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Biomedical subjects

M Baccarani-Contri

Publications and source records attributed to M Baccarani-Contri.

18 recordsLinked to original sources
PubMed2001

Characterization of pseudoxanthoma elasticum-like lesions in the skin of patients with beta-thalassemia.

BACKGROUND: Pseudoxanthoma elasticum (PXE), an inherited disorder of unknown pathogenesis, is characterized by elastic fiber mineralization, collagen fibril alterations, and accumulation of thread material in the extracellular space. PXE-like clinical lesions have been described in patients with beta-thalassemia. OBJECTIVE AND METHODS: Dermal lesions in these two genetic disorders were compared by light and electron microscopy and by immunocytochemistry. RESULTS: In both disorders, elastic fiber polymorphism, fragmentation, and mineralization were structurally identical. Elastic fiber mineralization in beta-thalassemia was associated with vitronectin, bone sialoprotein, and alkaline phosphatase, similar to what was observed in inherited PXE. Furthermore, abnormalities of collagen fibrils and filament aggregates were identical in both disorders. In both inherited and beta-thalassemia-associated PXE, unrelated gene defects seem to induce cell metabolic abnormalities that lead to identical clinical and structural phenotypes. CONCLUSION: Data indicate that patients with beta-thalassemia may undergo important alterations of connective tissues, a better understanding of which may help in preventing clinical complications.

Adult↗
PubMed1998

Inhibition of myogenesis by transforming growth factor beta is density-dependent and related to the translocation of transcription factor MEF2 to the cytoplasm.

Transforming growth factor beta (TGF-beta) was found to inhibit differentiation of myogenic cells only when they were grown to high density. Inhibition also occurred when myogenic cells were cocultured with other types of mesenchymal cells but not when they were cocultured with epithelial cells. It is therefore possible that some density-dependent signaling mediates the intracellular response to TGF-beta. Within 30 min of treatment, TGF-beta induced translocation of MEF2, but not MyoD, myogenin, or p21, to the cytoplasm of myogenic cells grown to high density. Translocation was reversible on withdrawal of TGF-beta. By using immune electron microscopy and Western blot analysis on subcellular fractions, MEF2 was shown to be tightly associated with cytoskeleton membrane components. To test whether MEF2 export from the nucleus was causally related to the inhibitory action of TGF-beta, we transfected C2C12 myoblasts with MEF2C containing the nuclear localization signal of simian virus 40 large T antigen (nlsSV40). Myogenic cells expressing the chimerical MEF2C/nlsSV40, but not wild-type MEF2C, retained this transcription factor in the nucleus and were resistant to the inhibitory action of TGF-beta. We propose a mechanism in which the inhibition of myogenesis by TGF-beta is mediated through MEF2 localization to the cytoplasm, thus preventing it from participating in an active transcriptional complex.

Animals↗
PubMed1997

Elastic fiber during development and aging.

Elastin molecules aggregate in the extracellular space where they are crosslinked by stable desmosine bridges. The resulting polymer is structurally organized as branched fibers and lamellae, which, in skin, are wider (a few microns) in the deep dermis and become progressively thinner (fraction of a micron) towards the papillary dermis. Several general and local factors seem to regulate elastin gene expression, deposition and degradation. In skin, the volume density of the elastin network increases from birth up to maturity, when it accounts for about 3-4% of the tissue. However, its amount and distribution depend on dermis areas, which are different among subjects and change with age. Several matrix molecules (glycosaminoglycans, decorin, biglycan, osteopontin) have been found to be associated with elastin into the normal fiber, and several others have been recognized within pathologic elastic fiber (osteonectin, vitronectin, alkaline phosphatase in PXE). With age, and in some pathologic conditions, skin elastin may undergo irreversible structural and compositional changes, which seem to progress from localized deposition of osmiophilic materials to the substitution of the great majority of the amorphous elastin with interwoven filaments negative for elastin specific antibodies.

Adult↗
PubMed1995

Osteopontin is a constitutive component of normal elastic fibers in human skin and aorta.

Osteopontin is an acidic matrix protein, mainly expressed in mineralized tissues, kidney and atherosclerotic vessels; its biological role is still largely undefined. In the present study, immunocytochemical approaches showed that osteopontin is localized within normal elastic fibers of human skin and aorta. Antibodies raised against human bone osteopontin (LF7) or against human osteopontin synthetic peptide (amino acids 1-10, LF19) recognized epitopes associated with the amorphous material within the elastic fibers. Elastic fiber-associated microfibrils were always negative. The positivity for osteopontin of the elastic fibers was independent of age and could be observed in fetal skin and aorta as well as in the same of children, young adults and old subjects. The altered elastic fibers in the skin of old individuals were only fairly positive for osteopontin. The presence of osteopontin within the elastic fibers suggests that it may play a role against the observed tendency of elastic fibers to favor mineral precipitation. A role of osteopontin in modulating crystal nucleation and growth in mineralizing tissues and, more generally, in conditions in which mineral precipitation should be controlled is also possible.

Adolescent↗
PubMed1995

Ultrastructure of elastin.

Almost all structural studies on elastin have been done in higher vertebrates, in which it is organized as an extracellular network of branched fibres which vary from fractions f microns to several microns in diameter. By conventional electron microscopy, elastin appears amorphous. By both freeze-fracture and negative staining on cryosections, it can be resolved as beaded filaments 5 nm in diameter forming a 3D meshwork that, upon stretching, becomes oriented in the direction of the force applied. This filamentous aggregation of elastin molecules is confirmed in vitro by the observation that its soluble precursor, tropoelastin, shows a strong tendency to associate into short 5 nm-thick filaments that, with time, become longer and aggregate into bundles of various dimensions. If chemically fixed and embedded, these aggregates appear amorphous and identical to natural elastin fibres. The tendency of tropoelastin to aggregate into 4-5 nm-thick beaded filaments, which then associate into 12 nm-thick filaments forming a 3D network, has been observed by atomic force microscopy for recombinant human tropoelastin. Therefore, the amorphous structure of elastin seems to be a technical artefact. Apart from elastin-associated microfibrils, which are always present at the periphery of growing elastic fibres and probably have a role more complex than being a scaffold for tropoelastin aggregation in vivo, the elastic fibres seem to be composed of several matrix constituents, which are different in different organs and change with age and in pathological conditions. This is demonstrated by immunocytochemical studies on ultrathin sections.

Animals↗
PubMed1994

Ultrastructural analysis of skin and aorta from a patient with Menkes disease.

Ultrastructural studies of the skin and aorta of a patient with Menkes disease, an X-linked recessive disorder of copper metabolism, are described. Dermal thickness was normal, while dermal collagen fibrils exhibited a heterogeneous size range, with a mean diameter smaller than normal. Long-spacing collagen was often observed near fibroblasts, the plasma membranes of which were decorated by aggregates of interwoven filaments. Dermal elastin fibers were scarce and consisted of thin strands of amorphous elastin associated with numerous microfibrils. In the aorta, the amount of collagen was normal, although the fibrils displayed a broader range of diameters than normal, with a slightly smaller mean. Elastin fibers showed considerable disruption, appearing fragmented and wider than normal, and displaying irregular contours. The inclusion of cationic dyes during tissue fixation gave rise to numerous electron-dense precipitates within the elastin fibers, suggesting the presence there of glycosaminoglycans or proteoglycans, among which unsulfated and sulfated chondroitins were demonstrated by immunoelectron microscopy to be prominent. Heparan sulfate, observed to be a constituent of normal elastin fibers, was much reduced in amount. Elastin was also found associated with glycosaminoglycans in the soluble matrix of the aortic wall.

Aorta↗
PubMed1994

Immunochemical identification of abnormal constituents in the dermis of pseudoxanthoma elasticum patients.

Pseudoxanthoma elasticum (PXE) is a connective tissue inherited disease characterized by dermal alterations and mineralization of the elastin fibres. To investigate its pathogenesis, which is still unknown, antibodies against the principal connective tissue components were assayed on ultrathin sections of dermis from 7 PXE subjects and 5 age matched controls. Both control and PXE elastin fibres were positive for heparan, dermatan and chondroitin 0-sulphates, decorin and biglycan. In PXE, elastin fibres were also highly positive for chondroitin 6-sulphate, vitronectin, fibronectin and serum amyloid antigen. Vitronectin and fibronectin were mostly concentrated in the areas of dense mineralization within the elastin fibres. The abnormal microfilament aggregates, often seen in PXE dermis, were positive for all the above mentioned molecular species as well as for collagen types I and III and fibrillin; on the contrary, they were always negative for elastin. The results suggest that PXE is a complex disorder, in which the whole extracellular matrix is deeply disturbed. Therefore, without excluding an elastin gene defect, the data seem rather to suggest that PXE is a disorder of the mechanisms controlling the production of matrix constituents and that elastin mineralization is caused by molecules abnormally produced and entrapped within the fibre during elastin fibrogenesis.

Actin Cytoskeleton↗
PubMed1993

A new method for the cytofluorimetric analysis of mitochondrial membrane potential using the J-aggregate forming lipophilic cation 5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolcarbocyanine iodide (JC-1).

A new method for the cytofluorimetric analysis of mitochondrial membrane potential in intact cells has been developed by using the lipophilic cationic probe 5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolcarbocyanine iodide (JC-1), whose monomer emits at 527 nm after excitation at 490 nm. Depending on the membrane potential, JC-1 is able of forming J-aggregates that are associated with a large shift in emission (590 nm). The color of the dye changes reversibly from green to greenish orange as the mitochondrial membrane becomes more polarized. In two human cell lines (K562 and U937), we have studied by flow cytometry the changes in membrane potential provoked by the K+ ionophor valinomycin, a drug known to affect mitochondrial membrane potential, while the K+/H+ ionophor nigericin, known to affect intracellular pH but not mitochondrial membrane potential, was used as control. The incubation with valinomycin for 10 min. at 37 degrees C in a low K+ medium provoked a marked and dose-dependent reduction in JC-1 greenish orange fluorescence, while nigericin had no effect.

Benzimidazoles↗
PubMed1993

A clinical, ultrastructural and immunochemical study of Dupuytren's disease.

Aponeurotic tissue from seven normal subjects and from apparently unaffected branches, nodules and cords of 16 Dupuytren's patients were compared. Control tissue was characterized by polymorphous cells, showing cytoplasmic microfilament bundles, numerous pinocytic vesicles, basement membrane-like structures, and a thick coat of interwoven filaments, and by type I- and III-positive heterogeneous collagen fibrils, fibronectin, vitronectin, decorin and proteoglycans. The clinically normal branches consisted of fibroblast-like cells, small type III-highly positive collagen fibrils, fibronectin and proteoglycans. Nodules and fibrotic cords contained fibroblast-like cells, type I and III collagen, fibronectin and proteoglycans. Myofibroblast-like cells in only five out of 16 patients were present. There was no relation between clinical stage and structural alterations; the whole aponeurosis always seemed to be involved; cord retraction would seem to depend on the interactions among fibroblast-like cells and matrix components and among matrix macromolecules themselves.

Adolescent↗
PubMed1992

Aging of the human synovium: an in vivo and ex vivo morphological study.

Age-associated changes of the human synovium have been investigated by microarthroscopy, optical and electron microscopy, immunohistochemistry, and cytochemistry. The knee joints of nineteen 15- to 56-year-old subjects, classified as normal by inspection, were carefully examined by microarthroscopy; small synovial tissue biopsy specimens from both the suprapatellar pouch and the medial tibiofemoral gutter were taken. Microarthroscopy showed that the villi were more numerous and the vascular network and cell distribution and profiles less regular in aged individuals. These data were confirmed by scanning electron microscopy, which also showed large areas of the synovial surface devoid of cells and collagen bundles in contact with the joint cavity in aged subjects. Light and transmission electron microscopy confirmed these data and allowed evaluation of the number, distribution, shape, and internal organization of cells as well as the distribution of vessels and the organization of the extracellular matrix in the full thickness of the synovium (down to 2 mm). Particular attention was paid to synovial lining cells, among which three main phenotypes could be recognized: synthetic type (present at all ages and hypertrophied in aged subjects), macrophagelike (increasing with age), and fibroblastlike. Collagen increased with age. Further studies are needed for comprehensive understanding of age-associated changes in the human synovium.

Adolescent↗
PubMed1990

Immunocytochemical localization of proteoglycans within normal elastin fibers.

Appreciable amounts of glycosaminoglycans have been found by immunocytochemistry within mature elastin fibers of human dermis. On thin sections, elastin fibers showed antigenic sites for monoclonal antibodies recognizing the unsaturated units remaining after digestion of hyaluronic acid with Streptomyces hyaluronidase and after digestion of dermatan and chondroitin sulfates with chondroitinase ABC. Moreover, sectioned elastin fibers were positive towards antibodies raised against synthetic peptides corresponding to amino acid sequences near the N-terminus of the protein core of small matrix proteoglycans PGI and PGII, respectively (Fisher et al., J. Biol. Chem. 262, 9702-9708 (1987)). This is the first demonstration that highly hydrophylic molecules are strictly associated with normally cross-linked elastin. The presence of highly hydrated molecules within the elastin polymer could greatly influence its physiological properties and behavior in pathology.

Adolescent↗
PubMed1989

Localization of human placenta lysyl oxidase on human placenta, skin and aorta by immunoelectronmicroscopy.

Polyclonal antibodies to human placenta lysyl oxidase (Kuivaniemi et al., 1984) were used to localize the enzyme at ultrastructural level in human placenta, skin and aorta, by using the indirect immunogold method. The antibodies were tested on thin sections of tissues fixed and embedded in various experimental conditions. With all methods employed, the immunoreaction was always positive on collagen fibers in all tissues examined, independently of the age of the subjects. In placenta, the reaction was also slightly positive on matrix microfilaments and cells. In dermis, fibroblasts and elastin were scarcely positive in a normal 5 day-old child, in a child with skin hyperelasticity, and in two babies with osteogenesis imperfecta type II; whereas they were negative in two 16 and 40 year-old normal subjects. In aorta, the immunoreaction was always positive on collagen, scarcely positive on cells and on elastin of a 24 week-old fetus, of a normal child, and of two babies who died of complications associated with O.I. type II; on the contrary, the reaction was negative on cells and elastin fibers of a 16 week-old fetus, and of a normal 19 year-old girl. When present on elastin, gold particles were localized mostly inside the fibers. Contrary to what was observed by Kagan and coworkers on bovine aorta by using antibodies against aortic lysyl oxidase (Kagan et al., 1986), no specific localization of gold particles could be observed on or adjacent to the elastin/associated microfibrils. The results indicate that antibodies raised against placenta lysyl oxidase recognize collagen-associated as well as elastin-associated epitopes.(ABSTRACT TRUNCATED AT 250 WORDS)

Adolescent↗
PubMed1987

Lysyl oxidase activity and elastin/glycosaminoglycan interactions in growing chick and rat aortas.

Hydrophobic tropoelastin molecules aggregate in vitro in physiological conditions and form fibers very similar to natural ones (Bressan, G. M., I. Pasquali Ronchetti, C. Fornieri, F. Mattioli, I. Castellani, and D. Volpin, 1986, J. Ultrastruct. Molec. Struct. Res., 94:209-216). Similar hydrophobic interactions might be operative in in vivo fibrogenesis. Data are presented suggesting that matrix glycosaminoglycans (GAGs) prevent spontaneous tropoelastin aggregation in vivo, at least up to the deamination of lysine residues on tropoelastin by matrix lysyl oxidase. Lysyl oxidase inhibitors beta-aminopropionitrile, aminoacetonitrile, semicarbazide, and isonicotinic acid hydrazide were given to newborn chicks, to chick embryos, and to newborn rats, and the ultrastructural alterations of the aortic elastic fibers were analyzed and compared with the extent of the enzyme inhibition. When inhibition was greater than 65% all chemicals induced alterations of elastic fibers in the form of lateral aggregates of elastin, which were always permeated by cytochemically and immunologically recognizable GAGs. The number and size of the abnormal elastin/GAGs aggregates were proportional to the extent of lysyl oxidase inhibition. The phenomenon was independent of the animal species. All data suggest that, upon inhibition of lysyl oxidase, matrix GAGs remain among elastin molecules during fibrogenesis by binding to positively charged amino groups on elastin. Newly synthesized and secreted tropoelastin has the highest number of free epsilon amino groups, and, therefore, the highest capability of binding to GAGs. These polyanions, by virtue of their great hydration and dispersing power, could prevent random spontaneous aggregation of hydrophobic tropoelastin in the extracellular space.

Amino Acid Oxidoreductases↗
PubMed1986

Aortic elastin abnormalities in osteogenesis imperfecta type II.

Skin and aortic samples from two patients who died by lethal perinatal Osteogenesis Imperfecta (O.I.) were studied by optical and electron microscopy and compared with similar samples from two normal human fetuses and one newborn child. No significant abnormalities were observed in the dermis of O.I. patients apart from small differences in the diameter of reticular collagen fibrils. On the contrary, in the aortas of both patients collagen fibrils were significantly smaller than in the controls; moreover, elastin lamellae were deeply altered and consisted of roundish aggregates of elastin, massively permeated by cytochemically recognizable glycosaminoglycans. As identical features were described in experimental lathyrism by using inhibitors of the enzyme lysyl oxidase (Pasquali Ronchetti et al., 1984), the conclusion is reached that in the two cases of lethal perinatal O.I. examined, a severe lysyl oxidase deficiency could account for the observed ultrastructural abnormalities of elastin and that, besides defects of collagen type I, additional alterations of cellular metabolism might be responsible for the clinical heterogeneity of the disease.

Aorta↗
PubMed1984

Elastin fiber-associated glycosaminoglycans in beta-aminopropionitrile-induced lathyrism.

Ruthenium red and toluidine blue O precipitates were described associated with lathyritic elastic fibers in aortas of chickens treated with beta-aminopropionitrile fumarate (I. Pasquali-Ronchetti, C. Fornieri, I. Castellani, G. M. Bressan, and D. Volpin (1981). Alterations of the connective tissue components induced by beta-aminopropionitrile. Exp. Mol. Pathol. 35, 42-56). In this report evidence is given that these precipitates reveal the presence of proteoglycans, as they are completely removed by 5 M guanidine-HCl incubation and by specific enzymatic digestions. In particular, proteoglycans associated with the poorly cross-linked lathyritic elastin can be removed by testicular hyaluronidase, chondroitinase ABC, heparitinase, and nitrous acid treatments, whereas they are rather resistant to streptococcal hyaluronidase and chondroitinase AC. On the contrary, proteoglycans of the matrix or associated with collagen fibers are particularly sensitive to these latter enzymatic treatments. The conclusion is reached that glycosaminoglycans associated with beta-aminopropionitrile-induced lathyritic elastin (i) are different from those of the matrix or associated with collagen, and (ii) include mainly dermatan and heparan sulfates.

Aminopropionitrile↗
PubMed1981

Pseudoxanthoma elasticum. Biochemical and ultrastructural studies.

Ultrastructural and biochemical studies of the dermis from patients with pseudoxanthoma elasticum are described. Elastin fibers showed mineral deposits that were associated with an elastin material exhibiting altered affinity for the electron microscopical stains and altered ultrastructural organization. In the most affected patient the elastin was shown to be increased and to have an abnormal amino acid composition. In some patients the collagen fibers were laterally fused. The ground substance was increased as evidenced by electron microscopy and confirmed by a slight increase in uronic acids, hexosamines, and neutral sugars.

Amino Acids↗