Connective-tissue macromolecules in Golgi chicken tendon organs and at their interface with muscle fibers and adjoining tendinous structures.

Tendon organs from leg and forearm muscles of white leghorn chickens were examined with a library of monoclonal antibodies to determine the composition of their connective-tissue framework and the types of connective-tissue macromolecules that occur at the sites where muscle fibers attach to the receptors. The capsules of the tendon organs were positive for connective-tissue macromolecules typical of basal lamina (collagen type IV, laminin, and heparin sulfate proteoglycan) and for tenascin, collagen types III and VI, and fibronectin. Connective-tissue bundles in the lumen of a receptor reacted primarily with antibodies against collagen type I and 4-chondroitin sulfate. The narrow partitions that divide each lumen into compartments stained for collagen type III. Toward its tendinous end, a receptor made few contacts with muscle fibers. Instead, the capsule and the collagenous bundles blended gradually with the intermuscular portions of tendons. At the muscular end, the connections were more complex. Muscle fibers that attached in series to tendon organs split to produce basal lamina-covered, finger-like extensions, which were separated from each other by fissures. Tongues of connective tissue containing tenascin, collagen types I and VI, and fibronectin extended into the fissures. Distally the tongues were continuous with the tenascin in the capsule and just internal to the capsule, fibronectin and basal lamina macromolecules in the capsule, and collagen type I in the collagenous bundles. The uninterrupted presence of these macromolecules around terminating muscle fibers and in the capsule and/or the intraluminal collagen bundles suggests that muscle fibers that attach in series at the muscular end exert a force during muscular contraction on the intraluminal collagen bundles and on the receptor capsule.     

Location of the integrin complex and extracellular matrix molecules at the chicken myotendinous junction

Summary The distribution of several extracellular matrix macromolecules was investigated at the myotendinous junction of adult chicken gastrocnemius muscle. Localization using monoclonal antibodies specific for 3 basal lamina components (type IV collagen, laminin, and a basement membrane form of heparan sulfate proteoglycan) showed strong fluorescent staining of the myotendinous junction for heparan sulfate proteoglycan and laminin, but not for type IV collagen. In addition, a strong fluorescent stain was observed at the myotendinous junction using a monoclonal antibody against the subunit of the chicken integrin complex (antibody JG 22). Neither fibronectin nor tenascin were concentrated at the myotendinous junction, but instead were present in a fibrillar staining pattern throughout the connective tissue which was closely associated with the myotendinous junction. Tenascin also gave bright fluorescent staining of tendon, but no detectable staining of the perimysium or endomysium. Type I collagen was observed throughout the tendon and in the perimysium, but only faintly in the endomysium. In contrast, type III collagen was present brightly in the endomysium and in the perimysium, but could not be detected in the tendon except when associated with blood vessels and in the epitendineum, which stained intensely. Type VI collagen was found throughout the tendon and in all connective tissue partitions of skeletal muscle. The results indicate that one or more molecules of the integrin family may play an important role in the attachment of muscle to the tendon. This interaction does not appear to involve extensive binding to fibronectin or tenascin, but may involve laminin and heparan sulfate proteoglycan.     

Immunohistochemistry of the intercellular matrix components and the epithelio-mesenchymal junction of the human tooth germ

Summary The immunohistochemical localization of heparan sulphate, collagen type I, III and IV, laminin, tenascin, plasma- and cellular fibronectin was studied in tooth germs from human fetuses. The lamina basalis ameloblastica or membrana preformativa, which separates the pre-ameloblasts from the pre-dentin and dentin, contained heparan sulphate, collagen type IV, laminin and fibronectin. Enamel reacted with antifibronectin, but the reaction varied depending on the type of fibronectin and the source of antibody. In early pre-dentin, collagen type I, laminin, tenascin and fibronectin were present. In late pre-dentin and dentin collagen type I was found in intertubular dentin and in the zone between enamel and dentin. The close relationship between collagen type I in dentin and fibronectin in immature enamel is interesting, as it may contribute to the stabilization of the amelodentinal interface. In dental pulp, collagen type IV and laminin were found in the endothelial basement membranes. Collagen type I and III, tenascin and fibronectin were localized to the mesenchymal intercellular matrix.The results of this study have supported the assumption that the lamina basalis ameloblastica is a basement membrane, and have lead to the suggestion that ameloblasts are producers of fibronectin or a fibronectin-like substance.     

Immunohistochemical study of subepidermal connective of molluscan integument

Sections of integument from gastropod, bivalve and cephalopod species were studied immunohistochemically to determine reactivity to antibody against the type I-like collagen from Sepia cartilage and antibodies against components of the extracellular matrix (ECM) of vertebrate connective tissue: type I, III, IV, V, and VI collagens, laminin, nidogen and heparan sulphate. All samples exhibited similar reactivities to the antibodies, although differences in the intensity and localization of the immunostaining were found that were clearly correlated with between-species differences in integumental ultrastructure. These findings indicate that the composition of the integumental ECM is similar in the three classes of molluscs examined and that several types of collagen are present. However molluscan subepidermal connective tissue differs from the ECM of vertebrate dermis: molluscan integumental ECM contains collagens similar to type I, V and VI collagens but has no type III-similar collagen. Furthermore molecules similar to the type IV collagen, laminin, nidogen and heparan sulphate of vertebrates were present ubiquitously in molluscan basement membrane, confirming the statement that the structure and composition of basement membrane have remained constant throughout the evolution of all animal phyla.     

Light microscopic immunolocalization of type IV collagen, laminin, heparan sulfate proteoglycan, and fibronectin in the basement membranes of a variety of rat organs

Immunohistochemical methods were used to determine whether type IV collagen, laminin, fibronectin, and heparan sulfate proteoglycan were present in diverse basement membranes. Antisera or antibodies against each substance were prepared, tested by enzyme-linked immunosorbent assay, and exposed to frozen sections of duodenum, trachea, kidney, spinal cord, cerebrum, and incisor tooth from rats aged 20 days to 34 months. Bound antibodies were then localized by indirect or direct peroxidase methods for examination in the light microscope. Immunostaining for type IV collagen, laminin, fibronectin, and heparan sulfate proteoglycan was observed in all of the basement membranes encountered. Fibronectin was also found in connective tissue. In general, the intensity of immunostaining was strong for type IV collagen and laminin, moderate for heparan sulfate proteoglycan, and weak for fibronectin. The pattern was similar in the age groups under study. Very recently the sulfated glycoprotein, entactin, was also detected in the basement membranes of the listed tissues in 20-day-old rats. It is accordingly proposed that, at least in the organs examined, type IV collagen, laminin, fibronectin, heparan sulfate proteoglycan, and entactin are present together in basement membranes.     

Macromolecular organization of bovine lens capsule

Rabbit antisera to type IV collagen, laminin, entactin, heparan sulfate proteoglycan and fibronectin were used to localize these proteins in cross-sections of bovine anterior lens capsule. The antisera were exposed to (a) 10-micron frozen-thawed sections of formaldehyde-fixed tissue for examination in the light microscope by the indirect immunofluorescence method and (b) formaldehyde-fixed and L. R. White plastic-embedded thin sections for electron microscopic examination by the protein A-gold technique. The intensity of immunofluorescence was both uniform and strong throughout for type IV collagen, laminin and entactin, but patchy and weak for fibronectin. Electron microscopic immunolabeling with protein A-gold showed that all five components were distributed throughout the full thickness of the membrane, albeit the density of gold particles was not identical for all basement membrane proteins. In general, the number of particles per micron2 was greatest for type IV collagen and entactin, moderate for laminin and heparan sulfate proteoglycan and low for fibronectin. The ultrastructure of the lens capsule as examined by the electron microscope revealed a relatively uniform parallel alignment of filaments, thought to be collagenous. Since the distribution of the filaments corresponds well with the observed immunocytochemical pattern it is concluded that type IV collagen, laminin, entactin, heparan sulfate proteoglycan and fibronectin co-localize throughout the cross-section of the anterior lens capsule.     

Localization of binding sites for laminin, heparan sulfate proteoglycan and fibronectin on basement membrane (type IV) collagen

Rotary shadowing electron microscopy was used to examine complexes formed by incubating combinations of the basement membrane components: type IV collagen, laminin, large heparan sulfate proteoglycan and fibronectin. Complexes were analyzed by length measurement from the globular (COOH) domain of type IV collagen, and by examination of the four arms of laminin and the two arms of fibronectin. Type IV collagen was found to contain binding sites for laminin, heparan sulfate proteoglycan and fibronectin. With laminin the most frequent site was centered approximately 81 nm from the carboxy end of type IV collagen. Less frequent sites appeared to be present at approximately 216 nm and approximately 291 nm, although this was not apparent when the sites were expressed as a fraction of the length of type IV collagen to which they were bound. For heparan sulfate proteoglycan the most frequent site occurred at approximately 206 nm with a less frequent site at approximately 82 nm. For fibronectin, a single site was present at approximately 205 nm. Laminin bound to type IV collagen through its short arms, particularly through the end of the lateral short arms and to heparan sulfate proteoglycan mainly through the end of its long arm. Fibronectin bound to type IV collagen through the free end region of its arms. Using a computer graphics program, the primary laminin binding sites of two adjacent type IV collagen molecules were found to align in the "polygonal" model of type IV collagen, whereas with the "open network" model, a wide meshed matrix is predicted. It is proposed that basement membrane may consist of a lattice of type IV collagen coated with laminin, heparan sulfate proteoglycan and fibronectin.     

Immunocytochemistry of extracellular matrix in the lamina propria of the rat testis: electron microscopic localization

The distribution of laminin, type IV collagen, heparan sulfate proteoglycan, and fibronectin was investigated in the rat testicular lamina propria by electron microscopic immunocytochemistry. Distinct patterns were observed for each antigen within the extracellular matrix (ECM) layers of the lamina propria. Laminin, type IV collagen, and heparan sulfate proteoglycan all localized to the seminiferous tubule basement membrane. Type IV collagen and heparan sulfate proteoglycan, but not laminin, localized to the seminiferous tubule side of the peritubular myoid cells. All four of the antigens were localized between the peritubular and lymphatic endothelial cells. Failure to localize fibronectin in the ECM layer between the Sertoli and peritubular myoid cells tends to support the concept that adult Sertoli cells do not produce this protein in vivo. Intracellular immunostaining was insufficient to allow unambiguous identification of the cellular source of any of the ECM molecules.     

Developmental expression of extracellular matrix components in intramuscular connective tissue of bovine semitendinosus muscle

 We have investigated the expression patterns of extracellular matrix components in intramuscular connective tissue during the development of bovine semitendinosus muscle by means of indirect immunofluorescence techniques. Types I, III, V, and VI collagen and fibronectin were located in the endomysium and the perimysium. Type IV collagen, laminin, and heparan sulfate proteoglycans (PGs) were exclusively located in the endomysium, and dermatan sulfate PGs existed only in the perimysium. The localization of these components in the intramuscular connective tissue of semitendinosus muscle remained unchanged throughout prenatal and postnatal growth of cattle, suggesting that they are essential for forming and maintaining structures of the endomysium and perimysium in bovine semitendinosus muscle. On the other hand, decorin was undetectable in the endomysium of neonates, although other matrix components were already expressed. It was expressed slightly in the endomysium of 2-month-old calves, and clearly detectable in the endomysium of cattle more than 6 months old. Chondroitin sulfate PGs were barely detectable in the perimysium of fetuses and neonatal calves, and progressively appeared during postnatal development of the muscle. It seems likely that these PGs are closely related to the postnatal development of the endomysium and perimysium. Accepted: 30 October 1996     

Amyloid precursor-like protein 2 promotes cell migration toward fibronectin and collagen IV.

Previous studies have established that in response to wounding, the expression of amyloid precursor-like protein 2 (APLP2) in the basal cells of migrating corneal epithelium is greatly up-regulated. To further our understanding of the functional significance of APLP2 in wound healing, we have measured the migratory response of transfected Chinese hamster ovary (CHO) cells expressing APLP2 isoforms to a variety of extracellular matrix components including laminin, collagen types I, IV, and VII, fibronectin, and heparan sulfate proteoglycans (HSPGs). CHO cells overexpressing either of two APLP2 variants, differing in chondroitin sulfate (CS) attachment, exhibit a marked increase in chemotaxis toward type IV collagen and fibronectin but not to laminin, collagen types I and VII, and HSPGs. Cells overexpressing APLP2-751 (CS-modified) exhibited a greater migratory response to fibronectin and type IV collagen than their non-CS-attached counterparts (APLP2-763), suggesting that CS modification enhanced APLP2 effects on cell migration. Moreover, in the presence of chondroitin sulfate, transfectants overexpressing APLP2-751 failed to exhibit this enhanced migration toward fibronectin. The APLP2-ECM interactions were also explored by solid phase adhesion assays. While overexpression of APLP2 isoforms moderately enhanced CHO adhesion to laminin, collagen types I and VII, and HSPGs lines, especially those overexpressing APLP2-751, exhibited greatly increased adhesion to type IV collagen and fibronectin. These observations suggest that APLP2 contributes to re-epithelialization during wound healing by supporting epithelial cell adhesion to fibronectin and collagen IV, thus influencing their capacity to migrate over the wound bed. Furthermore, APLP2 interactions with fibronectin and collagen IV appear to be potentiated by the addition of a CS chain to the core proteins.     

Lack of heparan sulfate proteoglycan in a discontinuous and irregular placental basement membrane

A discontinuous basement membrane of variable width that surrounds spongiotrophoblast cells of rat placenta was examined for the presence of type IV collagen, laminin, a heparan sulfate proteoglycan, entactin, and fibronectin using monospecific antibodies or antisera and the indirect peroxidase technique. At the level of the light microscope, the basement membrane was immunostained for type IV collagen, laminin, entactin, and fibronectin. Heparan sulfate proteoglycan immunostaining, however, was virtually absent even after pretreatment of sections with 0.1 N acetic acid, pepsin (0.1 microgram/ml) or 0.13 M sodium borohydride. Examination in the electron microscope confirmed the lack of immunostaining for heparan sulfate proteoglycan, whereas the other substances were mainly localized to the lamina densa part of the basement membrane. The absence of heparan sulfate proteoglycan in this discontinuous and irregular basement membrane even though type IV collagen, laminin, entactin, and fibronectin are present, suggests that heparan sulfate proteoglycan may have a structural role in the formation of basement membrane.     

Localization of laminin, type IV collagen, fibronectin, and heparan sulfate proteoglycan in chick retinal pigment epithelium basement membrane during embryonic development

Type IV collagen, laminin, heparan sulfate proteoglycan, and fibronectin were localized in the basement membrane (BM) of chick retinal pigment epithelium (RPE) during various stages of eye development. At different times over a 4-17 day period after fertilization, chick embryo eyes were dissected, fixed in periodate-lysine-paraformaldehyde, and 6 micron frozen sections through the central regions of the eye were prepared. Sections were postfixed in -20 degrees C methanol and stained immediately by indirect immunofluorescence using sheep anti-mouse laminin, sheep antimouse type IV collagen, rabbit anti-mouse heparan sulfate proteoglycan, and mouse monoclonal anti-porcine plasma fibronectin. Fluorescein-labeled F(ab')2 fragments of the appropriate immunoglobulins (IgGs) were used as secondary antibodies. Laminin could be readily demonstrated in the BM of the RPE during all stages of development. The staining for type IV collagen, fibronectin, and heparan sulfate proteoglycan HSPG) was less intense than that for laminin, but was also localized in the BM along the basal side of the RPE. In addition to staining the BM, antiserum to HSPG, gave a diffuse labeling from day 9 onward, above the RPE extending into the region of the photoreceptors. Whereas the intensity of staining generally increased between day 4 and day 17 of development, the distribution of the different BM components did not change. Hence the presence of type IV collagen, laminin, fibronectin, and HSPG in the BM of RPE in vivo during all the stages of development investigated supports the concept that these macromolecules are important basic components of this, and other, BMs. Furthermore, these results indicate that the composition of the BM of RPE cells in vivo is similar to the BM material deposited by RPE cells in vitro (Turksen K, Aubin JE, Sodek JE, Kalnins VI: Collagen Rel Res, 4:413-426, 1984) and that the in vitro cultures can therefore serve as a useful model for studying BM formation.     

Immunogold localization of basement membrane molecules in rat retinal capillaries

Vascular basement membrane contains laminin, fibronectin, proteoglycan and collagens. These molecules have been identified in various tissues by immunolabeling methods and biochemical analyses. We have previously localized laminin, fibronectin and type IV collagen to the basement membrane of rat retinal vessels at the ultrastructural level using an immunoperoxidase method. In this study, we use an immunogold method to re-examine the distribution of these molecules and also to study the localization of heparan sulfate proteoglycan and types I, III and V collagen in the retinal capillary basement membrane. Gold labeling for laminin, type IV collagen and proteoglycan were found diffusely on the basement membrane of the endothelium and pericyte, while that for fibronectin and type V collagen was spotty and variable and that for types I and III collagen was negligible. The segment of basement membrane between the endothelial cell and pericyte appeared less reactive to anti-laminin and anti-type IV collagen than the membrane between the pericyte and perivascular neuroretina. The immunogold method may be useful in quantitative studies of thickened basement membranes under abnormal conditions.     

The extracellular matrix of lip wounds in fetal, neonatal and adult mice.

Wound healing in the fetus occurs rapidly, by a regenerative process and without an inflammatory response, resulting in complete restitution of normal tissue function. By contrast, in the adult, wounds heal with scar formation, which may impair function and inhibit further growth. The cellular mechanisms underlying these differing forms of wound healing are unknown but the extracellular matrix (ECM), through its effects on cell function, may play a key role. We have studied the ECM in upper lip wounds of adult, neonatal and fetal mice at days 14, 16 and 18 of gestation. The spatial and temporal distribution of collagen types I, III, IV, V and VI, fibronectin, tenascin, laminin, chondroitin and heparan sulphates were examined immunohistochemically. Results from the fetal groups were essentially similar whilst there were distinct differences between fetus, neonate and adult. Fibronectin was present at the surface of the wound in all groups at 1 h post-wounding. Tenascin was also present at the wound surface but the time at which it was first present differed between fetus (1 h), neonate (12 h) and adult (24 h). The time of first appearance paralleled the rate of wound healing which was most rapid in the fetus and slowest in the adult. Tenascin inhibits the cell adhesion effect of fibronectin and during development the appearance of tenascin correlates with the initiation of cell migration. During wound healing the appearance of tenascin preceded cell migration and the rapid closure of fetal wounds may be due to the early appearance of tenascin in the wound. Collagen types I, III, IV, V and VI were present in all three wound groups but the timing and pattern of collagen deposition differed, with restoration of the normal collagen pattern in the fetus and a scar pattern in the adult. This confirms that lack of scarring in fetal wounds is due to the organisation of collagen within the wound and not simply lack of collagen formation. The distribution of chondroitin sulphate differed between normal fetal and adult tissues and between fetal and adult wounds. Its presence in the fetal wound may alter collagen fibril formation. No inflammatory response was seen in the fetal wounds. The differences in the ECM of fetal and adult wounds suggests that it may be possible to alter the adult wound so that it heals by a fetal-like process without scar formation, loss of tissue function or restriction of growth.     

Abnormal deposition of basement membrane and connective tissue components in dimethylbenzanthracene-induced rat mammary tumours: an immunocytochemical and ultrastructural study

Summary Dimethylbenzanthracene-induced rat mammary tumours consist of lobules of tumours cells surrounded by connective tissue. The interstitial connective tissue proteins, collagen types I, III and V, fibronectin and elastin are largely restricted to the interlobular connective tissue. The tumour lobules are surrounded by a basement membrane that stains with antiserum to laminin. Electron microscopy reveals a greatly thickened basement membrane to which striated interstitial collagen fibres are closely juxtaposed. The lumina within the tumour lobules are of two types. In the first type, the luminal surface is characterized by the presence of microvilli and tight junctions are reacts with antiserum to rat milk fat globule membrane. In the second type, the luminal surface is flattened and lined by a thickened basement membrane that stains with antiserum to laminin and type IV collagen. These abnormal patterns of growth and differentiation may be partly a consequence of the disorganization of extracellular matrix components at the interface between the tumour epithelial cells and the surrounding stroma.     

Structure, composition, and assembly of basement membrane

Basement membranes are thin layers of matrix separating parenchymal cells from connective tissue. Their ultrastructure consists of a three-dimensional network of irregular, fuzzy strands referred to as "cords"; the cord thickness averages 3-4 nm. Immunostaining reveals that the cords are composed of at least five substances: collagen IV, laminin, heparan sulfate proteoglycan, entactin, and fibronectin. Collagen IV has been identified as a filament of variable thickness persisting after the other components have been removed by plasmin digestion or salt extraction. Heparan sulfate proteoglycan appears as sets of two parallel lines, referred to as "double tracks," which run at the surface of the cords. Laminin is detected in the cords as diffuse material within which thin wavy lines may be distinguished. The entactin and fibronectin present within the cords have not been identified as visible structures. The ability of laminin, heparan sulfate proteoglycan, fibronectin, and entactin to bind to collagen IV has been demonstrated by visualization with rotary shadowing and/or biochemical studies. Incubation of three of these substances-collagen IV, laminin (with small entactin contamination), and proteoglycan-at 35 degrees C for 1 hr resulted in a precipitate that was sectioned for electron microscopic examination and processed for gold immunolabeling for each of the three incubated substances. Three structures are present in the precipitate: 1) a lacework, exclusively composed of heparan sulfate proteoglycan in the form of two parallel lines, similar to double tracks; 2) semi-solid, irregular accumulations, composed of the three initial substances distributed on a cord network; and 3) convoluted sheets, which are also composed of the three initial substances distributed on a cord network but which, in addition, have the uniform appearance and thickness of the lamina densa of basement membrane. Hence these sheets are closely similar to the main component of authentic basement membranes.     

Changes in the extracellular matrix of the normal human breast during the menstrual cycle

Summary The normal human mammary gland undergoes a well defined sequence of histological changes in both epithelial and stromal compartments during the menstrual cycle. Studies in vitro have suggested that the extracellular matrix surrounding the individual cells plays a central role in modulating a wide variety of cellular events, including proliferation, differentiation and gene expression. We therefore investigated the distribution of a number of extracellular matrix molecules in the normal breast during the menstrual cycle. By use of indirect immunofluorescence, with specific antibodies, we demonstrated that laminin, heparan sulphate proteoglycan, type IV collagen, type V collagen, chondroitin sulphate and fibronectin undergo changes in distribution during the menstrual cycle, whereas collagen types I, III, VI and VII remain unchanged. These changes were most marked in the basement membrane, sub-basement membrane zone and delimiting layer of fibroblasts surrounding the ductules where basement membrane markers such as laminin, heparan sulphate proteoglycan, and type IV and V collagens appear greatly reduced during the mid-cycle period (days 8 to 22). These results suggest that some extracellular matrix molecules may act as medittors in the hormonal control of the mammary gland, whereas others may have a predominantly structural role.     

Ontogenesis of the murine hepatic extracellular matrix: an immunohistochemical study

To define the role of the extracellular matrix (ECM) in hepatogenesis, we examined the temporal and spatial deposition of fibronectin, laminin and collagen types I and IV in 12.5-21.5 day fetal and 1, 7 and 14 day postnatal rat livers. In early fetal liver, discontinuous deposits of the four ECM components studied were present in the perisinusoidal space, with laminin being the most prevalent. All basement membrane zones contained collagen type IV and laminin, including those of the capsule (mesothelial), portal vein radicles and bile ductules. Fibronectin had a distribution similar to that of collagen type IV early in gestation. However, at later gestational dates, fibronectin distribution in the portal triads approached that of collagen type I, being present in the interstitial connective tissues; whereas, collagen type IV and laminin were restricted to vascular and biliary basement membrane zones in those regions. The cytoplasm of some sinusoidal lining cells and hepatocytes reacted with antibodies to extracellular matrix components. By electron microscopy the immunoreactive material was localized in the endoplasmic reticulum, indicating the ability of these cells to synthesize these ECM proteins. Biliary ductular cells had prominent intracytoplasmic staining for laminin and collagen type IV from day 19.5 gestation until 7 days of postnatal life, but lacked demonstrable fibronectin or collagen type I. These results demonstrate that by 12.5 days of gestation the rat liver anlage has deposited a complex extracellular matrix in the perisinusoidal space. The prevalence of laminin in the developing hepatic lobules suggests a possible role for this glycoprotein in hepatic morphogenesis. In view of the intimate association of the hepatic lobular extracellular matrix with the developing vasculature, we hypothesize that laminin provides a scaffold of the developing liver, but once the ontogenesis is complete, intrahepatic perisinusoidal laminin expression is suppressed.     

Connective tissue of the livers of newborn and adult marmosets (Callithrix jacchus)

Immunofluorescence microscopic and electron microscopic investigations revealed components of the matrix and of the basal lamina (collagen type I, III, IV and V, BL-heparan sulfate and fibronectin) in the sinus wall (Disse's space) of the livers of newborn and adult marmosets (Callithrix jacchus). Collagen type I was missing in both the two age groups. Small amounts of laminin were present in the livers of newborn and absent in those of adult animals, whereas collagen type III occurred in the form of delicate fibres. Light microscopic inspection showed a continuous distribution of all other components in the sinus wall. The amount of collagen type III and V increased depending on the age. Electron microscopic investigations revealed single or bundled fibrils (20-30 nm) and filaments (10-12 nm). After addition of tannic acid, plaques of a fine-filamentous network and incorporated granules were observed. After addition of resting Ruthenium Red, electron-dense granules (20-60 nm) were irregularly distributed in the structureless space, resting on collagenous fibrils and cell membranes. The fibrils were allocated to collagen type III, the filaments to collagen type V. The plaques were supposed to contain heparan sulfate, collagen type IV and fibronectin. The absence of a Lamina densa of the basal lamina was attributed to the absence of laminin which probably plays an important role in the formation of this layer. Differences in the distribution pattern of the matrix components and thus a functional mosaic of the permeability of Disse's space were assumed. The complete absence of collagen type I and laminin in the lobules makes the adult marmoset liver especially suited for studies on the importance of this collagen type under pathological conditions, since both components are expressed in this way.     

Structure and function of matrix components in the cruciate ligaments. An immunohistochemical, electron-microscopic, and immunoelectron-microscopic study.

In the present study, the matrix components of 100 cruciate ligaments were analyzed by conventional electron microscopy, immunohistology, morphometry, and immunoelectron microscopy. The anterior (ACL) and the posterior (PCL) cruciate ligaments contained collagen types III, IV, and VI. Several structural glycoproteins, like fibronectin, laminin, entactin, tenascin, and undulin were detected using monoclonal antibodies. Whereas laminin and entactin were higher concentrated in the PCL, type VI collagen was more frequently found in the ACL. The ACL had a critical nourishment in its distal and middle thirds. In all ligament parts the PCL revealed a better vascular supply with strong correlation to type IV collagen expression. The normal matrix of the cruciate ligaments represented a complicated regulatory network of proteins, glycoproteins, elastic systems, and glycosaminoglycans with multiple functional interactions.