Histochemical and morphological characterization of reticular fibers

Summary The results presented in this paper show that collagen fibers can be clearly distinguished from reticular fibers using the picrosirius-polarization method. A morphologic and morphometric study of these two types of fibers with electron microscopy shows that reticular fibers are characterized by the smaller diameter of their fibrillar components and the higher content of interfibrillar material, resulting in a loose arrangement of the fibrils. The evidences presented suggest that the amorphous matrix in which fibrils are embedded is responsible for the silver impregnation of reticular fibers. Our results show that the matrix of reticular fibers is characteristically rich in heparitin sulfate, and that the glycosaminoglycans present show a high interaction with the fibrillar component of these fibers.     

Ameloblastic secretion and calcification of the enamel layer in shark teeth

Tooth primordia at early stages of mineralization in the sharks Negaprion brevirostris and Triaenodon obesus were examined electron microscopically for evidence of ameloblastic secretion and its relation to calcification of the enamel (enameloid) layer. Ameloblasts are polarized with most of the mitochondria and all of the Golgi dictyosomes localized in the infranuclear end of the cell toward the squamous outer cells of the enamel organ. Endoplasmic reticular membranes and ribosomes are also abundant in this region. Ameloblastic vesicles bud from the Golgi membranes and evidently move through perinuclear and supranuclear zones to accumulate at the apical end of the cell. The vesicles secrete their contents through the apical cell membrane in merocrine fashion and appear to contribute precursor material both for the basal lamina and the enameline matrix. The enamel layer consists of four zones: a juxta-laminar zone containing newly polymerized mineralizing fibrils (tubules); a pre-enamel zone of assembly of matrix constituents; palisadal zones of mineralizing fibrils (tubules); and interpalisadal zones containing granular amorphous matrix, fine unit fibrils, and giant cross-banded fibers with a periodicity of 17.9 nm. It seems probable that amorphous, non-mineralizing fibrillar and mineralizing fibrillar constituents of the matrix are all products of ameloblastic secretion. Odontoblastic processes are tightly embedded in the matrix of the palisadal zones and do not appear to be secretory at the stages investigated. The shark tooth enamel layer is considered homologous with that of other vertebrates with respect to origin of its mineralizing fibrils from the innerental epithelium. The term enameloid is appropriate to connote the histological distinction that the enamel layer contains odontoblastic processes but should not signify that shark tooth enamel is a modified type of dentine. How amelogenins and/or enamelins secreted by amelo- blasts in the shark and other vertebrates are related to nucleation and growth of enamel crystallites is still not known.     

Fibronectin fibril alignment is established upon initiation of extracellular matrix assembly

The physical structure of the extracellular matrix (ECM) is tissue-specific and fundamental to normal tissue function. Proper alignment of ECM fibers is essential for the functioning of a variety of tissues. While matrix assembly in general has been intensively investigated, little is known about the mechanisms required for formation of aligned ECM fibrils. We investigated the initiation of fibronectin (FN) matrix assembly using fibroblasts that assemble parallel ECM fibrils and found that matrix assembly sites, where FN fibrillogenesis is initiated, were oriented in parallel at the cell poles. We show that these polarized matrix assembly sites progress into fibrillar adhesions and ultimately into aligned FN fibrils. Cells that assemble an unaligned meshwork matrix form matrix assembly sites around the cell periphery, but the distribution of matrix assembly sites in these cells could be modulated through micropatterning or mechanical stretch. While an elongated cell shape corresponds with a polarized matrix assembly site distribution, these two features are not absolutely linked, since we discovered that transforming growth factor beta (TGF-β1) enhances matrix assembly site polarity and assembly of aligned fibrils independent of cell elongation. We conclude that the ultimate orientation of FN fibrils is determined by the alignment and distribution of matrix assembly sites that form during the initial stages of cell–FN interactions.     

Development of tendon structure and function: regulation of collagen fibrillogenesis

In the tendon, the development of mature mechanical properties is dependent on the assembly of a tendon-specific extracellular matrix. This matrix is synthesized by the tendon fibroblasts and composed of collagen fibrils organized as fibers, as well as fibril-associated collagenous and non-collagenous proteins. All of these components are integrated, during development and growth, to form a functional tissue. During tendon development, collagen fibrillogenesis and matrix assembly progress through multiple steps where each step is regulated independently, culminating in a structurally and functionally mature tissue. Collagen fibrillogenesis occurs in a series of extracellular compartments where fibril intermediates are assembled and mature fibrils grow through a process of post-depositional fusion of the intermediates. Linear and lateral fibril growth occurs after the immature fibril intermediates are incorporated into fibers. The processes are regulated by interactions of extracellular macromolecules with the fibrils. Interactions with quantitatively minor fibrillar collagens, fibril-associated collagens and proteoglycans influence different steps in fibrillogenesis and the extracellular microdomains provide a mechanism for the tendon fibroblasts to regulate these extracellular interactions.     

Reticular meshwork of the spleen in rats studied by electron microscopy

The reticular meshwork of the rat spleen, which consists of both fibrous and cellular reticula, was investigated by transmission electron microscopy. The fibrous reticulum of the splenic pulp is composed of reticular fibers and basement membranes of the sinuses. These reticular fibers and basement membranes are continuous with each other. The reticular fibers are enfolded by reticular cells and are composed of two basic elements: 1) peripheral basal laminae of the reticular cells, and 2) central connective tissue spaces in which microfibrils, collagenous fibrils, elastic fibers, and unmyelinated adrenergic nerve fibers are present. The basement membranes of the sinuses are sandwiched between reticular cells and sinus endothelial cells and are composed of lamina-densalike material, microfibrils, collagenous fibrils, and elastic fibers. The presence of these connective tissue fibrous components indicates that there are connective tissue spaces in these basement membranes. The basement membrane is divided into three parts: the basal lamina of the reticular cell, the connective tissue space, and the basal lamina of the sinus endothelial cell. When the connective tissue space is very small or absent, the two basal laminae may fuse to form a single, thick basement membrane of the splenic sinus wall. The fibrous reticulum having these structures is responsible for support (collagenous fibrils) and rebounding (elastic fibers). The cells of the cellular reticulum--reticular cells and their cytoplasmic processes, which possess abundant contractile microfilaments, dense bodies, hemidesmosomes, basal laminae, and a well-developed, rough-surfaced endoplasmic reticulum, and Golgi complexes, which are characteristic of both fibroblasts and smooth muscle cells--are considered to be myofibroblasts. They may play roles in splenic contraction and in fibrogenesis of the fibrous reticulum. The contractile ability may be influenced by the unmyelinated adrenergic nerve fibers that pass through the reticular fibers. The three-dimensional reticular meshwork of the spleen consists of sustentacular fibrous reticulum and contractile myofibroblastic cellular reticulum. This meshwork not only supports the organ but also contributes to a contractile mechanism in circulation regulation, in collaboration with major contractile elements in the capsulo-trabecular system.     

Collagen types I, III, and V in human embryonic and fetal skin

The dermis of human skin develops embryonically from lateral plate mesoderm and is established in an adult-like pattern by the end of the first trimester of gestation. In this study the structure, biochemistry, and immunocytochemistry of collagenous matrix in embryonic and fetal dermis during the period of 5 to 26 weeks of gestation was investigated. The dermis at five weeks contains fine, individual collagen fibrils draped over the surfaces of mesenchymal cells. With increasing age, collagen matrix increases in abundance in the extracellular space. The size of fibril diameters increases, and greater numbers of fibrils associate into fiber bundles. By 15 weeks, papillary and reticular regions are recognized. Larger-diameter fibrils, larger fibers, denser accumulations of collagen, and fewer cells distinguish the deeper reticular region from the finer, more cellular papillary region located beneath the epidermis. The distribution of collagen types I, III, and V were studied at the light microscope level by immunoperoxidase staining and at the ultrastructural level by transmission (TEM) and scanning electron microscopy (SEM) with immunogold labeling. By immunoperoxidase, types I and III were found to be evenly distributed, regardless of fetal age, throughout the dermal and subdermal connective tissue with an intensification of staining at the dermal-epidermal junction (DEJ). Staining for types III and V collagen was concentrated around blood vessels. Type V collagen was also localized in basal and periderm cells of the epidermis. By immuno-SEM, types I and III were found associated with collagen fibrils, and type V was localized to dermal cell surfaces and to a more limited extent with fibrils. The results of biochemical analyses for relative amounts of types I, III, and V collagen in fetal skin extracts were consistent with immunoperoxidase data. Type I collagen was 70-75%, type III collagen was 18-21%, and type V was 6-8% of the total of these collagens at all gestational ages tested, compared to 85-90% type I, 8-11% type III, and 2-4% type V in adult skin. The enrichment of both types III and V collagen in fetal skin may reflect in part the proportion of vessel- and nerve-associated collagen versus dermal fibrillar collagen. The accumulation of dermal fibrillar collagen with increasing age would enhance the estimated proportion of type I collagen, even though the ratios of type III to I in dermal collagen fibrils may be similar at all ages.     

Type V collagen in splenic reticular fibers of the macaque monkey

The distribution of type I, III and V collagens in the monkey spleen was examined by indirect immunofluorescent microscopy and immunoelectron microscopy, and compared with that of reticular fibers revealed by a silver impregnation method. Type I collagen was localized on reticular fibers in the white pulps and on coarse reticular fibers in the splenic cords. Type III collagen was localized on the reticular fibers in the white pulps, and on the coarse reticular fibers and a limited number of fine reticular fibers, in the splenic cords. The anti-type V collagen antibody reacted with annular reticular fibers around the splenic sinuses, as well as with the reticular fibers in the white pulps and with the coarse and fine reticular fibers in the splenic cords. Thus, the distribution pattern of fibers that reacted with the anti-type V collagen antibody was very similar to that of the reticular fibers revealed by the silver impregnation method. Electron-microscopically, the fine reticular fibers in the splenic cords were composed of collagen fibrils, 30-50 nm in diameter, and amorphous substances. They were covered by reticular cell processes. By immunoperoxidase labeling with the anti-type V collagen antibody, electron-dense reaction products were found over the collagen fibrils with a banding pattern. These results indicate that type V collagen is an indispensable component of the reticular fibers.     

Structural variability of collagen fibers in the calcareous axial rod of a sea pen

Previous studies revealed that the organic matrix of the skeletal rod of the sea pen, Veretillum cynomorium, contained about 50% collagenous protein. The present ultrastructural study, based upon conventional staining methods, shows the existence of an abundant, longitudinally arranged nonbanded and fibrillar material separated by a reticular matrix. After incubation with ³H-proline, labeling is specifically localized on the fibrillar material. Some fibers occasionally display a transverse striation with a period of 11 to 14 nm which can be associated with a chevron striation. Infrequently, some other fibers display a more distinct banding of 55 to 70 nm or even yield a checkerboard pattern. However, a majority of fibers remain without a regular structure comparable to the periodic striations observed in the collagen of other animals. After treatment with 1% PTA in 70% ethanol, all the fibers show a clear banding of 14 nm and some of them possess two types of striations. The same result is obtained on fibers mechanically dissociated and negatively stained. As these methods show a periodic banding pattern on all the fibers, it is likely that all the fibers (striated or not) observed after routine electron microscopy correspond to collagen material. This collagen appears to be both polymorphic and completely new in comparison to that which is characteristic of the mesoglea. The polymorphic aspect is compared to that obtained from vertebrate collagens.     

The fiber components of insect connective tissue

Connective tissue around the nerve cord and heart have been studied in Calpodes ethlius. Four components at, distinguishable by selective staining and electron microscopy: matrix, collagen fine fibrils less than 60 Å in diameter and broad fibers about 400 Å in diameter after glutaraldehyde only the broad fibers react selectively for peroxidase and stain with phosphotungstic acid. These fibers are most abundant in connective tissue which is elastic. The fine fibrils are arranged parallel to and between the peroxidase reaciive fibers. It is suggested that the peroxidase activity of the fibers may be related to their stabilization. The collagen fibers have the narrow fibrillar form characteristic of Lepidoptera and Coleoptera and have a macroperiod of about 660 Å and a banding pattern matching that found in other insects.     

Revisiting the mystery of fibronectin multimers: The fibronectin matrix is composed of fibronectin dimers cross-linked by non-covalent bonds

Fibronectin (FN) matrix fibrils have long been thought to be formed by disulfide-bonded FN multimers, although there is no direct evidence that they are covalently linked with each other. To understand the biochemical properties of these fibrils, we extracted a crude FN matrix from FN-YPet transfected 3T3 cell culture using 0.2% deoxycholate and DNase. The insoluble extracted matrix preserved fibrillar structures and a major portion of the extracted proteins migrated as FN monomers on an SDS gel under reducing conditions. Under non-reducing conditions, some FN molecules appeared to be trapped at the top of the stacking gel. We tested this by mixing fluorescently labeled FN dimers with the extracted matrix just before loading on an SDS gel, and found that most of them were trapped with the extracted proteins at the top of the stacking gel. These results suggested that some components of the extracted matrix plugged the stacking gel and FN dimers were trapped with them. Rotary shadowing electron microscopy showed that the extracted matrix had some fibers that resembled fibrillin microfibrils. Peptide mass fingerprinting confirmed the presence of fibrillin in the extracted matrix. Fibrillin is known to form disulfide-bonded multimers and it is likely to be one of the components that plug the stacking gel and trap FN molecules in this system. The phenomenon by which FN molecules appear to migrate as multimers on SDS gels is thus an artifact rising from the presence of other large components in the extract. We conclude that FN matrix fibrils are made of FN dimers that are further cross-linked by non-covalent protein–protein bonds.     

Study on the fine structure of the central nervous system of Pholus labruscoe,L. (Lepidoptera)

Summary This is a preliminary electron microscope investigation in which the structure of insect neurons, neuropile, and interganglionic fibers are studied.Neurons of insect are pear-shaped and have an unique prolongation which ramifies into the neuropile. Their soma is surrounded by glial prolongations that exclude the possibility of nervous contacts. The neuronal cytoplasm is rich in granular material similar to the one described as R.N.A. by several authors; it is scattered at random or associated with endoplasmic reticulum cysternae. The latter does not adopt the regular array characterizing the vertebrate Nissl bodies.A large number of naked fibers is seen in the neuropile. The content of these fibers is different in fibers of different diameter. The thinner elements appear light and show a loose reticular matrix, few vesicles, and mitochondria. The thick fibers are characterized by a denser neuroplasm constituted by a reticular matrix and rows of tiny vesicles alternating with profils of tubuli. In some of these fibers the tubuli are seen in a central position.Three main types of contact relationships between fibers are described in the neuropile. These are; a) cross contacts; b) longitudinal contacts; and c) endknob contacts. The first type is in turn subdivided into subtypes, namely: minimum-area cross contacts and maximum-area cross contacts.A glial sheath enveloping each connective nerve fiber is described. Inside the cytoplasm of such cells there are bundles of dense, thin fibrils twisted along the nerve fibers.The criteria maintained by several authors in regard to the fine structure of the synaptic region are discussed and compared with facts reported in this paper.     

Identification of Histological Patterns in Clinically Affected and Unaffected Palm Regions in Dupuytren's Disease

Dupuytren''s disease is a fibro-proliferative disease characterized by a disorder of the extracellular matrix (ECM) and high myofibroblast proliferation. However, studies failed to determine if the whole palm fascia is affected by the disease. The objective of this study was to analyze several components of the extracellular matrix of three types of tissues—Dupuytren''s diseased contracture cords (DDC), palmar fascia clinically unaffected by Dupuytren''s disease contracture (NPF), and normal forehand fascia (NFF). Histological analysis, quantification of cells recultured from each type of tissue, mRNA microarrays and immunohistochemistry for smooth muscle actin (SMA), fibrillar ECM components and non-fibrillar ECM components were carried out. The results showed that DDC samples had abundant fibrosis with reticular fibers and few elastic fibers, high cell proliferation and myofibroblasts, laminin and glycoproteins, whereas NFF did not show any of these findings. Interestingly, NPF tissues had more cells showing myofibroblasts differentiation and more collagen and reticular fibers, laminin and glycoproteins than NFF, although at lower level than DDC, with similar elastic fibers than DDC. Immunohistochemical expression of decorin was high in DDC, whereas versican was highly expressed NFF, with no differences for aggrecan. Cluster analysis revealed that the global expression profile of NPF was very similar to DDC, and reculturing methods showed that cells corresponding to DDC tissues proliferated more actively than NPF, and NPF more actively than NFF. All these results suggest that NPF tissues may be affected, and that a modification of the therapeutic approach used for the treatment of Dupuytren''s disease should be considered.     

Surface structure of amyloid-beta fibrils contributes to cytotoxicity

Amyloid beta (Abeta) toxicity has been hypothesized to initiate the pathogenesis of Alzheimer's disease (AD). The characteristic fibrillar morphology of Abeta-aggregates, that constitute the main components of senile plaque, has long been considered to account for the neurotoxicity. But recent reports argue against a primary role for mature fibrils in AD pathogenesis because of the lack of a robust correlation between the severity of neurological impairment and the extent of amyloid deposition. Toxicity from the soluble prefibrillar intermediate entity of aggregates often called oligomer has recently proposed a plausible explanation for this inconsistency. An alternative explanation is based on the observation that certain amyloid fibril morphologies are more toxic than others, indicating that not all amyloid fibrils are equally toxic. Here, we report that it is not only the beta-sheeted fibrillar structure but also the surface physicochemical composition that affects the toxicity of Abeta fibrils. For the first time, colloidal gold was used to visualize by electron microscopy positive-charge clusters on Abeta fibrils. Chemical modifications as well as point-mutated Abeta synthesis techniques were applied to change the surface structures of Abeta and to show how local structure affects surface properties that are responsible for electrostatic and hydrophobic interactions with cells. We also report that covering the surface of Abeta fibers with myelin basic protein, which has surface properties contrary to those of Abeta, suppresses Abeta toxicity. On the basis of these results, we propose that the surface structure of Abeta fibrils plays an important role in Abeta toxicity.     

A comparison of the immunofluorescent localization of collagen types I,III, and V with the distribution of reticular fibers on the same liver sections of the snow monkey (Macaca fuscata)

Summary Localizations of collagen types I, III, and V in monkey liver, as determined by the indirect immunofluorescence method, were photographically superimposed on the fibers revealed by silver-staining in the same tissue sections. Immunofluorescence for type I collagen was found to correspond with the brown collagen fibers and with some of the coarse reticular fibers, while that for type III collagen was found to correspond with most, but not all, reticular fibers of the liver as well as with the brown collagen fibers. The distribution of type V collagen coincides not only with the collagen fibers in the stroma of portal triads and around the central veins, but also with the coarse and fine reticular fibers in the liver lobules. By immuno-electron microscopy, reaction products with anti-type III and V collagens antibodies were demonstrated on cross-striated collagen fibrils, about 45 nm in diameter, in the space of Disse. From these observations, it is concluded that: (1) the fine reticular fibers are mainly composed of type III and type V collagens, and (2) the collagen fibers and coarse reticular fibers in the periphery of liver lobules are composed of type I, type III and type V collagens.     

Organic structures of the hypercalcified peritubular matrix in horse dentine.

EDTA-insoluble organic structures of the hypercalcified peritubular matrix (PM) in horse dentine were observed by scanning electron microscopy. The PM was enveloped in double cylindrical structures composed of fibrillar sheaths in the inner and outer peripheries. Between the outer fibrillar sheath and intrinsic fibrils of the intertubular matrix, a calcified cementing membrane existed. Within the PM, warped cone-shaped structures of fibrillar sheaths, overlapping at intervals of 4-6 microns and semiconcentrically surrounding the dentinal tubule, extended from the inner fibrillar towards the outer fibrillar sheath. The cone-shaped fibrillar sheaths following the inner and outer fibrillar sheaths were identified as the incremental lines of the PM. Most of these fibrils may be collagen although it could not be confirmed, whereas non-collagenous organic materials in the lateral branches of the dentinal tubule are radially arranged in the PM. These EDTA-insoluble structures were three-dimensionally illustrated using an image-analysing system.     

Observing growth steps of collagen self-assembly by time-lapse high-resolution atomic force microscopy

Insights into molecular mechanisms of collagen assembly are important for understanding countless biological processes and at the same time a prerequisite for many biotechnological and medical applications. In this work, the self-assembly of collagen type I molecules into fibrils could be directly observed using time-lapse atomic force microscopy (AFM). The smallest isolated fibrillar structures initiating fibril growth showed a thickness of approximately 1.5 nm corresponding to that of a single collagen molecule. Fibrils assembled in vitro established an axial D-periodicity of approximately 67 nm such as typically observed for in vivo assembled collagen fibrils from tendon. At given collagen concentrations of the buffer solution the fibrils showed constant lateral and longitudinal growth rates. Single fibrils continuously grew and fused with each other until the supporting surface was completely covered by a nanoscopically well-defined collagen matrix. Their thickness of approximately 3 nm suggests that the fibrils were build from laterally assembled collagen microfibrils. Laterally the fibrils grew in steps of approximately 4 nm, indicating microfibril formation and incorporation. Thus, we suggest collagen fibrils assembling in a two-step process. In a first step, collagen molecules assemble with each other. In the second step, these molecules then rearrange into microfibrils which form the building blocks of collagen fibrils. High-resolution AFM topographs revealed substructural details of the D-band architecture of the fibrils forming the collagen matrix. These substructures correlated well with those revealed from positively stained collagen fibers imaged by transmission electron microscopy.     

A FIBRILLAR INTERCELLULAR MATERIAL BETWEEN REAGGREGATING EMBRYONIC CHICK CELLS

Lanthanum staining of embryonic chick cell reaggregates reveals an intercellular material composed of fibrils. Fibrillar arrays may be composed of parallel fibrils with a 35 A center-to-center spacing. Fibrils may also be disoriented, long, and tortuous. Newly dissociated cells show little lanthanum staining surface material, but appreciable amounts are present after 6 hr of reaggregation. Examination of intact tissue does not give the same clear evidence of a fibrillar matrix surrounding the cells, but treatment with a number of agents permits observation of intercellular fibrils, and in some cases there is evidence of orientation. Thus fibrillar material must be taken into account in considering mechanisms of cell aggregation.     

Innervation of the guinea pig spleen studied by electron microscopy

The innervation of the guinea pig spleen was investigated by electron microscopy. Unmyelinated nerve fibers in the capsulotrabecular and arterial systems were found to contain large and small granular and small agranular synaptic vesicles in their terminals and are thought to be sympathetic adrenergic in nature. They influence the contraction of the smooth muscle cells by diffusion innervation in these systems. These nerve terminals were also scattered in both the red and the white pulp. Pulp nerves wrapped by Schwann cells were further enclosed by myofibroblastic reticular cells. This condition revealed that the pulp nerves pass through the connective-tissue spaces of the reticular fibers, which contain elastic fibers, collagenous fibrils, and lamina densa-like materials of the usual basement laminae. One of the target cells for the pulp nerves is considered to be the myofibroblastic reticular cell in the reticular meshwork. Neurotransmitter substances released from the naked adrenergic nerve terminals travel through the reticular fibers and may play a role, by both close association innervation and diffusion innervation, in the contraction of reticular cells to expose the reticular fibers. At the exposed sides, connective-tissue elements of the reticular fibers are bathed with blood plasma, and the included naked nerve terminals, devoid of Schwann cells but with basement laminae of these cells, face free cells at some distance or are in close association with free cells, especially lymphocytes, macrophages, and plasma cells. The close ultrastructural relationship between the naked adrenergic nerve terminals and immunocytes strongly suggests that there is an intimate relationship between the immune system and the sympathetic nervous system through both close association innervation and diffusion innervation. Thus splenic adrenergic nerves of the guinea pig may play a triple role in 1) contraction of smooth muscle cells to regulate blood flow in the organ, 2) induction of the exposure of reticular fibers by contraction of the reticular cells in order to form a close relationship of the nerve terminals with the immunocytes, and 3) subsequent neuroimmunomodulation of the immunocytes.     

Collagen fibrils and elastic fibers in rat-tail tendon: an electron microscopic investigation

Earlier studies by the authors showed that the collagen fibrils in rat-tail tendon have a bi-modal distribution of fibril diameters from a time shortly after birth through to the onset of maturity at about 3–4 months. Present work has extended those observations for rats up to the age of 2 years. Histograms of the fibril diameter distributions for mature tail tendon and direct electron microscope observations show that the fibrils break down as the tendon ages. Further work on the constant diameter subfibrils of diameter 140 Å described previously, has confirmed that these are part of the elastic fibers present in tendon at all ages. It has been shown that there is relatively little variation in the collagen fibril diameter distribution as a function of the position of the specimen in the tail, and as the measured percentage of the area taken by the collagen fibrils present at any particular point. Estimation of the fibrillar collagen content of rat-tail tendon as a function of age indicates that it increases steadily from birth and reaches a maximum at the onset of maturity, beyond which the fibrillar collagen content appears to remain constant.     

Amianthoid (asbestoid) transformation: electron microscopical studies on aging human costal cartilage

The present study reports on the fine structure of human costal cartilage at different ages in order to obtain information on the morphogenesis of amianthoid fibers. Our results reveal an overall increase of collagen fibril diameter with increasing age, even in areas with no signs of amianthoid transformation. Ultrastructural evidence is presented that this increase in diameter is due to a gathering of the preexisting collagen fibrils. The age-related change in collagen fibril diameter is paralleled by changes in the composition and ultrastructural appearance of cartilage proteoglycans (as revealed by acridine orange staining). Acridine-orange-positive filaments indicative for proteoglycans are markedly reduced in size with advancing age in centrally located regions of costal cartilage. Treatment with testicular hyaluronidase previous to acridine-orange staining leaves these small proteoglycan filaments unaffected. By contrast, the filaments visible after acridine-orange staining in the extracellular matrix near to the perichondrium are susceptible to hyaluronidase treatment. Infrequently, a sharp increase in collagen fibril diameter can be observed in territorial matrix areas of degenerating chondrocytes. This observation is conspicuous at ages of 10 and 20 years. Amianthoid transformation is characterized by the appearance of collagen fibrils strictly arranged in parallel. These amianthoid fibers are embedded in a matrix rich in small acridine-orange-positive filaments similar to the proteoglycan filaments observed in centrally located matrix regions. It can be concluded that extensive remodelling not only of the collagen fibrils but also of the cartilage proteoglycans is involved in the development of amianthoid transformation.