Interactions between Cells and Collagen V Molecules or Single Chains Involve Distinct Mechanisms

Acid-soluble and pepsin-treated collagen V were prepared from fetal human bones or human placenta, respectively, to be tested for potential cell adhesion promoting activity. Out of 14 different collagen I-adhering cell lines, 10 showed distinct adhesion to collagen V. In all cases adhesion was followed by spreading. The activities of intact and pepsin-solubilized collagen V were similar, suggesting that the cell binding sites are restricted to the triple-helical domain of the molecules. Cell adhesion was also induced by the unfolded form of collagen V and after separation of the α chains by heparin affinity chromatography. Isolated α2(V) chains, rich in RGD sequences, were more efficient than isolated α1(V) chains. However, cell adhesion to native or denatured collagen V did not proceed by the same molecular mechanisms as shown by cell adhesion inhibition experiments. Cell adhesion to native collagen V was insensitive to the presence of RGD-containing synthetic peptides while adhesion to denatured collagen V was inhibited by the peptides. Furthermore, the results strongly suggested a major role for α1α1 and α2β1 integrins in the RGD-independent cell adhesion to native collagen V. These data indicate that collagen V is a specific adhesive substrate for different cell types. It also suggests that distinct sets of RGD-dependent and RGD-independent receptors mediate cell attachment to unfolded and native collagen V, respectively. This mechanism is shared by at least the interstitial collagens I and VI, which supports the hypothesis that when included in the triple-helical conformation of collagens, RGD sequences are either not accessible to cells or exhibit specific conformations recognized by different integrins.     

Interaction between collagens and glycosaminoglycans investigated using a surface plasmon resonance biosensor.

The interactions of glycosaminoglycans with collagens and other glycoproteins in extracellular matrix play important roles in cell adhesion and extracellular matrix assembly. In order to clarify the chemical bases for these interactions, glycosaminoglycan solutions were injected onto sensor surfaces on which collagens, fibronectin, laminin, and vitronectin were immobilized. Heparin bound to type V collagen, type IX collagen, fibronectin, laminin, and vitronectin; and chondroitin sulfate E bound to type II, type V, and type VII collagen. Heparin showed a higher affinity for type IX collagen than for type V collagen. On the other hand, chondroitin sulfate E showed the highest affinity for type V collagen. The binding of chondroitin sulfate E to type V collagen showed higher affinity than that of heparin to type V collagen. These data suggest that a novel characteristic sequence included in chondroitin sulfate E is involved in binding to type V collagen.     

Fluorescence microscopic distinction between elastin and collagen

Summary Distinction between elastin and collagen in arteriosclerotic lesions is difficult because immature and incompletely cross-linked collagen bind so-called elastica stains; furthermore, abnormal collagen can lack cross-striation and thus resemble elastin in electron microscopy. However, collagen and elastin differ significantly in their content of basic amino acids and hence in their affinity for heteropolyacids. This chemical difference was utilized for the development of a fluorescence microscopic method for distinction between collagen and elastin.Paraffin sections of human autopsy material were treated with a 1% aqueous solution of phosphomolybdic acid (PMA) for five minutes, rinsed in distilled water, dehydrated and mounted. Other series were treated with the PMA-molybdenum blue reaction and with various special stains.Only elastic membranes of aorta, the elastica interna and externa of sizable arteries, and true elastic fibers remained strongly fluorescent; the autofluorescence of collagen, reticulum fibers, basement membranes, pseudo-elastic fibers, and elastic membranes in small arteries was quenched. In other series PMA abolished the fluorescence of basic fluorochromes.Correlation of fluorescence and direct light microscopic observations with chemical and electron microscopic data showed that the PMA-fluorescence method permits distinction between elastin and various types of collagen.     

Forensic visualization of foreign matter in human tissue by near-infrared spectral imaging: methodology and data mining strategies.

BACKGROUND: Rapidity of data acquisition, high image fidelity and large field of view are of tremendous value when looking for chemical contaminants or for the proverbial "needle in the haystack" - in this case foreign inclusions in histologic sections of biopsy or autopsy tissues. Near infrared chemical imaging is one of three chemical imaging techniques (NIR, MIR and Raman) based on vibrational spectroscopy, and provides distinct technical advantages for this application. METHODS: We have chosen to utilize and evaluate near infrared (NIR) imaging for studies of foreign materials in tissue because the experimental configuration is relatively simple, data collection is rapid, and large sample areas can be screened with high image fidelity and spatial resolution. RESULTS: We have shown that NIR imaging can readily find and identify silicone gel inclusions in biological tissue samples. Additionally, preliminary results indicate that spectral signatures in the data set are also potentially sensitive to structural changes in the surrounding tissue that may be induced by the foreign body. CONCLUSIONS: NIR chemical imaging is a powerful, non-destructive tool for localization and identifying foreign contaminants in biological tissue. Preliminary results indicate that NIR imaging is also sensitive enough to differentiate tissue types (perhaps based on collagen structural differences), and provide data on the spatial localization of these components.     

Important data in understanding the interaction between the cell and collagen

In a collagen preparation obtained from rat tail tendon, besides triglycerides, cholesterol, cholesterol esters, and phosphatides whose presence have been reported before, the presence of plasmalogens and glycolipids including gangliosides was observed.     

Histochemical study of cardiac mast cells degranulation and collagen deposition: interaction with the cathecolaminergic system in the rat

Although their role in the cardiovascular system is still largely unknown, mast cells are present in the myocardium of both experimental animals and humans. Interestingly, cathecolaminergic nerve fibres and mast cells are often described in close morphological and functional interactions in various organs. In the present study we investigated the effects of chronic interference with beta-adrenergic receptors (via either sympathectomy or beta-blockade) on cardiac mast cell morphology/activation and on interstitial collagen deposition. In rats subjected to chemical sympathectomizy with the neurotoxin 6-hydroxydopamine (6-OHDA) we observed a significant increase of mast cell density, and in particular of degranulating mast cells, suggesting a close relationship between the cardiac catecholaminergic system and mast cell activation. In parallel, chronic 6-OHDA treatment was associated with increased collagen deposition. The influence of the beta-adrenergic receptor component was investigated in rats subjected to chronic propranolol administration, that caused a further significant increase in mast cell activation associated with a lower extent of collagen deposition when compared to chemical sympathectomy. These data are the first demonstration of a close relationship between rat cardiac mast cell activation and the catecholaminergic system, with a complex interplay with cardiac collagen deposition. Specifically, abrogation of the cardiac sympathetic efferent drive by chemical sympathectomy causes mast cell activation and interstitial fibrosis, possibly due to the local effects of the neurotoxin 6-hydroxydopamine. In contrast, beta-adrenergic blockade is associated with enhanced mast cell degranulation and a lower extent of collagen deposition in the normal myocardium. In conclusion, cardiac mast cell activation is influenced by beta-adrenergic influences.     

Three-dimensional spatial relationship between the collagen fibrils and the inorganic calcium phosphate crystals of pickerel (Americanus americanus) and herring (Clupea harengus) bone

High-voltage (1.0 MV) electron microscopy and stereomicroscopy, electron probe microanalysis, electron diffraction and three-dimensional computer reconstruction, have been used to examine the spatial relationship between the inorganic crystals of calcium phosphate and the collagen fibrils of pickerel and herring bone. High-voltage stereo electron-micrographs were obtained of cross-sections of the cylinder-shaped intramuscular bones in uncalcified regions, in regions where only one or only several crystals had been deposited in some of the fibrils, and in successive sections containing progressively more mineral crystals until the stage of full mineralization was reached. High-resolution electron probe microanalysis confirmed that the electron-dense particles contained calcium and phosphorus. In the earliest stages of mineralization and progressing throughout the mineralization process, the crystals are located only within the collagen fibrils; crystals are not observed free in the extracellular spaces between collagen fibrils. The progressive increase in the mass of mineral deposited in the bone tissue with time occurs, essentially, completely within the collagen fibrils including the stage of full mineralization. At this stage, cross-sectional profiles of collagen fibrils are completely obliterated by mineral. A small number of crystals that are located on or close to the surface of the fibrils appear to extend a very short distance into the spaces between the fibrils. These ultrastructural observations of the very onset of calcification in which nucleation of the calcium phosphate crystals is clearly shown to begin within specific volumes of collagen fibrils, and of the subsequent temporal and spatial sequences of this phenomenon, which shows that calcification continues wholly within the collagen fibrils until maximum calcification is achieved, add important information on the basic physical chemical mechanism of the calcification and the structural elements that are involved. The spatial and temporal independence of the sites where mineralization is initiated establishes that such ultrastructural locations within individual collagen fibrils represent independent, physical chemical nucleation loci. The findings are totally inconsistent with the proposal that crystals must first be deposited in matrix vesicles, or other components such as mitochondria, and subsequently released and propagated in the interfibrillar space, until they eventually reach and impregnate the hole zone regions of the collagen fibrils. Three-dimensional computer reconstruction of serial transverse and longitudinal sections demonstrates periodic swellings along the collagen fibrils, corresponding to the hole zone region of their axial period as mineralization proceeds.(ABSTRACT TRUNCATED AT 400 WORDS)     

Collagen-induced platelet aggregation and release. II critical size and structural requirements of collagen

To investigate the mechanisms governing collagen interaction with blood platelets, the effects of side-chain modifications on collagen-induced platelet aggregation and release of serotonin were studied. Since many chemical modifications alter the ability of collagen to form fibers that, according to current theory, may complicate interpretation of data, we eliminated this possibility by using collagen stabilized in a native-type fibrillar structure by treatment with either glutaraldehyde or ultraviolet irradiation. Acetylation, methylation, succinylation, treatment with 2,4-dinitrofluorobenzene, 2,4,6-trinitrobenzene sulfonic acid or 1,2-cyclohexanedione, and deguanidination with hypobromite were used to modify collagen side-chain reactive groups: amino, carboxyl, hydroxyl and guanidino. Both unmodified monomeric dispersed and fibrillar collagen preparations initiated platelet aggregation and release, although the kinetics and magnitude of the response were different. Monomeric collagen which had been modified by deguanidination, methylation or succinylation, failed to polymerize in physiological conditions and did not induce platelet aggregation and release. However, none of the chemical modifications of stabilized native-type collagen fibers, except treatment with hypobromite or cyclohexanedione, had an effect on collagen-induced platelet aggregation and release. Both hypobromite and cyclohexanedione modified guanidino groups of arginyl residues. Results showed that the ability of a collagen sample to induce platelet aggregation and release of serotonin is dependent on the arginine content of fibrillar collagen.These data demonstrate that manipulation of amino, carboxyl and hydroxyl groups is unimportant as long as the native-type fibrillar structure is maintained, and that arginyl residues are directly involved in collagen-platelet interaction. Moreover, the data suggest that only the arginyl residues in the Y position of the tripeptide unit Gly-X-Y of collagen are responsible.     

Collagen-induced platelet aggregation and release. I effects of side-chain modifications and role of arginyl residues

To investigate the mechanisms governing collagen interaction with blood platelets, the effects of side-chain modifications on collagen-induced platelet aggregation and release of serotonin were studied. Since many chemical modifications alter the ability of collagen to form fibers that, according to current theory, may complicate interpretation of data, we eliminated this possibility by using collagen stabilized in a native-type fibrillar structure by treatment with either glutaraldehyde or ultraviolet irradiation. Acetylation, methylation, succinylation, treatment with 2,4-dinitrofluorobenzene, 2,4,6-trinitrobenzene sulfonic acid or 1,2-cyclohexanedione, and deguanidination with hypobromite were used to modify collagen side-chain reactive groups: amino, carboxyl, hydroxyl and guanidino. Both unmodified monomeric dispersed and fibrillar collagen preparations initiated platelet aggregation and release, although the kinetics and magnitude of the response were different. Monomeric collagen which had been modified by deguanidination, methylation or succinylation, failed to polymerize in physiological conditions and did not induce platelet aggregation and release. However, none of the chemical modifications of stabilized native-type collagen fibers, except treatment with hypobromite or cyclohexanedione, had an effect on collagen-induced platelet aggregation and release. Both hypobromite and cyclohexanedione modified guanidino groups of arginyl residues. Results showed that the ability of a collagen sample to induce platelet aggregation and release of serotonin is dependent on the arginine content of fibrillar collagen.These data demonstrate that manipulation of amino, carboxyl and hydroxyl groups is unimportant as long as the native-type fibrillar structure is maintained, and that arginyl residues are directly involved in collagen-platelet interaction. Moreover, the data suggest that only the arginyl residues in the Y position of the tripeptide unit Gly-X-Y of collagen are responsible.     

Reversed phase chromatographic analysis of Nostoc and Euglena isoleucyl-tRNAs aminoacylated in vitro in homologous and heterologous systems.

To investigate the mechanisms governing collagen interaction with blood platelets, the effects of side-chain modifications on collagen-induced platelet aggregation and release of serotonin were studied. Since many chemical modifications alter the ability of collagen to form fibers that, according to current theory, may complicate interpretation of data, we eliminated this possibility by using collagen stabilized in a native-type fibrillar structure by treatment with either glutaraldehyde or ultraviolet irradiation. Acetylation, methylation, succinylation, treatment with 2,4-dinitrofluorobenzene, 2,4,6-trinitrobenzene sulfonic acid or 1,2-cyclohexanedione, and deguanidination with hypobromite were used to modify collagen side-chain reactive groups: amino, carboxyl, hydroxyl and guanidino. Both unmodified monomeric dispersed and fibrillar collagen preparations initiated platelet aggregation and release, although the kinetics and magnitude of the response were different. Monomeric collagen which had been modified by deguanidination, methylation or succinylation, failed to polymerize in physiological conditions and did not induce platelet aggregation and release. However, none of the chemical modifications of stabilized native-type collagen fibers, except treatment with hypobromite or cyclohexanedione, had an effect on collagen-induced platelet aggregation and release. Both hypobromite and cyclohexanedione modified guanidino groups of arginyl residues. Results showed that the ability of a collagen sample to induce platelet aggregation and release of serotonin is dependent on the arginine content of fibrillar collagen. These data demonstrate that manipulation of amino, carboxyl and hydroxyl groups is unimportant as long as the native-type fibrillar structure is maintained, and that arginyl residues are directly involved in collagen-platelet interaction. Moreover, the data suggest that only the arginyl residues in the Y position of the tripeptide unit Gly-X-Y of collagen are responsible.     

Three-dimensional collagen regulates collagen gene expression by a mechanism that requires serine/threonine kinases and is independent of mechanical contraction

Integrin alpha1beta1, one of the cellular collagen receptors, can participate in the regulation of collagen accumulation by acting as a negative feedback regulator. The molecular mechanism behind this phenomenon has been unknown. We have plated cells inside three-dimensional collagen and analyzed a set of chemical inhibitors for various signal transduction pathways. Only two wide-spectrum serine/threonine kinase inhibitors, H-7 and iso-H-7 could prevent the down-regulation of alpha1(I) collagen mRNA levels in cells exposed to three-dimensional collagen. In monolayer iso-H-7 slightly down-regulated collagen gene expression, indicating that inside collagen it affected integrin signaling rather than having a direct stimulatory effect on collagen mRNA levels. The effect of iso-H-7 was not dependent on its ability to inhibit protein kinases A, C, or G. H-7 and iso-H-7 could also inhibit collagen gel contraction, but this mechanism was independent of collagen gene regulation. Three-dimensional collagen could also up-regulate the mRNA levels of several matrix metalloproteinases (MMPs) but H-7 and iso-H-7 had no effect on the regulation of MMP genes. Our data indicate that three-dimensional collagenous matrix regulates distinct cellular signaling pathways and that collagen gene regulation is independent of the other effects of the matrix.     

Using sequence data to predict the self-assembly of supramolecular collagen structures

Collagen fibrils are the major constituents of the extracellular matrix, which provides structural support to vertebrate connective tissues. It is widely assumed that the superstructure of collagen fibrils is encoded in the primary sequences of the molecular building blocks. However, the interplay between large-scale architecture and small-scale molecular interactions makes the ab initio prediction of collagen structure challenging. Here, we propose a model that allows us to predict the periodic structure of collagen fibers and the axial offset between the molecules, purely on the basis of simple predictive rules for the interaction between amino acid residues. With our model, we identify the sequence-dependent collagen fiber geometries with the lowest free energy and validate the predicted geometries against the available experimental data. We propose a procedure for searching for optimal staggering distances. Finally, we build a classification algorithm and use it to scan 11 data sets of vertebrate fibrillar collagens, and predict the periodicity of the resulting assemblies. We analyzed the experimentally observed variance of the optimal stagger distances across species, and find that these distances, and the resulting fibrillar phenotypes, are evolutionary well preserved. Moreover, we observed that the energy minimum at the optimal stagger distance is broad in all cases, suggesting a further evolutionary adaptation designed to improve the assembly kinetics. Our periodicity predictions are not only in good agreement with the experimental data on collagen molecular staggering for all collagen types analyzed, but also for synthetic peptides. We argue that, with our model, it becomes possible to design tailor-made, periodic collagen structures, thereby enabling the design of novel biomimetic materials based on collagen-mimetic trimers.     

Polarized light microscopy in the study of the molecular structure of collagen and reticulin

Although collagen structure has been studied by polarized light microscopy since the early 19th century and continued since, modern studies and reviews failed to correlate the conclusions based on data obtained by the techniques with those of polarized light microscopy. Collagen I is intensely positively birefringent in respect to length of the fibres; the positive intrinsic birefringence indicates a quasi-crystalline alignment parallel to the fibre and molecule axis of the amino acid residues of the polypeptide chains. This would not have been compatible with a helical structure but has been achieved by similar tilt angles and opposite directions of the coiling and supercoiling. Birefringence characteristics of collagen are also affected by chemical treatments, extractions and staining procedures. Attachment of chemical groups to the anionic charges present on the surface of collagen molecules results in increased positive birefringence in the case of bipolar molecules attached to two or more anionic residues. Unipolar attachment to the same groups, or to the cationic groups of the associated proteoglycans, as well as sulfation or acetylation of hydroxyls of the protein and/or the carbohydrate, reduced or reversed the sign of birefringence. Increased birefringence caused by stretching cannot be due to intramolecular events and is caused by intermolecular changes. The same applies to changes in collagen during aging. Reticulin is a group of different substances which mostly contain collagen III. The pliability and deformability of this collagen is related to its weakly negative birefringence due to large side chains and presence of different and greater amounts of interstitial proteoglycans and other molecules. The so-called reticulin of healing wounds differs in its constitution from other reticulins but is also rich in intermolecular carbohydrate components.     

Cooperativity between Sertoli cells and testicular peritubular cells in the production and deposition of extracellular matrix components

We examined the synthesis and deposition of extracellular matrix (ECM) components in cultures of Sertoli cells and testicular peritubular cells maintained alone or in contact with each other. Levels of soluble ECM components produced by populations of isolated Sertoli cells and testicular peritubular cells were determined quantitatively by competitive enzyme-linked immunoabsorbent assays, using antibodies shown to react specifically with Type I collagen, Type IV collagen, laminin, or fibronectin. Peritubular cells in monoculture released into the medium fibronectin (432 to 560 ng/microgram cell DNA per 48 h), Type I collagen (223 to 276 ng/microgram cell DNA per 48 h), and Type IV collagen (350 to 436 ng/microgram cell DNA per 48 h) during the initial six days of culture in serum-free medium. In contrast, Sertoli cells in monoculture released into the medium Type IV collagen (322 to 419 ng/microgram cell DNA per 48 h) but did not form detectable amounts of Type I collagen or fibronectin during the initial six days of culture. Neither cell type produced detectable quantities of soluble laminin. Immunocytochemical localization investigations demonstrated that peritubular cells in monoculture were positive for fibronectin, Type I collagen, and Type IV collagen but negative for laminin. In all monocultures most of the ECM components were intracellular, with scant deposition as extracellular fibrils. Sertoli cells were positive immunocytochemically for Type IV collagen and laminin but negative for fibronectin and Type I collagen. Co-cultures of peritubular cells and Sertoli cells resulted in interactions that quantitatively altered levels of soluble ECM components present in the medium. This was correlated with an increased deposition of ECM components in extracellular fibrils. The data correlated with an increased deposition of ECM components in extracellular fibrils. The data presented here we interpret to indicate that the two cell types in co-culture act cooperatively in the formation and deposition of ECM components. Results are discussed with respect to the nature of interactions between mesenchymal peritubular cell precursors and adjacent epithelial Sertoli cell precursors in the formation of the basal lamina of the seminiferous tubule.     

A peptide study of the relationship between the collagen triple-helix and amyloid

Type XXV collagen, or collagen‐like amyloidogenic component, is a component of amyloid plaques, and recent studies suggest this collagen affects amyloid fibril elongation and has a genetic association with Alzheimer's disease. The relationship between the collagen triple helix and amyloid fibrils was investigated by studying peptide models, including a very stable triple helical peptide (Pro‐Hyp‐Gly)₁₀, an amyloidogenic peptide GNNQQNY, and a hybrid peptide where the GNNQQNY sequence was incorporated between (GPO)_n domains. Circular dichroism and nuclear magnetic resonance (NMR) spectroscopy showed the GNNQQNY peptide formed a random coil structure, whereas the hybrid peptide contained a central disordered GNNQQNY region transitioning to triple‐helical ends. Light scattering confirmed the GNNQQNY peptide had a high propensity to form amyloid fibrils, whereas amyloidogenesis was delayed in the hybrid peptide. NMR data suggested the triple‐helix constraints on the GNNQQNY sequence within the hybrid peptide may disfavor the conformational change necessary for aggregation. Independent addition of a triple‐helical peptide to the GNNQQNY peptide under aggregating conditions delayed nucleation and amyloid fibril growth. The inhibition of amyloid nucleation depended on the Gly‐Xaa‐Yaa sequence and required the triple‐helix conformation. The inhibitory effect of the collagen triple‐helix on an amyloidogenic sequence, when in the same molecule or when added separately, suggests Type XXV collagen, and possibly other collagens, may play a role in regulating amyloid fibril formation. © 2012 Wiley Periodicals, Inc. Biopolymers 97: 795–806, 2012.     

Sharing digital micrographs and other data files between computers

It ought to be easy to exchange digital micrographs and other computer data files with a colleague even on another continent. In practice, this often is not the case. The advantages and disadvantages of various methods that are available for exchanging data files between computers are discussed. When possible, data should be transferred through computer networking. When data are to be exchanged locally between computers with similar operating systems, the use of a local area network is recommended. For computers in commercial or academic environments that have dissimilar operating systems or are more widely spaced, the use of FTPs is recommended. Failing this, posting the data on a website and transferring by hypertext transfer protocol is suggested. If peer to peer exchange between computers in domestic environments is needed, the use of Messenger services such as Microsoft Messenger or Yahoo Messenger is the method of choice. When it is not possible to transfer the data files over the internet, single use, writable CD ROMs are the best media for transferring data. If for some reason this is not possible, DVD-R/RW, DVD+R/RW, 100 MB ZIP disks and USB flash media are potentially useful media for exchanging data files.     

Sharing digital micrographs and other data files between computers

It ought to be easy to exchange digital micrographs and other computer data files with a colleague even on another continent. In practice, this often is not the case. The advantages and disadvantages of various methods that are available for exchanging data files between computers are discussed. When possible, data should be transferred through computer networking. When data are to be exchanged locally between computers with similar operating systems, the use of a local area network is recommended. For computers in commercial or academic environments that have dissimilar operating systems or are more widely spaced, the use of FTPs is recommended. Failing this, posting the data on a website and transferring by hypertext transfer protocol is suggested. If peer to peer exchange between computers in domestic environments is needed, the use of Messenger services such as Microsoft Messenger or Yahoo Messenger is the method of choice. When it is not possible to transfer the data files over the internet, single use, writable CD ROMs are the best media for transferring data. If for some reason this is not possible, DVD-R/RW, DVD+R/RW, 100 MB ZIP disks and USB flash media are potentially useful media for exchanging data files.     

Testing quantitative traits for association and linkage in the presence or absence of parental data

Zhu and Elston developed a transmission disequilibrium test for quantitative traits by defining a linear transformation to condition out founder information. The method tests the null hypothesis of no linkage or association and can be applied to general pedigree structures. However, this method requires both genotype and phenotype parental information, which may be difficult to obtain. In this paper, we describe parametric and non-parametric methods to relax this requirement when only nuclear families are sampled. We show that neither method is affected by population stratification in the absence of linkage. The statistical power and validity of the tests are investigated by simulation. A simple simulation method to calculate the power of the nonparametric method is also discussed. In practice, the data may have some families with parental phenotype and genotype information available and some without. We briefly discuss how all the data may be analyzed jointly.     

Structural Insights into the Interactions between Platelet Receptors and Fibrillar Collagen

Collagen peptides have been used to identify binding sites for several important collagen receptors, including integrin α₂β₁, glycoprotein VI, and von Willebrand factor. In parallel, the structures of these collagen receptors have been reported, and their interactions with collagen peptides have been studied. Recently, the three-dimensional structure of the intact type I collagen fiber from rat tail tendon has been resolved by fiber diffraction. It is now possible to map the binding sites of platelet collagen receptors onto the intact collagen fiber in three dimensions. This minireview will discuss these recent findings and their implications for platelet activation by collagen.     

Ultrastructural localization of proteoglycans in tissue using Cuprolinic Blue according to the critical electrolyte concentration method: Comparison with biochemical data from the literature

Summary Several connective tissues were stained for proteoglycans using the cationic dye Cuprolinic Blue according to the critical electrolyte concentration method. With this method, proteoglycans are visualized as electron-dense filaments. In most tissues, two types of proteoglycan filaments are present: a small (maximum length 60 nm), thin, collagen fibril-associated filament, and a thick, heavily-staining filament which is predominantly localized between bundles of collagen fibrils. Cartilage contains very large (about 300 nm) proteoglycan filaments while in cornea they are very small. Comparison with biochemical data from the literature suggests that the appearance of the proteoglycan filaments may be indicative for the glycosaminoglycan—protein ratio and for the molecular weight of the part of the protein core to which glycosaminoglycans are attached. The data thus obtained on the localization and structure of a proteoglycan may be useful when planning a strategy for its isolation.