Muscle Attachment Sites Research Articles

Inferring internal anatomy from the trilobite exoskeleton: the relationship between frontal auxiliary impressions and the digestive system

Lerosey-Aubril, R., Hegna, T.A. & Olive, S. 2011: Inferring internal anatomy from the trilobite exoskeleton: the relationship between frontal auxiliary impressions and the digestive system. Lethaia, Vol. 44, pp. 166–184. The digestive system of trilobites is rarely preserved. As a result, many aspects of its organization remain unknown. Fortunately, the exoskeleton sometimes preserves evidence of soft-tissue attachment sites that can be used to infer internal anatomy. Among them are the frontal auxiliary impressions (FAIs), probable soft-tissue insertion sites located on the fronto-median glabellar lobe of some trilobites. FAIs are herein described in the Carboniferous trilobite Phillipsia belgicaOsmolska 1970– representing the only known example of such structures in the Proetida and their youngest occurrence. A taphonomic scenario is proposed to explain their variable preservation. Although particularly common in the Phacopina, FAIs or FAI-like structures are also found in several orders that differ greatly. Comparisons with modern analogues suggest that FAIs might represent attachment sites for extrinsic muscles associated with a differentiated crop within the foregut. A review of purported remains of the trilobite digestive system indicates that it usually consisted of a tube-like tract flanked by a variable number of metamerically paired diverticulae. Its anterior portion is not particularly individualized, except in a few specimens that might hint at the presence of a crop. This differentiation of a crop might have constituted a secondarily evolution of the foregut in trilobites, occurring independently in different clades. Accompanied by a strengthening of associated extrinsic muscles, this modification of the foregut might explain the presence of more conspicuous muscle insertion sites on the glabella. Study of FAIs might therefore provide new data on the anatomy of the foregut in trilobites and evidence of diverse feeding habits. □Arthropoda, digestive system, ecology, muscle scars, Proetida, Trilobita.

Morphometric pelvic muscle map analysis in extant and pleistocene carnivores

Recent studies in our laboratory have developed maps of pelvic musculature attachment sites (muscle maps) for Neofelis nebulosa (clouded leopard), Felis catis (domestic cat) and >Canis latrans (coyote). Correlations were made between actual muscle attachments and bone morphology at the attachment site. Based on this information we can map muscle attachment sites on bones from extinct closely related species. Pleistocene pelvis specimens were examined for bony muscle attachment sites, measured and photographed at the George C. Page Museum in Los Angeles, California. Specimens included Smilodon fatalis (n=9), Panthera atrox (n=4), Canis dirus (n=6), Canis latrans (n=5), and Arctodus simus (n=2). Muscle map surface areas for eight pelvic muscles were determined for each specimen along with pelvic measurements (ilium/ischium: width/length). The measurements were then compared between and within extant and extinct species listed above. Mm. gluteus medius, quadratus femoris, cranial and caudal gemellus surface areas differed significantly in comparisons between the felids, and between felids and non‐felid species in both extinct and extant groups. Pelvic measurements differed primarily between felid and non‐felids. These results suggest that muscle map surface area analysis may be useful for both phylogenetic and morphometric analysis.

Muscles Underlying the Elevation of the Hyolaryngeal Complex

IntroductionA critical event in the pharyngeal phase of swallowing is the elevation of the hyolaryngeal complex opening the upper esophageal sphincter. Current swallowing theory assigns this function to the suprahyoid (mylohyoid, geniohyoid, digastrics, and stylohyoid) and thyrohyoid muscles. The role of the long pharyngeal muscles (stylopharyngeus, palatopharyngeus, and salpingopharyngeus) that insert into the thyroid cartilage and distal pharynx is uninvestigated in humans. This study uses a cadaver model to document the structural properties of muscles to analyze their potential for elevating the hyolaryngeal complex.MethodsMeasurements to determine physiological cross‐sectional areas (PCSAs) of suprahyoid, thyrohyoid and long pharyngeal muscles were taken from hemisected head and neck formalin‐fixed cadaver specimens (n = 14). Coordinates of muscle attachment sites and PCSAs were used to calculate unit force vectors representing the relative maximum tetanic tension muscles in the superior direction. Force vectors of muscle samples were compared by an analysis of variance to evaluate their potential for force in elevating the hyolaryngeal complex.ResultsAn ANOVA with Tukey HSD post hoc analysis of unit force vectors in the superior direction showed the following results: suprahyoid (1.54±0.34 cm2) > long pharyngeal (.74±0.20 cm2) (p<.01), and long pharyngeal > thyrohyoid (.49±0.17 cm2) (p<.05).DiscussionThe morphology of these muscles suggests that while the suprahyoid muscles have the most potential for force in elevating the hyolaryngeal complex, the long pharyngeal muscles contribute significantly more to UES opening than the thyrohyoid. If verified by physiological data, this data would add to the current understanding of swallowing.Grant Funding Source: n/a

A central multifunctional role of integrin-linked kinase at muscle attachment sites

Integrin-linked kinase (ILK) is an essential component of a multiprotein complex that links actin to the plasma membrane. Here, we have used a genetic approach to examine the molecular interactions that are essential for the assembly of this ILK-containing complex at Drosophila muscle attachment sites (MASs). We show that, downstream of integrins, talin plays a decisive role in the recruitment of three proteins: ILK, PINCH and paxillin. The accumulation of ILK at MASs appears to follow an amplification mechanism, suggesting that numerous binding sites are generated by minimal levels of the upstream integrin and talin effectors. This property suggests that ILK functions as an essential hub in the assembly of its partner proteins at sites of integrin adhesion. We found that PINCH stability, and its subcellular localization at MASs, depends upon ILK function, but that ILK stability and localization is not dependent upon PINCH. An in vivo structure-function analysis of ILK demonstrated that each ILK domain has sufficient information for its independent recruitment at embryonic MASs, whereas at later developmental stages only the kinase domain was effectively recruited. Our data strengthen the view that the ILK complex is assembled sequentially at sites of integrin adhesion by employing multiple molecular interactions, which collectively stabilize the integrin-actin link.

Shape and size of the body vs. musculoskeletal stress markers

The objective of this paper is to assess the relationship between the degree of development of muscle attachment sites (musculoskeletal stress markers - MSM1) and the length and circumference measurements of long bones and the body build expressed with the reconstructed values of body height (BH) and body mass (BM). The bone material (102 male and 99 female skeletons) used in the study was collected in the medieval burial ground in Cedynia, Poland. The authors analyzed 10 musculoskeletal stress markers located on the scapula (2), humerus (2), radius (2), femur (2) and tibia (2). The frequency and the degree of expression of muscle attachment size was carried out using the scale prepared by Myszka (2007). The scale encompassed three degrees of expression of muscle attachment size. Only changes of robusticity type (nonpathological changes) were taken into account. The assessment of body build of individuals was carried out according to the method proposed by Vancata & Charvátová (2001). Body height was reconstructed from the length of the humerus and femur using eight equations. Body mass was reconstructed from the measurements of the breadth of the proximal and distal sections of the femur and tibia (mechanical method) using twenty one equations. The equations were developed for different reference populations. The same equations were used for men and women. The correlation between the MSM and the length and circumference measurements of the bones was analyzed using the principal components analysis and the Gamma correlation coefficient. The strength of the correlation between the reconstructed body build traits (BH, BM) and the moderate degree of musculoskeletal stress markers expression was studied based on the principal components method and the Pearson correlation coefficient. A linear correlation was found between musculoskeletal stress markers and the circumference measurements and the reconstructed body mass, but no relationship with body height and the length measurements of long bones was revealed. From previous research it is evident that the relationship between the MSM and metric skeletal traits does not occur in every population. Divergent findings necessitate further corroboration of results on diverse skeletal material.

Intertissue Mechanical Stress Affects Frizzled-Mediated Planar Cell Polarity in the Drosophila Notum Epidermis

Frizzled/planar cell polarity (Fz/PCP) signaling controls the orientation of sensory bristles and cellular hairs (trichomes) along the anteroposterior axis of the Drosophila thorax (notum). A subset of the trichome-producing notum cells differentiate as "tendon cells," serving as attachment sites for the indirect flight muscles (IFMs) to the exoskeleton. Through the analysis of chascon (chas), a gene identified by its ability to disrupt Fz/PCP signaling under overexpression conditions, and jitterbug (jbug)/filamin, we show that maintenance of anteroposterior planar polarization requires the notum epithelia to balance mechanical stress generated by the attachment of the IFMs. chas is expressed in notum tendon cells, and its loss of function disturbs cellular orientation at and near the regions where IFMs attach to the epidermis. This effect is independent of the Fz/PCP and fat/dachsous systems. The chas phenotype arises during normal shortening of the IFMs and is suppressed by genetic ablation of the IFMs. chas acts through jbug/filamin and cooperates with MyosinII to modulate the mechanoresponse of notum tendon cells. These observations support the notion that the ability of epithelia to respond to mechanical stress generated by one or more interactions with other tissues during development and organogenesis influences the maintenance of its shape and PCP features.

Muscle parameters for musculoskeletal modelling of the human neck

Background To study normal or pathological neuromuscular control, a musculoskeletal model of the neck has great potential but a complete and consistent anatomical dataset which comprises the muscle geometry parameters to construct such a model is not yet available. Methods A dissection experiment was performed on the left side of one 50th percentile male embalmed specimen. Geometrical data including muscle attachment sites were digitized using an Optotrak measurement system and laser diffraction was used to determine muscle sarcomere lengths. Bony landmarks were recorded and joint centres of rotation between different vertebrae were estimated using literature data. Findings A total of 34 muscle parts of the neck were divided in 129 elements per body side. Muscle attachment sites, mass, physiological cross sectional area, fibre length, tendon length and optimal fibre length for each element are supplied as digital annexes to the paper. Results are coherent with other studies and new data are provided for several smaller muscles not reported elsewhere. Interpretation Implementation of this dataset into a neck model is likely to improve the estimation of muscle forces and thus increase the model validity; this makes future neck models more suitable for the use as clinical tools.

Scientific Basis of Minimally Invasive Spine Surgery

Literature review. To describe the scientific basis of minimally invasive spine surgery as it relates to posterior lumbar surgery. Minimally invasive spine (MIS) surgery is predicated on several basic principles: (1) avoid muscle crush injury by self-retaining retractors; (2) do not disrupt tendon attachment sites of key muscles, particularly the origin of the multifidus muscle at the spinous process; (3) use known anatomic neurovascular and muscle compartment planes; and (4) minimize collateral soft tissue injury by limiting the width of the surgical corridor. Literature review. The conventional midline posterior approach for lumbar decompression and fusion violates these key principles of MIS surgery. The tendon origin of the multifidus muscle is detached, the surgical corridor is exceedingly wide, and significant muscle crush injury occurs through the use of powerful self-retaining retractors. The combination of these events leads to well-described changes in muscle physiology and function. MIS surgery is performed using table-mounted tubular retractors that focus the surgical dissection to a narrow corridor directly over the surgical target site. The path of the surgical corridor is selected on the basis of anatomic planes, specifically avoiding injury to the musculotendinous complex and the neurovascular bundle. With these relatively simple modifications to surgical technique, significant improvements in intraoperative blood loss, postoperative pain, surgical morbidity, return of function, among others, have been achieved. However, MIS techniques remain technically demanding and a significant complication rate has been observed during the initial learning curve of the procedures.

Evaluating the Structural Properties of Suprahyoid Muscles and their Potential for Moving the Hyoid

Superior and anterior hyoid movements are important events in pharyngeal deglutition. This cross-sectional study uses a cadaver model to document the structural properties of the muscles underlying these movements in an effort to understand how their morphology influences function. Measurements to determine physiological cross-sectional areas (PCSAs) of swallowing muscles were taken from hemisected head and neck formalin-fixed cadaver specimens (n = 13). Coordinates of muscle attachment sites and PCSAs were used to calculate î and ĵ unit force vectors, where î and ĵ represent anterior-posterior and superior-inferior directions, respectively. The suprahyoid muscle subsamples were grouped for analysis as follows: digastric (DG), geniohyoid (GH), mylohyoid (MH), and stylohyoid (SH). The ANOVA with Tukey HSD post hoc analysis of unit force vectors showed the following results: GH (-0.44 ± 0.15 cm(2)) >MH (-0.02 ± 0.21 cm(2)), DG (-0.05 ± 0.11 cm(2)), SH (0.14 ± 0.04 cm(2)), with negative values representing the anterior direction (p < 0.01); and MH (0.91 ± 0.28 cm(2)) >DG (0.29 ± 0.14 cm(2)), SH (0.22 ± 0.08 cm(2)), GH (12 ± 0.08 cm(2)), with positive values representing the superior direction (p < 0.01). The morphology of the suprahyoid muscles suggests that based on structural properties, the geniohyoid has the most potential to displace the hyoid in the anterior direction and the mylohyoid has the most potential to displace the hyoid in the superior direction. These data in complement with physiological findings may provide greater insight into these movements for those developing novel treatments for dysphagia.

The masticatory system under varying functional load. Part 2: effect of reduced masticatory load on the degree and distribution of mineralization in the rabbit mandible

A reduction in mechanical loading of the mandible brought about by mastication of soft food is assumed to decrease the remodelling rate of bone, which, in turn, might increase the degree of bone mineralization. The effect of a reduction in masticatory functional load on the degree and distribution of mineralization of mandibular bone was investigated in male juvenile New Zealand White rabbits. The experimental animals (n=8) had been raised on a diet of soft pellets from 8 to 20 weeks of age, while the controls (n=8) had been fed pellets of normal hardness. The degree of mineralization of bone (DMB) was assessed at the attachment sites of various jaw muscles, the condylar head, and the alveolar process. Differences between groups and among sites were tested for statistical significance using a Student's t-test and one-way analysis of variance, respectively. The DMB did not differ significantly between the experimental and control animals at any of the sites assessed. However, in the rabbits that had been fed soft pellets, both cortical bone at the attachment sites of the temporalis and digastric muscles and cortical bone in the alveolar process had a significantly higher DMB than cortical bone at the attachment site of the masseter muscle, while there were no significant differences among these sites in the control animals. The results suggest that a moderate reduction in masticatory functional load does not significantly affect the remodelling rate and the DMB in areas of the mandible that are loaded during mastication but might induce a more heterogeneous mineral distribution.

Dynamic patterns of mechanical stimulation co-localise with growth and cell proliferation during morphogenesis in the avian embryonic knee joint

Muscle contractions begin in early embryonic life, generating forces that regulate the correct formation of the skeleton. In this paper we test the hypothesis that the biophysical stimulation generated by muscle forces may be a causative factor for the changes in shape of the knee joint as it grows. We do this by predicting the spatial and temporal patterns of biophysical stimuli, where cell proliferation and rudiment shape changes occur within the emerging tissues of the joint over time. We used optical projection tomography (OPT) to create anatomically accurate finite element models of the embryonic knee at three time points (stages) of development. OPT was also used to locate muscle attachment sites and AFM was used to determine material properties. An association was found between the emergence of joint shape, cell proliferation and the pattern of biophysical stimuli generated by embryonic muscle contractions. Elevated rates of growth and cell proliferation in the medial condyle were found to co-localise with elevated patterns of biophysical stimuli including maximum principal stresses and fluid flow, throughout the time period studied, indicating that cartilage growth and chondrocyte proliferation in the epiphysis is potentially related to local patterns of biophysical stimuli. The development of the patella and articular cartilages, which is known to be affected by in ovo immobilisation, could be contributed to by specific patterns of fluid flow, pore pressure and stress in the joint interzone. This suggests that both cartilage growth and tissue differentiation in the embryonic joint is regulated by specific patterns of biophysical stimuli and that these stimuli are needed for the correct development of the joint.

Stereotypic boring behaviour inferred from the earliest known octopod feeding traces: Early Eocene, southern England

A bulk sample of 267 disarticulated valves of the bivalve Venericor clarendonensis (Wood) collected from the Lower Eocene London Clay (southern UK) yielded 38 individuals that had been perforated by small drill holes (0.70–2.14 mm in outer diameter). These drill holes had more or less circular plan views, with slightly irregular openings, and taper as they pass through the valve, conforming to the ichnotaxon Oichnus simplex Bromley. They show stereotypic positioning, being concentrated in the posterior region on the prey, moreover there is remarkable preference for perforating the sites of muscle attachment (principally the posterior adductor). We consider the most likely culprits to be octopods. As such these are the oldest octopod drill holes yet recorded. They provide the only evidence of these important top predators in this shallow marine community and also demonstrate that the sophisticated predatory behaviour shown by modern octopods had been evolved by at least the Early Eocene. □Eocene, octopod behaviour, Oichnus, stereotypic boring. Although predator prey interactions have arguably been a very important force in evolution (Vermeij 1987; Stanley 2008), quantitative data from the fossil record are frustratingly difficult to obtain. Many types of predatory techniques (such as swallowing prey whole or prising hardparts open) leave no discernible trace and even where the method used does cause damage to fossilizable hardparts, by crushing and biting, it may be difficult to distinguish such traces from post-mortem breakage or to attribute it to any particular predator. However, one of the more fruitful lines of research has come from the study of predatory drill holes in shelled prey (Leighton & Aronowsky 2003). In the case of drill holes it may be possible identify the potential predator and to collect data concerning aspects of predatory behaviour such as size selection of prey, stereotypic attack positioning and measures of success rates (see Harper 2003 for a review). Here we report on a large sample of an Early Eocene bivalve, Venericor clarendonensis, in which a number of valves have been punctured by distinctive small holes. We present evidence that they are likely to have been made by predatory octopods, which record a remarkable stereotypic behaviour.

Postcranial Functional Morphology of Hyracotherium (Equidae, Perissodactyla) and Locomotion in the Earliest Horses

A nearly complete skeleton of Hyracotherium grangeri is described from the early Wasatchian (early Eocene) of the Clarks Fork Basin in northwestern Wyoming. This specimen includes a virtually complete, well-preserved pre-caudal vertebral column allowing the first thorough investigation of the locomotory abilities of these early Eocene horses. The posterior thorax (T) and lumbus (L) are divided into a facultatively dorsostable region (T15-L3) and an obligately dorsostable region (L3-L7). The facultatively dorsostable region is characterized by robust, cranially-oriented neural spines and well-developed attachment sites for epaxial musculature, whereas the obligately dorsostable region is characterized by embracing zygapophyses and less developed spines and attachment sites. Limb morphology exhibits a mixture of primitive characters and derived features associated with cursoriality. As described by previous authors, limb mobility is restricted at the elbow and ankle due to interlocking humero-radial and tibio-astragalar articulations, respectively. In contrast, the presence of a rounded femoral head and well-developed pelvic and femoral attachment sites for hindlimb adductor muscles suggests a highly mobile hip joint. Limited inversion of the manus and pes may have been possible via differential movement between the proximal and distal carpal elements and at the transverse tarsal joint, respectively. The obligately dorsostable region of the vertebral column likely resisted variably-oriented torque applied by hip musculature, thereby preventing dislocation of the pelvis and lumbus. The facultatively dorsostable region would have facilitated dynamic positioning of the center of mass during rapid acceleration. Brief comparisons with Orohippus and Mesohippus postcrania are used to discuss the selective pressures imposed by changes in body mass and habitat on the postcranial evolution of Paleogene horses.

A novel stress distribution organ in the arthropod exoskeleton

The vertebrate endoskeleton possesses a massive internal network of load-distributing trabeculae that in most locations accounts for the vast majority of bone cross sectional area. In contrast, arthropods rely on the external cuticle and its intermittent outpocketings to distribute the daily stresses of physiological loading. One of the constraints of the arthropod exoskeleton is the necessity to house the musculature involved in locomotion, feeding and etc. Because of this lack of an extensive internal load-distributing trabecular network, any load-distributing mechanism in arthropods would necessarily have to incorporate the exoskeleton. Several authors have identified structural apophysi whose functions presumably have mechanical significance, but few have been identified using quantitative analyses. This study investigates a novel stress-reducing structure arising from the articulation sites in the exoskeleton of the blue crab, Callinectes sapidus. During dissection of the merus-carpus joint and leg cuticle of the blue crab, an unique system of internal strut-like members was found radiating, both longitudinally and laterally, from the articular surface of the proximal merus segment, tapering into the diaphyseal region. This strut system, an internal outpocketing of the exoskeleton and semi-circular in cross section, mirrors the trabecular pattern seen radiating from vertebrate joint surfaces. Earlier reports of this structural system described it as a muscle attachment site and made little or no reference to potential load distribution properties. Finite element analysis (FEA) models confirm the efficacy of stress distributing properties of this articular strut system in the blue crab leg. In the models, the struts significantly reduce stress concentrations, reduce localized strains and minimize the risk of failure via buckling. Models lacking this strut system generate 94.7% larger peak von Mises stress at the articulation site, 37% higher peak displacement and 4% greater equivalent strain. The model with the struts is capable of withstanding an applied physiological load of up to 16.6 N prior to buckling, more than twice that of the model without struts (7.8 N). We suggest that this novel arthropod strut system is likely utilized at many joint surfaces at locations of high skeletal stress concentrations, is an adaptation for minimizing skeletal failure via localized buckling, and may be present in other arthropod taxa.

Degree of Mineralization at the Attachment of Lateral Pterygoid

The degree of mineralization of bone (DMB) in the human mandibular condyle is heterogeneous, and differences in DMB have been related to variations in bone turnover caused by local strains. The lateral pterygoid muscle inserts at the anterior surface of the condyle. The aim of this study is to analyze the DMB at the attachment of this muscle as compared with a control region. It was hypothesized that, DMB at the attachment sites of lateral pterygoid muscles was lower than at the control regions, because of the larger number of loadings and subsequently higher remodeling rates. Also, as the human lateral pterygoid muscle is heterogeneous in its internal architecture, variations in DMB within the attachment sites were expected. 10 human mandibular condyles were scanned in a micro CT system. Within each condyle, two regions, that is, the pterygoid fovea and a posterior (control) region where no muscle was inserted, were selected to analyze regional differences in DMB. The attachment site was further divided into eight subregions to analyze subregional differences. At the pterygoid fovea the DMB of cortical bone was significantly lower than at the control region (p = 0.003) and increased in medio-lateral direction. The results of this study could suggest an influence of the lateral pterygoid muscle on bone turnover at this site.

Expression of intermediate filaments at muscle insertions in human fetuses

Desmin and vimentin are intermediate filaments that play crucial roles in the maturation, maintenance and recovery of muscle fibers and mesenchymal cells. The expression of these proteins has not been investigated extensively in human fetuses. In the present study, we examined the immunohistochemical expression of intermediate filaments in skeletal muscles of the head, neck and thorax in 12 mid-term human fetuses at 9-18 weeks of gestation. We also used immunohistochemistry to localize the expression of the myosin heavy chain and silver impregnation to identify the fetal endomysium. Expression of desmin and vimentin was already detectable in intercostal muscle at 9 weeks, especially at sites of muscle attachment to the perichondrium. At this stage, myosin heavy chain was expressed throughout the muscle fibers and the endomysium had already developed. Beginning with punctate expression, the positive areas became diffusely distributed in the muscle fibers. At 15-18 weeks, intermediate filament proteins were extensively expressed in all of the muscles examined. Expression at the bone-muscle interface was continuous with expression along the intramuscular tendon fibres. These results suggest that the development of intermediate filaments begins in areas of mechanical stress due to early muscle contraction. Their initially punctate distribution, as observed here, probably corresponds to the earliest stage of fetal enthesis formation.

Facioscapulohumeral muscular dystrophy region gene-1 (FRG-1) is an actin-bundling protein associated with muscle-attachment sites

In vertebrates, overexpression of facioscapulohumeral muscular dystrophy (FSHD) region gene 1 (FRG1) recapitulates the pathophysiology exhibited by FSHD patients, although the role of FRG1 in FSHD remains controversial and no precise function for FRG1 has been described in any organism. To gain insight into the function and potential role of FRG1 in FSHD, we analyzed the highly conserved Caenorhabditis elegans ortholog, frg-1. C. elegans body-wall muscles contain two distinct subcellular pools of FRG-1: nuclear FRG-1, concentrated in the nucleoli; and cytoplasmic FRG-1, associated with the Z-disk and costamere-like structures known as dense bodies. Functionally, we demonstrate that FRG-1 is an F-actin-bundling protein, consistent with its localization to dense bodies; this activity is conserved in human FRG1. This is particularly intriguing because it places FRG-1 along side the list of dense-body components whose vertebrate orthologs are involved in the myriad myopathies associated with disrupted costameres and Z-disks. Interestingly, overexpressed FRG-1 preferentially accumulates in the nucleus and, when overexpressed specifically from the frg-1 promoter, disrupts the adult ventral muscle structure and organization. Together, these data further support a role for FRG1 overexpression in FSHD pathophysiology and reveal the previously unsuspected direct involvement of FRG-1 in muscle structure and integrity.

The Rap1 Guanine Nucleotide Exchange Factor C3G Is Required for Preservation of Larval Muscle Integrity in Drosophila melanogaster

C3G is a guanine nucleotide exchange factor (GEF) and modulator of small G-protein activity, which primarily acts on members of the Rap GTPase subfamily. Via promotion of the active GTP bound conformation of target GTPases, C3G has been implicated in the regulation of multiple cellular and developmental events including proliferation, differentiation and apoptosis. The Drosophila C3G orthologue exhibits a domain organization similar to that of vertebrate C3G. Through deletion of the C3G locus, we have observed that loss of C3G causes semi-lethality, and that escaping adult flies are characterized by a reduction in lifespan and general fitness. In situ hybridization reveals C3G expression in the developing embryonic somatic and visceral muscles, and indeed analysis of C3G mutants suggests essential functions of C3G for normal body wall muscle development during larval stages. C3G mutants display abnormal muscle morphology and attachment, as well as failure to properly localize βPS integrins to muscle attachment sites. Moreover, we show that C3G stimulates guanine nucleotide exchange on Drosophila Rap GTPases in vitro. Taken together, we conclude that Drosophila C3G is a Rap1-specific GEF with important functions in maintaining muscle integrity during larval stages.

Ontogenetic change in skull morphology and mechanical advantage in the spotted hyena (Crocuta crocuta)

Weaning represents a challenging transition for young mammals, one particularly difficult for species coping with extreme conditions during feeding. Spotted hyenas (Crocuta crocuta) experience such extreme conditions imposed by intense feeding competition during which the ability to consume large quantities of food quickly is highly advantageous. As adult spotted hyenas have massive skulls specialized for durophagy and can feed very rapidly, young individuals are likely at a competitive disadvantage until that specialized morphology is completely developed. Here we document developmental changes in skull size, shape, and mechanical advantage of the jaws. Sampling an ontogenetic series of Crocuta skulls from individuals ranging in age from 2 months to 18 years, we use linear measurements and geometric morphometrics to test hypotheses suggesting that size, limited mechanical advantage of the jaws, and/or limited attachment sites for jaw muscles might constrain the feeding performance of juveniles. We also examine skull development in relation to key life history events, including weaning and reproductive maturity, to inquire whether ontogeny of the feeding apparatus is slower or more protracted in this species than in carnivores not specialized for durophagy. We find that, although mechanical advantage reaches maturity in hyenas at 22 months, adult skull size is not achieved until 29 months of age, and skull shape does not reach maturity until 35 months. The latter is nearly 2 years after mean weaning age, and more than 1 year after reproductive maturity. Thus, skull development in Crocuta is indeed protracted relative to that in most other carnivores. Based on the skull features that continue to change and to provide additional muscle attachment area, protracted development may be largely due to development of the massive musculature required by durophagy. These findings may ultimately shed light on the adaptive significance of the unusual "role-reversed" pattern of female dominance over males in this species.

Endostatin Affects Osteoblast Behavior In Vitro, but Collagen XVIII/Endostatin Is Not Essential for Skeletal Development In Vivo

Endostatin, a fragment of collagen XVIII, can inhibit vascular endothelial growth factor (VEGF) signaling. VEGF is known to be crucial for bone development. The aims of this study were to investigate the influences of endostatin on osteoblast behavior in vitro and the roles of collagen XVIII/endostatin on bone development in vivo. For the in vitro experiments, MC3T3-E1 osteoblasts were treated with VEGF-A, 2 microg/ml endostatin, 20 microg/ml endostatin, VEGF-A + 2 microg/ml endostatin, or VEGF-A + 20 microg/ml endostatin. Osteoblast proliferation and matrix mineralization were analyzed. Faxitron, pQCT, and histological analyses were performed on hindleg bones of transgenic mice overexpressing endostatin (ES-tg) and mice lacking collagen XVIII (Col18a1 (-/-)) to study bone development in vivo. Treatment of cells with endostatin decreased osteoblast proliferation. Moreover, VEGF-A together with endostatin (2 microg/ml) decreased osteoblast proliferation and matrix mineralization. In vivo, Col18a1 (-/-) and ES-tg mice displayed no differences in bone density or mineral content during bone development, but ES-tg bones grew in length more slowly compared to the controls. The formation of secondary ossification centers was delayed in Col18a1 (-/-) mice. Immunohistochemistry revealed collagen XVIII in basement membranes of periosteal and bone marrow vessels and at muscle attachment sites. In conclusion, endostatin affects osteoblast behavior in vitro, the effects being boosted by simultaneous treatment with VEGF. In vivo, Col18a1 (-/-) and ES-tg mice show mild delays in bone development. These changes are transitory and suggest that collagen XVIII/endostatin does not play an indispensable role in skeletal development.