Spatial arrangement of white muscle fibers and myoseptal tendons in fishes

We describe the arrangement of white muscle fibers and tendinous myoseptal structures and the relation of these structures to each other in order to provide an anatomical framework for discussions and experimental research on fish swimming mechanics. For the three major craniate groups, the petromyzontids, myxinids and gnathostomes, we identify three conditions that differ remarkably. Myxinids are characterized by asymmetrical myosepta with long cones. Within a single myoseptum these are connected by collagenous fibers that are almost oriented longitudinally. Distinct tendons are absent in myxinid myosepta. Petromyzontid myosepta lack cones and distinct myoseptal tendons, whereas gnathostomes bear cones and distinct tendinous structures: the lateral band, epineural (epipleural) tendon and myhabdoid tendon. Myoseptal fibers of petromyzontids and myoseptal tendons of gnathostome myosepta are firmly anchored in the skin. Myxinids lack firm myoseptal-skin-connections. Their muscular arrangement is neither comparable to that of petromyzontids nor to that of gnathostomes. The latter two bear archlike arrangements of muscle fibers spanning several segments that are hypothesized to play a role during bending. In gnathostomes, archlike helical muscle fiber arrangements (HMFAs) are present that span the length of several body segments and are multiply intersected by myosepta. Hence, a series of tendinous lateral bands of myosepta is embedded in HMFAs. The posterodorsally oriented HMFAs are underlain by posteroventrally oriented crossing muscle fibers (CMFs). Bending may be generated by contraction of the muscle fibers belonging to an HMFA and the simultaneous counteraction of CMFs. Moving caudally, this anterior muscle fiber arrangement gradually changes, eventually becoming the posterior muscle fiber arrangement. This pattern suggests that the function of the myomeres will also change. Three additional putative roles of myoseptal tendons can be deduced from their relations to white muscle fibers in gnathostomes (and in part in petromyzontids): (1) Posterior transmission of anteriorly generated muscular forces via lateral bands and/or myorhabdoid tendons. These tendons are more robust posteriorly. Anterior and posterior cones appear to play an important role in force transmission. (2) Pulling on collagen fibers of the skin via lateral bands and myorhabdoid tendons, suggesting a transmission of muscular forces that puts the skin into tension. (3) Resisting radial expansion of contracting muscle fibers by epineural (epipleural) tendons. By the latter two mechanisms modulation of body stiffness is likely to be achieved.     

Myoseptal architecture of sarcopterygian fishes and salamanders with special reference to Ambystoma mexicanum

During axial undulatory swimming in fishes and salamanders muscular forces are transmitted to the vertebral axis and to the tail. One of the major components of force transmission is the myoseptal system. The structure of this system is well known in actinopterygian fishes, but has never been addressed in sarcopterygian fishes or salamanders. In this study we describe the spatial arrangement and collagen fiber architecture of myosepta in Latimeria, two dipnoans, and three salamanders in order to gain insight into function and evolution of the myoseptal system in these groups. Salamander myosepta lack prominent cones, and consist of homogenously distributed collagen fibers of various orientations that never form distinct tendons. Fiber orientations are difficult to homologize with those of fish myosepta. The myosepta of Latimeria and dipnoans (Protopterus and Neoceratodus) illustrate that major changes in architecture occurred in the sarcopterygian clade (loss of horizontal septum), in the rhipidistian (dipnoans + tetrapods) clade (loss of epineural and epipleural tendon), and in tetrapods (loss of lateral tendons and myoseptal folding). When compared to fishes, the myosepta of wholly aquatic salamanders (Ambystoma mexicanum, Amphiuma tridactylum, Necturus maculosus) do not have the lateral tendons we suppose serve to transfer muscular forces posteriorly. We propose that alternative structures (most conspicuously present in Ambystoma) perform this function: posteriorly the relative amount of connective tissue increases considerably, and myosepta are disintegrated to horizontal lamellae of connective tissue. The structures thought to be involved in modulation of body stiffness in fishes during swimming are also absent in salamanders. Our data also have implications for the hypothesis that salamander hypaxial myosepta are designed to increase shortening amplification of the hypaxial muscle fibers. The posterior hypaxial myosepta of all three salamander species possess only mediolaterally directed collagen fibers, which would indeed amplify the shortening of the associated muscle.     

Tendon exhibits complex poroelastic behavior at the nanoscale as revealed by high-frequency AFM-based rheology

Tendons transmit load from muscle to bone by utilizing their unique static and viscoelastic tensile properties. These properties are highly dependent on the composition and structure of the tissue matrix, including the collagen I hierarchy, proteoglycans, and water. While the role of matrix constituents in the tensile response has been studied, their role in compression, particularly in matrix pressurization via regulation of fluid flow, is not well understood. Injured or diseased tendons and tendon regions that naturally experience compression are known to have alterations in glycosaminoglycan content, which could modulate fluid flow and ultimately mechanical function. While recent theoretical studies have predicted tendon mechanics using poroelastic theory, no experimental data have directly demonstrated such behavior. In this study, we use high-bandwidth AFM-based rheology to determine the dynamic response of tendons to compressive loading at the nanoscale and to determine the presence of poroelastic behavior. Tendons are found to have significant characteristic dynamic relaxation behavior occurring at both low and high frequencies. Classic poroelastic behavior is observed, although we hypothesize that the full dynamic response is caused by a combination of flow-dependent poroelasticity as well as flow-independent viscoelasticity. Tendons also demonstrate regional dependence in their dynamic response, particularly near the junction of tendon and bone, suggesting that the structural and compositional heterogeneity in tendon may be responsible for regional poroelastic behavior. Overall, these experiments provide the foundation for understanding fluid-flow-dependent poroelastic mechanics of tendon, and the methodology is valuable for assessing changes in tendon matrix compressive behavior at the nanoscale.     

Biomimetic approaches to tendon repair

The linear organization of collagen fibers in tendons results in optimal stiffness and strength at low strains under tensile load. However, this organization makes repairing ruptured or lacerated tendons extremely difficult. Current suturing techniques to join split ends of tendons, while providing sufficient mechanical strength to prevent gapping, are inadequate to carry normal loads. Immobilization protocols necessary to restore tendon congruity result in scar formation at the repair site and peripheral adhesions that limit excursion. These problems are reviewed to emphasize the need for novel approaches to tendon repair, one of which is the development of biomimetic tendons. The objective of the empirical work described here was to produce biologically-based, biocompatible tendon replacements with appropriate mechanical properties to enable immediate mobilization following surgical repair. Nor-dihydroguaiaretic acid (NDGA), a di-catechol from creosote bush, caused a dose dependent increase in the material properties of reconstituted collagen fibers, achieving a 100-fold increase in strength and stiffness over untreated fibers. The maximum tensile strength of the optimized NDGA treated fibers averaged 90 MPa; the elastic modulus of these fibers averaged 580 MPa. These properties were independent of strain rates ranging from 0.60 to 600 mm/min. Fatigue tests established that neither strength nor stiffness were affected after 80 k cycles at 5% strain. Treated fibers were not cytotoxic to tendon fibroblasts. Fibroblasts attached and proliferated on NDGA treated collagen normally. NDGA-fibers did not elicit a foreign body response nor did they stimulate an immune reaction during six weeks in vivo. The fibers survived 6 weeks with little evidence of fragmentation or degradation. The polymerization scheme described here produces a fiber-reinforced NDGA-polymer with mechanical properties approaching an elastic solid. The strength, stiffness and fatigue properties of the NDGA-treated fibers are comparable to those of tendon. These fibers are biocompatible with tendon fibroblasts and elicit little rejection or antigenic response in vivo. These results indicate that NDGA polymerization may provide a viable approach for producing collagenous materials that can be used to bridge gaps in ruptured or lacerated tendons. The tendon-like properties of the NDGA-fiber would allow early mobilization after surgical repair. We predict that timely loading of parted tendons joined by this novel biomaterial will enhance mechanically driven production of neo-tendon by the colonizing fibroblasts and result in superior repair and rapid return to normal properties.     

A novel fibrocartilaginous<Emphasis Type="Italic"> tendon</Emphasis> from an elasmobranch fish (<Emphasis Type="Italic">Rhinoptera bonasus</Emphasis>)

Tendons of the jaw adductor muscles of a hard prey crushing stingray exhibit similar adaptations to compressive and shear loads as those seen in mammalian tendons. Ventral intermandibular tendon from the cownose ray, Rhinoptera bonasus, has a prominent fibrocartilaginous pad that lies between a fibrous region of the tendon and the mineralized tissue of the jaw. Histologically the pad is similar to the fibrocartilaginous meniscus of mammals, and these tissues also share some biochemical traits. Proteoglycan (PG) content in the fibrocartilaginous pad is nearly four times higher than in the linearly arrayed tendinous tissue. The predominant PGs appear to be an aggrecan-like molecule and a decorin-like molecule. The decorin-like molecule is quite small when compared to mammalian decorin (20-80 kDa vs. 100-200 kDa). This study is the first to document adaptations to compressive/shear loading in tendon from a cartilaginous fish, and the similarities to the mammalian condition argue for the early evolution of this reactive ability of tendinous tissue.     

Programmable mechanical stimulation influences tendon homeostasis in a bioreactor system

Identification of functional programmable mechanical stimulation (PMS) on tendon not only provides the insight of the tendon homeostasis under physical/pathological condition, but also guides a better engineering strategy for tendon regeneration. The aims of the study are to design a bioreactor system with PMS to mimic the in vivo loading conditions, and to define the impact of different cyclic tensile strain on tendon. Rabbit Achilles tendons were loaded in the bioreactor with/without cyclic tensile loading (0.25 Hz for 8 h/day, 0–9% for 6 days). Tendons without loading lost its structure integrity as evidenced by disorientated collagen fiber, increased type III collagen expression, and increased cell apoptosis. Tendons with 3% of cyclic tensile loading had moderate matrix deterioration and elevated expression levels of MMP‐1, 3, and 12, whilst exceeded loading regime of 9% caused massive rupture of collagen bundle. However, 6% of cyclic tensile strain was able to maintain the structural integrity and cellular function. Our data indicated that an optimal PMS is required to maintain the tendon homeostasis and there is only a narrow range of tensile strain that can induce the anabolic action. The clinical impact of this study is that optimized eccentric training program is needed to achieve maximum beneficial effects on chronic tendinopathy management. Biotechnol. Bioeng. 2013; 110: 1495–1507. © 2012 Wiley Periodicals, Inc.     

The influence of fiber orientation on the equilibrium properties of neutral and charged biphasic tissues

Constitutive models facilitate investigation into load bearing mechanisms of biological tissues and may aid attempts to engineer tissue replacements. In soft tissue models, a commonly made assumption is that collagen fibers can only bear tensile loads. Previous computational studies have demonstrated that radially aligned fibers stiffen a material in unconfined compression most by limiting lateral expansion while vertically aligned fibers buckle under the compressive loads. In this short communication, we show that in conjunction with swelling, these intuitive statements can be violated at small strains. Under such conditions, a tissue with fibers aligned parallel to the direction of load initially provides the greatest resistance to compression. The results are further put into the context of a Benninghoff architecture for articular cartilage. The predictions of this computational study demonstrate the effects of varying fiber orientations and an initial tare strain on the apparent material parameters obtained from unconfined compression tests of charged tissues.     

Age-dependent influence of strain rate on the tensile failure of rat-tail tendon

Sensitivity of tensile strength, failure strain, and failure energy density to strain rate was studied for rat-tail tendon (RTT), a collagen-rich connective tissue. Tendons from animals aged 1-27 months were stretched at a high (720 percent/s) and low (3.6 percent/s) strain rate. Each failure parameter increased with strain rate. However, the sensitivity of tendon failure to rate of strain decreased rapidly during growth and sexual maturation of the animal. The study provides basic data on the rate-sensitive strength of collagen fibers using RTT.     

PDGF-BB, IGF-I and mechanical load stimulate DNA synthesis in avian tendon fibroblasts in vitro

Resident cells in the surface epitenon and internal compartment of flexor tendons are subjected to cyclic mechanical load as muscle contracts to move limbs or digits. Tendons are largely tensile load bearing tissues and are highly matrix intensive with nondividing cells providing maintenance functions. However, when an injury occurs, tendon cells are stimulated to divide by activated endogenous growth factors and those from platelets and plasma. We hypothesize that tendon cells detect mechanical load signals but do not interpret such signals as mitogenic unless an active growth factor is present. We have used an in vitro mechanical load model, application of cyclic strain to cells cultured on flexible bottomed culture plates, to test the hypothesis that tendon cells require platelet-derived growth factor (PDGF-BB) and insulin-like growth factor-I (IGF-I) in addition to mechanical load to stimulate DNA synthesis. In addition, we demonstrate that in avian tendon cells, load and growth factors stimulate phosphorylation of tyrosine residues in multiple proteins, including pp₆₀src, a protein kinase that phosphorylates receptor protein tyrosine kinases. A lack of mitogenic responsiveness to mechanical load alone by tendon cells may be a characteristic of a regulatory pathway that modulates cell division.     

Actin stress fibres and cell-cell adhesion molecules in tendons: organisation in vivo and response to mechanical loading of tendon cells in vitro.

Tendons consist of parallel longitudinal rows of cells separated by collagen fibres. The cells are in intimate contact longitudinally within rows, and laterally via sheet-like lateral cell processes between rows. At points of contact, they are linked by gap junctions. Since tendons stretch under load, such cell contacts require protection. Here we describe the organisation of the actin cytoskeleton and actin-based cell-cell interactions in vivo and examine the effect of cyclic tensile loading on tendon cells in vitro. Cells within longitudinal rows contained short longitudinally running actin stress fibres. Each fibre was aligned with similar fibres in the cells longitudinally on either side, and fibres appeared to be linked via adherens junctions. Overall, these formed long oriented rows of stress fibres running along the rows of tendon cells. In culture, junctional components n-cadherin and vinculin and the stress fibre component tropomyosin increased in strained cultures, whereas actin levels remained constant. These results suggest that: (1) cells are linked via actin-associated adherens junctions along the line of principal strain; and (2) under load, cells appear to attach themselves more strongly together, and assemble more of their cytoplasmic actin into stress fibres with tropomyosin. Taken together, this suggests that cell-cell contacts are protected during stretch, and also that the stress fibres, which are contractile, may provide an active mechanism for recovery from stretch. In addition, stress fibres are ideally oriented to monitor tensile load and thus may be important in mechanotransduction and the generation of signals passed via the gap junction network.     

Structural and Ultrastructural Characteristics of Bone-Tendon Junction of the Calcaneal Tendon of Adult and Elderly Wistar Rats

Tendons are transition tissues that transfer the contractile forces generated by the muscles to the bones, allowing movement. The region where the tendon attaches to the bone is called bone-tendon junction or enthesis and may be classified as fibrous or fibrocartilaginous. This study aims to analyze the collagen fibers and the cells present in the bone-tendon junction using light microscopy and ultrastructural techniques as scanning electron microscopy and transmission electron microscopy. Forty male Wistar rats were used in the experiment, being 20 adult rats at 4 months-old and 20 elderly rats at 20 months-old. The hind limbs of the rats were removed, dissected and prepared to light microscopy, transmission electron microscopy and scanning electron microscopy. The aging process showed changes in the collagen fibrils, with a predominance of type III fibers in the elderly group, in addition to a decrease in the amount of the fibrocartilage cells, fewer and shorter cytoplasmic processes and a decreased synthetic capacity due to degradation of the organelles involved in synthesis.     

Tribological characteristics of healthy tendon

Tendons transfer muscular forces efficiently and painlessly, facilitating joint motion. Whilst the tribology of articular cartilage is constantly explored, a poorer understanding remains of tendon lubrication and friction. This study reports experimental data describing the tribological characteristics of tendon and its surrounding tissue, before presenting an arithmetic solution to facilitate numerical modelling. The experimental characteristics of the tensile (i.e. mid-substance) and compressive (i.e. fibrocartilaginous) regions of bovine flexor tendon were investigated using a pin-on-plate tribometer, with immunofluroscence analysis describing the relative intensity and distribution of surface-bound lubricin. Arithmetic analysis considering the digital extensor tendon determined that, in physiological conditions, the tensile tendon region was able to generate elastohydrodynamic lubrication (EHL). The equivalent region of compressive tendon exhibited a higher intensity of surface-bound lubricin which, it is hypothesised, serves to minimise the increased frictional resistance due to generating only mixed or boundary lubrication regimes. Arithmetic analysis indicates that, given a more favourable biomechanical environment, this region can also generate EHL. Whilst acknowledging the limitations of transferring data from an animal model to a clinical environment, by providing the first data and equations detailing the film thicknesses and lubrication regime for these two tendon regions it is hoped that clinicians, engineers and scientists can consider improved clinical strategies to tackle both tendinopathy and tendon rupture.     

Collagen fiber orientation at the tendon to bone insertion and its influence on stress concentrations

The tendon to bone insertion serves the mechanical role of transferring loads from a relatively compliant tendon to a relatively rigid bone. The details of the mechanism of load transfer are of great importance, since current surgical procedures for tendon reattachment have high failure rates. We hypothesized that the microscopic structure of the insertion is optimized to minimize stress concentrations associated with this load transfer. To explore this, collagen fiber orientation distributions were measured in the supraspinatus tendons of rats. The angular deviation of fibers was fairly uniform across the insertion, and the mean angles of the local distributions deviated mildly from the tendon axis. To explore how these observed property distributions could influence load transfer, these distributions were used to derive material properties for an idealized two-dimensional mechanical model of an insertion. Comparison between stress concentrations in this idealized model and those in three comparison models suggests that the microstructure serves to (1) simultaneously reduce stress concentrations and material mass, and (2) shield the insertion's outward splay from the highest stresses.     

Allometry of the leg muscles of birds

The musculoskeletal components of the hindlimbs of 20 species of birds, considered non-runners, were examined. Allometry was used to compare these data with previously published information on the limbs of running birds. In non-runners the digital flexor muscle and tendon areas scaled approximately isometrically, in contrast to running birds where tendon areas had a lower exponent. Non-running birds had muscle fibre areas approximately half that of runners of equal mass. In both groups, the muscle:tendon area ratio for m. gastrocnemius increased as M^0.13, suggesting factors other than elastic energy storage are important. Runners exhibited relatively longer tibiotarsi and tarsometatarsi and shorter toes. With very few exceptions, the linear dimensions of bones, tendon cross-sectional areas, and muscle masses and fibre areas in the legs of the non-running birds scaled closely according to the requirements for geometric similarity, but the confidence limits are often wide. Deviations from geometric similarity in birds reflect differences in locomotor behaviours and abilities.     

From head to tail: the myoseptal system in basal actinopterygians

Experimental studies indicated that myomeres play several functional roles during swimming. Some of the functions in question are thought to change rostrocaudally, e.g., anterior myomeres are thought to generate forces, whereas posterior myomeres are thought to transmit forces. In order to determine whether these putative functions are reflected in myoseptal morphology we carried out an analysis of the myoseptal system that includes epaxial and hypaxial myosepta of all body regions for the first time. We combined clearing and staining, microdissections, polarized light microscopy, SEM technique, and length measurements of myoseptal parts to reveal the spatial arrangement, collagen fiber architecture, and rostrocaudal gradients of myosepta. We included representatives of the four basal actinopterygian clades to evaluate our findings in an evolutionary and in a functional context. Our comparison revealed a set of actinopterygian groundplan features. This includes a set of specifically arranged myoseptal tendons (epineural, epipleural, lateral, and myorhabdoid tendons) in all epaxial and postanal hypaxial myosepta. Only preanal hypaxial myosepta lack tendons and exclusively consist of mediolateral fibers. Laterally, myosepta generally align with the helically wound fibers of the dermis in order not to limit the body's maximum curvature. Medially, the relationship of myosepta to vertebrae clearly differs from a 1:1 relationship: a myoseptum attaches to the anterior margin of a vertebra, turns caudally, and traverses at least three vertebrae in an almost horizontal orientation in all body regions. By this arrangement, horizontal multiple layers of myosepta are formed along the trunk dorsal and ventral to the horizontal septum. Due to their reinforcement by epineural or epipleural tendons, these multiple layers are hypothesized to resist the radial expansion of underlying muscle fibers and thus contribute to modulation of body stiffness. Rostrocaudally, a dorsoventral symmetry of epaxial and hypaxial myosepta in terms of spatial arrangement and collagen fiber architecture is gradually developed towards the postanal region. Furthermore, the rostrocaudal extension of myosepta measured between anterior and posterior cones gradually increases. This myoseptal region is reinforced by longitudinal fibers of lateral tendons. Furthermore, the percentage of connective tissue in a cross section increases. These morphological data indicate that posterior myosepta are equipped for multisegmental force transmission towards the caudal fin. Anteriormost myosepta have reinforced and elongated dorsal posterior cones. They are adequately designed to transmit epaxial muscular forces to the neurocranium in order to cause its elevation during suction feeding.     

Organization of collagen bundles during tendon healing in rats treated with L-NAME

The Achilles tendon can support high tension forces and may experience lesions. The damaged tissue does not regenerate completely, with the organization and mechanical properties of the repaired tendon being inferior to those of a healthy tendon. Nitric oxide (NO) plays an important role in wound repair. We have examined the structural reorganization and repair in Achilles tendon after injury in rats treated with the NO synthase inhibitor L-NAME. The right Achilles tendon of male Wistar rats was partially transected. One group of rats was treated with L-NAME (~300 mg/kg per day, given in drinking water) for 4 days prior to tendon sectioning and throughout the post-operative period. Control rats received water without L-NAME. The tendons were excised at 7, 14, and 21 days post-injury and used to quantify hydroxyproline and for mechanical tests. Tendons were also processed for histomorphological analysis by polarized light microscopy, which showed that the collagen fibers were disorganized by day 7 in non-treated and L-NAME-treated rats. In non-treated rats, the organization of the extracellular matrix was more homogeneous by days 14 and 21 compared with day 7, although this homogeneity was less than that in normal tendon. In contrast, in injured tendons from L-NAME-treated rats, the collagen fibers were still disorganized on day 21. Tendons from treated rats had more hydroxyproline but lower mechanical properties compared with those from non-treated rats. Thus, NO modulates tendon healing, with a reduction in NO biosynthesis delaying reorganization of the extracellular matrix, especially collagen. T.C.T. and W.R.N were supported by studentships from CAPES, and S.H. was supported by a research fellowship from Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq).     

Evidence for receptor nerve endings in tendons and related tissues of an electric teleost, Gnathonemus petersii

Tendons and muscle aponeurosis associated with muscle lodged in the mental appendage of a weakly electric mormyrid fish, Gnathonemus petersii, have been examined for nerve terminations in them, using methods of silver impregnation, histology and electron microscopy. Large myelinated afferent nerve fibres innervate the tendinous tissues and form nerve terminals in the perimysium, the muscle-tendinous junctions and the tendons, the last recalling the tendon organs of higher vertebrates. Nerve terminals are characterized by loss of myelin, Schwann sheath and basal lamina, the neurolemma being freely exposed to either fibroblasts and their laminar processes or the collagen fibres and the tendon intercellular matrix. Nerve terminals lack synaptic specialisation of their membrane. This study is the first demonstration of sensory nerve terminals (tension receptors) associated with tendons and related tissue in a teleost fish.     

The myoseptal system in Chimaera monstrosa: collagenous fiber architecture and its evolution in the gnathostome stem lineage

Recent studies have revealed the 3D morphology and collagen fiber architecture of myosepta in teleostome fishes. Here we present the first data set on the myoseptal structure of a representative of the chondrichthyan clade. We investigate the series of myosepta in the ratfish Chimaera monstrosa (Holocephali) from the anterior to the posterior body using microdissections of cleared and stained specimens, polarized light microscopy of excised myosepta, and histology. The features of the myoseptal system of Chimaera are compared to data from closely related vertebrate groups and are mapped onto a phylogenetic tree to further clarify the characteristics of the myoseptal series in the gnathostome ancestor. The 3D morphology and collagen fiber architecture of the myoseptal series in C. monstrosa resembles that of Teleostomi (Actinopterygii+Sarcopterygii) with regard to several features. Our comparative analysis reveals that some of them have evolved in the gnathostome stem lineage. (1) A series of epineural and epaxial lateral tendons (LTs) along the whole body, and a series of epipleural and hypaxial LTs in the postanal region evolved in the gnathostome stem lineage. (2) The LTs increase in length towards the posterior body (three-fold in Chimaera). Data on Chimaera and some comparative data on actinopterygian fishes indicate that LTs also increase in thickness towards the posterior body, but further data are necessary to test whether this holds true generally. (3) Another conspicuous apomorphic gnathostome feature is represented by multi-layer structures of myosepta. These are formed along the vertebral column by converging medial regions of successive sloping parts of myosepta. (4) The dorsalmost and ventralmost flanking parts of myosepta bear a set of mediolaterally oriented collagen fibers that are present in all gnathostomes but are lacking in outgroups. Preanal hypaxial myosepta are clearly different from epaxial myosepta and postanal hypaxial myosepta in terms of their collagen fiber architecture. In Chimaera, preanal hypaxial myosepta consist of an array of mediolaterally oriented collagen fibers closely resembling the condition in other gnathostome groups and in petromyzontids. Only one series of tendons, the myorhabdoid tendons of the flanking parts of myosepta, have evolved in the stem lineage of Myopterygii (Gnathostomata+Petromyzontida). Similar to LTs, the tendons of this series also increase in length towards the posterior body. In combination with other studies, the present study provides a framework for the design of morphologically based experiments and modeling to further address the function of myosepta and myoseptal tendons in gnathostomes.     

The role of tenascin-C in adaptation of tendons to compressive loading

Although most tendon regions are subjected primarily to high tensile loads, selected regions, primarily those that directly contact bones that change the direction of the tendon, must withstand high compressive loads as well. Compressed tendon regions differ from regions subjected to primarily tensile loads: they have a fibrocartilaginous structure with spherical cells surrounded by a matrix containing aggrecan and collagen types I and II, in contrast regions not exposed to compression have a fibrous structure with spindle shaped fibroblasts surrounded by a matrix of dense, longitudinally oriented type I collagen fibrils. The spherical shape of cells in fibrocartilagenous regions indicates these cells are more loosely attached to the matrix than their spindle-shaped counterparts in fibrous regions, a feature that may help to minimize cell deformation during tendon compression. We hypothesized that expression of tenascin-C, an anti-adhesive protein, is part of the adaptation of tendon cells to compression that helps establish and maintain fibrocartilaginous regions. To test this hypothesis we compared tenascin-C content and expression in compressed (distal) versus uncompressed (proximal) segments of bovine flexor tendons. Immunohistochemistry and immunoblot analyses showed that tenascin-C content was increased in the distal tendon where it co-distributed with type II collagen and aggrecan. Tendon cells from the distal segments expressed more tenascin-C than did cells from the proximal segments for up to four days in cell culture, indicating that increased tenascin-C expression is a relatively stable feature of the distal cells. These observations support the hypothesis that tenascin-C expression is a cellular adaptation to compression that helps establish and maintain fibrocartilagenous regions of tendons.