Palaeohistology of the bones of pterosaurs (Reptilia: Archosauria): anatomy, ontogeny, and biomechanical implications

Thin sections from long bones of specimens representing pterosaurs ranging from the Early Jurassic to the latest Cretaceous provide a profile of bone histology across a range of sizes, skeletal elements, growth stages, and phylogenetic positions. Most pterosaur bone is fibro-lamellar, organized in an unusual way that suggests high growth rates through ontogeny. Fibro-lamellar deposits are finished by a relatively abrupt deceleration or cessation of growth represented by lamellar, poorly vascularized subperiosteal bone in what appear to be adults. Pterosaurs had the thinnest bone walls of any tetrapods; they complemented high rates of periosteal deposition with almost equally high rates of endosteal erosion. Pterosaurs show a great variety of histologic features that include articular calcified cartilage, sub-chondral bone plates, trabecular bone struts and related internal supports, and secondary deposition and remodeling of bone. They remodeled their bones internally by (1) depositing endosteal bone coatings on the inner cortex and over struts of pre-existing internal bone, (2) secondarily filling bone spaces, and (3) Haversian reworking. The construction of these struts reflects both developmental patterns of bone construction and biomechanical function. Alternating plywood-like layers of bone, heretofore undescribed in tetrapods, provided strength, as did the obliquely oriented system of reticular blood vessels in the bones. The distribution and ontogenetic features of pterosaur bone tissues, when combined with other evidence, suggest generally high growth rates, high metabolic levels, altricial birth, and extended parental care.     

Sequencing three crocodilian genomes to illuminate the evolution of archosaurs and amniotes

The International Crocodilian Genomes Working Group (ICGWG) will sequence and assemble the American alligator (Alligator mississippiensis), saltwater crocodile (Crocodylus porosus) and Indian gharial (Gavialis gangeticus) genomes. The status of these projects and our planned analyses are described.     

Development of an image-based technique to examine joint congruency at the elbow

Identifying joint contact in articular joints is important for both the biomechanical investigation of joint mechanics and the study of osteoarthritis. The purpose of this study is to develop a proximity mapping technique to non-invasively determine joint congruency, as a surrogate of joint contact. To illustrate the capabilities of this algorithm, a cadaveric upper extremity was positioned at varying degrees of elbow flexion. This technique was validated using a gold standard experimental casting technique. The pattern of the cast showed an excellent agreement with the generated proximity map using the inter-bone distance algorithm. The results from this study agree with the results of previous studies examining joint contact at the elbow both in the location and in the tracking of the joint contact throughout elbow flexion. Ultimately, this technique will lead to an increased understanding of the effect of malalignment and instability of the joint on contact mechanics.     

Effect of surface roughness on squeeze film poroelastic bearings with special reference to synovial joints

A simplified mathematical model has been developed for understanding the combined effects of surface roughness and couple stresses on lubrication aspects of synovial joints. The modified Reynolds equation which incorporates the elastic as well as randomized surface roughness structure of cartilage with couple-stress fluid as lubricant is derived. The mean pressure, load carrying capacity and time of approach as functions of film thickness during normal articulation of joints are obtained by using Christensen stochastic theory with the assumption that the roughness asperity heights are to be small compared to the film thickness. The effects of surface roughness and elasticity are considerably pronounced for the poroelastic bearings with couple-stress fluid as lubricant compared with classical case.     

Biomechanical implications of degenerative joint disease in the apophyseal joints of human thoracic and lumbar vertebrae

An experimental technique for quantifying load-sharing in cadaveric spines is used to test the hypothesis that degenerative changes in human apophyseal joints are directly related to high levels of compressive load-bearing by these joints. About 36 cadaveric thoraco-lumbar motion segments aged 64-92 years were subjected to a compressive load of 1.5 kN. The distribution of compressive stress was measured in the intervertebral discs using a miniature pressure transducer, and stress measurements were summed over area to give the compressive force resisted by the disc. This was subtracted from the applied 1.5 kN to indicate compressive load-bearing by the apophyseal joints. The cartilage of each apophyseal joint surface was then graded for degree of degeneration. After maceration, each joint surface was scored for degenerative joint disease (DJD) affecting the bone. Results demonstrated that the apophyseal joints resisted 5-96% (mean 45%) of the applied compressive force. A significant positive correlation was demonstrated between age and cartilage degeneration, age and DJD bone score, apophyseal joint load-bearing and bone score, and cartilage score and load-bearing. The latter correlation was strongest for load-bearing above 50%. Ordinal regression showed that the variables describing bone DJD (marginal osteophytes, pitting, bony contour change, and eburnation) were significantly correlated with degree of cartilage degeneration. It is concluded that in elderly individuals apophyseal joint load-bearing above a threshold of 50% is associated with severe degenerative changes in cartilage and bone, and that markers of DJD observed palaeopathologically may be used as predictors of such loadingin life.     

Phenotypic disparity of the elbow joint in domestic dogs and wild carnivores*

In this article, I use geometric morphometrics in 2D from a sample of 366 elbow joints to quantify phenotypic disparity in domestic dog breeds, in wild canids, and across the order Carnivora. The elbow joint is a well‐established morphological indicator of forearm motion and, by extension, of functional adaptations toward locomotor or predatory behavior in living carnivores. The study of the elbow joint in domestic dogs allows the exploration of potential convergences between (i) pursuit predators and fast‐running dogs, and (ii) ambush predators and fighting breeds. The results indicate that elbow shape disparity among domestic dogs exceeds than in wolves; it is comparable to the disparity of wild Caninae, but is significantly lower than the one observed throughout Canidae and Carnivora. Moreover, fast‐running and fighting breeds are not convergent in elbow joint shape with extreme pursuit and ambush wild carnivores, respectively. The role of artificial selection and developmental constraints in shaping limb phenotypic disparity through the extremely fast evolution of the domestic dog is discussed in the light of this new evidence.     

Tissue engineering of cartilage using a mechanobioreactor exerting simultaneous mechanical shear and compression to simulate the rolling action of articular joints

The effect of dynamic mechanical shear and compression on the synthesis of human tissue‐engineered cartilage was investigated using a mechanobioreactor capable of simulating the rolling action of articular joints in a mixed fluid environment. Human chondrocytes seeded into polyglycolic acid (PGA) mesh or PGA–alginate scaffolds were precultured in shaking T‐flasks or recirculation perfusion bioreactors for 2.5 or 4 weeks prior to mechanical stimulation in the mechanobioreactor. Constructs were subjected to intermittent unconfined shear and compressive loading at a frequency of 0.05 Hz using a peak‐to‐peak compressive strain amplitude of 2.2% superimposed on a static axial compressive strain of 6.5%. The mechanical treatment was carried out for up to 2.5 weeks using a loading regime of 10 min duration each day with the direction of the shear forces reversed after 5 min and release of all loading at the end of the daily treatment period. Compared with shaking T‐flasks and mechanobioreactor control cultures without loading, mechanical treatment improved the amount and quality of cartilage produced. On a per cell basis, synthesis of both major structural components of cartilage, glycosaminoglycan (GAG) and collagen type II, was enhanced substantially by up to 5.3‐ and 10‐fold, respectively, depending on the scaffold type and seeding cell density. Levels of collagen type II as a percentage of total collagen were also increased after mechanical treatment by up to 3.4‐fold in PGA constructs. Mechanical treatment had a less pronounced effect on the composition of constructs precultured in perfusion bioreactors compared with perfusion culture controls. This work demonstrates that the quality of tissue‐engineered cartilage can be enhanced significantly by application of simultaneous dynamic mechanical shear and compression, with the greatest benefits evident for synthesis of collagen type II. Biotechnol. Bioeng. 2012; 109:1060–1073. © 2011 Wiley Periodicals, Inc.     

In vitro action of combinations of selected antimicrobial agents and adult bovine articular cartilage (sesamoid bone)

Anatomically intact articular cartilage in form of sesamoid bones from metacarpophalangeal joints of 2-year-old cows was tested for its influence on the microbicidal effect of the iodophore Betaisodona, the bispyridinamine Octenisept, and the biguanide Lavasept. Comparisons were carried out in Ham's F12 medium with and without 0.2% bovine serum albumin as organic matter loading. The expected abolition of the microbicidal effect of these antiseptics against the test organisms Escherichia coli or Staphylococcus aureus in the presence of sesamoid bone was not evident. Furthermore, sesamoid bone alone demonstrated antibacterial activity against Staphylococcus aureus, which may involve adherence of bacteria to surface constituents of articular cartilage. Final concentrations of 2.5-5% Betaisodona, 5% Octenisept as well as 0.025% Lavasept are effective in killing of 10(8)-10(9) cfu/ml Escherichia coli or Staphylococcus aureus in the presence of sesamoid bone without the reduction of antimicrobial activity expected from binding to CS, which has previously been demonstrated for CS in solution.     

Comparison of two methods for calculating the frictional properties of articular cartilage using a simple pendulum and intact mouse knee joints

In attempts to better understand the etiology of osteoarthritis, a debilitating joint disease that results in the degeneration of articular cartilage (AC) in synovial joints, researchers have focused on joint tribology, the study of joint friction, lubrication, and wear. Several different approaches have been used to investigate the frictional properties of articular cartilage. In this study, we examined two analysis methods for calculating the coefficient of friction (μ) using a simple pendulum system and BL6 murine knee joints (n=10) as the fulcrum. A Stanton linear decay model (Lin μ) and an exponential model that accounts for viscous damping (Exp μ) were fit to the decaying pendulum oscillations. Root mean square error (RMSE), asymptotic standard error (ASE), and coefficient of variation (CV) were calculated to evaluate the fit and measurement precision of each model. This investigation demonstrated that while Lin μ was more repeatable, based on CV (5.0% for Lin μ; 18% for Exp μ), Exp μ provided a better fitting model, based on RMSE (0.165° for Exp μ; 0.391° for Lin μ) and ASE (0.033 for Exp μ; 0.185 for Lin μ), and had a significantly lower coefficient of friction value (0.022±0.007 for Exp μ; 0.042±0.016 for Lin μ) (p=0.001). This study details the use of a simple pendulum for examining cartilage properties in situ that will have applications investigating cartilage mechanics in a variety of species. The Exp μ model provided a more accurate fit to the experimental data for predicting the frictional properties of intact joints in pendulum systems.     

Division and death of cells in developing synovial joints and long bones

Articular chondrocytes are a unique set of cells from the time the cellular condensations that become the anlagen of the long bones develop in the embryo. In the presumptive joint the cells of the opposing bones are packed very closely together, but at cavitation, the central, flattened cells move apart to form the articular surfaces. As the articular cartilage develops the cells are pushed further apart by the cartilaginous matrix. To determine the contributions of cell proliferation and death to cavitation and the subsequent development and growth of articular cartilage, direct observations were made to identify mitotic cells and those with apoptotic bodies in haematoxylin-stained sections of developing joints, and growing and ageing articular cartilage of the rabbit knee. These observations were extended using antibodies to the proliferating cell nuclear antigen (PCNA) and TdT-mediated dUTP nick end labelling (TUNEL) on corresponding sections. Low levels of cell division do occur in the articular cartilage up to 6 weeks postnatally, but matrix formation makes the major contribution to the increase in size of the cartilage. Cell death is not observed during cavitation, nor during the development of the articular cartilage proper. Apoptosis is essential, however, for the removal of the epiphyseal cartilage during ossification of the epiphyses and in the growth plate.     

The 'fabrosaurid' ornithischian dinosaurs of the Upper Elliot Formation (Lower Jurassic) of South Africa and Lesotho

The Upper Elliot Formation of South Africa and Lesotho contains the world's most diverse fauna of early Jurassic ornithischian dinosaurs. Nevertheless, despite four decades of work on this fauna there remains significant taxonomic confusion and many important specimens remain undescribed. A review of the non-heterodontosaurid ('fabrosaurid') ornithischians of the Upper Elliot Formation is presented, following re-examination of all known ornithischian material from the Elliot Formation. ' Fabrosaurus australis' is based upon a single undiagnostic dentary, and is here considered a nomen dubium . Lesothosaurus diagnosticus is considered to be valid and is rediagnosed based upon a unique combination of plesiomorphic and derived characteristics. Stormbergia dangershoeki gen. et. sp. nov. is described from three partial skeletons including numerous postcranial material. Stormbergia dangershoeki is significantly larger than previously described Elliot Formation ornithischians, and can be recognized on the basis of a unique combination of characters, the most important of which is the possession of a distinctive tab-shaped obturator process on the ischium. A preliminary systematic analysis is presented, the results of which differ significantly from other recent ornithischian phylogenies.  © 2005 The Linnean Society of London, Zoological Journal of the Linnean Society , 2005, 145 , 175–218.     

The internal mechanical functioning of intervertebral discs and articular cartilage,and its relevance to matrix biology

Degeneration of intervertebral discs and articular cartilage can cause pain and disability. Risk factors include genetic inheritance and age, but mechanical loading also is important. Its influence has been investigated using miniature pressure transducers to measure the distribution of compressive stress (force per unit area) within loaded tissue. The technique quantifies stress concentrations, and detects regions that behave in a fluid-like manner.Intervertebral discs demonstrate a central fluid-like region which normally extends beyond the anatomical nucleus pulposus so that the whole disc functions like a “water bed”. With increasing age, the fluid region shrinks and pressure within it falls. Stress concentrations appear in the surrounding anulus fibrosus, with location depending on posture. Stress concentrations become large in degenerated discs, and are intensified by sustained loading or injury. Articular cartilage never exhibits an internal fluid pressure: stress gradients and concentrations normally occur within it, and are intensified by sustained loading.Excessive matrix stresses can cause pain and progressive damage. They also inhibit matrix synthesis and stimulate production of matrix-degrading enzymes. In this way, injury to chondroid tissues can initiate a ‘vicious circle’ of abnormal matrix stresses, abnormal metabolism, weakened matrix, and further injury, which explains many features of their degeneration.     

Palaeohistology and external microanatomy of rauisuchian osteoderms (Archosauria: Pseudosuchia)

Abstract: The presence of postcranial dermal armour is plesiomorphic for Archosauria. Here, we survey the external microanatomy and histology of postcranial osteoderms (i.e. dorsal paramedian and caudal osteoderms) of rauisuchians, a widely distributed assemblage of extinct predatory pseudosuchians from the Triassic. The osteoderms of eight rauisuchian taxa were found to be rather compact bones, which usually lack significant bone remodelling or large areas of cancellous bone. The presence of highly vascularized woven or fibrolamellar bone tissue deposited in the core areas indicates higher growth rates during earlier life stages, whereas a more compact parallel‐fibred bone matrix indicates reduced growth rates in later development. This pattern of change corroborates earlier studies on long bone histology. With the exception of a bone tissue found in the sample of Batrachotomus kupferzellensis, which might be the result of metaplastic ossification, the general mode of skeletogenesis is comparable with intramembraneous ossification. The lack of cancellous bone tissue and remodelling processes associated with bone ornamentation, as well as the predominantly intramembraneous mode of ossification, indicates that rauisuchian osteoderm formation differs profoundly from that of the osteoderms of the only extant pseudosuchian lineage, the crocodylians.     

Esophageal cancer related gene 4 (ECRG4) is a marker of articular chondrocyte differentiation and cartilage destruction

With the aim of identifying novel genes regulating cartilage development and degeneration, we screened a cartilage-specific expressed sequence tag database. Esophageal cancer related gene 4 (ECRG4) was selected, based on the criteria of ‘chondrocyte-specific’ and ‘unknown function.’ ECRG4 expression was particularly abundant in chondrocytes and cartilage, compared to various other mouse tissues. ECRG4 is a secreted protein that undergoes cleavage after secretion. The protein is specifically expressed in chondrocytes in a manner dependent on differentiation status. The expression is very low in mesenchymal cells, and dramatically increased during chondrogenic differentiation. The ECRG4 level in differentiated chondrocytes is decreased during hypertrophic maturation, both in vitro and in vivo, and additionally in dedifferentiating chondrocytes induced by interleukin-1β or serial subculture, chondrocytes of human osteoarthritic cartilage and experimental mouse osteoarthritic cartilage. However, ectopic expression or exogenous ECRG4 treatment in a primary culture cell system does not affect chondrogenesis of mesenchymal cells, hypertrophic maturation of chondrocytes or dedifferentiation of differentiated chondrocytes. Additionally, cartilage development and organization of extracellular matrix are not affected in transgenic mice overexpressing ECRG4 in cartilage tissue. However, ectopic expression of ECRG4 reduced proliferation of primary culture chondrocytes. While the underlying mechanisms of ECRG4 expression and specific roles remain to be elucidated in more detail, our results support its function as a marker of differentiated articular chondrocytes and cartilage destruction.     

Effect of exercise on bone and articular cartilage in heterozygous manganese superoxide dismutase (SOD2) deficient mice

Abstract

Reactive oxygen species (ROS) are involved in both bone and cartilage physiology and play an important role in the pathogenesis of osteoporosis and osteoarthritis. The present study investigated the effect of running exercise on bone and cartilage in heterozygous manganese superoxide dismutase (SOD2)-deficient mice. It was hypothesized that exercise might induce an increased production of ROS in these tissues. Heterozygous SOD2-deficient mice should exhibit an impaired capability to compensate, resulting in an increased oxidative stress in cartilage and bone. Thirteen female wild type and 20 SOD2^+/? mice (aged 16 weeks) were randomly assigned to a non-active wild type (SOD2^+/+Con, n = 7), a trained wild type (SOD2^+/+Run, n = 6), a non-active SOD2^+/? (SOD2^+/?Con, n = 9) and a trained SOD2^+/? (SOD2^+/?Run, n = 11) group. Training groups underwent running exercise on a treadmill for 8 weeks. In SOD2^+/? mice elevated levels of 15-F_2t-isoprostane and nitrotyrosine were detected in bone and articular cartilage compared to wild type littermates. In osteocytes the elevated levels of these molecules were found to be reduced after exercise while in chondrocytes they were increased by aerobic running exercise. The observed changes in oxidative and nitrosative stress did neither affect morphological, structural nor mechanical properties of both tissues. These results demonstrate that exercise might protect bone against oxidative stress in heterozygous SOD2-deficient mice.     

The trackmaker of Apatopus (Late Triassic, North America): implications for the evolution of archosaur stance and gait

Abstract:  For some decades, a major focus of research has been on how locomotor modes changed in some archosaurian reptiles from a more or less 'sprawling' to an 'erect' posture, whether there were discrete intermediate stages, and how many times 'erect' posture evolved. The classic paradigm for the evolution of stance and gait in archosaurs, a three-stage transition from sprawling to 'semi-erect' to erect posture, has been replaced by a subtler understanding of a continuum of changing limb joint angles. We suggest a further separation of terminology related to stance vs. gait so as not to entail different processes: 'sprawling' and 'erect' should refer to continua of stance; 'rotatory' and 'parasagittal' are more appropriate ends of a continuum that describes the motions of gait. We show that the Triassic trackway Apatopus best fits the anatomy and proportions of phytosaurs, based on a new reconstruction of their foot skeleton; it is less likely to have been made by another pseudosuchian or non-archosaurian archosauromorph. Moreover, the trackmaker was performing the high walk. A phytosaurian trackmaker would imply that the common ancestor of pseudosuchians, and therefore archosaurs could approximate the high walk (depending on phylogeny), and if so, erect stance and parasagittal gait did not evolve independently in pseudosuchians and ornithosuchians, although the kinematic mechanisms differed in the two groups. It remains to be seen how far outside Archosauria, if at all, more or less erect posture and parasagittal gait may have evolved.