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1.
Extracellular matrix components that flank the fissura prima, a primary surface infolding of the cerebellum in birds and mammals, were examined in the embryonic chick using light and transmission electron microscopy. Cerebella dissected from Day 10 embryos were perfused with a paraformaldehyde-glutaraldehyde-tannic acid primary fixative and sectioned in the sagittal plane through the mid-vermis. Ultrastructural analysis revealed a distinct, continuous basal lamina separating the organ parenchyma (epithelia) from pia mater (mesenchyme) at the fissure surface (arbitrarily labeled; fissure floor, folia wall, and folia apex). The basal lamina was significantly thicker (P < 0.001) at the fissure floor compared to that found at the folia wall, which was significantly thicker (P < 0.001) than that observed at the folia apex. Folds in the basal lamina were observed exclusively at the fissure floor. Surface-associated collagen fibrils were distributed in an aligned, relatively dense manner at the fissure floor, compared with fibrils observed in various orientations and widely separated or absent at the folia wall and folia apex. Metachromasia was more pronounced in the fissure floor than in either the folia wall or folia apex in methylene blue-stained tissue sections. Together, the thicker, folded basal lamina and densely aligned collagen fibrils at the fissure floor provide a chemical rationale for this color change. These findings suggest that the differential accumulation of extracellular matrix at the fissura prima is positioned to play a structural and/or biochemical role in the maintenance of this fold. 相似文献
2.
Kadler K 《Birth defects research. Part C, Embryo today : reviews》2004,72(1):1-11
Nothing in biology stimulates the imagination like the development of a single fertilized egg into a newborn child. Consequently, a major focus of biomedical research is aimed at understanding cell differentiation, proliferation, and specialization during child health and human development. However, the fact that the increase in size and shape of the growing embryo has as much to do with the extracellular matrix (ECM) as with the cells themselves, is largely overlooked. Cells in developing tissues are surrounded by a fiber-composite ECM that transmits mechanical stimuli, maintains the shape of developing tissues, and functions as a scaffold for cell migration and attachment. The major structural element of the ECM is the collagen fibril. The fibrils, which are indeterminate in length, are arranged in different tissues in exquisite supramolecular architectures, including parallel bundles, orthogonal lamellae, and concentric weaves. This article reviews our current understanding of the synthesis and assembly of collagen fibrils, and discusses challenging questions about how cells assemble an organized ECM during embryogenesis. 相似文献
3.
Judith Aggeler 《In vitro cellular & developmental biology. Plant》1988,24(7):633-638
Summary Specific interactions between cells and the extracellular matrix (ECM) in which they are embedded play a vital role in tissue organization. In recent years, many of the individual components of the extracellular matrix have been isolated and their molecular structures elucidated, but the detailed topography of most extracellular matrices, as they are deposited by cells, is still largely unknown. In this study, the insoluble extracellular matrix produced by cultured rat vascular smooth muscle cells has been characterized morphologically using high-resolution electron microscopy of rotary platinum replicas. These cells grew as flat sheets in culture, secreting their matrix laterally and basally. The matrix was composed of a cross-linked fibrillar meshwork. Some fine fibers (10 to 15 nm in diameter) were naked, but most of the filamentous mesh was covered with coarse granular material. Limited digestion with trypsin or pancreatic elastase removed most of this coating, indicating that the granules were glycoproteins and proteoglycans. Another subset of matrix fibrils (20 to 40 nm in diameter) was identified as type I collagen by direct comparison with purified bovine skin collagen. In addition to exposing the underlying filamentous substructure of the matrix, protease treatment also revealed large, straight fiber bundles and globules of amorphous material suspended in the filamentous web. This novel view of a complex matrix promises to provide spatial information that will be useful in future studies of cell interactions with the ECM. These studies were supported in part by NIH Biomedical Research Support grant S07-RR-05684. 相似文献
4.
Attachment and extracellular matrix differences between tendon and synovial fibroblastic cells 总被引:3,自引:0,他引:3
M. A. Riederer-Henderson A. Gauger L. Olson C. Robertson T. K. Greenlee Jr. 《In vitro cellular & developmental biology. Plant》1983,19(2):127-133
Summary Fibroblasts of the synovium of sheathed tendons were isolated, and their biochemical properties were compared with those of
the fibroblasts of the remaining tendon. The synovial cells had a lower attachment efficiency than did the tendon cells. On
the day of cell isolation the synovial cells synthesized collagen as 10% of their total protein, whereas the tendon cells
synthesized 30% collagen. After growth in fetal bovine serum (FBS), the percentage of collagen synthesized by both populations
decreased; however, the synovial cells still made less collagen than did the tendon cells (5 versus 11%). On the basis of
cyanogen bromide peptide analysis, the synovial cells were found to synthesize Types I and III collagen in primary culture,
whereas the tendon cells synthesized only Type I. The synovial cells aslo synthesized two to three times less sulfated glycosaminoglycans
in culture than did the tendon cells. Thus, the two cell, populations differed in attachment efficiency and in their biosynthesis
of collagen and sulfated glycosaminoglycans. These differences reflect extracellular matrix differences that have been observed
in the tendon in vivo. In addition, the results augment existing data showing that not all fibroblasts have identical phenotypes.
This investigation was supported by National Institutes of Health Grant AM 25749. 相似文献
5.
Uchena N. G. Wudebwe Alistair Bannerman Pola Goldberg-Oppenheimer Jennifer Z. Paxton Richard L. Williams Liam M. Grover 《Philosophical transactions of the Royal Society of London. Series B, Biological sciences》2015,370(1661)
Progress in tissue engineering is now impacting beyond the field of regenerative medicine. Engineered tissues are now used as tools to evaluate the toxicity of compounds or even to enable the modelling of disease. While many of the materials that are used to facilitate tissue growth are designed to enable cell attachment, many researchers consider that the contraction and modification of these matrices by attached cells is not desirable and take measures to prevent this from occurring. Where substantial alignment of the molecules within tissues, however, is a feature of structure the process of contraction can be exploited to guide new matrix deposition. In this paper, we will demonstrate how we have used the cell contraction process to generate tissues with high levels of organization. The tissues that have been grown in the laboratory have been characterized using a suite of analytical techniques to demonstrate significant levels of matrix organization and mechanical behaviour analogous to natural tissues. This paper provides an overview of research that has been undertaken to determine how tissues have been grown in vitro with structuring from the molecular, right through to the macroscopic level. 相似文献
6.
Zhang G Ezura Y Chervoneva I Robinson PS Beason DP Carine ET Soslowsky LJ Iozzo RV Birk DE 《Journal of cellular biochemistry》2006,98(6):1436-1449
Tendon function involves the development of an organized hierarchy of collagen fibrils. Small leucine-rich proteoglycans have been implicated in the regulation of fibrillogenesis and decorin is the prototypic member of this family. Decorin-deficient mice demonstrate altered fibril structure and mechanical function in mature skin and tail tendons. However, the developmental role(s) of decorin needs to be elucidated. To define these role(s) during tendon development, tendons (flexor digitorum longus) were analyzed ultrastructurally from postnatal day 10 to 90. Decorin-deficient tendons developed abnormal, irregularly contoured fibrils. Finite mixture modeling estimated that the mature tendon was a three-subpopulation mixture of fibrils with characteristic diameter ranges. During development, in each subpopulation the mean diameter was consistently larger in mutant mice. Also, diameter distributions and the percentage of fibrils in each subpopulation were altered. Biomechanical analyses demonstrated that mature decorin-deficient tendons had significantly reduced strength and stiffness; however, there was no reduction in immature tendons. Expression of decorin and biglycan, a closely related family member, was analyzed during development. Decorin increased with development while biglycan decreased. Spatially, both had a comparable localization throughout the tendon. Biglycan expression increased substantially in decorin-deficient tendons suggesting a potential functional compensation. The accumulation of structural defects during fibril growth, a period associated with decorin expression and low biglycan expression, may be the cause of compromised mechanical function in the absence of decorin. Our findings indicate that decorin is a key regulatory molecule and that the temporal switch from biglycan to decorin is an important event in the coordinate regulation of fibrillogenesis and tendon development. 相似文献
7.
Collagen fibrils in the corneal stroma have been recognised to have a high degree of uniformity of diameter and spatial arrangement compared with those in other mature connective tissues. The precision of this lateral size control has been determined in this study by mass per unit length measurements on fibrils isolated from adult bovine corneal stroma. At the molecular level, however, there are substantial variations in lateral size, both between fibrils and along individual fibrils. The mean mass per unit length was measured to be 304 kDa nm(-1), equivalent to 347 collagen molecules in transverse section and had a standard deviation of 8.3%. The variation of lateral size along individual fibrils was measured as a mass slope over approximately 7 microm lengths (100 D-periods) and had a mean mass slope equivalent to 0.56 molecules per D-period. Smoothly tapered tips of length approximately 7 microm were also observed with a mass slope of about approximately three molecules per D-period. The frequency of these tips was used to estimate a mean fibril length of approximately 940 microm in the sample tissue. Observations of molecular polarity within the fibril shafts and tips were used to consider possible models of fibril assembly. 相似文献
8.
Type I collagen is a fibril‐forming protein largely responsible for the mechanical stability of body tissues. The tissue level properties of collagen have been studied for decades, and an increasing number of studies have been performed at the fibril scale. However, the mechanical properties of collagen at the molecular scale are not well established. In the study presented herein, the persistence length of pepsin digested bovine type I collagen is extracted from the conformations assumed when deposited from solution onto two‐dimensional surfaces. This persistence length is a measure of the flexibility of the molecule. Comparison of the results for molecules deposited from different solvents allows for the study of the effect of the solutions on the flexibility of the molecule and provides insight into the molecule's behavior in situ. © 2013 Wiley Periodicals, Inc. Biopolymers 101: 329–335, 2014. 相似文献
9.
T Aigner L Hambach S S?der U Schl?tzer-Schrehardt E P?schl 《Biochemical and biophysical research communications》2002,290(2):743-748
In articular cartilage, type VI collagen is concentrated in the pericellular matrix compartment. During protein synthesis and processing at least the alpha3(VI) chain undergoes significant posttranslational modification and cleavage. In this study, we investigated the processing of type VI collagen in articular cartilage. Immunostaining with a specific polyclonal antiserum against the C5 domain of alpha3(VI) showed strong cellular staining seen in nearly all chondrocytes of articular cartilage. Confocal laser-scanning microscopy and immunoelectron microscopy allowed localization of this staining mainly to the cytoplasm and the immediate pericellular matrix. Double-labeling experiments showed a narrow overlap of the C5 domain and the pericellular mature type VI collagen. Our results suggest that at least in human adult articular cartilage the C5 domain of alpha3(VI) collagen is synthesized and initially incorporated into the newly formed type VI collagen fibrils, but immediately after secretion is cut off and is not present in the mature pericellular type VI matrix of articular cartilage. 相似文献
10.
Lai-Man Chan Carroll Hatier Gordon Parry Zena Werb Mina J. Bissell 《In vitro cellular & developmental biology. Plant》1987,23(4):308-314
Summary Using gelatin, casein, and fibronectin as substrates and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), we have identified protein-degrading enzymes in both normal and Rous sarcoma virus-transformed primary avian tendon cells. Although there are some consistent differences in the profile of the gelatinolytic activities (mainly metalloproteinases) between normal and transformed cells, the amounts of fibronectin-degrading activities seem to be comparable. In vitro studies reported here demonstrate that the degradation of fibronectin is partially and specifically inhibited by gelatin and collagen. We therefore propose that the abundant collagen present in normal tendon cells protects fibronectin against degradation. Conversely, in transformed cells, where collagen levels are drastically reduced, fibronectin may be more accessible to degradation. Thus differences in the steady-state levels of fibronectin on normal and transformed cells may be, at least in part, a consequence of changes in collagen levels. This work was supported by the Office of Health and Environmental Research, Office of Energy Research, U.S. Department of Energy, Washington, D.C., under contracts DE-AC03-76-SF00098 and DE-AC03-76-SF01012. 相似文献
11.
E.F. Eikenberry B. Brodsky D.A.D. Parry 《International journal of biological macromolecules》1982,4(6):322-328
The low angle equatorial X-ray diffraction () from hydrated embryonic chick metatarsal tendon contains minima and maxima that are not seen in mature tendons. This diffraction derives from the disordered array of parallel, cylindrical fibrils of collagen of small, uniform diameter that comprise the major part of this tissue. Comparison of the positions of the minima and maxima with those expected from an array of cylinders allows estimation of the mean diameter of the cylinders and the average centre-to-centre nearest neighbour separation. It was found that in the age range from 13 to 19 days fetal, the mean diameter increased from ~ 46 to ~ 58 nm, whereas the mean nearest neighbour separation remained constant at ~ 90 nm. Detailed analysis of the X-ray intensity profile of a 17 day fetal tendon indicated the presence of a paucidisperse distribution of fibril diameters with two or more discrete populations of preferred diameters separated by 10 to 12 nm. 相似文献
12.
The extracellular matrix (ECM) represents a major barrier for delivery of therapeutic drugs, and the transport is determined by the ECM composition, structure, and distribution. Because of the high interstitial fluid pressure in tumors, diffusion becomes the main transport mechanism through ECM. The purpose of this work was to study the impact of the structure of the collagen network on diffusion, by studying to what extent the orientation and chemical modification of the collagen network influenced diffusion. Collagen gels with a concentration of 0.2-2.0% that is comparable with the amount of collagen in the tumor ECM were used as a model system for ECM. Collagen gels were aligned in a low-strength magnetic field and geometrical confinement, and chemically modified by adding decorin or hyaluronan. Diffusion of dextran 2-MDa molecules in the collagen gels was measured using fluorescence recovery after photobleaching. Alignment of the collagen fibers in our gels was found to have no impact on the diffusion coefficient. Adding decorin reduced the diameter of the collagen fibers, but no effect on diffusion was observed. Hyaluronan also reduced the fiber diameter, and high concentration of hyaluronan (2.5 mg/ml) increased the diffusion coefficient. The results indicate that the structure of the collagen network is not a major factor in determining the diffusion through the ECM. Rather, increasing the concentration of collagen was found to reduce the diffusion coefficient. Concentration of the collagen network is more important than the structure in determining the diffusion coefficient. 相似文献
13.
Neil A. Duncan Sabina B. Bruehlmann Christopher J. Hunter Xinxin Shao Elizabeth J. Kelly 《Computer methods in biomechanics and biomedical engineering》2014,17(1):39-47
Designing biomaterials to mimic and function within the complex mechanobiological conditions of connective tissues requires a detailed understanding of the micromechanical environment of the cell. The objective of our study was to measure the in situ cell–matrix strains from applied tension in both tendon fascicles and cell-seeded type I collagen scaffolds using laser scanning confocal microscopy techniques. Tendon fascicles and collagen gels were fluorescently labelled to simultaneously visualise the extracellular matrix and cell nuclei under applied tensile strains of 5%. There were significant differences observed in the micromechanics at the cell–matrix scale suggesting that the type I collagen scaffold did not replicate the pattern of native tendon strains. In particular, although the overall in situ tensile strains in the matrix were quite similar (~2.5%) between the tendon fascicles and the collagen scaffolds, there were significant differences at the cell–matrix boundary with visible shear across cell nuclei of >1 μm measured in native tendon which was not observed at all in the collagen scaffolds. Similarly, there was significant non-uniformity of intercellular strains with relative sliding observed between cell rows in tendon which again was not observed in the collagen scaffolds where the strain environment was much more uniform. If the native micromechanical environment is not replicated in biomaterial scaffolds, then the cells may receive incorrect or mixed mechanical signals which could affect their biosynthetic response to mechanical load in tissue engineering applications. This study highlights the importance of considering the microscale mechanics in the design of biomaterial scaffolds and the need to incorporate such features in computational models of connective tissues. 相似文献
14.
Collagens present in the connective tissues of the extracellular matrix of fibrosarcoma were isolated and characterized. The fibrosarcoma was induced in rats by the administration of 3-methylcholanthrene. The results obtained were compared with normal muscle. An excess amount of type V collagen was found to be produced by the fibrosarcoma tissue compared to the normal muscle. Type V collagen from fibrosarcoma was characterized on the basis of solubility behavior in sodium chloride solutions, electrophoretic mobility on SDS-polyacrylamide gels, elution pattern of phosphocellulose chromatography and amino acid composition. 相似文献
15.
Ryan S. Lim Adelheid Kratzer Nicholas P. Barry Shinobu Miyazaki-Anzai Makoto Miyazaki William W. Mantulin Moshe Levi Eric O. Potma Bruce J. Tromberg 《Journal of lipid research》2010,51(7):1729-1737
We characterized several cellular and structural features of early stage Type II/III atherosclerotic plaques in an established model of atherosclerosis—the ApoE-deficient mouse—by using a multimodal, coregistered imaging system that integrates three nonlinear optical microscopy (NLOM) contrast mechanisms: coherent anti-Stokes Raman scattering (CARS), second harmonic generation (SHG), and two-photon excitation fluorescence (TPEF). Specifically, the infiltration of lipid-rich macrophages and the structural organization of collagen and elastin fibers were visualized by CARS, SHG, and TPEF, respectively, in thick tissue specimens without the use of exogenous labels or dyes. Label-free CARS imaging of macrophage accumulation was confirmed by histopathology using CD68 staining. A high-fat, high-cholesterol Western diet resulted in an approximate 2-fold increase in intimal plaque area, defined by CARS signals of lipid-rich macrophages. Additionally, analysis of collagen distribution within lipid-rich plaque regions revealed nearly a 4-fold decrease in the Western diet–fed mice, suggesting NLOM sensitivity to increased matrix metalloproteinase (MMP) activity and decreased smooth muscle cell (SMC) accumulation. These imaging results provide significant insight into the structure and composition of early stage Type II/III plaque during formation and allow for quantitative measurements of the impact of diet and other factors on critical plaque and arterial wall features. 相似文献
16.
S Florim da Silva M Taffarel S Allodi 《Biology of the cell / under the auspices of the European Cell Biology Organization》2001,93(5):293-299
Glial cells in higher invertebrate groups are usually recognized on the basis of their location and general morphological or functional criteria. In this study of the crustacean visual system, we have approached the analysis of the relations between glial cells and the extracellular matrix by classical histochemical methods for carbohydrates at the light and electron microscopic levels, carbonic anhydrase histochemistry and by the biochemical characterization of sulphated polysaccharides. Periodic acid-Schiff-positive glial cells and processes were observed in the retina, basement membrane below the retina and in the optic ganglia. Carbonic anhydrase was not detected in the retina but it was demonstrated in all optic ganglia. The biochemical analysis of the extracellular matrix confirmed the alcian blue reaction and showed that sulphated polysaccharides are not abundant in the optic neuropils. This article describes into more details the crustacean visual system glial cells classification, and the relation between them and the extracellular matrix. In addition, they show that glial cells are the main components of the retinal basement membrane. 相似文献
17.
Patrizia Sini Antonella Denti M. Enrica Tira Cesare Balduini 《Glycoconjugate journal》1997,14(7):871-874
Tendon and corneal decorins are differently iduronated dermatan sulphate/proteoglycan (DS/PG) and the biochemical parameter
that differentiates type I collagens is the hydroxylysine glycoside content. We have examined the effect of tendon and corneal
decorins on the individual phases (tlag, dA/dt) of differently glycosylated type I collagens fibril formation, at molar ratios
PG:collagen monomer ranging from 0.15 : 1 to 0.45 : 1. The results obtained indicate that decorins exert a different effect
on the individual phases of fibril formation, correlated to the degree of glycosylation of collagen: at the same PG:collagen
ratio the fibril formation of highly glycosylated corneal collagen is more efficiently inhibited than that of the poorly glycosylated
one (tendon). Moreover tendon and corneal decorins exert a higher control on the fibrillogenesis of homologous collagen with
respect to the heterologous one. These data suggest a possible tissue-specificity of the interaction decorin/type I collagen
correlated to the structure of the PG and collagen present in extracellular matrices.
This revised version was published online in November 2006 with corrections to the Cover Date. 相似文献
18.
Tobias Maa? Christopher P. Bayley Matthias M?rgelin Sandra Lettmann Paolo Bonaldo Mats Paulsson Clair Baldock Raimund Wagener 《The Journal of biological chemistry》2016,291(10):5247-5258
Collagen VI, a collagen with uncharacteristically large N- and C-terminal non-collagenous regions, forms a distinct microfibrillar network in most connective tissues. It was long considered to consist of three genetically distinct α chains (α1, α2, and α3). Intracellularly, heterotrimeric molecules associate to form dimers and tetramers, which are then secreted and assembled to microfibrils. The identification of three novel long collagen VI α chains, α4, α5, and α6, led to the question if and how these may substitute for the long α3 chain in collagen VI assembly. Here, we studied structural features of the novel long chains and analyzed the assembly of these into tetramers and microfibrils. N- and C-terminal globular regions of collagen VI were recombinantly expressed and studied by small angle x-ray scattering (SAXS). Ab initio models of the N-terminal globular regions of the α4, α5, and α6 chains showed a C-shaped structure similar to that found for the α3 chain. Single particle EM nanostructure of the N-terminal globular region of the α4 chain confirmed the C-shaped structure revealed by SAXS. Immuno-EM of collagen VI extracted from tissue revealed that like the α3 chain the novel long chains assemble to homotetramers that are incorporated into mixed microfibrils. Moreover, SAXS models of the C-terminal globular regions of the α1, α2, α4, and α6 chains were generated. Interestingly, the α1, α2, and α4 C-terminal globular regions dimerize. These self-interactions may play a role in tetramer formation. 相似文献
19.
Majsterek I McAdams E Adachi E Dhume ST Fertala A 《Protein science : a publication of the Protein Society》2003,12(9):2063-2072
Recombinant collagens are attractive proteins for a number of biomedical applications. To date, significant progress was made in the large-scale production of nonmodified recombinant collagens; however, engineering of novel collagen-like proteins according to customized specifications has not been addressed. Herein we investigated the possibility of rational engineering of collagen-like proteins with specifically assigned characteristics. We have genetically engineered two DNA constructs encoding multi-D4 collagens defined as collagen-like proteins, consisting primarily of a tandem of the collagen II D4 periods that correspond to the biologically active region. We have also attempted to decrease enzymatic degradation of novel collagen by mutating a matrix metalloproteinase 1 cleavage site present in the D4 period. We demonstrated that the recombinant collagen alpha-chains consisting predominantly of the D4 period but lacking most of the other D periods found in native collagen fold into a typical collagen triple helix, and the novel procollagens are correctly processed by procollagen N-proteinase and procollagen C-proteinase. The nonmutated multi-D4 collagen had a normal melting point of 41 degrees C and a similar carbohydrate content as that of control. In contrast, the mutant multi-D4 collagen had a markedly lower thermostability of 36 degrees C and a significantly higher carbohydrate content. Both collagens were cleaved at multiple sites by matrix metalloproteinase 1, but the rate of hydrolysis of the mutant multi-D4 collagen was lower. These results provide a basis for the rational engineering of collagenous proteins and identifying any undesirable consequences of altering the collagenous amino acid sequences. 相似文献
20.
According to previous studies, the nonlinear susceptibility tensor ratio χ33/χ31 obtained from polarization‐resolved second harmonic generation (P‐SHG) under the assumption of cylindrical symmetry can be used to distinguish between fibrillar collagen types. Discriminating between collagen fibrils of types I and II is important in tissue engineering of cartilage. However, cartilage has a random organization of collagen fibrils, and the assumption of cylindrical symmetry may be incorrect. In this study, we simulated the P‐SHG response from different collagen organizations and demonstrated a possible method to exclude areas where cylindrical symmetry is not fulfilled and where fibrils are located in the imaging plane. The χ33/χ31‐ratio for collagen type I in tendon and collagen type II in cartilage was estimated to be 1.33 and 1.36, respectively, using this method. These ratios are now much closer than what has been reported previously in the literature, and the larger reported differences between collagen types can be explained by variation in the structural organization. 相似文献