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C Hadházy

Publications and source records attributed to C Hadházy.

At least 19 recordsLinked to original sources
PubMed1989

Exogenous glycosaminoglycans modulate chondrogenesis, cyclic AMP level and cell growth in limb bud mesenchyme cultures.

Effects of hyaluronate, heparin and chondroitin-6-sulfate were studied on micromass cultures of chick limb bud mesenchyme (Hamburger and Hamilton stages 23-24). Histochemical, electron microscopical, biochemical and radiochemical investigations of day 4 cultures revealed dose-dependent inhibitory effects of these glycosaminoglycans on chondrogenesis, cyclic AMP level and growth of cells. In addition, hyaluronate with 100 micrograms/ml dose caused a displacement of newly formed proteoglycan from cultures into the medium. It is supposed that exogenous glycosaminoglycans influence ionic equilibrium in the immediate vicinity of cells and disturb the organization of the prechondrogenic extracellular matrix resulting in alterations of cell membrane--cytoskeleton associations. These alterations may provoke a reduction in cyclic AMP level and DNA synthesis. It is suggested that a reduction in cyclic AMP level preceding the expression of cartilage phenotype results in the inhibition of chondrogenesis.

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PubMed1988

Effect of vitamin D3 and 25 hydroxyvitamin D3 on glycosaminoglycans in micro high density culture.

Vitamin D3 and 25-OH-D3 were tested for their effect on glycosaminoglycan (GAG) production in micro high density cultures of stage 22-24 chicken limb bud mesenchyme. Vitamin D3 (at concentration of 5 x 10(-6)M) and its biologically active metabolite, the 25-OH-D3 (at concentration of 5 x 10(-8)M) stimulated the synthesis of GAGs indicating the increase of radioactive sulfate incorporation and uronic acid content in a dose-dependent way. The findings obtained after the treatment with 25-OH-D3 are comparable with earlier studies performed on chondrocyte cultures (Corvol et al. 1978, 1980), while the results observed after the vitamin D3 treatment are the first indications that vitamin D3 can also influence the cartilage matrix composition.

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PubMed1987

The effect of OH(.) radicals generated by Fenton reaction on the growth and cartilage differentiation in limb bud cell culture.

A consistent chondrogenesis takes place in micro high-density cultures of chick limb bud mesenchyme cells stage 22-24. The effect of an increased generation of OH(.) free radicals by Fenton reaction was tested in these cultures. Components of Fenton reaction (i.e., ferrous iron in form of ADP-Fe2+ complex in 0.1 mM concentration, or 0.2 mM H2O2, or the combination of these components with each other, as well as with 0.2 mM ascorbate) were supplemented to the culture medium after the first 24 h. ADP-Fe2+ complex resulted in a drastic decrease of the frequency and confluence of the cartilage nodules seen in light microscope, accompanied electron microscopically by a strikingly increased frequency of occurrence of lipofuscin-like, residual bodies in the cells. Biochemical methods revealed a significant decrease of both the DNA and glycosaminoglycan contents (to 48.6 and 20.7% of the controls, respectively), in day 6 cultures. H2O2 alone caused similar alterations of the cultures, whereas the combination of it with ADP-Fe2+ complex proved to be lethal for the cells. Ascorbate when added to the ADP-Fe2+-treated cultures displayed a slight protective effect for the glycosaminoglycan content but not for DNA. The results are interpreted in terms of free radical theory of aging.

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PubMed1985

Appearance and persistence of fibronectin in cartilage. Specific interaction of fibronectin with collagen type II.

Binding of fibronectins (FN) to collagen types I-IV were studied using polyclonal antibodies against human and chicken FNs, proteoglycan monomers, collagen type II and monoclonal antibodies reacting with both soluble and insoluble forms of human FN. Plasma fibronectin and type II collagen were shown to interact specifically in a homologous system. Type II collagen, however, proved to be less effective in inhibition assays compared to other types of collagen. In high density cultures of chicken limb bud cells, fibronectin was first localized within the fibroblast-like cells of 4 hr cultures and an extensive extracellular filamentous network developed by the end of day 1. Fibronectin was present in the newly formed cartilage nodules although it seemed to disappear by day 6, when the proteoglycan accumulation became more intensive. Enzyme treatments (testicular hyaluronidase, chondroitinase ABC) helped to localize FN at this stage of development of chicken cartilage, in microdroplet high density cultures of human fetal chondrocytes and in articular cartilage. Fibronectin was localized only in the pericellular ring of intact human articular cartilage using monoclonal antibodies with the biotin-avidin system.

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PubMed1983

Changes in cyclic AMP and cyclic GMP levels during in vitro chondrogenesis.

cAMP and cGMP levels were measured in micro high-density cultures of chick limb bud mesenchyme cells stages 22-24 after 1, 2, 4 and 6 days of culturing. In these cultures, a consistent cartilage differentiation proceeds parallel to the progressive accumulation of cells in the G0 phase. The cAMP level increased by 45% by the time of the onset of cartilage phenotype expression, and significantly decreased thereafter. The cGMP level gradually diminished by a total of 39% during the period examined. It is suggested that the decrease in cell proliferation may be the consequence of the reduction of the cGMP level, and that a short-term marked elevation of cAMP level induces cartilage differentiation in the limb mesenchymal cells which are able to select only from a limited number of differentiation programmes.

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PubMed1983

In vitro cartilage differentiation: modification of division cycle and effect of some polyanions.

A modification of division cell cycle under conditions of in vitro cartilage differentiation could be demonstrated. The frequency distribution of histograms showed that the G1 cells represented 50% of total cells number at zero time (i.e. in suspension before inoculation), 69% on day 2 and about 90% in following period of days 4, 6 and 14. Dextran sulfate and hyaluronic acid produced a conspicuous inhibition and the chondroitin sulfate as well as heparin exerted a moderate inhibiting effect on the cell proliferation.

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PubMed1983

Modification of the cell cycle of limb bud mesenchyme during in vitro cartilage differentiation.

A consistent chondrogenesis takes place in micro high-density cultures derived from limb mesenchymal cells of chick embryos of stages 23-24. Flow-cytometric measurements of DNA content showed that cells in the phase of G1 or G0 made up 51% of the dispersed cell suspensions. The proportion of these cells increased to 71% by the onset of cartilage differentiation in day-2 cultures. This ratio was 84% when the voluminous matrix formation began on the 4th day of culturing. Thereafter, it increased to 90% by the 6th day, and to 93% by the 14th day. The results suggest that cartilage differentiates from G0 mesenchymal cells of the limb. In our measurements, however, the G0 phase includes all non-proliferative cell population which have identical DNA content with G1 cells. Therefore, the G0 phase contains also an increasing number of chondroblasts and chondrocytes as the chondrogenesis proceeds.

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PubMed1980

Studies on cartilage formation. XXII. Investigations of certain oxidative metabolic processes in regenerating articular cartilage.

The distal articular surface of the femur was surgically removed in 57 dogs. Succinate dehydrogenase and cytochrome oxidase activities were assayed on postoperative days 7, 20, 26, 33 and 70 in the regenerating, chondrifying articular surface and in the granulation tissue adhering to the capsule. In the 70-day samples, the cyanide-induced inhibition of oxygen consumption was determined and enzyme histochemical reactions (cytochrome oxidase, monoamine oxidase, xanthine oxidase, peroxidase and "catalase") were performed. The succinate dehydrogenase activity was the highest in the early postoperative stage in both tissues. This was followed by a definite decrease and a subsequent significant increase in activity when chondrification took place. Measurement of cytochrome oxidase activity could not reveal any convincing result, presumably because of the properties of the tissues studied. The oxygen consumption by the chondrifying articular surface at 70 days was inhibited to about 50% by cyanide, and about 90% inhibition was observed in the tissue adhering to the capsule. The cells of the regenerating articular surface possess cytochrome oxidase and a cyanide- (and sodium azide-) resistant oxidase activity. The enzyme activity of the cartilaginous islets exceeded that of their connective tissue environment. The cytochrome oxidase activity increased in the cells during cartilage differentiation. Presumably, some further cyanide-sensitive and cyanide-resistant oxidases are present in chondroblasts and young chondrocytes.

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PubMed1979

Studies on cartilage formation XXL Activity of enzymes belonging to the pentose-phosphate cycle in the regenerating articular surface.

The distal articular surface of the femur was removed operatively in 36 dogs. In the regenerating chondrifying articular surface and in the granulation tissue adhering to the capsule glucose-6-phosphate dehydrogenase and 6-phosphogluconate dehydrogenase activities were determined 7, 33 and 70 days after operation. In both tissues the activity of these enzymes characteristic of the pentose phosphate cycle ws the highest in the early postoperative stage. This initial increase in activity was followed by a marked reduction in the regenerating articular surface and by a moderate decrease in the tissue adhering to the capsule. For the loss in activity occurring in the chondrifying articular surface, the connective tissue cells (fibroblasts) are responsible. Cartilage precursors and young chondrocytes show a high glucose-6-phosphate dehydrogenase and 6-phosphogluconate activity. Presumably, in the given case of the functions of the pentose-phosphate cycle the NADPH generation and supply of building stones prevail. The activity of these enzymes ws determined in the articular cartilage and in the synovial membrane of the knee joint in further 18 dogs. The activity in the articular cartilage was very slight as compared to that in the synovial membrane.

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PubMed1977

The localization of proteoglycans and glycoproteins in the hyaline cartilage.

Antibodies to proteoglycan (PG) and glycoprotein of bovine nasal cartilage were conjugated with fluorescein isothiocyanate and with horseradish peroxidase. Hyaluronidase digestion of cartilage tissue-specimens increased the intensity of immune reactions; pronase digestion or extraction with 4 M guanidinium chloride abolished the staining. In the intercellular matrix fine filaments beaded with small granules were seen forming an irregular network. The interstices of the network are filled with collagen fibers linked together by the filaments and granules. In view of the linear conformation of core proteins of PGs and the globular conformation of glycoproteins (link proteins), it may be supposed that the granules and filaments represent these two protein components of PG-aggregates. In chondrocytes a homogeneous staining was recorded in the endoplasmic reticulum, in the juxtanuclear areas and in several smooth-walled vesicles and elongated areas situating subjacent to the cell membrane. In contrast to the extracellular immune reactions, this homogeneous intracellular staining was never enhanced by hyaluronidase digestion. This is interpreted in the sense that conformation changes of molecules secreted, and the aggregation of PGs, occur extracellularly.

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PubMed1976

Studies on cartilage formation XIX. Oxygen and glucose supply of the regenerating articular surface.

Complete removal of the articular cartilage in dogs is followed by regeneration of the articular surface. At the site of the bone wound, granulation tissue develops, which later differentiates into cartilage. The O2 and glucose supply of the regenerating articular surface is ensured by the synovial fluid, by the large exposed surface of the medullary cavity, and by the capillary network of the granulation tissue. Oxygen and glucose supply of the articular surface in different stages of differentiation has been statistically analyzed. It is suggested that in the early stage of regeneration O2 supply comes predominantly from the capillaries of the granulation tissue. Later on, as capillarization regresses, the oxygen supply, originating from the synovia and medullary cavity, assumes a more important role. In the stage of cartilage regeneration an oxygen-deficient state can be supposed in the entire articular surface, but areas differing in oxygen supply may be formed owing to local differences (due mainly to the extent of vascularization and degree of generation of the subchondral bone layer). At the site of chondrogenesis, conditions allowing aerobic metabolism of cells with reduced O2 requirements seem to be ensured. Glucose supply deriving from the above-mentioned sources satisfies the highest glucose requirements of the cells in the regenerating articular surface.

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PubMed1975

Studies on cartilage formation. XVI. Chemical and histochemical assay of lipids in the regenerating articular cartilage.

The articular surface of the distal part of the femur was removed operatively in dogs, and the regenerating articular surface and the GTC were investigated at different stages from the 7th to the 70th postoperative days. During this period cartilage islets arose in the GTAS, while the GTC transformed to connective tissue. At 7 days the lipid content of the tissue was markedly higher than at the other stages studied. Lipids, predominantly triglycerides, were present in extracellular form as well. From the 20th to the 70th day the PL fraction became predominant and, in addition to the pre-existing lecithin, relatively large quantities of lysolecithin, sphingomyelin, phosphatidyl-ethanolamine, phosphatidyl-serine and phosphatidyl-inositol could be gradually demonstrated. Differences were noted in the time of appearance and binding of PLs between the two types of granulation tissue. As time proceeded, the proportion of saturated fatty acids decreased in favour of unsaturated ones. At 70 days, the GTAS contained fatty acids up to C18. About 50% of the fatty acids consisted of C16:1, C18:2 and C18:1. At the same stage, in the GTC C16:1, C18:1 and C20:1 were present in larger amounts. Of the free fatty acids C16:1, C16 and C18 were in predominance in the GTAS and the proportion of fatty acids having more then one double bonds increased with time. In the GTC C16 and C18:1 were in great majority. According to histochemical evidence, the tissues did not contain extracellular lipids from the 20th postoperative day. In the cells, the presence of glycerides, PLs, lipoproteins and cholesterol was demonstrated. In addition, in cartilage precursors of more advanced maturity, a considerable fatty acid positivity was noted.

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PubMed1975

Studies on cartilage formation. XVIII. Changes of the composition of glycosaminoglycans in the regenerating articular surface.

The cartilaginous articular surface of the distal part of the femur of adult dogs was removed and the composition of GAGs was determined in the granulation tissue adhering to the bone wound and in that adhering to the articular capsule 7, 33, and 70 days after operation. The articular cartilage and the synovial layer of the articular capsule of intact adult dogs were also studies. The materials were digested with papain and the released GAGs were fractionated according to Svejcar and Robertson's method. The articular cartilage of non-operated dogs contained, on the average, 65.3% ChS, 13% KS, 5.8% HA and 15.8% GAG of lower molecular weight. The synovial layer of the capsule contained 41.1% HA, 15.3% Ch4-S and Ch6-S, 13.7% DS, 21.7% KS, 2% H and 6% GAG of lower molecular weight. The granulation tissue of the articular surface and that adhering to the capsule show a different developmental course. The former differentiates into cartilage, whereas the latter is simply added to the tissue of the capsule. The two tissues are different in GAG composition as early as on the 7th postoperative day. With time an increase of Ch4-S, Ch6-S and KS can be observed in the tissue of the articular surface. The tissue adhering to the capsule is characterized by a high HA and an increasing DS content. From the study of the composition of GAG's (proportion of GAG building stones) a deeper insight can be obtained into the details of GAG biosynthesis characteristic of cartilage than from the analysis of quantitative data of ChS. In the development of GAG composition characteristic of the tissue, the epimerase reactions participating in GAG biosynthesis, and the mechanisms regulating their activities seem to play a primary role.

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PubMed1975

Studies on cartilage formation. XX. Histochemical investigation of some enzymes of glycogen metabolsim in regenerative articular surfaces.

In 28 dogs the distal articular cartilage of the femur was removed and the regenerating articular surface on the 70th postoperative day was studied histochemically for hexokinase, glucose-6-phosphatase, phosphohexose-isomerase, fructose-1, 6-diphosphatase, aldolase, glyceraldehyde-3-phosphate dehydrogenase, lactate dehydrogenase, lactate dehydrogenase isoenzymes, phosphoglucomutase, phosphorylase, glycogen synthetase, UDP--glucose dehydrogenase, and UDP-glucuronic acid-4-epimerase. The articular surface consisted of fibrous tissue and of cartilage islets. The latter contained cells differentiating into cartilage and young chondrocytes. The glycolytic enzymes reacted positively in the regenerative articular surface. Enzyme activities were higher in the cells (particularly the chondroblasts and young chondrocytes) of the cartilage islets than in the connective tissue. In the cells differentiations into cartilage, beside the LDH isoenzymes characteristic of glycolysis, a significant LDH1 and LDH2 activity was observed. At the same site the presence of fructose-1, 6-diphosphatase-activity could be assumed, but there was no glucose-6-phosphatase activity. Glycogen synthesis proceeded in the cells of the cartilage islets and UDP-glucuronic acid-4-epimerase activity was observed in the differentiated cells. UDP-glucose dehydrogenase activity was positive in every section of the articular surface.

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