Tendon cell array isolation reveals a previously unknown fibrillin-2-containing macromolecular assembly

Within tendon, between collagen fascicles, cells are organized in linear arrays surrounded by a specialized environment of extracellular matrix (ECM) proteins that are largely unidentified. Our goal was to identify interfascicular, pericellular ECM components and provide additional resolution to the organization of the pericellular matrix. To this end, we employed a combination of enzymatic digestion, mechanical disruption, and differential sedimentation to demonstrate for the first time that it possible to liberate living linear tendon cell arrays from whole tendon. Here, we identify type VI collagen, versican, and fibrillin-2 as components of the immediate pericellular ECM of linearly arrayed tendon cells. Additionally, a unique fibrillin-2-containing macromolecular assembly is described in detail for the first time. This new structure is unlike any previously described fibrillin-containing macromolecular assembly. Having a largely constant diameter, it runs axially along tendon cell arrays and can exceed 1000 microm in length.     

Neutralisation of TGF beta or binding of VLA-4 to fibronectin prevents rat tendon adhesion following transection

Following tendon injury, severe loss of function often occurs either as a result of obliteration of the synovial canal with fibrous scar tissue or from rupture of the repaired tendon. The role of cell engineering in tendon repair is to promote strong and rapid healing of tendon whilst at the same time facilitating rapid reconstitution of the synovial canal. Modification of the immediate inflammatory response around healing tendon has been found to be of value. Experimentally this has been achieved by neutralisation of transforming growth factor-beta over the first 3 days following injury, or by blockade of inflammatory cell binding to the CS-1 locus on fibronectin with an anti-VLA-4 antibody, or with the synthetic VLA-4 inhibitor, CS-1 peptide, in a rat model of tendon transection. It is concluded from this pilot study that the treatments described hold promise in improving outcomes of the common clinical problem of tendon injury in man.     

Structural modifications involved in the fore- and hind limb grip of some flying foxes (Chiroptera: Pteropodidae)

A mechanism which enables flying foxes to lock their hind limb digits and thumbs in flexion is described. The deep digital tendons of the hind limbs have roughened fibrocartilage surfaces. The adjacent flexor tendon sheath supports ridges which interact with the rough tendon surface, temporarily 'locking' the two structures together. This tendon locking mechanism is of importance as it enables bats to reduce the energetic cost of hanging from branches. It does this by reducing, or eliminating, the need for digital flexor muscle activity.     

Trapdoor pulley repair for chronic subluxation of the digital extensor tendon

A case of chronic subluxation of the digital extensor tendon is reported, and a new surgical technique for repair is described. A partial-thickness trapdoor three-sided flap is made out of the dorsal joint capsule, brought through an incision in the ulnar sagittal band, and sutured back down. This centers the tendon and creates a new, stable tunnel through which it may glide.     

Tendons from non-diabetic humans and rats harbor a population of insulin-producing, pancreatic beta cell-like cells

Diabetes mellitus is a risk factor for various types of tendon disorders. The mechanisms underlying diabetes associated tendinopathies remain unclear, but typically, systemic factors related to high blood glucose levels are thought to be causally involved. We hypothesize that tendon immanent cells might be directly involved in diabetic tendinopathy. We therefore analyzed human and rat tendons by immunohistochemistry, laser capture microdissection, and single cell PCR for pancreatic β-cell associated markers. Moreover, we examined the short term effects of a single injection of streptozotocin, a toxin for GLUT2 expressing cells, in rats on insulin expression of tendon cells, and on the biomechanical properties of Achilles tendons. Tendon cells, both in the perivascular area and in the dense collagenous tissue express insulin and Glut2 on both protein and mRNA levels. In addition, glucagon and PDX-1 are present in tendon cells. Intraperitoneal injection of streptozotocin caused a loss of insulin and insulin mRNA in rat Achilles tendons after only 5 days, accompanied by a 40% reduction of mechanical strength. In summary, a so far unrecognized, extrapancreatic, insulin-producing cell type, possibly playing a major role in the pathophysiology of diabetic tendinopathy is described. In view of these data, novel strategies in tendon repair may be considered. The potential of the described cells as a tool for treating diabetes needs to be addressed by further studies.     

Moment arm of the patellar tendon in the human knee

The moment arm of the knee-extensor mechanism is described by the moment arm of the patellar tendon calculated with respect to the screw axis of the tibia relative to the femur. The moment arm may be found once the line of action of the patellar tendon and the position and orientation of the screw axis are known. In this study, the orientation of the patellar tendon and the position and orientation of the finite screw axis of the tibia relative to the femur were calculated from measurements of the three-dimensional positions of the bones obtained from fresh cadaver specimens. Peak values of the patellar tendon moment arm ranged from 4-6 cm for the six knees tested; the moment arm was maximum near 45 degrees of knee flexion. The moment arm of the patellar tendon was nearly equal to the shortest (perpendicular) distance between the line of action of the patellar tendon and the axis of rotation of the knee at all flexion angles, except near full extension. Near full extension, the angle between the patellar tendon and the screw axis was significantly less than 90 degrees, and the magnitude of the moment arm was then less than the perpendicular distance between these two lines. The patellar tendon moment arm remained roughly constant across individuals when normalized by femoral condyle width, suggesting that anatomical differences play a large role in determining the moment arm of the extensor mechanism.     

The tensile and stress relaxation responses of human patellar tendon varies with specimen cross-sectional area.

In order to provide insight into the mechanical response of the collagen fascicle structures in tendon, a series of constant strain rate and constant displacement, stress relaxation mechanical tests were performed on sequentially sectioned human patellar tendon specimens (protocol 1) and specimens with both small (approximately 1 mm2) and large (approximately 20 mm2) cross-sectional areas (protocol 2). These data described the stress relaxation and constant strain rate tensile responses as a function of cross-sectional area and water content. The experimental data suggested that small portions of tendon exhibit a higher tensile modulus, a slower rate of relaxation and a lower amount of relaxation in comparison to larger specimens from the same location in the same tendon. The decrease in relaxation response and the increase in tensile modulus with decreasing cross-sectional area was nonlinear. These data suggest that there may be structures other than the subfascicle, such as the epitenon and other connective tissue components, which influence the tensile and stress relaxation responses in tendon.     

Insertions and functions of certain flexor muscles in the hind leg of rodents

This survey includes 58 genera of rodents from 26 families. The medial tarsal bone is probably unique to the order. Its presence, nature, and constant relationship with M. tibialis posterior are discussed. This muscle inverts and supinates the pes at the astragulo-navicular joint and moves the ankle. The M. flexor tibialis inserts on the medial sesamoid, on this sesamoid and the integument, on the sesamoid and the tendon of M. flexor fibularis, on the latter tendon only, or on the integument only. The occurrence, nature, and cam-like action of the sesamoid are described. A distal segment of the tendon of M. flexor tibialis usually extends from the sesamoid to either the first phalanx of the first digit or to fascia of an adjacent muscle. Functions of the medial sesamoid include (1) stabilization of the tendon of M. flexor tibialis, (2) deflection of this tendon to benefit flexion of the first phalanx, (3) winching of the medial tarsal ligament to flex the first metatarsal, (4) control of the angle of insertion of the tendon to provide flexion or abduction of the first digit as appropriate during swimming, (5) mechanical multiplication of the tension in the tendon between the segments proximal and distal to the sesamoid, and (6) longitudinal folding of the sole of the pes to grip the substrate, as in climbing.     

Evolution of the digital tendon locking mechanism in bats and dermopterans: A phylogenetic perspective

A tendon locking mechanism (TLM) in the digits of the feet has been described previously only in bats and birds. In bats, this mechanism typically consists of a patch of tuberculated fibrocartilage cells on the plantar surface of the proximal flexor tendons, and a corresponding plicated portion of the adjacent flexor tendon sheath. The two components mesh together like parts of a ratchet, locking the digit in a flexed position until the mechanism is disengaged. This system apparently allows bats to hang for long periods of time with reduced muscular activity. In this study, we document for the first time the presence of a similar tendon lock in dermopterans, an occurrence that provides additional support for the hypothesis that dermopterans and bats are sister taxa. The present work also includes observations on the morphology of the digital tendon system in chiropteran species not previously examined, including members of the Craseonycteridae, Mystacinidae and Kerivoulinae. Unlike other bats that have a TLM,Craseonycteris andKerivoula have a plicated proximal tendon sheath but lack distinct tubercles on the flexor tendon. This condition may be related to small body size or may represent an evolutionary intermediate between the presence of a well-developed TLM and the complete absence of this structure. Phyllostomids apparently lack the ratchet-like TLM typical of other bats, instead exhibiting modifications of the tendon sheath that may contribute to its function as a friction lock. Consideration of the distribution of TLM structures in the context of previous phylogenetic hypotheses suggests that a ratchet-type tendon lock was lost and reexpressed at least once and perhaps several times within Microchiroptera. The friction lock is an autapomorphy of Phyllostomidae.     

Understanding cellular and molecular mechanisms of pathogenesis of diabetic tendinopathy

There is accumulating evidence of an increased incidence of tendon disorders in people with diabetes mellitus. Diabetic tendinopathy is an important cause of chronic pain, restricted activity, and even tendon rupture in individuals. Tenocytes and tendon stem/progenitor cells (TSPCs) are the dominant cellular components associated with tendon homeostasis, maintenance, remodeling, and repair. Some previous studies have shown alterations in tenocytes and TSPCs in high glucose or diabetic conditions that might cause structural and functional variations in diabetic tendons and even accelerate the development and progression of diabetic tendinopathy. In this review, the biomechanical properties and histopathological changes in diabetic tendons are described. Then, the cellular and molecular alterations in both tenocytes and TSPCs are summarized, and the underlying mechanisms involved are also analyzed. A better understanding of the underlying cellular and molecular pathogenesis of diabetic tendinopathy would provide new insight for the exploration and development of effective therapeutics.     

Determination of biomechanical characteristics of restrictive adhesions and of functional impairment after flexor tendon surgery: a methodological study of rabbits.

Formation of restrictive adhesions is one of the main obstacles in rehabilitation following hand surgery. Most experimental work, however, involves only a macroscopic and/or histologic evaluation of the amount of adhesions, and their functional characteristics are poorly described. The aim of this study was to develop an experimental technique for characterization of the biomechanical properties of the finger-tendon unit. An instrument was developed for continuous and simultaneous recording of tensile load, tendon excursion and angular rotation in the distal interphalangeal joint of rabbit digits. Utilizing this instrument, it was revealed that the first 50 degrees of flexion required virtually no tensile load either in unoperated digits or immediately after tenorrhaphy. Thereafter, the load required to obtain further flexion was progressively increased. The strength of adhesions, determined 2 weeks after tenorrhaphy, was best expressed as the maximum tensile load recorded before 50 degrees of flexion was reached. This measurement could also be used to register the strength of the tendon repair and to detect partial tendon rupture during the measurement. The technique allows both adequate measurements of the strength of the adhesions and of the tendon gliding ability after flexor tendon surgery.     

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.     

Determination of the compressive material properties of the supraspinatus tendon

A methodology was developed for determining the compressive properties of the supraspinatus tendon, based on finite element principles. Simplified three-dimensional models ure re reated based on anatomical thickness measurements of unloaded supraspinatus tendons over 15 points. The tendon material was characterized as a composite structure of' longitudinally arranged collagen fibers within an extrafibrillar matrix. The matrix was formulated as a hyperelastic material described by the Ogden form of the strain energy potential. The hyperelastic material parameters were parametrically manipulated until the analytical load-displacement results were similar to the results obtaizned from indentation testinrg. In the geometrically averaged tendon, the average ratio of experimental to theoretical maximum indentation displacement was 1.00 (SD: 0.01). The average normalization of residuals was 2.1 g (SD: 0.9 g). Therefjore, the compressive material properties of the supraspinatus tendo'n extrafibrillar matrix were adequately derived with a first-order hyperelastic formulation. The initial comnpressive elastic modulus ranged from 0.024 to 0.090 MPa over the tendon surface and increased nonlinearly with additional compression. Using these material properties, the stresses induced during acromional impingement can be analyzed.     

An ultrastructural investigation of muscle attachment in the opercular filament of a polychaete annelid

The ultrastructure of peduncle muscle attachment to the cuticular flange in the opercular filament of the serpulid Pomatoceros lamarckii Quatrefages is described. The cuticular flange is composed of layers of orthogonally arranged fibres. Specialized epithelial cells (tendon cells) and a collagenous matrix intervene between the peduncle muscles and the cuticular flange. The tendon cells are characterized by hemidesmosomes at both apical and basal ends, connected by thick bundles of tonofilaments. Apically long specialized microvilli from the tendon cells penetrate the cavities in the orthogonally arranged layers of fibres of the cuticular flange. The basal surfaces of the tendon cells and the terminal ends of the peduncle muscles anchor independently of one another in the collagenous matrix. The peduncle muscles appear to be smooth muscles which contain thin filaments, 5 nm in diameter, and thick filaments, 40-100 nm in diameter, with a faint axial periodicity 12-14 nm. The method of peduncle muscle attachment in the opercular filament is compared with those of other invertebrates.     

Use of extensor pollicis longus tendon as a distal extension for an opponens transfer.

We describe the use of the extensor pollicis longus tendon as an extension of the motor unit in an opponens transfer. Its advantages, and its successful application in 4 cases, are illustrated. To the best of our knowledge, this method has not been described before. We believe it to be a satisfactory addition to the other methods described.     

Migration of radio-opaque markers injected into tendon grafts: a study using roentgen stereophotogrammetric analysis (RSA)

An increase in anterior laxity following reconstruction of the anterior cruciate ligament (ACL) can result from lengthening of the graft construct either at the sites of fixation and/or between the sites of fixation (i.e., graft substance). Roentgen stereophotogrammetric analysis (RSA), which requires that radio-opaque markers be attached to the graft, has been shown to be a useful technique in determining lengthening in these regions. Previous methods have been used for attaching radio-opaque markers to the graft, but they all have limitations particularly for single-loop grafts. Therefore, the objective of this study was to evaluate injecting markers directly into the substance of a tendon as a viable method for measuring lengthening of single-loop graft constructs by determining the maximum amount of migration after cyclic loading. Tantalum spheres of 0.8 mm diameter were used as tendon markers. Ten single-loop tendon grafts were passed through tibial tunnels drilled in calf tibias and fixed with a tibial fixation device. Two tendon markers were inserted in one tendon bundle of each graft and the grafts were cyclically loaded for 225,000 cycles from 20 N to 170 N. At specified intervals, simultaneous radiographs were obtained of the tendon markers. Marker migration was computed as the change in distance between the two tendon markers parallel to the axis of the tibial tunnel. Marker migration had a root mean square (RMS) value of less than 0.1 mm. Because the RMS value indicates the error introduced into measurements of lengthening and because this error is negligible, the method described for attaching markers to single-loop ACL grafts has the potential to be useful for determining lengthening of single-loop ACL graft constructs in in vivo studies in humans.     

Mechanism for the regulation of post-translational modifications of procollagens synthesized by matrix-free cells from chick embryos

Three possible mechanisms are considered to account for the variations of post-translational modifications in different collagen types. 1) The cells have different amounts of post-translational modifying enzymes, 2) the rate of prolylhydroxylation of different procollagen types is varied, and 3) the rate of chain association of pro-alpha chains of different collagen types is modulated. In an attempt to examine the three possibilities, we have determined the activities of prolyl hydroxylase and lysyl hydroxylase, and we have examined the kinetics of the secretion of procollagens and the kinetics of pro-gamma chain formation of different procollagen types in matrix-free cells isolated from tissues of 17-day-old chick embryos. Type II collagen synthesized by cartilage cells contains more hydroxylysine than type I collagen synthesized by tendon and cornea cells. It was found, however, that cartilage cells contain significantly less lysyl hydroxylase than tendon and cornea cells. In contrast, we found only a small difference in the amount of prolyl hydroxylase in tendon, cornea, and cartilage cells. The secretion of type I procollagen by tendon and cornea cells can be described by two first order processes. In contrast, the secretion of type II procollagen by cartilage cells, type IV procollagen by lens cells, and type V procollagen by cornea cells can be described by single first order processes. Examination of the formation of pro-gamma components of procollagen types I and II revealed that it occurs via intermediate dimers of two pro-alpha chains. The formation or pro-gamma(I) chains in tendon and cornea cells is about three times faster than the formation of pro-gamma(II) chains in cartilage cells. These results are consistent with the hypothesis that the rate of association of pro-alpha chains regulates the synthesis of procollagens with different degrees of post-translational modifications.     

Chick myotendinous antigen. I. A monoclonal antibody as a marker for tendon and muscle morphogenesis

Extracellular matrix components are likely to be involved in the interaction of muscle with nonmuscle cells during morphogenesis and in adult skeletal muscle. With the aim of identifying relevant molecules, we generated monoclonal antibodies that react with the endomysium, i.e., the extracellular matrix on the surface of single muscle fibers. Antibody M1, which is described here, specifically labeled the endomysium of chick anterior latissimus dorsi muscle (but neither the perimysium nor, with the exception of blood vessels and perineurium, the epimysium ). Endomysium labeling was restricted to proximal and distal portions of muscle fibers near their insertion points to tendon, but absent from medial regions of the muscle. Myotendinous junctions and tendon fascicles were intensely labeled by M1 antibody. In chick embryos, " myotendinous antigen" (as we tentatively call the epitope recognized by M1 antibody) appeared first in the perichondrium of vertebrae and limb cartilage elements, from where it gradually extended to the premuscle masses. Around day 6, tendon primordia were clearly labeled. The other structures labeled by M1 antibody in chick embryos were developing smooth muscle tissues, especially aorta, gizzard, and lung buds. In general, tissues labeled with M1 antibody appeared to be a subset of the ones accumulating fibronectin. In cell cultures, M1 antibody binds to fuzzy, fibrillar material on the substrate and cell surfaces of living fibroblast and myogenic cells, which confirms an extracellular location of the antigenic site. The appearance of myotendinous antigen during limb morphogenesis and its distribution in adult muscle and tendon are compatible with the idea that it might be involved in attaching muscle fibers to tendon fascicles. Its biochemical characterization is described in the accompanying paper ( Chiquet , M., and D. Fambrough , 1984, J. Cell Biol. 98:1937-1946).     

Analysis of tendinous actuation in balancing the maximal fingertip force for normal and abnormal forefinger system

This work displayed the force capabilities of the musculoskeletal system of the forefinger under external loading. Different states of normal and pathological fingers are studied. We evaluated the impact of losing musculo-tendon unit strength capacities in terms of maximal output fingertip force and tendon tensions distribution. A biomechanical model for a static force analysis is developed through anatomical and kinematic studies. An optimisation approach is then used to determine tendon tension distribution when performing an isometric task. Furthermore, pathological fingers with common cases of injured flexors and extensors are analysed. The method of simulation for forefinger abnormities is described. Furthermore, the simulation results are interpreted.     

Development of collagenous fibres in autologous and preserved homologous tendon grafts.

Collagen synthesis has been studied in preserved autologous and homologous tendon grafts, applied in dogs. The stages of collagen morphogenesis observed in the synthetizing fibroblasts appearing in the decomposing original structure of the transplants as well as in the extracellular ground substance are described. The results suggest that autologous tendon grafts and homologous ones preserved by irradiation, stored in cold physiological saline and beta-propiolactone, are rebuilt within eight weeks; meanwhile the original collagenous fibre structure is decomposed. When other preserving procedures such as deep-freezing or gamma irradiation are applied recomposition of the graft is retarded or fails to occur; the result is a cicatrizing adhesion.