Methods for the optimal preservation of blow fly intra-puparial forms for morphological analysis in forensic casework.

Accurate minimum post-mortem interval (minPMI) estimations often rely on a precise age determination of insect developmental stages, which is significantly influenced by environmental temperature. An optimal preservation of the entomological samples collected at crime scenes is pivotal for a reliable aging of immature insect samples. For blow flies (Diptera: Calliphoridae), the most widely used insect indicators in forensic investigations, an appropriate preservation of tissues is particularly important in the case of puparial samples because aging methods for intra-puparial forms usually depend on morphological analyses; however, although informative soft tissues and structures could be discoloured and/or distorted if they are not properly fixed, there is a lack of studies to assess different methods for the optimal preservation of intra-puparial forms collected in forensic investigations. The present study compares three preservation methods for intra-puparial forms of the blow fly Calliphora vicina Robineau-Desvoidy, 1830: (i) direct immersion into 80% ethanol, (ii) puncturing of the puparium and hot water killing (HWK) prior to preservation in 80% ethanol, and (iii) HWK without puncturing before preservation in 80% ethanol. External and internal morphological analyses of intra-puparial forms of different ages were conducted to assess the quality of preservation. The results indicate that direct immersion in ethanol led to poor preservation, affecting both external and internal tissues. Both methods with HWK resulted in a better preservation, but puncturing resulted, in some cases, in physical damage of the specimens. HWK without puncturing emerged as the optimal preservation method, consistently yielding high preservation scores for both external and internal morphological analyses. These findings have practical implications for forensic practitioners and emphasise the need for updating some published guidelines and protocols in forensic entomology.

Larval anatomy of the digestive and excretory systems of the pharyngeal bot fly, Pharyngomyia picta (Diptera: Oestridae).

Oestrid flies (Diptera: Oestridae) are obligate parasites of mammals during their larval stage and show anatomical adaptations for the infestation of host tissues. Unlike the species that parasitize domestic mammals, those oestrid species that infest wild mammal hosts remain poorly known. With the use of x-ray micro-computed tomography, we describe for the first time the anatomy of the digestive and excretory systems of the second and third larval instars of Pharyngomyia picta (Meigen), a parasite of cervids that, like other species within the subfamily Oestrinae, causes nasopharyngeal myiasis. Both larval instars of P. picta show a pair of remarkably large salivary glands arranged in a characteristic 'glandular band', a convoluted and thickly uniform midgut and a greatly enlarged distal region of the anterior pair of Malpighian tubules. These anatomical features also have been described in other species within the subfamily Oestrinae, whereas they differ from the observations in other oestrid subfamilies. We discuss the potential functional significance of the anatomy of the digestive and excretory systems of Oestrinae larvae as specific adaptations to parasitize the nasopharyngeal cavities of mammal hosts.

The Application of 3base™ Technology to Diagnose Eight of the Most Clinically Important Gastrointestinal Protozoan Infections.

Globally, over 3.5 billion people are infected with intestinal parasites each year, resulting in over 200,000 deaths. Three of the most common protozoan pathogens that affect the gastrointestinal tract of humans are Cryptosporidium spp., Giardia intestinalis, and Entamoeba histolytica. Other protozoan agents that have been implicated in gastroenteritis in humans include Cyclospora cayetanensis, Dientamoeba fragilis, Blastocystis hominis, and the microsporidia Enterocytozoon bieneusi and Encephalitozoon intestinalis. Genetic Signatures previously developed a 3base™ multiplexed Real-Time PCR (mRT-PCR) enteric protozoan kit (EP001) for the detection of Giardia intestinalis/lamblia/duodenalis, Cryptosporidium spp., E. histolytica, D. fragilis, and B. hominis. We now describe improvements to this kit to produce a more comprehensive assay, including C. cayetanensis, E. bieneusi, and E. intestinalis, termed EP005. The clinical performance of EP005 was assessed using a set of 380 clinical samples against a commercially available PCR test and other in-house nucleic acid amplification tests where commercial tests were not available. All methods provided at least 90% agreement. EP005 had no cross-reactivity against 82 organisms commonly found in the gut. The EP005 method streamlines the detection of gastrointestinal parasites and addresses the many challenges of traditional microscopic detection, resulting in cost savings and significant improvements in patient care.

Ocular lens morphology is influenced by ecology and metamorphosis in frogs and toads.

The shape and relative size of an ocular lens affect the focal length of the eye, with consequences for visual acuity and sensitivity. Lenses are typically spherical in aquatic animals with camera-type eyes and axially flattened in terrestrial species to facilitate vision in optical media with different refractive indices. Frogs and toads (Amphibia: Anura) are ecologically diverse, with many species shifting from aquatic to terrestrial ecologies during metamorphosis. We quantified lens shape and relative size using 179 micro X-ray computed tomography scans of 126 biphasic anuran species and tested for correlations with life stage, environmental transitions, adult habits and adult activity patterns. Across broad phylogenetic diversity, tadpole lenses are more spherical than those of adults. Biphasic species with aquatic larvae and terrestrial adults typically undergo ontogenetic changes in lens shape, whereas species that remain aquatic as adults tend to retain more spherical lenses after metamorphosis. Further, adult lens shape is influenced by adult habit; notably, fossorial adults tend to retain spherical lenses following metamorphosis. Finally, lens size relative to eye size is smaller in aquatic and semiaquatic species than other adult ecologies. Our study demonstrates how ecology shapes visual systems, and the power of non-invasive imaging of museum specimens for studying sensory evolution.

Effect of an Individualized Audit and Feedback Intervention on Rates of Musculoskeletal Diagnostic Imaging Requests by Australian General Practitioners: A Randomized Clinical Trial.

Importance: Audit and feedback can improve professional practice, but few trials have evaluated its effectiveness in reducing potential overuse of musculoskeletal diagnostic imaging in general practice. Objective: To evaluate the effectiveness of audit and feedback for reducing musculoskeletal imaging by high-requesting Australian general practitioners (GPs). Design, Setting, and Participants: This factorial cluster-randomized clinical trial included 2271 general practices with at least 1 GP who was in the top 20% of referrers for 11 imaging tests (of the lumbosacral or cervical spine, shoulder, hip, knee, and ankle/hind foot) and for at least 4 individual tests between January and December 2018. Only high-requesting GPs within participating practices were included. The trial was conducted between November 2019 and May 2021, with final follow-up on May 8, 2021. Interventions: Eligible practices were randomized in a 1:1:1:1:1 ratio to 1 of 4 different individualized written audit and feedback interventions (n = 3055 GPs) that varied factorially by (1) frequency of feedback (once vs twice) and (2) visual display (standard vs enhanced display highlighting highly requested tests) or to a control condition of no intervention (n = 764 GPs). Participants were not masked. Main Outcomes and Measures: The primary outcome was the overall rate of requests for the 11 targeted imaging tests per 1000 patient consultations over 12 months, assessed using routinely collected administrative data. Primary analyses included all randomized GPs who had at least 1 patient consultation during the study period and were performed by statisticians masked to group allocation. Results: A total of 3819 high-requesting GPs from 2271 practices were randomized, and 3660 GPs (95.8%; n = 727 control, n = 2933 intervention) were included in the primary analysis. Audit and feedback led to a statistically significant reduction in the overall rate of imaging requests per 1000 consultations compared with control over 12 months (adjusted mean, 27.7 [95% CI, 27.5-28.0] vs 30.4 [95% CI, 29.8-30.9], respectively; adjusted mean difference, -2.66 [95% CI, -3.24 to -2.07]; P < .001). Conclusions and Relevance: Among Australian general practitioners known to frequently request musculoskeletal diagnostic imaging, an individualized audit and feedback intervention, compared with no intervention, significantly decreased the rate of targeted musculoskeletal imaging tests ordered over 12 months. Trial Registration: ANZCTR Identifier: ACTRN12619001503112.

Ontogenetic development of the holocephalan dentition: Morphological transitions of dentine in the absence of teeth.

Among the cartilaginous fishes (Chondrichthyes), the Holocephali are unique in that teeth are absent both in ontogeny and adult regenerative growth. Instead, the holocephalan dentition of ever-growing nonshedding dental plates is composed of dentine, trabecular in arrangement, forming spaces into which a novel hypermineralized dentine (whitlockin) is deposited. These tissue features form a variety of specific morphologies as the defining characters of dental plates in the three families of extant holocephalans. We demonstrate how this morphology changes through ontogenetic development with continuity between morphologies, through successive growth stages of the dentition represented by the dental plate. For example, rod-shaped whitlockin appears early, later transformed into the tritoral pad, including a regular arrangement of vascular canals and whitlockin forming with increasing mineralization (95%-98%). While the tritoral pads develop lingually, stacks of individual ovoids of whitlockin replace the rods in the more labial parts of the plate, again shaped by the forming trabecular dentine. The ability to make dentine into new, distinctive patterns is retained in the evolution of the Holocephali, despite the lack of teeth forming in development of the dentition. We propose that developmentally, odontogenic stem cells, retained through evolution, control the trabecular dentine formation within the dental plate, and transition to form whitlockin, throughout lifetime growth. Our model of cellular activity proposes a tight membrane of odontoblasts, having transformed to whitloblasts, that can control active influx of minerals to the rapidly mineralizing dentine, forming whitlockin. After the reduced whitloblast cells transition back to odontoblasts, they continue to monitor the levels of minerals (calcium, phosphate and magnesium) and at a slower rate of growth in the peritubate 'softer' dentine. This model explains the unique features of transitions within the holocephalan dental plate morphology.

The multiplicative impacts of working hours and fine particulate matter concentration on life expectancy: A longitudinal analysis of US States.

This study contributes to interdisciplinary research on the social and environmental determinants of population health, with a focus on the interaction between working hours and fine particulate matter (PM2.5) concentration. The authors estimate longitudinal models of the relationship between US state-level average life expectancy and both average working hours and PM2.5 concentration for the 2005-2014 period. Results obtained from two-way fixed effects models indicate that average life expectancy is negatively associated with both average working hours and fine particulate matter concentration. Findings also indicate clear moderating relationships: the negative association between life expectancy and working hours is amplified as PM2.5 concentration increases, and the negative relationship between life expectancy and fine particulate matter concentration is amplified when average working hours increase. The results of this study underscore the need for additional research on the multiplicative impacts of socioeconomic factors and environmental factors in the modeling of population health.

Holocephalan (Chondrichthyes) dental plates with hypermineralized dentine as a substitute for missing teeth through developmental plasticity.

All extant holocephalans (Chimaeroidei) have lost the ability to make individual teeth, as tooth germs are not part of the embryonic development of the dental plates or of their continuous growth. Instead, a hypermineralized dentine with a unique mineral, whitlockin, is specifically distributed within a dentine framework into structures that give the dental plates their distinctive, species-specific morphology. Control of the regulation of this distribution must be cellular, with a dental epithelium initiating the first outer dentine, and via contact with ectomesenchymal tissue as the only embryonic cell type that can make dentine. Chimaeroids have three pairs of dental plates within their mouth, two in the upper jaw and one in the lower. In the genera Chimaera, Hydrolagus and Harriotta, the morphology and distribution of this whitlockin within each dental plate differs both between different plates in the same species and between species. Whitlockin structures include ovoids, rods and tritoral pads, with substantial developmental changes between these. For example, rods appear before the ovoids and result from a change in the surrounding trabecular dentine. In Harriotta, ovoids form separately from the tritoral pads, but also contribute to tritor development, while in Chimaera and Hydrolagus, tritoral pads develop from rods that later are perforated to accommodate the vasculature. Nevertheless, the position of these structures, secreted by the specialized odontoblasts (whitloblasts), appears highly regulated in all three species. These distinct morphologies are established at the aboral margin of the dental plate, with proposed involvement of the outer dentine. We observe that this outer layer forms into serially added lingual ridges, occurring on the anterior plate only. We propose that positional, structural specificity must be contained within the ectomesenchymal populations, as stem cells below the dental epithelium, and a coincidental occurrence of each lingual, serial ridge with the whitlockin structures that contribute to the wear-resistant oral surface.

3D Super-Resolution US Imaging of Rabbit Lymph Node Vasculature in Vivo by Using Microbubbles.

Background Variations in lymph node (LN) microcirculation can be indicative of metastasis. The identification and quantification of metastatic LNs remains essential for prognosis and treatment planning, but a reliable noninvasive imaging technique is lacking. Three-dimensional super-resolution (SR) US has shown potential to noninvasively visualize microvascular networks in vivo. Purpose To study the feasibility of three-dimensional SR US imaging of rabbit LN microvascular structure and blood flow by using microbubbles. Materials and Methods In vivo studies were carried out to image popliteal LNs of two healthy male New Zealand white rabbits aged 6-8 weeks. Three-dimensional, high-frame-rate, contrast material-enhanced US was achieved by mechanically scanning with a linear imaging probe. Individual microbubbles were identified, localized, and tracked to form three-dimensional SR images and super-resolved velocity maps. Acoustic subaperture processing was used to improve image contrast and to generate enhanced power Doppler and color Doppler images. Vessel size and blood flow velocity distributions were evaluated and assessed by using Student paired t test. Results SR images revealed microvessels in the rabbit LN, with branches clearly resolved when separated by 30 µm, which is less than half of the acoustic wavelength and not resolvable by using power or color Doppler. The apparent size distribution of most vessels in the SR images was below 80 µm and agrees with micro-CT data, whereas most of those detected with Doppler techniques were larger than 80 µm in the images. The blood flow velocity distribution indicated that most of the blood flow in rabbit popliteal LN was at velocities lower than 5 mm/sec. Conclusion Three-dimensional super-resolution US imaging using microbubbles allows noninvasive nonionizing visualization and quantification of lymph node microvascular structures and blood flow dynamics with resolution below the wave diffraction limit. This technology has potential for studying the physiologic functions of the lymph system and for clinical detection of lymph node metastasis. Published under a CC BY 4.0 license. Online supplemental material is available for this article.

Aquaculture and the displacement of fisheries captures.

In modern aquaculture, animal-production technology is used to increase aquatic food sources. Such controlled rearing of seafood can, in principle, shift the pressure off wild stocks and aquatic ecosystems by reducing fishing activities, which may advance marine conservation goals. We examined resource displacement-the reduced consumption of a resource due to its replacement with a more environmentally benign substitute-in fisheries. We employed panel regression techniques in an analysis of time-series data from 1970 through 2014 to assess the extent to which aquaculture production displaced fisheries captures for all nations for which data were available. We estimated 9 models to assess whether aquaculture production suppresses captures once other factors related to demand have been controlled for. Only 1 model predicted significant suppression of fisheries captures associated with aquaculture systems within nations over time. These results suggest that global aquaculture production does not substantially displace fisheries capture; instead, aquaculture production largely supplements fisheries capture.

La Acuacultura y el Desplazamiento de Capturas de las Pesquerías Resumen En la acuacultura moderna, la tecnología de producción animal se usa para incrementar las fuentes acuáticas de alimento. Tal crianza controlada de peces puede, en principio, modificar la presión que afecta los stocks silvestres y los ecosistemas acuáticos al reducir las actividades de pesca, lo que podría acercarnos a la obtención de las metas de conservación. Examinamos el desplazamiento de recursos - el consumo reducido de un recurso debido a su reemplazo con un sustituto más benigno con el ambiente - en las pesquerías. Empleamos técnicas de regresión de paneles en un análisis de datos de series de tiempo de 1970 a 2014 para evaluar hasta dónde ha llegado el desplazamiento de las capturas de las pesquerías en todos los países que tenían datos disponibles. Estimamos nueve modelos para evaluar si la producción de la acuacultura suprime las capturas una vez que otros factores relacionados con la demanda han sido controlados. Solamente un modelo pronosticó la supresión de las capturas de las pesquerías asociadas con los sistemas de acuacultura en los países a lo largo del tiempo. Estos resultados sugieren que la producción acuícola no desplaza sustancialmente la captura de las pesquerías; en su lugar, la producción acuícola complementa en gran parte a estas capturas.

Modernizing our way out or digging ourselves in? Reconsidering the impacts of efficiency innovations and affluence on residential energy consumption, 2005-2015.

This study scrutinizes the impacts of efficiency innovations as well as affluence on residential energy consumption, which is a major driver of greenhouse gas emissions. The study draws on the ecological-modernization perspective, which is optimistic about how technological innovations and affluence can help societies overcome environmental challenges associated with production and consumption, and the political-economy perspective, which raises doubts about whether these factors are beneficial to the environment, given their tendency to drive more consumption. Analysis of nationally representative longitudinal data reveals mixed relationships between efficiency innovations and residential energy consumption: while some measures of efficiency innovations, generally those not requiring human-technology interactions, are negatively related to residential energy consumption, others are either unrelated to it or drive more consumption. These findings suggest efficiency innovations offer only minimal opportunities for conserving energy, and may depend on the nature of the innovation. Raising doubts about the potential for rising affluence to promote environmental protection, this study reveals positive relationships between our measures of affluence and residential energy consumption.

Development and evolution of tooth renewal in neoselachian sharks as a model for transformation in chondrichthyan dentitions.

A defining feature of dentitions in modern sharks and rays is the regulated pattern order that generates multiple replacement teeth. These are arranged in labio-lingual files of replacement teeth that form in sequential time order both along the jaw and within successively initiated teeth in a deep dental lamina. Two distinct adult dentitions have been described: alternate, in which timing of new teeth alternates between two adjacent files, each erupting separately, and the other arranged as single files, where teeth of each file are timed to erupt together, in some taxa facilitating similarly timed teeth to join to form a cutting blade. Both are dependent on spatiotemporally regulated formation of new teeth. The adult Angel shark Squatina (Squalomorphii) exemplifies a single file dentition, but we obtained new data on the developmental order of teeth in the files of Squatina embryos, showing alternate timing of tooth initiation. This was based on micro-CT scans revealing that the earliest mineralised teeth at the jaw margin and their replacements in file pairs (odd and even jaw positions) alternate in their initiation timing. Along with Squatina, new observations from other squalomorphs such as Hexanchus and Chlamydoselachus, together with representatives of the sister group Galeomorphii, have established that the alternate tooth pattern (initiation time and replacement order) characterises the embryonic dentition of extant sharks; however, this can change in adults. These character states were plotted onto a recent phylogeny, demonstrating that the Squalomorphii show considerable plasticity of dental development. We propose a developmental-evolutionary model to allow change from the alternate to a single file alignment of replacement teeth. This establishes new dental morphologies in adult sharks from inherited alternate order.

Reengineering cyt b562 for hydrogen production: A facile route to artificial hydrogenases.

Bioinspired, protein-based molecular catalysts utilizing base metals at the active are emerging as a promising avenue to sustainable hydrogen production. The protein matrix modulates the intrinsic reactivity of organometallic active sites by tuning second-sphere and long-range interactions. Here, we show that swapping Co-Protoporphyrin IX for Fe-Protoporphyrin IX in cytochrome b562 results in an efficient catalyst for photoinduced proton reduction to molecular hydrogen. Further, the activity of wild type Co-cyt b562 can be modulated by a factor of 2.5 by exchanging the coordinating methionine with alanine or aspartic acid. The observed turnover numbers (TON) range between 125 and 305, and correlate well with the redox potential of the Co-cyt b562 mutants. The photosensitized system catalyzes proton reduction with high efficiency even under an aerobic atmosphere, implicating its use for biotechnological applications. This article is part of a Special Issue entitled Biodesign for Bioenergetics--the design and engineering of electronic transfer cofactors, proteins and protein networks, edited by Ronald L. Koder and J.L. Ross Anderson.

RBM5 is a male germ cell splicing factor and is required for spermatid differentiation and male fertility.

Alternative splicing of precursor messenger RNA (pre-mRNA) is common in mammalian cells and enables the production of multiple gene products from a single gene, thus increasing transcriptome and proteome diversity. Disturbance of splicing regulation is associated with many human diseases; however, key splicing factors that control tissue-specific alternative splicing remain largely undefined. In an unbiased genetic screen for essential male fertility genes in the mouse, we identified the RNA binding protein RBM5 (RNA binding motif 5) as an essential regulator of haploid male germ cell pre-mRNA splicing and fertility. Mice carrying a missense mutation (R263P) in the second RNA recognition motif (RRM) of RBM5 exhibited spermatid differentiation arrest, germ cell sloughing and apoptosis, which ultimately led to azoospermia (no sperm in the ejaculate) and male sterility. Molecular modelling suggested that the R263P mutation resulted in compromised mRNA binding. Within the adult mouse testis, RBM5 localises to somatic and germ cells including spermatogonia, spermatocytes and round spermatids. Through the use of RNA pull down coupled with microarrays, we identified 11 round spermatid-expressed mRNAs as putative RBM5 targets. Importantly, the R263P mutation affected pre-mRNA splicing and resulted in a shift in the isoform ratios, or the production of novel spliced transcripts, of most targets. Microarray analysis of isolated round spermatids suggests that altered splicing of RBM5 target pre-mRNAs affected expression of genes in several pathways, including those implicated in germ cell adhesion, spermatid head shaping, and acrosome and tail formation. In summary, our findings reveal a critical role for RBM5 as a pre-mRNA splicing regulator in round spermatids and male fertility. Our findings also suggest that the second RRM of RBM5 is pivotal for appropriate pre-mRNA splicing.

RAB-like 2 has an essential role in male fertility, sperm intra-flagellar transport, and tail assembly.

A significant percentage of young men are infertile and, for the majority, the underlying cause remains unknown. Male infertility is, however, frequently associated with defective sperm motility, wherein the sperm tail is a modified flagella/cilia. Conversely, a greater understanding of essential mechanisms involved in tail formation may offer contraceptive opportunities, or more broadly, therapeutic strategies for global cilia defects. Here we have identified Rab-like 2 (RABL2) as an essential requirement for sperm tail assembly and function. RABL2 is a member of a poorly characterized clade of the RAS GTPase superfamily. RABL2 is highly enriched within developing male germ cells, where it localizes to the mid-piece of the sperm tail. Lesser amounts of Rabl2 mRNA were observed in other tissues containing motile cilia. Using a co-immunoprecipitation approach and RABL2 affinity columns followed by immunochemistry, we demonstrated that within developing haploid germ cells RABL2 interacts with intra-flagella transport (IFT) proteins and delivers a specific set of effector (cargo) proteins, including key members of the glycolytic pathway, to the sperm tail. RABL2 binding to effector proteins is regulated by GTP. Perturbed RABL2 function, as exemplified by the Mot mouse line that contains a mutation in a critical protein-protein interaction domain, results in male sterility characterized by reduced sperm output, and sperm with aberrant motility and short tails. Our data demonstrate a novel function for the RABL protein family, an essential role for RABL2 in male fertility and a previously uncharacterised mechanism for protein delivery to the flagellum.

An essential role for katanin p80 and microtubule severing in male gamete production.

Katanin is an evolutionarily conserved microtubule-severing complex implicated in multiple aspects of microtubule dynamics. Katanin consists of a p60 severing enzyme and a p80 regulatory subunit. The p80 subunit is thought to regulate complex targeting and severing activity, but its precise role remains elusive. In lower-order species, the katanin complex has been shown to modulate mitotic and female meiotic spindle dynamics and flagella development. The in vivo function of katanin p80 in mammals is unknown. Here we show that katanin p80 is essential for male fertility. Specifically, through an analysis of a mouse loss-of-function allele (the Taily line), we demonstrate that katanin p80, most likely in association with p60, has an essential role in male meiotic spindle assembly and dissolution and the removal of midbody microtubules and, thus, cytokinesis. Katanin p80 also controls the formation, function, and dissolution of a microtubule structure intimately involved in defining sperm head shaping and sperm tail formation, the manchette, and plays a role in the formation of axoneme microtubules. Perturbed katanin p80 function, as evidenced in the Taily mouse, results in male sterility characterized by decreased sperm production, sperm with abnormal head shape, and a virtual absence of progressive motility. Collectively these data demonstrate that katanin p80 serves an essential and evolutionarily conserved role in several aspects of male germ cell development.

Transfer learning for auto-segmentation of 17 organs-at-risk in the head and neck: Bridging the gap between institutional and public datasets.

BACKGROUND: Auto-segmentation of organs-at-risk (OARs) in the head and neck (HN) on computed tomography (CT) images is a time-consuming component of the radiation therapy pipeline that suffers from inter-observer variability. Deep learning (DL) has shown state-of-the-art results in CT auto-segmentation, with larger and more diverse datasets showing better segmentation performance. Institutional CT auto-segmentation datasets have been small historically (n < 50) due to the time required for manual curation of images and anatomical labels. Recently, large public CT auto-segmentation datasets (n > 1000 aggregated) have become available through online repositories such as The Cancer Imaging Archive. Transfer learning is a technique applied when training samples are scarce, but a large dataset from a closely related domain is available. PURPOSE: The purpose of this study was to investigate whether a large public dataset could be used in place of an institutional dataset (n > 500), or to augment performance via transfer learning, when building HN OAR auto-segmentation models for institutional use. METHODS: Auto-segmentation models were trained on a large public dataset (public models) and a smaller institutional dataset (institutional models). The public models were fine-tuned on the institutional dataset using transfer learning (transfer models). We assessed both public model generalizability and transfer model performance by comparison with institutional models. Additionally, the effect of institutional dataset size on both transfer and institutional models was investigated. All DL models used a high-resolution, two-stage architecture based on the popular 3D U-Net. Model performance was evaluated using five geometric measures: the dice similarity coefficient (DSC), surface DSC, 95th percentile Hausdorff distance, mean surface distance (MSD), and added path length. RESULTS: For a small subset of OARs (left/right optic nerve, spinal cord, left submandibular), the public models performed significantly better (p < 0.05) than, or showed no significant difference to, the institutional models under most of the metrics examined. For the remaining OARs, the public models were inferior to the institutional models, although performance differences were small (DSC ≤ 0.03, MSD < 0.5 mm) for seven OARs (brainstem, left/right lens, left/right parotid, mandible, right submandibular). The transfer models performed significantly better than the institutional models for seven OARs (brainstem, right lens, left/right optic nerve, left/right parotid, spinal cord) with a small margin of improvement (DSC ≤ 0.02, MSD < 0.4 mm). When numbers of institutional training samples were limited, public and transfer models outperformed the institutional models for most OARs (brainstem, left/right lens, left/right optic nerve, left/right parotid, spinal cord, and left/right submandibular). CONCLUSION: Training auto-segmentation models with public data alone was suitable for a small number of OARs. Using only public data incurred a small performance deficit for most other OARs, when compared with institutional data alone, but may be preferable over time-consuming curation of a large institutional dataset. When a large institutional dataset was available, transfer learning with models pretrained on a large public dataset provided a modest performance improvement for several OARs. When numbers of institutional samples were limited, using the public dataset alone, or as a pretrained model, was beneficial for most OARs.

14-3-3σ mediates G2-M arrest produced by 5-aza-2'-deoxycytidine and possesses a tumor suppressor role in endometrial carcinoma cells.

OBJECTIVES: To determine the effect of 5-aza-2'-deoxycytidine (DAC) on human endometrial carcinoma cell (HECC) oncogenicity and demonstrate a molecular mechanism by which DAC modulates HECC oncogenicity. METHODS: The effect of DAC was tested on HECC RL95-2, AN3, Ishikawa and ECC1 cells. The role of 14-3-3σ on HECC oncogenicity in response to DAC treatment was evaluated in RL95-2 and AN3 cells after forced expression or silencing of 14-3-3σ gene expression. RESULTS: Treatment of HECC with DAC produced non-cytotoxic cell growth inhibition and G2/M cell cycle arrest. This effect was strongly correlated with increased expression of p21 and 14-3-3σ. Silencing of 14-3-3σ induced cellular proliferation and reduced the effect of DAC on cell cycle arrest in G2/M phases. Conversely, forced expression of 14-3-3σ showed the opposite effect. Furthermore, forced expression of 14-3-3σ in human endometrial cell lines reduced cell growth and colony formation. CONCLUSIONS: We suggest that 14-3-3σ in HECC suppresses cell proliferation and mediates DAC induced G2/M arrest and inhibition of cell proliferation in HECC.

Bricks, trusses and superstructures: Strategies for skeletal reinforcement in batoid fishes (rays and skates).

Crushing and eating hard prey (durophagy) is mechanically demanding. The cartilage jaws of durophagous stingrays are known to be reinforced relative to non-durophagous relatives, with a thickened external cortex of mineralized blocks (tesserae), reinforcing struts inside the jaw (trabeculae), and pavement-like dentition. These strategies for skeletal strengthening against durophagy, however, are largely understood only from myliobatiform stingrays, although a hard prey diet has evolved multiple times in batoid fishes (rays, skates, guitarfishes). We perform a quantitative analysis of micro-CT data, describing jaw strengthening mechanisms in Rhina ancylostoma (Bowmouth Guitarfish) and Rhynchobatus australiae (White-spotted Wedgefish), durophagous members of the Rhinopristiformes, the sister taxon to Myliobatiformes. Both species possess trabeculae, more numerous and densely packed in Rhina, albeit simpler structurally than those in stingrays like Aetobatus and Rhinoptera. Rhina and Rhynchobatus exhibit impressively thickened jaw cortices, often involving >10 tesseral layers, most pronounced in regions where dentition is thickest, particularly in Rhynchobatus. Age series of both species illustrate that tesserae increase in size during growth, with enlarged and irregular tesserae associated with the jaws' oral surface in larger (older) individuals of both species, perhaps a feature of ageing. Unlike the flattened teeth of durophagous myliobatiform stingrays, both rhinopristiform species have oddly undulating dentitions, comprised of pebble-like teeth interlocked to form compound "meta-teeth" (large spheroidal structures involving multiple teeth). This is particularly striking in Rhina, where the upper/lower occlusal surfaces are mirrored undulations, fitting together like rounded woodworking finger-joints. Trabeculae were previously thought to have arisen twice independently in Batoidea; our results show they are more widespread among batoid groups than previously appreciated, albeit apparently absent in the phylogenetically basal Rajiformes. Comparisons with several other durophagous and non-durophagous species illustrate that batoid skeletal reinforcement architectures are modular: trabeculae can be variously oriented and are dominant in some species (e.g. Rhina, Aetobatus), whereas cortical thickening is more significant in others (e.g. Rhynchobatus), or both reinforcing features can be lacking (e.g. Raja, Urobatis). We discuss interactions and implications of character states, framing a classification scheme for exploring cartilage structure evolution in the cartilaginous fishes.

Evaluation and imaging of an untreated grade III hamstring tear: a case report.

BACKGROUND: Muscle strains are one of the most common complaints treated by physicians. High-force lengthening contractions can produce very high forces resulting in pain and tissue damage; such strains are the most common cause of muscle injuries. The hamstring muscles are particularly susceptible as they cross two joints and regularly perform lengthening contractions during running. We describe a patient with return to full function after a large hamstring tear. CASE DESCRIPTION: We report the case of a 26-year-old man who presented 1 year after a noncontact, left-sided proximal hamstring tear incurred while sprinting. He received no medical treatment or formal rehabilitation. He was able to return to all sports and activities 1 to 2 months after injury, but noted a persistent deformity of the proximal thigh, which led him to seek evaluation. Physical examination, MRI functional tests, and specific muscle tests 1 year after his injury documented a major hamstring tear at the musculotendinous junction with muscle retraction, but no avulsion of the proximal tendon attachment. LITERATURE REVIEW: Surgery often is recommended for major proximal hamstring tendon tears, especially when more than one tendon of origin is ruptured from the ischial tuberosity. Myotendinous tears are treated nonoperatively, but may be associated with decreased strength, prolonged recovery, and recurrence. PURPOSE AND CLINICAL RELEVANCE: We describe the case of a young man who sustained a hamstring tear, with retraction, at the proximal myotendinous junction, where the biceps femoris and semitendinosus arise from the conjoint tendon. He achieved full functional recovery without medical attention, but had a persistent cosmetic deformity and slight hamstring tightness. This case suggests a benign natural history for this injury and the appropriateness of noninvasive treatment.