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Biomedical subjects

L Robert

Publications and source records attributed to L Robert.

At least 217 records · Page 12Linked to original sources
PubMed1984

Incorporation of radioactive proline and fucose in gastric mucosal biopsies. Correlation with histological findings.

Human gastric mucosal biopsies incorporate in vitro radioactive proline and fucose into macromolecular glycoproteins (mucin). Differences were found between the incorporation pattern of antral and fundic mucosae according to their pathology, confirmed by histology. Antral mucosae with abnormal histology showed a significantly higher incorporation of proline than normal samples. Fucose incorporation was also increased. Similar results were found with fundic biopsies. The ratio of total fucose to total proline incorporated into mucin secreted during incubation also increased significantly in these samples. Biochemical analyses on the other hand showed no significant change. The results suggest a breakdown in the processing, storage and secretory processes of mucin-type glycoproteins in pathological mucosae, but not in their biosynthesis. The mucosal samples could be classified as A or B types according to their proline and fucose incorporation: A mucosae have a lower proline incorporation than B mucosae (greater than 380 cpm/micrograms DNA for the antrum and greater than 200 cpm/micrograms DNA for the fundus). These results confirm the possibility of studying abnormal mucus secretion using gastric biopsy samples.

Adult↗
PubMed1984

Appearance of fibronectin in putative preneoplastic lesions and in hepatocellular carcinoma during chemical hepatocarcinogenesis in rats and in human hepatomas.

The distribution of a high-molecular-weight glycoprotein, fibronectin (FN), was studied by immunohistochemical methods in putative preneoplastic lesions, as well as in fully developed hepatocellular carcinomas, formed in rats during chemical hepatocarcinogenesis, and in 10 human hepatomas. All but two preneoplastic lesions contained FN in their intercellular matrix. FN was present around groups of transformed liver cells, as well as on the surfaces and in the cytoplasms of certain cells. A similar, typically altered, distribution pattern was seen in the majority of the carcinomas studied. Six out of seven human hepatocellular carcinomas were found to contain FN. In three cases of cholangiocarcinoma, FN was, however, confined to the reactive stroma only. The presence of FN in hepatomas, which is in contrast to carcinomas of other organs, may serve as a marker for the demonstration of preneoplastic lesions of experimental carcinogenesis and for diagnostic differentiation of human hepatomas.

Adenoma, Bile Duct↗
PubMed1984

Differentiation of the elastase-type protease of platelets from other elastases.

Elastase activities were determined near neutral pH on several specific substrates using platelet-derived preparations mixed with decreasing amounts of leukocytes. Activities were extrapolated to zero leukocyte content enabling the estimation of intrinsic platelet elastase activity. In contrast to human leukocyte elastase, metal chelating agents inhibited partly the elastase activity of the platelet extract and soybean trypsin inhibitor did not modify its activity. Serine active site titrants (phenylmethane sulfonyl fluoride) as well as acetyl-di-L-alanyl-L-propyl-L-valine chloromethylketone completely abolished the activity of platelet lysates. The platelet protease was purified from Triton X-100 platelet lysates. No cross-reactivity could be demonstrated by immunoelectrophoresis with either porcine pancreatic elastase or human leukocyte elastase using monospecific antisera. Applying gel electrophoresis, most of the elastase activity of the platelet protease migrated towards the anode, whereas the pancreatic and leukocyte elastases migrated towards the cathode. The anionic character of the platelet enzyme might explain its capacity to degrade better elastin treated with cationic detergents in contradistinction to other elastases which act better on anionic detergent-treated elastins.

Animals↗
PubMed1983

Properties and subcellular localization of elastase-like activities of arterial smooth muscle cells in culture.

The properties and subcellular localization of the elastase-like activities of smooth muscle cells cultured from pig aortas have been investigated. Homogenates of the cells hydrolysed N-succinyl-L-alanyl-L-alanyl-L-alanine-p-nitroanilide, a synthetic substrate for elastases, with a distinct pH optimum of 8.2 and hydrolysed insoluble elastin with a distinct pH optimum of 8.5. Both enzyme activities were directly proportional to the concentration of homogenate in the assay mixture. The activities toward both substrates were inhibited by phenylmethylsulphonyl fluoride and were therefore probably due to a serine peptidase(s). The activities were also inhibited by EDTA and, in a dose-related manner, by alpha 1-antiprotease. Pepstatin, which inhibits cathepsin D, and leupeptin, which inhibits cathepsin B, did not significantly inhibit the elastase-like activities in these cells. The cells were homogenized and a post-nuclear supernatant subjected to sucrose density gradient centrifugation. The distribution of elastase-like activity toward both substrates was similar to that of the plasma membrane marker 5'-nucleotidase, and distinct from those of marker enzymes for the other organelles. Cells were also homogenized with digitonin, which selectively increases the equilibrium density of the plasma membrane. The equilibrium densities of both 5'-nucleotidase and of the elastase-like activities were increased considerably, confirming the plasma membrane localization of the elastase-like activities. The subcellular localization of the elastase-like activities of arterial smooth muscle cells is therefore consistent with a role for them in the degradation of elastin in the normal arterial wall and in atherosclerotic lesions.

Animals↗
PubMed1983

Structural glycoproteins from rabbit aortic media.

Rabbit aortic intima-media fragments were incubated with [14C]mannose and [3H]fucose for 6 h to detect glycoproteins synthesized in situ. The radioactively labelled and the non-labelled samples were extracted with 0.2 mM-CaCl2/0.5 mM-dithiothreitol/0.5 mM-ATP and chloroform/methanol/water (4:4:1, by vol.). The delipidated residue was extracted with 5 M-guanidinium chloride/0.05 M-dithiothreitol/0.1 M-Tris/0.4% Na2EDTA, pH 7.5, before (extract 1) and after hydrolysis with collagenase (extract 2). The proteins in extracts 1 and 2 were S-carboxamidomethylated and separated by molecular-sieve chromatography, polyacrylamide-gel electrophoresis and isoelectric focusing in sucrose gradients in urea. The apparent molecular weights of glycoproteins were 36 000 (glycoprotein I) from extract 1, 50 000 (glycoprotein II) and 130 000 (glycoprotein III) from extract 2. The molecular weights of the non-labelled and radioactively labelled glycoproteins were identical. Glycoproteins I, II and III contain large amounts of polar amino acids and methionine. They contain neither hydroxyproline nor 3-methylhistidine. A hydroxyproline-containing component of 160 000-apparent-mol.wt. relatively rich in polar amino acids and labelled with incorporated sugars was isolated from extract 1. The incorporation in vitro of radioactive sugars into glycoproteins I, II, III and collagenous glycoproteins indicates that they are synthesized in the surviving aorta by the smooth-muscle cells.

Animals↗
PubMed1983

Distribution of fibronectin in diabetic skin.

Distribution of the fibronectin (FN) present in the forearm skin was studied in 23 controls and 34 insulin dependent diabetics. All the subjects were lean males under 50 years. After biopsy FN was studied by an indirect immunofluorescence technic. A semi-quantitative evaluation was attempted by giving a score to the fluorescent intensity read at the three following sites: vascular basement membranes, papillary dermis, and the dermo-epidermal basement membranes. We found an increased amount of FN-immunofluorescence in diabetic skins. This increase was seen not only in vascular basement membranes but also at the two other sites. No correlation was found between apparent FN levels and the duration or equilibration of diabetes. These results are consistent with other findings of the literature concerning the involvement of the intercellular matrix in diabetes.

Adult↗
PubMed1983

Elastase-type activity of human serum. Its variation in chronic obstructive lung diseases and atherosclerosis.

Human serum was found to contain enzyme activities hydrolyzing succinyl trialanine paranitroanilide and 3H-kappa-elastin Sepharose substrates. Both types of activities could be partly abolished by serine active site titrants (phenylmethanesulfonylfluoride, diisopropylphosphorofluoridate) and partly by neutral chelating agents (EDTA; 1-10-phenanthroline). The combination of phenylmethanesulfonylfluoride and EDTA gave a complete inhibition of human serum elastase-type activities indicating the presence of at least two different types of elastases (serine and metalloproteases) in human serum. In nonsmokers, the average serum elastase-type activity on succinyl trialanine paranitroanilide was found equal to 78.1 ng/ml porcine pancreatic elastase equivalents and on 3H-kappa-elastin sepharose beads equal to 688.8 ng/ml. No statistically significant differences were observed in elastase levels in the sera of individuals presenting clinical symptoms of atherosclerosis. The sera of patients suffering from chronic obstructive lung diseases contained, however, higher amounts of elastase-type activities, respectively equal to 237.2 ng/ml on succinyl trialanine paranitroanilide and 1,096 ng/ml on 3H-kappa-elastin Sepharose beads and was quantitatively significant when compared with control subjects.

Adult↗
PubMed1983

Plasma binding of an alpha-blocking agent, nicergoline--affinity for serum albumin and native and modified alpha 1-acid glycoprotein.

The binding of nicergoline, an alpha-blocking drug, by human plasma proteins was studied using gel filtration, polyacrylamide gel electrophoresis, and equilibrium dialysis techniques. 3H-labeled nicergoline added to plasma was eluted together with two major protein fractions, one containing mainly serum albumin, the other glycoproteins such as alpha 1-acid glycoprotein (alpha 1-AG). Equilibrium dialysis experiments with pure human serum albumin and alpha 1-AG as well as with its chemically modified forms, desialylated, carboxymethylated, and both desialylated and carboxymethylated alpha 1-AG gave the following results: nicergoline has about a 4-fold higher affinity for alpha 1-AG than for serum albumin. There are two binding sites per molecule on serum albumin and one on alpha 1-AG. The binding parameters of alpha 1-AG were not significantly modified by desialylation or carboxymethylation. Only desialylated and carboxymethylated alpha 1-AG showed a decreased binding for nicergoline, suggesting conformational modifications induced by these combined treatments. The fact that desialylated alpha 1-AG keeps its affinity for nicergoline suggests the possibility of a selective introduction of this drug in cells possessing the Ashwell-type specific receptor for desialylated alpha 1-AG, for instance hepatocytes. Increased serum alpha 1-AG concentration induced by inflammatory reactions will also modify the distribution of bound nicergoline between serum albumin and alpha 1-AG and as a consequence its half-life and cell distribution.

Binding Sites↗
PubMed1982

Isolation and characterization of lung connective-tissue glycoproteins.

1. Glycoproteins of hamster, rat and baboon lung parenchyma were investigated by using [14C]glucosamine incorporation in vitro followed by sequential extraction of the macromolecular components and characterization of the glycoproteins in the extracts. 2. Slices of lung parenchyma maintained in vitro incorporated [U-14C]glucosamine linearly with time into non-diffusible macromolecules for up to 5h. All the macromolecule-associated 14C label was present as [14C]glucosamine. 3. These 14C-labelled macromolecules were extracted from previously delipidated and salt-extracted lung by 5M-guanidinium chloride in the presence of dithiothreitol and proteinase inhibitors before (extract A1) and after (extract A2) hydrolysis of the collagen by collagenase. The [14C]glucosamine-labelled glycoproteins in extracts A1 and A2 contained 55 and 5% respectively of the total [14C]glucosamine incorporated in the lung of all three species studied. 4. The [14C]glucosamine-labelled glycoproteins were analysed by gel-filtration chromatography, sodium dodecyl sulphate/polyacrylamide-gel electrophoresis and isoelectric focusing. The major [14C]glucosamine-labelled glycoproteins of baboon lung parenchyma had apparent mol.wts. of about 400 000, 140 000 and 65 000 with isoelectric points respectively of 4.8, 5.4 and 5.4. The hamster lung glycoproteins with isoelectric points of 4.1 and 5.8 were devoid of hydroxyproline and contained galactose, mannose and N-acetylglucosamine. These experiments indicate that several distinct glycoproteins are synthesized in situ by the cells of pulmonary parenchyma and may well play a role in its structure and function.

Amino Acids↗
PubMed1982

Isolation and partial characterization of an elastase-type protease in human vulva fibroblasts: its possible involvement in vulvar elastic tissue destruction of patients with lichen sclerosus et atrophicus.

A complete disappearance of orcein positive material was observed in the superficial dermis of patients suffering from Lichen sclerosus et atrophicus. An elastase-type protease was isolated and partially purified from Triton X-100 extracts of human vulvar fibroblasts by gel permeation chromatography. It presents the characteristics of a metalloenzyme hydrolyzing Succinoyl-tri-alanine paranitroanilide maximally at pH 8.0 and is also active towards insoluble elastin. When partially purified enzyme is directly applied on to rabbit skin sections or when injected intradermally to young rabbits, it produces appreciable degradation of elastic fibers. The involvement of this protease in the disappearance of elastic fibers in Lichen sclerosus et atrophicus is postulated.

Adult↗
PubMed1982

Radiation therapy in primary carcinoma of the vagina.

The experiences with irradiation of 25 patients with primary carcinoma of the vagina are presented. The 3-year survival was 36 per cent despite the fact that most of the cases were advanced. Further improvements in survival may probably be achieved with extension of the radium implantation to paravaginal tissues. Choice and technique of treatment must be individualized. Surgery is limited to the more advanced cases with involvement of the bladder and rectum.

Adult↗
PubMed1982

Variation of elastase-type protease activity and elastin biosynthesis in rabbit aorta induced by cholesterol diet and immunization with elastin peptides.

Elastase-type proteases were shown to be produced by arterial smooth muscle cells and fibroblasts in culture and are probably involved in the development of the arterio-atherosclerotic process (1-4). The present investigation was aimed at the quantitative determination of the elastase-type enzyme activity in the aortas of rabbits submitted to two different athero-arteriosclerosis inducing treatments: high cholesterol diet and immunization with kappa-elastin peptides. Simultaneously we determined the incorporation of 14C-lysine in cross-linked elastin peptides by the surviving aorta extracts, determined with a synthetic substrate (Suc-(Ala)3-pNA) increased two fold after 1.5 month cholesterol diet, and three fold after 8 weeks of immunization with kappa-elastin in complete Freund's adjuvant. The incorporation of 14C-lysine in cross-linked elastin slightly increased (+20%) in cholesterol-fed aorta-explants and strongly decreased (-65%) in the immunized aorta-explants, on a DNA basis. These results confirm our contention that atherogenic stimuli produce an increase of elastase-type enzyme activity in the arterial wall. This increase appears to be correlated with elastic fibers degradation. It may also be accompanied by a decrease of elastin biosynthesis as in the kappa-elastin induced immuno-arteriosclerosis model.

Animals↗
PubMed1982

[Distribution of fibronectin in the skin of patients with scleroderma].

The distribution of fibronectin (FN), a major glycoproteic component of extra-cellular matrix, has been studies by an indirect immunofluorescence technique in the skin of 50 normal controls and 19 sclerodermic patients. In the normal skin, FN was present mainly in the papillary dermis, as thin strips and less abundant in reticular dermis, bound to collagen bundles. In scleroderma skins, FN was increased in the deep dermis of extensive and evolutive lesions (11 cases). In an other hand, the distribution of FN was not modified in stabilized lesions (8 cases). We conclude that the detection of FN in the scleroderma skin is an useful marker of the activity of the systemic sclerosis process and we discuss the possible role of FN as a primary matrix for organization of the collagenous connective tissue during the sclerosing process.

Extracellular Space↗
PubMed1982

[Distribution of fibronectin in renal pathology (author's transl)].

The distribution of fibronectin (FN), a major glycoproteic component of extracellular matrix, has been detected in the human kidney by an indirect immunofluorescence technique using a rabbit anti-human FN serum on 6 embryonic renal tissues, 5 normal kidneys and 120 renal biopsies. In the metanephros, FN was seen within tubular basement membranes and in embryonic glomeruli. There was a progressive loss of tubular basement membrane staining during the differentiation of renal parenchyma. In the normal kidney and in kidneys with nephritis without endo- and/or extracapillary proliferation, FN was localized only in the mesangium and faintly in capillary walls of glomeruli. In glomerulonephritis with endo- and/or extracapillary proliferation, FN was increased around mesangial cells. FN was also bound to fibrin in epithelial crescents, fibrinoid necrosis and in thrombi of thrombotic microangiography. FN was increased in the mesangium of diabetic glomeruli without endocapillary proliferation. FN has not been found in amyloid deposits and in sclerosed glomeruli. We therefore conclude that FN is a good mesangial marker and is probably involved in the inflammatory process.

Diabetic Nephropathies↗