examinations showed that the newly formed osteoid did not stain with acidic toluidine blue or with alcian blue, but stained intensively with the van Gieson stain and with Periodic acid-Schiff (PAS). The osteoid reacted with antibodies against type I collagen but not with antibodies against type II collagen. Electron microscopic examinations showed that the mineralization appeared to be associated with collagen fibers in bone rather than with matrix vesicles in the cartilage. The process of bone formation progressed with time and by the 10th day new bone replaced almost the entire cartilage, thus forming an expanded layer of membrane bone. This in vitro system represents an experimental model whereby undifferentiated precursor cells transform into osteoblasts with the subsequent formation of a typical membrane bone.

Animals↗
PubMed1982

Landmarks in chondrocyte differentiation and maturation as envisaged by changes in the distribution of calcium complexes: an ultrastructural histochemical study.

Organ cultures of fetal condylar cartilages have been examined using the K-pyroantimonate-osmium fixation method to follow alterations in cellular calcium distribution in maturing cartilage. Special attention has been given to analyzing the temporal and spatial relationships among intracellular calcium accumulation, cell maturation, and matrix mineralization. The findings of this study suggest that concomitant with the change in cellular metabolism from aerobic respiration to glycolysis there is a distinctive change in the distribution of cellular and matrical calcium complexes. This ultrastructural histochemical method can thus be used to evaluate the maturational state of chondrocytes actively involved in endochondral ossification.

Animals↗