scholarly journals Collagenous constituents of amniotic fluid.

1998 ◽  
Vol 45 (4) ◽  
pp. 1037-1046
Author(s):  
T Gogiel ◽  
D A Bielecki ◽  
E Bańkowski
Keyword(s):  
Amniotic Fluid ◽  
Classical Method ◽  
Degradation Products ◽  
Gel Filtration ◽  
Molecular Forms ◽  
Monomeric Form ◽  
Alpha Subunits ◽  
Bacterial Collagenase ◽  
Sds Page ◽  
Collagen Degradation Products

The amniotic fluid (AF) was fractionated by dialysis, gel filtration and SDS/PAGE, and submitted to the assay of collagenous constituents. The collagenous character of peptides and proteins of amniotic fluid was confirmed by hydroxyproline (Hyp) assay and treatment with bacterial collagenase followed by electrophoresis and gel filtration of the digestion products. It was found that AF contains collagen degradation products but the classical method of Hyp determination described by Woessner (Arch. Biochem. Biophys., 1961, 93, 440-447) gives overestimated values due to the interference with other AF components. Fractionation of AF on Sephadex G-100 column allowed to remove the interfering material and to estimate the actual Hyp content which equals to approx. 6.2 microg/ml. About 70% of Hyp was found in low molecular dialyzable products and the rest (about 30%) appears to be a constituent of nondialyzable collagenous polypeptides of the molecular mass of about 7.9-26.3 kDa. It is suggested that such collagenous polypeptides may be the products of proteolytic conversion of collagen precursor (procollagen) into the monomeric form of this protein. No high molecular forms of collagen, corresponding to alpha-subunits, were found.

TUMOR-ASSOCIATED FIBRINOLYSIS IN OVARIAN CARCINOMA - HPLC AND N-TERMINAL AMINO ACID ANALYSIS REVEAL THE PATHWAY OF DEGRADATION OF CROSSLINKED FIBRIN

1987 ◽  
Author(s):  
O Wilhelm ◽  
A Henschen ◽  
R Hafter ◽  
H Graeff
Keyword(s):  
Ovarian Cancer ◽  
Ovarian Carcinoma ◽  
Ascitic Fluid ◽  
High Performance ◽  
Degradation Products ◽  
Gel Filtration ◽  
Advanced Ovarian Cancer ◽  
Reversed Phase ◽  
Fibrin Degradation Products ◽  
Sds Page

Crosslinked fibrin has been demonstrated by immunohistochemi-cal tests to occur around tumor plugs, on the surface and in the stroma of the tumor in ovarian cancer. High levels of D-Dimer (200-800μg/ml), the characteristic terminal degradation product of crosslinked fibrin, are found in ascitic fluid of patients with advanced ovarian cancer. These findings suggest that fibrin polymerisation and degradation are related to and even may influence tumor growth. The kind of proteases which are responsible for degradation of crosslinked fibrin is, however, unknown.lt was the aim of this study to evaluate whether plasmin and/or other proteases are involved in tumor-associated fibrinolysis. Therefore the total high-molecular-weight fibrin degradation products in ascitic fluid were purified by protamine sulfate precipitation, gel filtration, immunoadsorption and compared with the components of plasmin-degraded crosslinked fibrin, i.e. DD,DY,YX,DXD and DXY, by direct SDS-PAGE in the absence of mercaptoethanol and after excision of the bands, mercaptolysis and re-electrophoresis. Pronounced similarity between the two sets of fragments was observed. For further information the fragments from the two sources were mercaptolysed and their polypeptide chain components separated by reversed-phase high-performance liquid chromatography, the components being identified by N-terminal sequence analysis and SDS-PAGE. Highly similar patterns were obtained and components corresponding to γ-γ ,γ-γ1, β, β2 and α1 could be recognized. The findings provide strong evidence for plasmin being the primary protease involved in ovarian carcinoma-related fibrinolysis, (supported by Deutsche Forschungsgemeinschaft.SFB 207, A2).

scholarly journals Analysis of the protein and glycan parts of CA125 antigen from human amniotic fluid

2007 ◽  
Vol 59 (2) ◽  
pp. 97-103 ◽  
Author(s):  
Bojana Milutinovic ◽  
Miroslava Jankovic
Keyword(s):  
Amniotic Fluid ◽  
Lectin Binding ◽  
Gel Filtration ◽  
Placental Tissue ◽  
Peptide Mass Fingerprinting ◽  
Chain Structure ◽  
Exchange Chromatography ◽  
Molecular Forms ◽  
Human Amniotic Fluid ◽  
Peptide Mass

CA125 antigen is a mucin-type molecule with a complex protein backbone and oligosaccharide chain structure. In this study, we characterized CA125 antigen from human amniotic fluid by gel filtration, ion-exchange chromatography, peptide mass fingerprinting and lectin-binding assays. The obtained results indicate CA125 to be structurally heterogeneous, existing in different glycoisoforms with subtle differences in the profile of molecular forms in comparison to placental tissue-derived and cancer-derived CA125 antigen. The complexity of CA125 structure suggests that it can act as a multifunctional molecule. Further investigation is therefore needed in order for the biological meaning of the tissue-specific structural forms to be comprehended fully. .

scholarly journals Molecular forms and activities of glycosidases in cultures of amniotic-fluid cells

1976 ◽  
Vol 155 (3) ◽  
pp. 599-605 ◽  
Author(s):  
B Hultberg ◽  
J Lindsten ◽  
S Sjöblad
Keyword(s):  
Ion Exchange ◽  
Amniotic Fluid ◽  
Lysosomal Enzymes ◽  
Gel Filtration ◽  
Ion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Molecular Forms ◽  
Cell Type ◽  
Amniotic Fluid Cells ◽  
Total Enzyme

Ion-exchange chromatography of gel filtration demonstrated the presence of different molecular forms of nine lysosomal enzymes in cultured amniotic-fluid cells. The patterns of molecular forms were similar to those known from skin fibroblasts and liver tissue. During cultivation total enzyme activities fluctuated with the number of passages, without any consistent trend of increase or decrease, and without correlation to the dominating cell type in the culture.

scholarly journals Role of plasmin in the degradation of the stroma-derived fibrin in human ovarian carcinoma

Blood ◽  
1990 ◽  
Vol 75 (8) ◽  
pp. 1673-1678 ◽  
Author(s):  
O Wilhelm ◽  
R Hafter ◽  
A Henschen ◽  
M Schmitt ◽  
H Graeff
Keyword(s):  
Ovarian Cancer ◽  
High Performance ◽  
Degradation Products ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Gel Filtration ◽  
Fibrin Degradation Products ◽  
Sds Page ◽  
Human Ovarian Carcinoma

Abstract The aim of this study was to evaluate the type of enzymes involved in tumor-associated fibrinolysis of the stroma component fibrin in ovarian cancer patients. For this purpose, the high-molecular-mass fibrin degradation products (HMM-XDP) were isolated from malignant ascitic fluid by protamine sulfate precipitation and further purified by gel filtration and acid precipitation. After reduction with 2- mercaptoethanol, the peptide chain components were separated by reverse- phase high-performance liquid chromatography (RP-HPLC). The nature of these components was elucidated by sodium dodecyl sulfate- polyacrylamide gel electrophoresis (SDS-PAGE) and N-terminal amino acid sequence analysis and compared with fibrin-derived fragments formed in vitro. The results indicate that plasmin is the essential protease involved in the degradation of the stroma-derived fibrin portion found in ovarian cancer ascites.

Fibrin/Fibrinogen Degradation Products in Amniotic Fluid

1975 ◽  
Author(s):  
D. W. Purdie ◽  
P. W. Howie ◽  
W. Edgar ◽  
G. R. M. Prentice
Keyword(s):  
Amniotic Fluid ◽  
Degradation Products ◽  
Polyacrylamide Gel Electrophoresis ◽  
Haemagglutination Inhibition ◽  
Gel Filtration ◽  
Normal Group ◽  
Double Immunodiffusion ◽  
Fibrinogen Degradation Products ◽  
The Mean

Fibrin/fibrinogen degradation products (F. D. P.) have been studied in amniotic fluid from normal pregnancies and in pregnancies complicated by hypertension, anencephaly, and hydramnios without foetal anomaly.Using the haemagglutination inhibition immunoassay, F. D. P. were identified in 51 of 53 (mean 3.75 mcg/ml, S.D.±2.80) samples of amniotic fluid from healthy patients with normal pregnancies between 14 and 40 weeks gestation. In the hypertensive group of 16 patients the mean F. D. P. level of 5.00 mcg/ml (S.D.±3.25), was not significantly different from the normal group. In 7 patients with anencephalic foetuses the amniotic fluid F. D. P. level was significantly elevated (mean 17.5 mcg/ml, S.D.±5.75), while in 6 patients with hydramnios without foetal anomaly the F. D. P. level (mean 6.00 mcg/ml ±3.25) was not significantly different from the normal group.Characterization of the amniotic fluid F. D. P. was carried out using double immunodiffusion polyacrylamide gel electrophoresis, and gel filtration on Sephadex G-200. The F. D. P. were present in two distinct components. The major fraction was fragment X (M. W. 240,000) and in a lower concentration, fragment D was identified.The detection of raised amniotic fluid F. D. P. levels in early pregnancy may provide a diagnostic test for open central nervous system foetal abnormalities.

Isolation and ultrastructural analysis of microfibrillar structures from foetal bovine elastic tissues. Relative abundance and supramolecular architecture of type VI collagen assemblies and fibrillin

1991 ◽  
Vol 99 (4) ◽  
pp. 797-807
Author(s):  
C.M. Kielty ◽  
C. Cummings ◽  
S.P. Whittaker ◽  
C.A. Shuttleworth ◽  
M.E. Grant
Keyword(s):  
Relative Abundance ◽  
Gel Filtration ◽  
Supramolecular Architecture ◽  
Collagen Vi ◽  
Type Vi Collagen ◽  
Bacterial Collagenase ◽  
New Information ◽  
Sds Page ◽  
Complex Polymers ◽  
Nuchal Ligament

Extensive intact assemblies of matrix macromolecules have been solubilized from foetal calf skin, nuchal ligament and aorta by a new procedure that includes bacterial collagenase digestion under non-reducing, non-denaturing conditions and gel filtration chromatography. Type VI collagen was identified as the major microfibrillar element of these tissues by SDS-PAGE analysis and Western blotting. Rotary shadowing electron microscopy of these preparations revealed by far the most abundant and extensive arrays of intact collagen VI microfibrils isolated to date. The distinct microfibrillar species, fibrillin, which was identified on the basis of its periodicity and morphology, was also solubilized in abundance by this protocol. Analysis of these complex polymers has generated new information on their supramolecular architecture and relative abundance in these tissues. The protocol also demonstrates that the release of intact collagen VI microfibrils from these tissues is largely dependent on the removal of the major collagen fibrils.

scholarly journals Purification of human interleukin 2 to apparent homogeneity and its molecular heterogeneity.

1982 ◽  
Vol 156 (2) ◽  
pp. 454-464 ◽  
Author(s):  
K Welte ◽  
C Y Wang ◽  
R Mertelsmann ◽  
S Venuta ◽  
S P Feldman ◽  
...  
Keyword(s):  
T Cell ◽  
Isoelectric Point ◽  
Interleukin 2 ◽  
Gel Filtration ◽  
Molecular Forms ◽  
Cytotoxic T Cell ◽  
Molecular Heterogeneity ◽  
Apparent Homogeneity ◽  
Sds Page ◽  
Daudi Cells

Interleukin 2 (IL-2), produced with and without co-stimulation by the Burkitt's lymphoma line Daudi, was purified 37,000-fold to apparent homogeneity from lymphocyte conditioned medium by (NH4)2SO4 precipitation, DEAE-cellulose ion-exchange chromatography, gel filtration, and chromatography on blue agarose and on Procion-red agarose. The purified IL-2 showed a 10(6) U/mg protein sp act. IL-2 produced in the absence of Daudi cells had a mol wt of 26,000 as measured by gel filtration and an isoelectric point of 6.7. This IL-2 showed a 16,000 and 17,000 mol wt in sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). IL-2, produced in the presence of Daudi cells (10(6)/ml), showed a mol wt of approximately 14,000, as measured by both gel filtration and SDS-PAGE, and an isoelectric point of 8.1. The purified IL-2 lacked detectable interferon (alpha and gamma), granulocyte-macrophage colony-stimulating factor, B cell growth factor, T cell-replacing factor, and thymocyte-differentiating activity and was free of any contaminating proteins as judged by silver staining in SDS-PAGE. All three molecular forms of IL-2 were biologically active at concentrations of 10(-11) - 10(-10) M, supporting the growth of human and murine cytotoxic T cell lines.

scholarly journals Role of plasmin in the degradation of the stroma-derived fibrin in human ovarian carcinoma

Blood ◽  
1990 ◽  
Vol 75 (8) ◽  
pp. 1673-1678
Author(s):  
O Wilhelm ◽  
R Hafter ◽  
A Henschen ◽  
M Schmitt ◽  
H Graeff
Keyword(s):  
Ovarian Cancer ◽  
High Performance ◽  
Degradation Products ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Gel Filtration ◽  
Fibrin Degradation Products ◽  
Sds Page ◽  
Human Ovarian Carcinoma

The aim of this study was to evaluate the type of enzymes involved in tumor-associated fibrinolysis of the stroma component fibrin in ovarian cancer patients. For this purpose, the high-molecular-mass fibrin degradation products (HMM-XDP) were isolated from malignant ascitic fluid by protamine sulfate precipitation and further purified by gel filtration and acid precipitation. After reduction with 2- mercaptoethanol, the peptide chain components were separated by reverse- phase high-performance liquid chromatography (RP-HPLC). The nature of these components was elucidated by sodium dodecyl sulfate- polyacrylamide gel electrophoresis (SDS-PAGE) and N-terminal amino acid sequence analysis and compared with fibrin-derived fragments formed in vitro. The results indicate that plasmin is the essential protease involved in the degradation of the stroma-derived fibrin portion found in ovarian cancer ascites.

scholarly journals Purification and Properties of a Polyester Polyurethane-Degrading Enzyme from Comamonas acidovorans TB-35

1998 ◽  
Vol 64 (1) ◽  
pp. 62-67 ◽  
Author(s):  
Yukie Akutsu ◽  
Toshiaki Nakajima-Kambe ◽  
Nobuhiko Nomura ◽  
Tadaatsu Nakahara
Keyword(s):  
Degradation Products ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Gel Filtration ◽  
Water Soluble ◽  
Degrading Enzyme ◽  
Comamonas Acidovorans ◽  
Water Soluble Compound ◽  
Sds Page ◽  
Hydrophobic Adsorption

ABSTRACT A polyester polyurethane (PUR)-degrading enzyme, PUR esterase, derived from Comamonas acidovorans TB-35, a bacterium that utilizes polyester PUR as the sole carbon source, was purified until it showed a single band in sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). This enzyme was bound to the cell surface and was extracted by addition of 0.2%N,N-bis(3-d-gluconamidopropyl)deoxycholamide (deoxy-BIGCHAP). The results of gel filtration and SDS-PAGE showed that the PUR esterase was a monomer with a molecular mass of about 62,000 Da. This enzyme, which is a kind of esterase, degraded solid polyester PUR, with diethylene glycol and adipic acid released as the degradation products. The optimum pH for this enzyme was 6.5, and the optimum temperature was 45°C. PUR degradation by the PUR esterase was strongly inhibited by the addition of 0.04% deoxy-BIGCHAP. On the other hand, deoxy-BIGCHAP did not inhibit the activity whenp-nitrophenyl acetate, a water-soluble compound, was used as a substrate. These observations indicated that this enzyme degrades PUR in a two-step reaction: hydrophobic adsorption to the PUR surface and hydrolysis of the ester bond of PUR.

Collagen degradation products modulate matrix metalloproteinase expression in cultured articular chondrocytes

2005 ◽  
Vol 24 (1) ◽  
pp. 63-70 ◽  
Author(s):  
M. Fichter ◽  
U. Körner ◽  
J. Schömburg ◽  
L. Jennings ◽  
A. A. Cole ◽  
...  
Keyword(s):  
Matrix Metalloproteinase ◽  
Articular Chondrocytes ◽  
Degradation Products ◽  
Collagen Degradation ◽  
Collagen Degradation Products
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