Grasping convergent evolution in syngnathids: a unique tale of tails.

Seahorses and pipehorses both possess a prehensile tail, a unique characteristic among teleost fishes, allowing them to grasp and hold onto substrates such as sea grasses. Although studies have focused on tail grasping, the pattern of evolutionary transformations that made this possible is poorly understood. Recent phylogenetic studies show that the prehensile tail evolved independently in different syngnathid lineages, including seahorses, Haliichthys taeniophorus and several types of so-called pipehorses. This study explores the pattern that characterizes this convergent evolution towards a prehensile tail, by comparing the caudal musculoskeletal organization, as well as passive bending capacities in pipefish (representing the ancestral state), pipehorse, seahorse and H. taeniophorus. To study the complex musculoskeletal morphology, histological sectioning, µCT-scanning and phase contrast synchrotron scanning were combined with virtual 3D-reconstructions. Results suggest that the independent evolution towards tail grasping in syngnathids reflects at least two quite different strategies in which the ancestral condition of a heavy plated and rigid system became modified into a highly flexible one. Intermediate skeletal morphologies (between the ancestral condition and seahorses) could be found in the pygmy pipehorses and H. taeniophorus, which are phylogenetically closely affiliated with seahorses. This study suggests that the characteristic parallel myoseptal organization as already described in seahorse (compared with a conical organization in pipefish and pipehorse) may not be a necessity for grasping, but represents an apomorphy for seahorses, as this pattern is not found in other syngnathid species possessing a prehensile tail. One could suggest that the functionality of grasping evolved before the specialized, parallel myoseptal organization seen in seahorses. However, as the grasping system in pipehorses is a totally different one, this cannot be concluded from this study.

EDART, a discrete algebraic reconstructing technique for experimental data obtained with high resolution computed tomography.

A novel reconstruction method is presented to improve the image quality of three dimensional (3D) datasets for samples consisting of only one material and surrounding air, obtained with high resolution X-ray Computed Tomography (µCT). It combines discrete tomography with iterative reconstruction algorithms, it is applicable for routine µCT applications and is referred to as the Experimental Discrete Algebraic Reconstruction Technique (EDART). A fast and intuitive method to estimate the attenuation coefficient and segmentation threshold, in case these are unknown, is included. Experimental results illustrate that EDART allows the improvement of the reconstruction quality as compared to standard iterative reconstruction when few projections are available, without significantly increasing the reconstruction time.

In silico Mechanics of Stem Cells Intramyocardially Transplanted with a Biomaterial Injectate for Treatment of Myocardial Infarction.

PURPOSE: Biomaterial and stem cell delivery are promising approaches to treating myocardial infarction. However, the mechanical and biochemical mechanisms underlying the therapeutic benefits require further clarification. This study aimed to assess the deformation of stem cells injected with the biomaterial into the infarcted heart. METHODS: A microstructural finite element model of a mid-wall infarcted myocardial region was developed from ex vivo microcomputed tomography data of a rat heart with left ventricular infarct and intramyocardial biomaterial injectate. Nine cells were numerically seeded in the injectate of the microstructural model. The microstructural and a previously developed biventricular finite element model of the same rat heart were used to quantify the deformation of the cells during a cardiac cycle for a biomaterial elastic modulus (Einj) ranging between 4.1 and 405,900 kPa. RESULTS: The transplanted cells' deformation was largest for Einj = 7.4 kPa, matching that of the cells, and decreased for an increase and decrease in Einj. The cell deformation was more sensitive to Einj changes for softer (Einj ≤ 738 kPa) than stiffer biomaterials. CONCLUSIONS: Combining the microstructural and biventricular finite element models enables quantifying micromechanics of transplanted cells in the heart. The approach offers a broader scope for in silico investigations of biomaterial and cell therapies for myocardial infarction and other cardiac pathologies.

An exploratory study of contrast agents for soft tissue visualization by means of high resolution X-ray computed tomography imaging.

High resolution X-ray computed tomography (CT), or microCT, is a promising and already widely used technique in various scientific fields. Also for histological purposes it has great potential. Although microCT has proven to be a valuable technique for the imaging of bone structures, the visualization of soft tissue structures is still an important challenge due to their low inherent X-ray contrast. One way to achieve contrast enhancement is to make use of contrast agents. However, contrary to light and electron microscopy, knowledge about contrast agents and staining procedures is limited for X-ray CT. The purpose of this paper is to identify useful X-ray contrast agents for soft tissue visualization, which can be applied in a simple way and are also suited for samples larger than (1 cm)(3) . And 28 chemical substances have been investigated. All chemicals were applied in the form of concentrated aqueous solutions in which the samples were immersed. First, strips of green Bacon were stained to evaluate contrast enhancement between muscle and adipose tissue. Furthermore it was also tested whether the contrast agents remained fixed in the tissue after staining by re-immersing them in water. Based on the results, 12 contrast agents were selected for further testing on postmortem mice hind legs, containing a variety of different tissues, including muscle, fat, bone, cartilage and tendons. It was evaluated whether the contrast agents allowed a clearer distinction between the different soft tissue structures present. Finally also penetration depth was measured. And 26 chemicals resulted in contrast enhancement between muscle and adipose tissue in the Bacon strips. Mercury(II)chloride (HgCl2 ), phosphotungstic acid (PTA), phosphomolybdic acid (PMA) and ammonium orthomolybdate ((NH4 )2 MoO4 ) remained fixed after re-immersion in water. The penetration tests showed that potassium iodide (KI) and sodium tungstate can be most efficiently used for large samples of the order of several tens of cm(3) . PMA, PTA, HgCl2 and also to a lesser extent Na2 WO4 and (NH4 )2 MoO4 allowed a clearer distinction between the different soft tissue structures present.

Effect of biomaterial stiffness on cardiac mechanics in a biventricular infarcted rat heart model with microstructural representation of in situ intramyocardial injectate.

Intramyocardial delivery of biomaterials is a promising concept for treating myocardial infarction. The delivered biomaterial provides mechanical support and attenuates wall thinning and elevated wall stress in the infarct region. This study aimed at developing a biventricular finite element model of an infarcted rat heart with a microstructural representation of an in situ biomaterial injectate, and a parametric investigation of the effect of the injectate stiffness on the cardiac mechanics. A three-dimensional subject-specific biventricular finite element model of a rat heart with left ventricular infarct and microstructurally dispersed biomaterial delivered 1 week after infarct induction was developed from ex vivo microcomputed tomography data. The volumetric mesh density varied between 303 mm-3 in the myocardium and 3852 mm-3 in the injectate region due to the microstructural intramyocardial dispersion. Parametric simulations were conducted with the injectate's elastic modulus varying from 4.1 to 405,900 kPa, and myocardial and injectate strains were recorded. With increasing injectate stiffness, the end-diastolic median myocardial fibre and cross-fibre strain decreased in magnitude from 3.6% to 1.1% and from -6.0% to -2.9%, respectively. At end-systole, the myocardial fibre and cross-fibre strain decreased in magnitude from -20.4% to -11.8% and from 6.5% to 4.6%, respectively. In the injectate, the maximum and minimum principal strains decreased in magnitude from 5.4% to 0.001% and from -5.4% to -0.001%, respectively, at end-diastole and from 38.5% to 0.06% and from -39.0% to -0.06%, respectively, at end-systole. With the microstructural injectate geometry, the developed subject-specific cardiac finite element model offers potential for extension to cellular injectates and in silico studies of mechanotransduction and therapeutic signalling in the infarcted heart with an infarct animal model extensively used in preclinical research.

Electromagnetic stimulation to optimize the bone regeneration capacity of gelatin-based cryogels.

One of the key challenges in reconstructive bone surgery is to provide living constructs that possess the ability to integrate in the surrounding host tissue. Bone graft substitutes and biomaterials have already been widely used to heal critical-size bone defects due to trauma, tumor resection and tissue degeneration. In the present study, gelatin-based cryogels have been seeded with human SAOS-2 osteoblasts followed by the in vitro culture of the cells. In order to overcome the drawbacks associated with static culture systems, including limited diffusion and in homogeneous cell-matrix distribution, the present work describes the application of a bioreactor to physically enhance the cell culture in vitro using an electromagnetic stimulus. The results indicate that the physical stimulation of cell-seeded gelatin-based cryogels upregulates the bone matrix production. We anticipate that the scaffolds developed consisting of human bone proteins and cells could be applied for clinical purposes related to bone repair.

An extensin-rich matrix lines the carinal canals in Equisetum ramosissimum, which may function as water-conducting channels.

BACKGROUND AND AIMS: The anatomy of Equisetum stems is characterized by the occurrence of vallecular and carinal canals. Previous studies on the carinal canals in several Equisetum species suggest that they convey water from one node to another. METHODS: Cell wall composition and ultrastructure have been studied using immunocytochemistry and electron microscopy, respectively. Serial sectioning and X-ray computed tomography were employed to examine the internode-node-internode transition of Equisetum ramosissimum. KEY RESULTS: The distribution of the LM1 and JIM20 extensin epitopes is restricted to the lining of carinal canals. The monoclonal antibodies JIM5 and LM19 directed against homogalacturonan with a low degree of methyl esterification and the CBM3a probe recognizing crystalline cellulose also bound to this lining. The xyloglucan epitopes recognized by LM15 and CCRC-M1 were only detected in this lining after pectate lyase treatment. The carinal canals, connecting consecutive rings of nodal xylem, are formed by the disruption and dissolution of protoxylem elements during elongation of the internodes. Their inner surface appears smooth compared with that of vallecular canals. CONCLUSIONS: The carinal canals in E. ramosissimum have a distinctive lining containing pectic homogalacturonan, cellulose, xyloglucan and extensin. These canals might function as water-conducting channels which would be especially important during the elongation of the internodes when protoxylem is disrupted and the metaxylem is not yet differentiated. How the molecularly distinct lining relates to the proposed water-conducting function of the carinal canals requires further study. Efforts to elucidate the spatial and temporal distribution of cell wall polymers in a taxonomically broad range of plants will probably provide more insight into the structural-functional relationships of individual cell wall components or of specific configurations of cell wall polymers.

Development and evaluation of injection-molded sustained-release tablets containing ethylcellulose and polyethylene oxide.

PURPOSE: It was the aim of the present study to develop sustained-release matrix tablets by means of injection molding of ethylcellulose (EC) and polyethylene oxide (PEO) mixtures and to evaluate the influence of process temperature, matrix composition, and viscosity grade of EC and PEO on processability and drug release. METHODS: Formulations consisting of metoprolol tartrate (MPT, concentration: 30%), EC plasticized by dibutyl sebacate, and PEO were extruded and consequently injection molded into tablets. The influence of process temperature (120°C and 140°C), matrix composition, viscosity grade of EC (4, 10, 20, 45, and 100 mPa·s) and PEO (7 × 10(6), 1 × 10(6), and 1 × 10(5) Mw) on processability and drug release was determined. RESULTS: Formulations consisting of 70% EC and 30% MPT showed incomplete drug release, whereas drug release was too fast for formulations without EC. Higher PEO concentrations increased drug release. Formulations containing 30% metoprolol, EC, and different concentrations of PEO showed first-order release rates with limited burst release. Drug release from direct compressed tablets showed faster drug release rates compared to injection-molded formulations. There was no clear relationship between the molecular weight of EC and drug release. The melting endotherm (113.9°C) of MPT observed in the differential scanning calorimeter thermogram of the tablets indicated that a solid dispersion was formed which was confirmed by X-ray diffractogram. X-ray tomography demonstrated a difference in pore structure between tablets processed at 120°C and 140°C. CONCLUSION: It was concluded that injection molding can be applied successfully to develop sustained-release PEO/EC matrix tablets.

Musculoskeletal structure of the feeding system and implications of snout elongation in Hippocampus reidi and Dunckerocampus dactyliophorus.

A thorough morphological description of the feeding apparatus in Hippocampus reidi, a long-snouted seahorse, and Dunckerocampus dactyliophorus, an extremely long-snouted pipefish, revealed specialized features that might be associated with the fast and powerful suction feeding, like the two ligamentous connections between the lower jaw and the hyoid, the saddle joint of the latter with the suspensorium and the vertebro-pectoral fusion that articulates on three points with the cranium. Despite the conserved morphology of the feeding apparatus, it was found that in H. reidi the orientation of the occipital joint is ventrocaudal, the sternohyoideus and epaxial muscles are more bulky and both have a short tendon. In D. dactyliophorus, on the other hand, the protractor hyoidei muscle is enclosed by the mandibulo-hyoid ligament, the sternohyoideus and epaxial tendons are long and a sesamoid bone is present in the latter. These features were compared to other syngnathid species with different snout lengths to evaluate the implications of snout elongation on the musculoskeletal structure of the cranium. The arched path of the adductor mandibulae and the greater rigidity of the lower jaw might be related to elongation of the snout, as it yields an increased mechanical advantage of the lower jaw system and a reduced torque between the elements of the lower jaw during protractor hyoidei muscle contraction, respectively. Nevertheless, most observed features did not seem to be related to snout length, but might be associated with different force-generating strategies.

A LabVIEW® based generic CT scanner control software platform.

UGCT, the Centre for X-ray tomography at Ghent University (Belgium) does research on X-ray tomography and its applications. This includes the development and construction of state-of-the-art CT scanners for scientific research. Because these scanners are built for very different purposes they differ considerably in their physical implementations. However, they all share common principle functionality. In this context a generic software platform was developed using LabVIEW® in order to provide the same interface and functionality on all scanners. This article describes the concept and features of this software, and its potential for tomography in a research setting. The core concept is to rigorously separate the abstract operation of a CT scanner from its actual physical configuration. This separation is achieved by implementing a sender-listener architecture. The advantages are that the resulting software platform is generic, scalable, highly efficient, easy to develop and to extend, and that it can be deployed on future scanners with minimal effort.

A new preparation method to study fresh plant structures with X-ray computed tomography.

Since the development of X-ray computed tomography as a medical diagnostic tool, it was adapted and extended for many scientific applications, including plant structure research. As for many biological studies, sample preparation is of major importance to obtain good-quality images. Therefore, we present a new preparation method for fresh material which includes critical point drying and heavy metal staining. This technique enhances the contrast of fresh tissues, prevents artefacts such as tissue compression, and requires no embedding.

The paradox of scored tablets: a cost-saving risk.

One of the cornerstones of pharmacotherapy is the proper dose of medicine, which should ideally be tailored to the individual patient. However, even if clinically possible, this is economically not feasible as a too large number of different dosage strengths would be required. Therefore, a balance is required between the patient's benefit/risk and the cost to the individual and society on the other hand. Scored or splitted tablets were, and still are, often used strategies to these opposite interests, enabling more dose-flexibility, but also at the same time increasing the dose-variability as a consequence of the breaking process. The question of how to deal with this paradox was investigated by exploring the prevalence and classification of scored tablets as well as the cost-benefits. A strategy for clinical pharmacologists is presented to improve the outcome of this paradox.

Virtual histology by means of high-resolution X-ray CT.

Micro-CT is a non-destructive technique for 3D tomographic investigation of an object. A 3D representation of the internal structure is calculated based on a series of X-ray radiographs taken from different angles. The spatial resolution of current laboratory-used micro-CT systems has come down over the last years from a few tens of microns to a few microns. This opens the possibility to perform histological investigations in 3D on a virtual representation of a sample, referred to as virtual 3D histology. The advantage of micro-CT based virtual histology is the immediate and automated 3D visualization of the sample without prior slicing, sample preparation like decalcification, photographing and aligning. This not only permits a drastic reduction in preparation time but also offers the possibility to easily investigate objects that are difficult to slice. This article presents results that were obtained on punch biopsies of horse skin, (dental) alveolus of ponies and chondro-osseous samples from the tarsus of foals studied with the new high resolution micro-CT set-up (HRXCT) at the Ghent University (Belgium) (http://www.ugct.ugent.be). This state-of-the-art set-up provides a 1 micron resolution and is therefore ideally suited for a direct comparison with standard light microscopy-based histology.

3D printing of high drug loaded dosage forms using thermoplastic polyurethanes.

It was the aim of this study to develop high drug loaded (>30%, w/w), thermoplastic polyurethane (TPU)-based dosage forms via fused deposition modelling (FDM). Model drugs with different particle size and aqueous solubility were pre-processed in combination with diverse TPU grades via hot melt extrusion (HME) into filaments with a diameter of 1.75 ± 0.05 mm. Subsequently, TPU-based filaments which featured acceptable quality attributes (i.e. consistent filament diameter, smooth surface morphology and good mechanical properties) were printed into tablets. The sustained release potential of the 3D printed dosage forms was tested in vitro. Moreover, the impact of printing parameters on the in vitro drug release was investigated. TPU-based filaments could be loaded with 60% (w/w) fine drug powder without observing severe shark skinning or inconsistent filament diameter. During 3D printing experiments, HME filaments based on hard TPU grades were successfully converted into personalized dosage forms containing a high concentration of crystalline drug (up to 60%, w/w). In vitro release kinetics were mainly affected by the matrix composition and tablet infill degree. Therefore, this study clearly demonstrated that TPU-based FDM feedstock material offers a lot of formulation freedom for the development of personalized dosage forms.

Porous hydroxyapatite tablets as carriers for low-dosed drugs.

The present study evaluated an innovative technique for the manufacturing of low-dosed tablets. Tablets containing hydroxyapatite and a pore forming agent (50% (w/w) Avicel PH 200/20, 37.5% and 50% corn starch/37.5% sorbitol) were manufactured by direct compression followed by sintering. The influence of pore forming agent (type and concentration), sinter temperature and sinter time on tablet properties was investigated. Sintering (1250 degrees C) revealed tablets with an acceptable friability (<1%). Using 50% (w/w) Avicel PH 200 as pore forming agent resulted in tablets combining the highest porosity (50%) and the highest median pore diameter (5 microm). Aqueous drug solutions (metoprolol tartrate, riboflavin sodium phosphate) were spiked on the tablet surface. The maximum volume of drug solution absorbed was limited (2x100 microl), revealing that these porous carriers were ideal for low dosed formulations. Drug release from the tablets was slow, independent of the drug. To accelerate drug release, tablets were manufactured using a modified gelcasting technique yielding tablets with a median pore size of 60 and 80 microm. Release from these tablets was drastically increased indicating that the permeability of the tablets was influenced by the pore size, shape and connectivity of the porous network. Changing and controlling these parameters made it possible to obtain drug delivery systems providing different drug delivery behaviour.

A comparative study between melt granulation/compression and hot melt extrusion/injection molding for the manufacturing of oral sustained release thermoplastic polyurethane matrices.

During this project 3 techniques (twin screw melt granulation/compression (TSMG), hot melt extrusion (HME) and injection molding (IM)) were evaluated for the manufacturing of thermoplastic polyurethane (TPU)-based oral sustained release matrices, containing a high dose of the highly soluble metformin hydrochloride. Whereas formulations with a drug load between 0 and 70% (w/w) could be processed via HME/(IM), the drug content of granules prepared via melt granulation could only be varied between 85 and 90% (w/w) as these formulations contained the proper concentration of binder (i.e. TPU) to obtain a good size distribution of the granules. While release from HME matrices and IM tablets could be sustained over 24h, release from the TPU-based TSMG tablets was too fast (complete release within about 6h) linked to their higher drug load and porosity. By mixing hydrophilic and hydrophobic TPUs the in vitro release kinetics of both formulations could be adjusted: a higher content of hydrophobic TPU was correlated with a slower release rate. Although mini-matrices showed faster release kinetics than IM tablets, this observation was successfully countered by changing the hydrophobic/hydrophilic TPU ratio. In vivo experiments via oral administration to dogs confirmed the versatile potential of the TPU platform as intermediate-strong and low-intermediate sustained characteristics were obtained for the IM tablets and HME mini-matrices, respectively.

Comparison of the blood supply to the articular-epiphyseal growth complex in horse vs. pony foals.

REASONS FOR PERFORMING STUDY: To increase understanding of why the prevalence of clinical/radiographic osteochondrosis (OC) dissecans is high in horses and low in ponies. OBJECTIVES: To investigate whether the clinical difference in OC occurrence between horses and ponies could partly be explained by a difference in: 1) number of patent vessels in the epiphyseal growth cartilage; 2) duration of the presence of patent cartilage canals; or 3) growth cartilage thickness at predilection sites for OC. The hypothesis was that pony foals would have fewer cartilage canals, shorter duration of blood supply and thinner growth cartilage than horse foals. STUDY DESIGN: Observational, cross-sectional study. METHODS: Nine Standardbred foals (horse group) 1-49 days old and 11 Norwegian Fjord foals (pony group) 1-62 days old were included. A total of 15 anatomical locations in the tarsocrural and metatarsophalangeal joints were examined by one or more of the following techniques: arterial perfusion; photography of cleared specimens; microcomputed tomography; radiography; and histology. The number of cartilage canals was counted. Cartilage thickness was measured. Duration of blood supply was assessed in histological sections. RESULTS: Of the 3 common predilection sites for OC investigated, there were significantly fewer vessels (P = 0.003) and thinner cartilage (P = 0.002) at the distal lateral trochlear ridge of the talus in the pony group. There was no difference in the duration of presence of cartilage canals between the groups. CONCLUSION: The hypothesis that pony foals would have fewer cartilage canals and thinner growth cartilage than horse foals was confirmed for the lateral trochlear ridge of the talus. The current results may contribute towards an explanation for the low prevalence of OC at the distal lateral trochlear ridge of the talus in pony foals.

Fatty acids for controlled release applications: A comparison between prilling and solid lipid extrusion as manufacturing techniques.

The aim of the present study was to evaluate the solid state characteristics, drug release and stability of fatty acid-based formulations after processing via prilling and solid lipid extrusion. Myristic acid (MA), stearic acid (SA) and behenic acid (BA) were used as matrix formers combined with metoprolol tartrate (MPT) as model drug. The prilling process allowed complete dissolution of MPT in the molten fatty acid phase, generating semi-crystalline MPT and the formation of hydrogen bonds between drug and fatty acids in the solid prills. In contrast, as solid lipid extrusion (SLE) induced only limited melting of the fatty acids, molecular interaction with the drug was inhibited, yielding crystalline MPT. Although the addition of a low melting fatty acid allowed more MPT/fatty acid interaction during extrusion, crystalline MPT was detected after processing. Mathematical modeling revealed that the extrudates exhibited a higher apparent drug/water mobility than prills of the same composition, probably due to differences in the inner systems' structure. Irrespective of the processing method, mixed fatty acid systems (e.g. MA/BA) exhibited a lower matrix porosity, resulting in a slower drug release rate. Solid state analysis of these systems indicated that the crystalline structure of the fatty acids was maintained after SLE, while prilling generated a reduced MA crystallinity. Binary MPT/fatty acid systems processed via extrusion showed better stability during storage at 40 °C than the corresponding prills. Although mixed fatty acid systems were stable at 25 °C, stability problems were encountered during storage at 40 °C: a faster release was obtained from the prills, whereas drug release from the extrudates was slower.

Holistic approach of pre-existing flaws on the decay of two limestones.

This study aims to understand the influence of the microfacies and the determination of pre-existing flaws on the weathering behavior of two types of limestones. Therefore, both Lede and Noyant limestones were independently weathered by strong acid tests and freeze-thaw cycles. In order to characterize the weathering patterns inside the stones, a combination of high resolution X-ray CT, SEM-EDS and thin section microscopy was used. The advantage of high resolution X-ray CT is its non-destructive character and the obtained 3D structural information. By using this technique, a time-lapse sequence of the weathering patterns was obtained for both gypsum crust formation as well as crack formation due to freezing and thawing. This way, a clear link could be made with the initial non-weathered state. Thin section microscopy and SEM-EDS provided additional chemical information. The focus of this study lies in the processes that occur in the bioclast fragments in the stone and the influence of the surrounding cement or matrix. The results show that weathering patterns vary for both limestones although the causes of weathering were similar. In case of the Noyant stone, the weathering by crystallizing gypsum was mainly restricted to the microporous matrix of the stone, while in case of the Lede stone, several foraminifera and shell fragments were preferentially recrystallized. In general, the underlying microstructure determines the weathering pattern of the stone.