Search PubMed⌕ Search

Biomedical subjects

B Nusgens

Publications and source records attributed to B Nusgens.

At least 55 records · Page 3Linked to original sources
PubMed1988

Laminin and type III procollagen peptide in human preovulatory follicular fluid.

The levels of laminin P1 fragment, a marker of basement membrane, and of the aminoterminal sequence of type III procollagen, a marker of interstitial connective tissue, were measured in human preovulatory follicular fluids. The concentrations of these peptides correlated with progesterone levels but not with those of estradiol or testosterone. Immunocytochemical studies confirmed the remodeling of the perifollicular basement membrane and interstitial matrix during oocyte maturation. The studies suggest that monitoring of the ovarian connective tissue macromolecules could be useful for estimating follicular maturation.

Basement Membrane↗
PubMed1988

Fibroblasts induce the assembly of the macromolecules of the basement membrane.

The mechanism regulating the deposition of basement membrane components (BMCs) in a polymeric structure at the junction with the connective tissues is not yet understood. Cultures and cocultures of epithelial BMC-producing cells (L2 or PER cells) and fibroblasts were prepared in several experimental conditions and the organization of BMCs was studied by immunofluorescence. The pattern of BMCs in pure cultures of L2 or pulmonary epithelial rat (PER) cells consisted of intra- and extracellular granular deposits. At very high density, the cell contours were also underlined by a disrupted network of BMC deposits. A different fibrillar plexus--containing laminin, collagen type IV, and heparan-sulfate proteoglycan resistant to deoxycholate treatment and distant from the cell membrane--was observed in cocultures of L2 or PER cells with fibroblasts. Fibrils of fibronectin and/or collagen type I were most often dissociated from this plexus of BMCs. Similar results were obtained by adding a conditioned medium of L2 or PER cells to confluent fibroblasts, even when the cells were killed. Pure laminin also bound to the fibroblast layer. A coated film of fibronectin or polymeric collagen type I was unable to bind BMC provided by a conditioned medium. It is suggested that molecule(s) synthesized by fibroblasts and deposited in the pericellular matrix are involved in the assembly of BMCs.

Animals↗
PubMed1988

Effect of EGF on human skin fibroblasts is modulated by the extracellular matrix.

The aim of this work was to clarify the reason why a discrepancy exists between the effects of epidermal growth factor (EGF) on fibroblasts in culture repressing collagen biosynthesis and in vivo stimulating wound healing. The effect of EGF on the biosynthetic activity of fibroblasts was measured in various conditions of cultures: on plastic, on plastic coated with various macromolecules of the extracellular matrix, on top of a type I collagen gel, and within a three-dimensional collagen lattice. While the noncollagen protein (NCP) synthesis was not affected by the interactions of the cell with the various coated matrices, collagen synthesis was inhibited. At the surface of a collagen gel, protein synthesis was reduced, while collagen synthesis and degradation were slightly stimulated. When embedded in a lattice, the overall biosynthetic activity of fibroblasts was largely depressed. The addition of EGF to cultures on plastic and on the various coated macromolecules resulted in a further repression of collagen synthesis while cell multiplication was slightly stimulated. On the contrary, the addition of EGF to fibroblasts in a collagen lattice resulted in a stimulation of both NCP and collagen synthesis as observed in vivo. These opposite effects of EGF in a two- or three-dimensional culture system are not related to modification in number or affinity of the EGF receptors at the cell surface. These results further support the similarity in the state of differentiation of fibroblasts in a three-dimensional lattice and in vivo.

Adolescent↗
PubMed1987

Reconstituted basement-membrane matrix modulates fibroblast activities in vitro.

Cellular growth and collagen biosynthesis were compared in dermal calf fibroblasts cultured on plastic or on a reconstituted basement membrane gel, termed matrigel. This matrix, extracted from Engelbreth-Holm-Swarm tumors, consists mainly of laminin, entactin, type IV collagen, and heparan sulfate proteoglycan. The multiplication rate of fibroblasts grown on matrigel was stimulated compared to that of monolayered cells cultured on plastic, and these cells formed multilayers after 4 days. Protein and collagen biosynthesis was reduced in fibroblasts cultured on matrigel. A higher proportion of the newly synthesized collagen (40%) was incorporated to the extracellular matrix in cultures grown on matrigel than in those grown on plastic (14%). Type III collagen was the preferential collagen type deposited on matrigel, and the ratio of type III:type I collagens secreted in the medium was also slightly higher in cultures grown on matrigel. Partially processed collagen was more abundant in fibroblasts grown on matrigel than in cells cultured on plastic. Finally, cells grown on matrigel exhibited a higher catabolic activity than cells grown on plastic. In this experimental model, the reconstituted basement-membrane matrix seems to influence the activities of fibroblasts significantly.

Animals↗
PubMed1987

Collagen metabolism and basement membrane formation in cultures of mouse mammary epithelial cells. Induction of 'assembly' on fibrillar type I collagen substrata.

Collagen metabolism was compared in cultures of mouse mammary epithelial cells maintained on plastic or fibrillar type I collagen gel substrata. The accumulation of dialysable and non-dialysable [3H]hydroxyproline and the identification of the collagens produced suggest no difference between substrata in the all over rates of collagen synthesis and degradation. The proportion of the [3H]collagen which accumulates in the monolayers of cultures on collagen, however, markedly exceeds that of cultures on plastic. Cultures on collagen deposit a sheet-like layer of extracellular matrix materials on the surface of the collagen fibres. Immunoprecipitation of the labelled extracts, electrophoresis, indirect immunofluorescence and immunoperoxidase techniques reveal the presence of type IV collagen, along with laminin and heparan sulfate proteoglycan in this layer, in excess over the amounts detectable on cells cultured on plastic. Transformed cells on collagen produce and accumulate more [3H]collagen, yet are less effective in basement membrane formation than normal cells, indicating that the accumulation of collagen alone and the effect of interstitial collagen thereupon do not suffice. Thus, exogenous fibrillar collagen appears to enhance, but is not sufficient for proper assembly of collagenous basement membrane components near the basal epithelial cell surface.

Animals↗
PubMed1987

Acidic glycosaminoglycans, isolation and structural analysis of a proteodermatan sulfate from dermatosparactic calf skin.

Dermatan sulfate and hyaluronic acid are the major glycosaminoglycan components of dermatosparactic and normal calf skin but the ratio of hyaluronic acid to dermatan sulfate is 50-70% higher in dermatosparactic calf skin than that in normal calf skin. Hyaluronic acid in normal calf skin could be extracted to 93% of the total by NaCl and guanidine hydrochloride, while in dermatosparactic calf skin 57% of hyaluronic acid remained in the insoluble material after NaCl and guanidine hydrochloride extraction. The recovery of proteodermatan sulfate in NaCl extracts was similar in normal and dermatosparactic skin. The amount of proteodermatan sulfate extracted in guanidine hydrochloride form normal calf skin was 2-4 fold higher than that from dermatosparactic calf skin. No significant difference between dermatosparactic and normal calf skin proteodermatan sulfate could be detected in terms of size of the core protein (Mr = 55,000) by sodium dodecyl sulfate polyacrylamide gel electrophoresis, amino acid analysis, immunological cross-reactivity using polyclonal antibodies against either dermatosparactic or normal calf skin core protein, tryptic peptides or size of dermatan sulfate chains (Mr = 17,000-18,000) on gel chromatography.

Animals↗
PubMed1986

Bone remodeling during the development of osteoporosis in paraplegia.

Osteoporosis developing during the first weeks after the onset of traumatic paraplegia was studied with cortical and cancellous samples of iliac crest and tibia of 14 patients, and compared to normals. We used a procedure of bone particle fractionation (according to degree of mineralization) that allowed us to establish a profile reflecting the metabolic remodeling of bone and to analyze the organic matrix of the newly synthesized tissue. In paraplegics, we observed a large increase in the proportion of little calcified bone in the cortical as well as in the cancellous bone. Based on amino acid analyses, we found a decreased number of hydroxyproline residues in the newly synthesized organic matrix from paraplegia bone resulting either from an alteration of the prolyl hydroxylation or from the presence of an excess of noncollagen polypeptides. These results, together with previously published data reporting increased urinary hydroxyproline and calcium kinetic parameters, suggest an enhanced rate of skeletal remodeling in acute paraplegia. When investigated 2 years after injury, the patterns of distribution approach that of normal subjects.

Adolescent↗
PubMed1986

Metabolic changes in the extracellular matrix during differentiation of myoblasts of the L6 line and of a Myo- non-fusing mutant.

In the present study we have characterized by biochemical and immunochemical methods the changes which take place in collagen, laminin and fibronectin biosynthesis during the differentiation of clonal skeletal myoblasts of the L6 line. Time-course experiments showed that the relative rate of synthesis of collagen increased significantly during the cell-cell contact step of myogenesis and decreased later on. The major collagens synthesized were types I and III, found mainly as soluble precursors in the culture medium. Types IV and V collagens were detected exclusively in the cell layer. The relative amounts of types I and III collagens remained unchanged during myogenesis, while types IV and V collagens increased as the cells of the L6 line fused. In a non-fusing alpha-amanitin-resistant mutant of the L6 line (Ama 102), the rate of collagen synthesis was largely depressed and its rate of degradation was increased as compared with the fusing wild type. The synthesis of laminin was very low in cells of the fusing wild type, but abundant and associated with the cell layer of the Myo- mutant. The appearance of a muscle-specific extracellular matrix is a complex process involving changes in the organization, the biosynthesis and remodelling of its macromolecules of the extracellular matrix.

Animals↗
PubMed1986

Antibodies to laminin in preeclampsia.

Laminin is a large basement membrane glycoprotein localized in the trophoblast, glomerular basement membrane and in the mesangial matrix of human glomeruli. It promotes the attachment of epithelial cells to basement membrane collagen. We have found that 14 sera from 52 patients with severe preeclampsia or eclampsia contain IgG and IgM antibodies which react with placental and kidney basement membranes. These antibodies were specific for laminin and did not react with other basement membrane proteins. They were able to fix complement. They have been demonstrated by radial immunodiffusion, radioimmunoassay and immunofluorescence blocking studies. In primary cultures they were shown to impair the attachment of trophoblast cells to basement membrane collagen. High levels of circulating immune complexes were detected only in sera from preeclamptic patients with circulating antibodies to laminin. The auto-antibodies to laminin could play a major role in the pathogenesis of severe preeclampsia by impairing the attachment of trophoblast cells to placental basement membranes and by fixation to the glomerular basement membranes and mesangial matrix.

Antibody Specificity↗
PubMed1986

Characterization of a spontaneously transformed pulmonary embryonic rat (PER) epithelial cell line.

A spontaneously transformed pulmonary embryonic rat epithelial cell line (PER) is described in terms of growth, tumorigenicity, growth factor responsiveness and biosynthetic capacity. At low-passage subcultures, PER cells grew as a monolayer and did not form colonies in soft agar. After long-term subcultivation, they lost contact inhibition, became anchorage-independent and formed tumours in nude mice. Low concentrations of foetal calf serum permit the maximum growth rate. The multiplication and metabolic activity, assessed by 2-deoxy-D-glucose uptake, was significantly stimulated by growth factors. PER cells synthesized collagen types I, III, IV and V, laminin and fibronectin, and organized a pericellular matrix made up of only basement membrane components (type IV collagen and laminin) and fibronectin. These data enabled us to define PER cells as a transformed epithelial cell line evolving towards malignancy with long-term subcultivation. These cells appeared to be a valuable tool in studies of tumour cell-matrix interactions and regulation of growth factor receptors in tumorigenesis.

Animals↗
PubMed1985

Synthesis of basement membrane components by differentiated thyroid cells.

Morphological studies indicate that basement membrane formation or maintenance can be achieved in cultures of thyroid cells. In the present investigation we have studied the biosynthesis of this extracellular matrix by differentiated porcine thyroid cells in culture. They were prepared by two procedures: (1) thyroid cells isolated by dispase digestion of the thyroid gland were maintained in serum-free medium on poly(L-lysine) coated dishes; (2) thyroid follicles released by collagenase treatment of the gland were isolated by differential filtration and cultured in suspension on agarose-coated dishes. In both cases, functional follicular-like structures were obtained as shown by their ability to organify Na125I and to respond to thyrotropin stimulation (250 microU/ml). After incubating the cells with radiolabeled proline or methionine, collagen synthesis was observed with the two types of culture, as shown by the formation of radioactive hydroxyproline and by the synthesis of peptides with electrophoretic properties identical to those of authentic collagen molecules and susceptible to collagenase. Besides variable amounts of type I and type III collagen-like peptides, significant proportions of labeled peptides migrated with type IV collagen chains and were precipitated by anti-type IV collagen antibody; thyrotropin had no significant effect either on the total collagen synthesis or on the relative amounts of the different collagen peptides. When thyroid cells were incubated with [35S]sulfate, a labeled glycosaminoglycan with chromatographic properties analogous to that of heparan sulfate could be obtained in both culture conditions; here again, no effect of thyrotropin was observed. The ability of differentiated porcine thyroid cells to synthesize basement membrane was suggested by their production of type IV collagen and heparan sulfate, two of its potential components. Thyrotropin, which drastically enhanced the functional property of the cells, did not seem to regulate this synthesis.

Animals↗
PubMed1985

[The composition of connective tissue in the normal and fibrocystic breast].

A recent immunofluorescent study has tried to define the precise nature of the macromolecules present in the connective tissue of normal and fibrocystic breasts. Nothing abnormal has been found in the distribution of the different types of collagen and of the two glycoproteins, namely fibronectin and laminin. A quantitative approach to the research into the composition of the organ has brought us to realise that there are different amounts of the constituents of the connective tissue to be found in 80 samples which have been taken from 4 specific regions of the breast. These have been taken from 16 women aged between 28 and 82 with a mean age of 57 years. Their breasts could be ascribed to different clinical and radiological categories. We have been able to show that there is a significant rise in the water content as well as in the collagen content and, to a lesser degree, in the total proteins of the body of the breast as compared with the fatty layer in front of the breast. This increase is compensated for by a significant lessening in the percentage of lipids. It was not possible to demonstrate a statistically significant difference between these two regions as far as the concentration of proteoglycans and glycoproteins. Fibrous types of breast have, in the main body of the breast, a significant increase in the percentage of water, collagen, proteoglycans and glycoproteins as compared with fatty breasts.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1984

Collagen biosynthesis by cells in a tissue equivalent matrix in vitro.

Collagen biosynthesis by fibroblasts and subsequent processing and polymerization have been studied in conventional monolayer cultures and in a new model system in vitro in which cells organize into a tissue like structure. While the bulk of the newly made collagen becomes tightly and selectively bound to the matrix of the tissue equivalent model, in monolayer cultures most of the collagen passes into the culture medium. Collagen biosynthesis appears to be regulated differently in cells of the tissue equivalent model as compared with monolayered cells. In the former there is a 6- to 8-fold decrease in collagen output even though overall protein synthesis per unit of DNA is twice as great as in monolayered cells. Cells grown in tissue lattices exhibit much higher collagenolytic activity than cells in monolayer suggesting the model may also be of special use for studying collagen turnover and matrix remodeling.

Cell Line↗
PubMed1983

The capacity of retracting a collagen matrix is lost by dermatosparactic skin fibroblasts.

When cultivated within a matrix made of reconstituted collagen fibers, fibroblasts derived from skin, tendon, vena cava, and aorta of a normal (N) calf retract the lattice. This effect progresses with time and is related to the density of the cells included in the lattice. Under similar conditions, fibroblasts derived from the skin of 2 dermatosparactic (D) calves do not contract the lattice. Fibroblasts from D tendon and cells from D vena cava and aorta contract the lattice at the same rate and to the same extent as do their normal counterparts. In the lattice, N skin fibroblasts are elongated along the collagen fibers while D skin fibroblasts remain round and develop little cell processes. N skin fibroblasts do not multiply in the lattice while D skin fibroblasts increase in number by a factor of 3 in 5 days. The addition of N skin fibroblasts, in an amount insufficient to retract the lattice, to D skin fibroblasts does not correct their defective capacity. It is suggested that the disturbed relationship between the D skin fibroblasts and collagen fibers is responsible for the lack of architectural organization of the bundles of collagen polymers in the D skin.

Animals↗
PubMed1983

The reconstitution of living skin.

A living-skin equivalent useful as a skin replacement and as a model system for basic studies has been fabricated and tested extensively. It consists of two components: (1) a dermal equivalent made up of fibroblasts in a collagen matrix that is contracted and modified by the resident cells, and (2) an epidermis that develops from keratinocytes "plated" on the dermal equivalent. A multilayered keratinizing epidermis with desmosomes, tonofilaments, and hemidesmosomes forms. Basement lamella formation occurs within 2 weeks in vitro when rat cells are used. With human cells, crypt or pseudofollicular morphogenesis is observed in vitro within 3 weeks after plating cells on the dermal equivalent. Autografts and isografts of rat-skin equivalents made with cultured cells from biopsies are rapidly vascularized, block wound contraction, and persist essentially for the lifespan of the host. Seven to 9 days after grafting, donor cells become activated biosynthetically and mitotically. By 1 year, the dermal population decreases to a normal level and the matrix has been extensively remodeled. The grafts remain free of hair and sebaceous glands. Grafts to rats have been in place for over 2 years. Now, allografts of dermal equivalents have been made across a major histocompatibility barrier and are not rejected. The persistence of cellular elements of the grafts is monitored by use of a genetic marker. Challenge of the allograft with a second skin-equivalent graft after 1 month does not result in rejection of the original graft or of the second skin-equivalent graft. We propose that allografts of tissue equivalents are tolerated because cells with class II antigens are selected against during in vitro cultivation and are excluded from the graft. Thus the fabrication of skin-equivalent tissues or of other equivalent tissues with parenchymal cells that do not bear class II antigens may render transplants of such tissues immunologically acceptable despite the presence of allogeneic cells. The capacity to graft across major histocompatibility barriers using living tissue equivalents may have important clinical significance.

Animals↗