Showing papers in "Connective Tissue Research in 1980"
TL;DR: Positive effects on the tensile characteristics of swine digital extensors were found following twelve months of exercise training, and the tendons from the exercised animals became stronger as a material and exhibited hypertrophy.
Abstract: Positive effects on the tensile characteristics of swine digital extensors were found following twelve months of exercise training. Compared to sedentary controls, the tendons from the exercised animals became stronger as a material and exhibited hypertrophy. These biomechanical results were supported by biochemical analyses of tendon composition. Exercise increased the concentration of collagen as well as the total weights of the tendons. For determining stress and strain in tendon material, we used specially designed instruments to measure the tendon cross-sectional area, and a video dimensional analyzer system to measure accurately its "non-contact" tensile strain. With these newly developed apparatus, the mechanical properties of the tendons were accurately determined so that the effects of exercise training could be compared.
333 citations
TL;DR: A new device, built and tested in the laboratory enabled us to obtain isometric tension curves by measuring a "swelling" phenomenon observed in other directions, and suggests that these parameters depend not only on the nature but also on the density of the collagen crosslinks.
Abstract: Isometric tensions developed in connective tissue under the influence of temperature have previously been measured by their effect on a “shrinkage” phenomenon observed along the main axis of the collagen fibers. A new device, built and tested in our laboratory enabled us to obtain isometric tension curves by measuring a “swelling” phenomenon observed in other directions. Curves recorded in the two systems have been compared. Shapes were unchanged and the parameters chosen to define these morphologies showed the same values in both types of experiments.The parameters, which are independent of maximal tension, were also found to be independent of the total collagen content of the sample. Interpretation of the results suggests that these parameters depend not only on the nature but also on the density of the collagen crosslinks.The new device is applicable to minute tissue samples, whatever their fragility and fiber orientation.
172 citations
TL;DR: A direct view has been obtained of the manner in which the fibrous components components and chondrocytes in hyaline cartilage respond to the application of uniaxial tensile loading and plane-strain compressive loading.
Abstract: A direct view has been obtained of the manner in which the fibrous components and chondrocytes in hyaline cartilage respond to the application of uniaxial tensile loading and plane-strain compressive loading.A micro-mechanical testing device has been developed which inserts directly into the stage of a high-resolution optical microscope fitted with Nomarski interference contrast and this has permitted simultaneous morphological and mechanical observations to be conducted on articular cartilage maintained in its wet functional condition.Aligned and crimped fibrous arrays surround the deeper chondrocytes and can be observed to undergo well-defined geometric changes with applied stress. It is thought that these arrays may act as displacement or strain sensors transmitting mechanical information from the bulk matrix to their associated cells thus inducing a specific metabolic response.The process of tissue recovery following sustained high levels of compressive loading can also be observed with this experimen...
103 citations
TL;DR: A radioimmunoassay developed for the determination of desmosine was injected into rabbits which developed useful titers of serum antibodies after six months, and the antibody is highly selective for Desmosine, reacting less than 1% with other known crosslinks.
Abstract: A radioimmunoassay was developed for the determination of desmosine. Desmosine conjugated to albumin was injected into rabbits which developed useful titers of serum antibodies after six months. A radioactive probe was prepared with desmosine using [125I]-Bolton-Hunter reagent. Bound desmosine was separated from free desmosine by cellulose acetate filter binding. The sensitivity of the assay is 1-50 picomoles of desmosine. The antibody is highly selective for desmosine, reacting less than 1% with other known crosslinks. Some prepurification may be necessary with complex samples which contain trace amounts of elastin peptides.
100 citations
TL;DR: A method was developed to quantitate the collagen-proteoglycan interaction in tissue sections and the initial results show a variability of this interaction in different tissues, suggesting a reduction of collagen- Proteoglycans interaction in atherosclerotic lesions.
Abstract: Since the dye Sirius Red reacts with the basic groups of collagen, and it is possible to hydrolyze the proteoglycans bound to collagen by enzymatic digestion, a method was developed to quantitate the collagen-proteoglycan interaction in tissue sections. The method consists of measuring, with a spectrophotometer, the amount of dye bound to control and papain-digested tissue sections. The difference observed between the results obtained in control and digested sections is considered to be due to the unmasking of basic groups of collagen originally bound to proteoglycans. The initial results show a variability of this interaction in different tissues. They also suggest a reduction of collagen-proteoglycan interaction in atherosclerotic lesions.
50 citations
TL;DR: Findings indicate that mechanical stress plays a primary role in maturation of regenerating tendon, and emphasizes the importance of close approximation of tendon ends in the management of tendon ruptures and lacerations.
Abstract: Regenerated tendon tissue replacing partial-thickness defects produced in rabbit calcaneal tendons was seen to undergo almost complete maturation between three and sixteen weeks after surgery. At this stage, most collagen fibers had large diameters as in normal tendon, and the majority of elastic fibers were mature. This findings, along with those of a previous study on completely severed and unsutured tendons, indicate that mechanical stress plays a primary role in maturation of regenerating tendon. This emphasizes the importance of close approximation of tendon ends in the management of tendon ruptures and lacerations. This is possibly by surgical repair.
47 citations
TL;DR: Investigation of eggshell membrane by amino acid analysis and radioimmunoassay established the presence of the crosslinking amino acids desmosine and isodesmosine at levels of 28 microgram/100 mg dry weight.
Abstract: Investigation of eggshell membrane by amino acid analysis and radioimmunoassay established the presence of the crosslinking amino acids desmosine and isodesmosine at levels of 28 μg/100 mg dry weig...
46 citations
TL;DR: Large variations were found in the ability of six species of sera to support growth of rabbit, human and dog articular chondrocytes in monolayer culture and there was no consistent relation between the response of chonds from a given species and its homologous serum.
Abstract: Large variations were found in the ability of six species of sera to support growth of rabbit, human and dog articular chondrocytes in monolayer culture. In most instances the DNA content of the cell pellets increased directly as the serum concentration rose from 10 to 30%. Indications of inhibitory as well as growth-promoting actions were found in some sera. Stimulation of rabbit chondrocyte proliferation by increasing concentrations of serum was accompanied by a reduction of radiosulfate incorporation and cell protein content. There was no consistent relation between the response of chondrocytes from a given species and its homologous serum. However, the growth of human chondrocytes was greatly potentiated by human serum provided that interference with initial attachment of the cells to the culture flask by homologous serum was overcome by priming with fetal calf serum.
44 citations
TL;DR: The simultaneous appearance of abnormal collagen fibrils and matrix vesicles containing lysosomal enzymes in the majority of electron photomicrographs suggests an alteration in the ground-substance as a main cause of non-hereditary collagen dysplasia.
Abstract: Quite unrelated diseases affecting different organs can lead to the appearance of abnormal collagen fibrils, which may be due to genetic disorders, transformation of mesenchyme cells, increased collagen synthesis or a change in the ground-substance. The simultaneous appearance of abnormal collagen fibrils and matrix vesicles containing lysosomal enzymes in the majority of our electron photomicrographs suggests an alteration in the ground-substance as a main cause of non-hereditary collagen dysplasia.
41 citations
TL;DR: The periarticular connective tissue contracture process, therefore, is different from the intraarticular process in which fibrofatty connectedive tissue proliferation is prominent.
Abstract: The distribution of collagen types was studied in control and immobilized periarticular connective tissue to test the hypothesis that inflammatory tissue is the basis of the contracture process. The hypothesis derives from the fact that Type III collagen is prominent in inflammatory processes. The content of Type I and Type III collagen was determined by standard differential salt precipitation of the pepsin solubilized collagen, followed by resolution of the subunits on CM-cellulose. Analysis of the peptide fragments produced by cyanogen bromide was performed and their distribution determined by SDS gel electrophoresis. The collagen of control and immobilized periarticular connective tissue proved to be entirely Type I. Type III was not present. The periarticular connective tissue contracture process, therefore, is different from the intraarticular process in which fibrofatty connective tissue proliferation is prominent.
39 citations
TL;DR: The upper dermis (the top three layers) consisting of fine collagen fibers had a higher ratio of hyaluronic acid/dermatan sulfate than the lower layers where coarse collagen fibers are mainly located, and possible involvement of these macromolecular constituents in collagen fibrillogenesis in vivo is also discussed.
Abstract: Distribution of type I and type III collagens, glycosaminoglycans and non-collagenous glycoprotein(s) in calf skin has been investigated in five horizontally sliced layers of 250 microns thickness from the surface to the bottom. The upper dermis (the top three layers) consisting of fine collagen fibers had a higher ratio of hyaluronic acid/dermatan sulfate than the lower layers where coarse collagen fibers are mainly located. The ratios of total glycosaminoglycans/collagen and glycoprotein(s) collagen were higher in the upper dermis, while dermatan sulfate/collagen remained constant throughout the dermis. Relative content of type III collagen/total collagen (type I + III collagens) was also higher in the upper dermis. Possible involvement of these macromolecular constituents in collagen fibrillogenesis in vivo is also discussed.
TL;DR: A novel steady-state gel-matrix model system is described which facilitates the quantitative kinetic study of the influence of media and matrix composition on a precipitation process, and its significance for the study of in vitro and in vivo precipitation processes in general is discussed.
Abstract: A novel steady-state gel-matrix model system is described which facilitates the quantitative kinetic study of the influence of media and matrix composition on a precipitation process. The potentialities of the system are illustrated in experiments in which the precipitation of calcium phosphate in a polyacrylamide film is studied as a function of the calcium and phosphate concentration in the solutions flowing along opposite sides of the film.Addition of pyrophosphate to the reactant solutions was found to diminish the (calcium) × (phosphate) millimolar product at which precipitation starts, indicating a positive effect on nucleation. The slope of the curve was found to decrease, which points to a negative influence of pyrophosphate on the crystal growth process. Incorporation of collagen in the matrix did not change the curve, but incorporation of chondroitin sulfate decreased the formation product since the intercept of the curve was reduced. The usefulness of the system compared with test-tube and non-...
TL;DR: The AGAG components of cartilage proteoglycans were distributed with a certain regularity in the fractions of CsCl density gradients, but underwent changes with increasing age and in arthritic process.
Abstract: Proteoglycans extracted from normal and arthritic bovine articular cartilage of various ages were fractionated and purified under associative and dissociative conditions. After proteolytic digestion, the composition of the acidic glycosaminoglycans (AGAG) in the proteoglycans was determined enzymatically by digestion with chondroitinase-AC II, chondroitinase-ABC, Streptomyces hyaluronidase and keratanase.Under both associative and dissociative conditions, uniform distribution of chondroitin sulfate (CS) isomers from proteoglycans of different ages was observed: With increasing age, the relative proportion of 4-sulfated disaccharide units in total AGAG decreased, whereas that of 6-sulfated disaccharide units increased. The relative proportion of 4-sulfated disaccharide units in total CS and the ratio of 4-sulfated disaccharide units to 6-sulfated disaccharide units were greater in arthritic cartilages than in normal cartilages of the same ages. At all three ages studied, the relative proportion of 4-sulfat...
TL;DR: The elastin radioimmunoassay can be modified to detect bovine or other elastins depending upon the choice of radiolabeled antigen and permits determination and quantification of elast in synthesis and degradation in a wide range of species without having to raise specific antibodies to each.
Abstract: By radioimmunologic techniques, it has been possible to detect two serologically distinct elastin antibody fractions in antiserum prepared against bovine ligamentum nuchae elastin; a species specific fraction which bound only bovine elastin, and a population of antibodies with affinity for elastin from porcine aorta, bovine ligamentum nuchae and aorta, rabbit and hamster lung, and human aorta. Bovine specific antibodies were detected when bovine alpha-elastin was used as specific ligand whereas interspecies reactivity was detected when radiolabeled nonbovine solubilized elastins were used in the assay. Two distinct antibody populations were confirmed using immunoabsorption techniques. Thus, the elastin radioimmunoassay can be modified to detect bovine or other elastins depending upon the choice of radiolabeled antigen. The assay permits determination and quantification of elastin synthesis and degradation in a wide range of species without having to raise specific antibodies to each.
TL;DR: A study in which aorta was exhaustively extracted with agents previously claimed to extract SGP yielded mixtures of proteins which, upon separation by polyacrylamide gel electrophoresis, were found to consist largely of actin, collagen, myosin and other known proteins.
Abstract: It has been repeatedly claimed that structural glycoprotein or SGP is a major component of many connective tissues, particularly aorta. SGP is usually defined as material extractable from tissue with chaotropic agents such as urea and having an amino acid composition matching that of other, similar extracts. It has been claimed that SGP comprises 10-60% of the dry weight of aortic tissue. This paper describes a study in which aorta was exhaustively extracted with agents previously claimed to extract SGP. All the extracts yielded mixtures of proteins which, upon separation by polyacrylamide gel electrophoresis, were found to consist largely of actin, collagen, myosin and other known proteins. No significant quantity of any one unidentified protein was found which could have been SGP. This study casts grave doubt upon the existence of SGP as a major component of aorta.
TL;DR: It is suggested that collagen-bound collagenase is not an artifact of the extraction procedure but rather a physiological reality, probably corresponding in the living animal to the enzyme closely associated with extracellular collagen fibers, revealed by immunohistochemical methods.
Abstract: The presence of collagenase bound to collagen extracted and purified from several animal and human sources by a standard procedure has been confirmed by different methods. Polyacrylamide (10%) gel electrophoresis at pH 8.1 of intact or "spontaneously"degraded neutral salt soluble collagen results in the separation of two components: the upper one says at the origin and represents collagen or collagen ragments, whereas the lower protein component contains no collagen, often preserves specific collagenolytic activity, and migrates as a single band in SDS/polyacrylamide electrophoresis. With lower polyacrylamide gel concentration the electrophoretic separation of the two components is less clear. Removal of the lower protein component from collagen solutions by two different methods (TCA-ethanol purification cycles and pepsin digestion) results in concomitant loss of their "spontaneous" instability. Eluates of the lower protein component stimulate the heterologous production of a monospecific antibody capable of inhibiting the collagenolytic activity of homologous crude collagenase preparations. It is suggested that collagen-bound collagenase is not an artifact of the extraction procedure but rather a physiological reality, probably corresponding in the living animal to the enzyme closely associated with extracellular collagen fibers, revealed by immunohistochemical methods.
TL;DR: Cells cultured from calf ligamentum nuchae produce extracellular microfibrils identical to those of intact elastic tissue as determined by ultrastructural appearance, degradative enzyme susceptibilities and amino acid composition, but the cultures do not appear to synthesize the amorphous component, elastin.
Abstract: Cells cultured from calf ligamentum nuchae produce extracellular microfibrils identical to those of intact elastic tissue as determined by ultrastructural appearance, degradative enzyme susceptibilities and amino acid composition. A method to isolate large amounts of the microfibrillar component using fluorescein mercuric acetate is described. The cultures do not appear to synthesize the amorphous component, elastin, in that radioactive desmosine and isodesmo-sine were not detected after incubating cultures with labeled lysine nor was elastin seen ultrastructurally.
TL;DR: Rabbit vitreous contains minor components that migrate on polyacrylamide gel electrophoresis more slowly than the minor 1 alpha and 2 alpha chains of rabbit cartilage.
Abstract: Where similar techniques are employed, the properties of collagen peptides extracted from rabbit vitreous by pepsin treatment resemble those described in published reports of bovine vitreous. Further, rabbit vitreous also contains minor components that migrate on polyacrylamide gel electrophoresis more slowly than the minor la and 2α chains of rabbit cartilage. Properties of the major vitreous components suggest a mixture of the closely-related cartilage collagen chains al(II) and 3α.
TL;DR: The 70,000 dalton protein from the elastic cartilage is identified as tropoelastin, a relatively heavily labeled protein that was precipitated by elastin-specific antibody prepared against hamster insoluble aorta Elastin.
Abstract: Elastic ear cartilage and aortae from nine-day-old hamsters were incubated in Krebs-Ringer medium containing ascor-bate, β-aminopropionitrile and either [14C]proline or [3H] valine. The [14C] proline-labeled proteins were separated by chromatography on Agarose A-5 m in sodium dodecyl sulfate and their [14C]-hydroxyproline content measured. A fraction having an approximate molecular weight of 70,000 measured by polyacrylamide gel electrophoresis had a [14C] hydroxyproline content of 9.1% in the cartilage and 11.8% in the aorta. This fraction was also relatively heavily labeled with [3H]valine. The 70,000 dalton, [3H]valine-labeled protein of both the aorta and ear cartilage was precipitated by elastin-specific antibody prepared against hamster insoluble aorta elastin, but no higher molecular weight protein was immunoprecipitated. Based on these results we identify the 70,000 dalton protein from the elastic cartilage as tropoelastin.
TL;DR: Elastin peptides were fractionated on a SE-Sephadex C-50 column by sequential elution with the following buffers and only polar fractions containing desmosine and isodesmosine proved to be antigenic when assayed by double immunodiffusion test.
Abstract: Elastin peptides were fractionated on a SE-Sephadex C-50 column by sequential elution with the following buffers: a) 0. 01 N acetic acid, b) linear gradient from 0. 05 N sodium acetate buffer, pH 4. 0 to 0. 4 N sodium acetate buffer, pH 4. 6, c) 0. 4 N sodium acetate buffer, pH 4. 6, d) 0. 5 N sodium hydroxide. Based on the elution profile, tubes were pooled into 12 fractions. Amino acid analysis of these fractions showed that the first 7 fractions contained mostly nonpolar peptides which lacked desmosine and isodesmosine residues. The other 5 fractions contained more polar peptides with desmosine and isodesmosine. Only polar fractions containing desmosine and isodesmosine proved to be antigenic when assayed by double immunodiffusion test. These antigenic peptide fractions were subjected to further analysis. Separation by gel filtration on Bio-Gel P-150 showed that each of the antigenic fractions consisted of at least 3 polypeptides with apparent molecular weights of 57, 000, 46, 000 and 9, 000. The two h...
TL;DR: A biochemical study of skin biopsies of young normal subjects and two groups of aged subjects revealed an increase in ground substance and fragmentation of collagen bundles in the dermis of older subjects, agreeing in part with the changes observed by light and electron microscopy.
Abstract: A biochemical study of skin biopsies of young normal subjects and two groups of aged subjects, one active and the other confined to bed, was made and the results compared with histological observations. Skin biopsies were incubated with 3H proline and 14C glucosamine and connective tissue components were sequentially extracted. In each extract the specific activity of total and collagenous protein was determined. With age a decrease was observed in both the amount of protein and the 3H proline incorporation of collagenous extracts. A parallel increase in 14C glucosamine incorporation was also noticed in extracts containing glycosaminoglycans and structural proteins. These results agree in part with the changes observed by light and electron microscopy, which revealed an increase in ground substance and fragmentation of collagen bundles in the dermis of older subjects.
TL;DR: The ratio of type I collagen/type III collagen was relatively constant, regardless of the size of the fibrous materials, and the ratios of glycosaminoglycans/collagen and non-collagenous glycoproteins/Collagen were significantly higher in the thinner fiber fractions.
Abstract: Collagen fibrils, fibers and fiber bundles have been isolated from insoluble calf skin chips by disaggregation with a neutral salt buffer followed by differential centrifugation. The ratio of type I collagen/type III collagen was relatively constant, regardless of the size of the fibrous materials. On the other hand, the ratios of glycosaminoglycans/collagen and non-collagenous glycoproteins/collagen were significantly higher in the thinner fiber fractions. However, the ratio of hyaluronic acid to dermatan sulfate was unchanged. The role of the non-collanenous constituents in collagen fibrillogenesis in vivo is discussed.
TL;DR: It is clear from these wet tissue studies that the fibrous layout in hyaline cartilage is considerably more "disciplined" than has been previously recognized from morphological data obtained using more indirect experimental techniques involving prepared histological sections or scanning and transmission electron microscopy.
Abstract: The structural features of hyaline cartilage maintained in its wet functional condition have been examined using the technique of Nomarski interference microscopy. The collagenous arrays and associated chondrocytes in both the extreme superficial layers and in the deeper subsurface zones were satisfactorily imaged with this technique. Most significantly the collagen fibers were observed to possess a geometric waveform or "crimp" of varying acuteness and the role of this crimp is discussed in relation to the mechanical and biological function of the tissue. It is clear from these wet tissue studies that the fibrous layout in hyaline cartilage is considerably more "disciplined" than has been previously recognized from morphological data obtained using more indirect experimental techniques involving prepared histological sections or scanning and transmission electron microscopy.
TL;DR: Analysis of the chemical composition of the matrix of the medullary bone indicates that proteoglycans are present in large amounts and the calcium binding glycoprotein appears to be a compound present in different calcified tissues.
Abstract: The discovery in calcifying cartilage of a glycoprotein, endowed with high calcium affinity and alkaline phosphatase activity, has prompted the investigation of the presence of this compound in other calcified tissues. From medullary bone, a tissue which is highly mineralized under estrogen stimulus, a glycoprotein has been extracted which had the properties described. Besides the high calcium affinity (Kd = 10−7M), this protein shows phosphatase activity and rate of hydrolysis of ATP, GTP and pyrophosphate was measured. Analysis of the chemical composition of the matrix of the medullary bone indicates that proteoglycans are present in large amounls. The calcium binding glycoprotein appears to be a compound present in different calcified tissues.
TL;DR: Hydroxyproline analyses of tissue hydrolysates indicated that chondrosarcoma samples contained significantly less collagen than normal articular cartilage, yet were incorporating significantly greater amounts of 3H-proline into 3 H-hydroxyproline.
Abstract: Protein synthesis by human chondrosarcoma tissue and normal articular cartilage was studied in organ and primary monolayer cell culture systems. Protein synthesis by cell cultures was evaluated at 2,7, and 21 days after plating. When compared to normal, incorporation of 3H-proline and 35S-methionine into proteins was elevated in chondrosarcoma samples under both culture conditions. Hydroxyproline analyses of tissue hydrolysates indicated that chondrosarcoma samples contained significantly less collagen than normal articular cartilage, yet were incorporating significantly greater amounts of 3H-proline into 3H-hydroxyproline. Collagen production by cell cultures was assessed by digestion of samples with purified clostridial collagenase. Chondrosarcoma cells produced more collagenase-sensitive protein than normal cells at all intervals. SDS polyacrylamide gel analyses of all preparations showed two collagenase-sensitive proteins with apparent molecular weights of 165,000 and 175,000. Decreased synthesis of a...
TL;DR: Results indicate that hyaluronidase can degrade matrix by releasing proteoglycan monomers, and indicate that endogenous enzymes degrade matrix proteoglycans into smaller units.
Abstract: Degradation of cartilage matrix was studied by characterizing the size and amount of products released from cultures of fetal rat long bones into the medium. Na35SO4 was administered to pregnant rats and the fetal radii and ulnae were explanted 24 hours later and cultured in chemically defined medium. Under these conditions, 35S-activity is released from the cartilagenous ends of the explants into the culture medium gradually over 72 hours. The 35S-labeled molecules in the medium were smaller than proteoglycan monomers and larger than papain-released products when chromatographed on Sepharose 2B. The amount of 35S-activity released into the medium decreased when heat-inactivated rat serum or plasma or heparin was added to the medium. Adding proteases or hyaluronidases or freeze-thawing the tissue enhanced release of 35S-activity.Streptomyces hyaluronidase was chromatographed on CM-cellulose and Ultrogel to remove protease activity. The protease-free enzyme released 35S-labeled molecules, similar in size t...
TL;DR: Effect on thermal coacervation, solubilization of lipophilic dye, difference spectra measurements and fluorescence probe indicate that the detergents examined interact with α-elastin below critical micelle concentration in two different ways.
Abstract: Coacervation is known to be a fundamental step in elastogenesis that is influenced by various naturally occurring substances in connective tissue. Therefore, interaction of α-elastin with ionic detergents (sodium dodecyl sulfate, dodecylbenzyldimethylammonium bromide, cetyltrimethylammonium bromide, pentadecacarbethoxytrimethylammonium bromide and cetylpyridinium bromide) was studied as a model for the elastin interaction with amphiphilic substances. The course of the interactions was followed by the effect on thermal coacervation, solubilization of lipophilic dye, difference spectra measurements and fluorescence probe. The results indicate that the detergents examined interact with α-elastin below critical micelle concentration in two different ways. In the first case, a mixed micelle can be formed from the detergent molecules (SDS, DBD) and a-elastin; alternatively mixed micelles are not formed, but the detergent molecules (CT, PCT, CP) do interact with elastin. The interaction of α-elastin at the isoel...
TL;DR: Results of this study indicate that inhibition is due to an IgG antibody to the porcine elastase that binds to activated Sepharose and the resulting immunoadsorbent system allows purification of pancreaticElastase by affinity chromatography.
Abstract: By affinity chromatography on porcine pancreatic elastase bound to Sepharose an anti-elastase antibody was isolated from the serum of rabbits who had been injected with porcine pancreatic elastase and fed a light cholesterol diet. The protein (molecular weight 150,000 daltons) was recovered with both a high degree of purity and intact biological properties. It specifically inhibited elastolysis by a mechanism distinct from that of known elastase inhibition by alpha1 antiprotease and alpha2 macroglobulin. Results of this study indicate that inhibition is due to an IgG antibody to the porcine elastase. The antigen-antibody reaction appears to involve the active site of elastase. The isolated IgG in turn binds to activated Sepharose and the resulting immunoadsorbent system allows purification of pancreatic elastase by affinity chromatography.
TL;DR: Cells from a rat chondrosarcoma in primary suspension culture were used for labeling studies and isolation of RNA, and a distinct fraction of poly(A+)RNA sedimented in the region of 26S on denaturing sucrose gradients.
Abstract: Cells from a rat chondrosarcoma in primary suspension culture were used for labeling studies and isolation of RNA. With extended labeling, a distinct fraction of poly(A+)RNA. sedimented in the region of 26S on denaturing sucrose gradients. Similar profiles were obtained with labeled poly(A+)RNA isolated from normal cartilage tissues, as well as from chondrocytes in culture. This mRNA coded for collagenous protein when translated in a cell-free system derived from wheat germ. Among the polypeptides synthesized in vitro, one distinctive component of approximately 150,000 daltons was detected by gel electrophoresis. Polypeptides of identical size, sensitive to bacterial collagenase, were also synthesized in vitro by polysomes isolated from the chondrosarcoma.
TL;DR: The results initially suggested a substrate specificity by human elastase for humanElastin, but additional data indicated that the method of purification of elastins significantly affected the rates of solubilization by both human and porcine pancreaticElastase.
Abstract: Elastase from human pancreas was purified by ion-exchange and affinity chromatography with an 85% yield and to approximately 95% homogenicity as judged by discontinuous slab-gel electrophoresis. The human enzyme differed in chemical and enzymatic properties in comparison to porcine pancreatic elastase. Human elastase antiserum exhibited no cross-reactivity with porcine elastase, but precipitated the human enzyme. The human elastase possessed 25% of the enzymatic activity of porcine elastase in solubilizing alkaline-extracted bovine elastin, but solubilized human aortic elastin at a much higher rate. The results initially suggested a substrate specificity by human elastase for human elastin, but additional data indicated that the method of purification of elastins significantly affected the rates of solubilization by both human and porcine pancreatic elastase.