Visualizing the Human Body Using an Artistic Approach.

This chapter describes an innovative approach to the cross-disciplinary study of anatomy and art to facilitate visualization of the human body. We draw upon the literature, together with our own experience of designing, delivering and researching a cross-disciplinary art and anatomy course, to indicate the critical elements of the approach that foster students' visualization of the anatomy of the human body.Visual arts have been linked with anatomy for centuries, but typically biomedical science has existed in a utilitarian relationship with art only used as an aid. In this chapter, we discuss the rationale underpinning a cross-disciplinary anatomy and art course and describe our experience of devising activities and assessment that create a stimulating and mutually beneficial environment for visualizing the experience and physicality of the human body. We describe the structure of the course which integrates art and anatomy to train students in the language of anatomy and visual representation, by engaging them in a process of attempting their own visual communication. The cross-disciplinary nature of our approach creates a unique social environment that offers a supportive environment for exploration and experimentation without fear of failure. Students' personal growth in resilience, tolerance for uncertainty and creativity prepares them for the inclusion of these values in their career.

Technology enhanced assessment: Ottawa consensus statement and recommendations.

INTRODUCTION: In 2011, a consensus report was produced on technology-enhanced assessment (TEA), its good practices, and future perspectives. Since then, technological advances have enabled innovative practices and tools that have revolutionised how learners are assessed. In this updated consensus, we bring together the potential of technology and the ultimate goals of assessment on learner attainment, faculty development, and improved healthcare practices. METHODS: As a material for the report, we used the scholarly publications on TEA in both HPE and general higher education, feedback from 2020 Ottawa Conference workshops, and scholarly publications on assessment technology practices during the Covid-19 pandemic. RESULTS AND CONCLUSION: The group identified areas of consensus that remained to be resolved and issues that arose in the evolution of TEA. We adopted a three-stage approach (readiness to adopt technology, application of assessment technology, and evaluation/dissemination). The application stage adopted an assessment 'lifecycle' approach and targeted five key foci: (1) Advancing authenticity of assessment, (2) Engaging learners with assessment, (3) Enhancing design and scheduling, (4) Optimising assessment delivery and recording learner achievement, and (5) Tracking learner progress and faculty activity and thereby supporting longitudinal learning and continuous assessment.

A method for gene knockdown in the retina using a lipid-based carrier.

Purpose: The use of small non-coding nucleic acids, such as siRNA and miRNA, has allowed for a deeper understanding of gene functions, as well as for development of gene therapies for complex neurodegenerative diseases, including retinal degeneration. For effective delivery into the eye and transfection of the retina, suitable transfection methods are required. We investigated the use of a lipid-based transfection agent, Invivofectamine® 3.0 (Thermo Fisher Scientific), as a potential method for delivery of nucleic acids to the retina. Methods: Rodents were injected intravitreally with formulations of Invivofectamine 3.0 containing scrambled, Gapdh, Il-1ß, and C3 siRNAs, or sterile PBS (control) using a modified protocol for encapsulation of nucleic acids. TdT-mediated dUTP nick-end labeling (TUNEL) and IBA1 immunohistochemistry was used to determine histological cell death and inflammation. qPCR were used to determine the stress and inflammatory profile of the retina. Electroretinography (ERG) and optical coherence tomography (OCT) were employed as clinical indicators of retinal health. Results: We showed that macrophage recruitment, retinal stress, and photoreceptor cell death in animals receiving Invivofectamine 3.0 were comparable to those in negative controls. Following delivery of Invivofectamine 3.0 alone, no statistically significant changes in expression were found in a suite of inflammatory and stress genes, and ERG and OCT analyses revealed no changes in retinal function or morphology. Injections with siRNAs for proinflammatory genes (C3 and Il-1ß) and Gapdh, in combination with Invivofectamine 3.0, resulted in statistically significant targeted gene knockdown in the retina for up to 4 days following injection. Using a fluorescent Block-It siRNA, transfection was visualized throughout the neural retina with evidence of transfection observed in cells of the ganglion cell layer, inner nuclear layer, and outer nuclear layer. Conclusions: This work supports the use of Invivofectamine 3.0 as a transfection agent for effective delivery of nucleic acids to the retina for gene function studies and as potential therapeutics.

Obesity-induced metabolic disturbance drives oxidative stress and complement activation in the retinal environment.

Purpose: Systemic increases in reactive oxygen species, and their association with inflammation, have been proposed as an underlying mechanism linking obesity and age-related macular degeneration (AMD). Studies have found increased levels of oxidative stress biomarkers and inflammatory cytokines in obese individuals; however, the correlation between obesity and retinal inflammation has yet to be assessed. We used the leptin-deficient (ob/ob) mouse to further our understanding of the contribution of obesity to retinal oxidative stress and inflammation. Methods: Retinas from ob/ob mice were compared to age-matched wild-type controls for retinal function (electroretinography) and gene expression analysis of retinal stress (Gfap), oxidative stress (Gpx3 and Hmox1), and complement activation (C3, C2, Cfb, and Cfh). Oxidative stress was further quantified using a reactive oxygen species and reactive nitrogen species (ROS and RNS) assay. Retinal microglia and macrophage migration to the outer retina and complement activation were determined using immunohistochemistry for IBA1 and C3, respectively. Retinas and sera were used for metabolomic analysis using QTRAP mass spectrometry. Results: Retinal function was reduced in ob/ob mice, which correlated to changes in markers of retinal stress, oxidative stress, and inflammation. An increase in C3-expressing microglia and macrophages was detected in the outer retinas of the ob/ob mice, while gene expression studies showed increases in the complement activators (C2 and Cfb) and a decrease in a complement regulator (Cfh). The expression of several metabolites were altered in the ob/ob mice compared to the controls, with changes in polyunsaturated fatty acids (PUFAs) and branched-chain amino acids (BCAAs) detected. Conclusions: The results of this study indicate that oxidative stress, inflammation, complement activation, and lipid metabolites in the retinal environment are linked with obesity in ob/ob animals. Understanding the interplay between these components in the retina in obesity will help inform risk factor analysis for acquired retinal degenerations, including AMD.

670nm light treatment following retinal injury modulates Müller cell gliosis: Evidence from in vivo and in vitro stress models.

Photobiomodulation (PBM) with 670 nm light has been shown to accelerate wound healing in soft tissue injuries, and also to protect neuronal tissues. However, little data exist on its effects on the non-neuronal components of the retina, such as Müller cells (MCs), which are the principal macroglia of the retina that play a role in maintaining retinal homeostasis. The aim of this study was to explore the effects of 670 nm light on activated MCs using in vivo and in vitro stress models. Adult Sprague-Dawley rats were exposed to photo-oxidative damage (PD) for 24 h and treated with 670 nm light at 0, 3 and 14 days after PD. Tissue was collected at 30 days post-PD for analysis. Using the in vitro scratch model with a human MC line (MIO-M1), area coverage and cellular stress were analysed following treatment with 670 nm light. We showed that early treatment with 670 nm light after PD reduced MC activation, lowering the retinal expression of GFAP and FGF-2. 670 nm light treatment mitigated the production of MC-related pro-inflammatory cytokines (including IL-1ß), and reduced microglia/macrophage (MG/MΦ) recruitment into the outer retina following PD. This subsequently decreased photoreceptor loss, slowing the progression of retinal degeneration. In vitro, we showed that 670 nm light directly modulated MC activation, reducing rates of area coverage by suppressing cellular proliferation and spreading. This study indicates that 670 nm light treatment post-injury may have therapeutic benefit when administered shortly after retinal damage, and could be useful for retinal degenerations where MC gliosis is a feature of disease progression.

Photobiomodulation with 670 nm light ameliorates Müller cell-mediated activation of microglia and macrophages in retinal degeneration.

Müller cells, the supporting cells of the retina, play a key role in responding to retinal stress by releasing chemokines, including CCL2, to recruit microglia and macrophages (MG/MΦ) into the damaged retina. Photobiomodulation (PBM) with 670 nm light has been shown to reduce inflammation in models of retinal degeneration. In this study, we aimed to investigate whether 670 nm light had an effect on Müller cell-initiated inflammation under retinal photo-oxidative damage (PD) in vivo and in vitro. Sprague-Dawley rats were pre-treated with 670 nm light (9J/cm2) once daily over 5 days prior to PD. The expression of inflammatory genes including CCL2 and IL-1ß was analysed in retinas. In vitro, primary Müller cells dissociated from neonatal rat retinas were co-cultured with 661W photoreceptor cells. Co-cultures were exposed to PD, followed by 670 nm light treatment to the Müller cells only, and Müller cell stress and inflammation were assessed. Primary MG/MΦ were incubated with supernatant from the co-cultures, and collected for analysis of inflammatory activation. To further understand the mechanism of 670 nm light, the expression of COX5a and mitochondrial membrane potential (ΔΨm) were measured in Müller cells. Following PD, 670 nm light-treated Müller cells had a reduced inflammatory activation, with lower levels of CCL2, IL-1ß and IL-6. Supernatant from 670 nm light-treated co-cultures reduced activation of primary MG/MΦ, and lowered the expression of pro-inflammatory cytokines, compared to untreated PD controls. Additionally, 670 nm light-treated Müller cells had an increased expression of COX5a and an elevated ΔΨm following PD, suggesting that retrograde signaling plays a role in the effects of 670 nm light on Müller cell gene expression. Our data indicates that 670 nm light reduces Müller cell-mediated retinal inflammation, and offers a potential cellular mechanism for 670 nm light therapy in regulating inflammation associated with retinal degenerations.

The broad-spectrum chemokine inhibitor NR58-3.14.3 modulates macrophage-mediated inflammation in the diseased retina.

BACKGROUND: The activity of macrophages is implicated in the progression of retinal pathologies such as atrophic age-related macular degeneration (AMD), where they accumulate among the photoreceptor layer and subretinal space. This process is aided by the local expression of chemokines, which furnish these cells with directional cues that augment their migration to areas of retinal injury. While these qualities make chemokines a potential therapeutic target in curtailing damaging retinal inflammation, their wide variety and signalling redundancy pose challenges in broadly modulating their activity. Here, we examine the efficacy of the broad-spectrum chemokine inhibitor NR58-3.14.3-a suppressor of Ccl- and Cxcl- chemokine pathways-in suppressing macrophage activity and photoreceptor death, using a light-induced model of outer retinal atrophy and inflammation. METHODS: Photo-oxidative damage was induced in SD rats via exposure to 1000 lux of light for 24 h, after which animals were euthanized at 0- or 7-day post-exposure time points. Prior to damage, NR58-3.14.3 was injected intravitreally. Retinas were harvested and evaluated for the effect of NR58-3.14.3 on subretinal macrophage accumulation and cytokine expression profile, as well as photoreceptor degeneration. RESULTS: We report that intravitreal administration of NR58-3.14.3 reduces the accumulation of macrophages in the outer retina following exposure to light damage, at both 0- and 7-day post-exposure time points. Injection of NR58-3.14.3 also reduced the up-regulation of inflammatory markers including of Il6, Ccl3, and Ccl4 in infiltrating macrophages, which are promoters of their pathogenic activity in the retina. Finally, NR58-3.14.3-injected retinas displayed markedly reduced photoreceptor death following light damage, at both 0 and 7 days post-exposure. CONCLUSIONS: Our findings indicate that NR58-3.14.3 is effective in inhibiting subretinal macrophage accumulation in light-induced retinal degeneration and illustrate the potential of broad-spectrum chemokine inhibitors as novel therapeutic agents in thwarting retinal inflammation. Although broad-spectrum chemokine inhibitors may not be appropriate for all retinal inflammatory conditions, our results suggest that they may be beneficial for retinal dystrophies in which chemokine expression and subretinal macrophage accumulation are implicated, such as advanced AMD.

A model of progressive photo-oxidative degeneration and inflammation in the pigmented C57BL/6J mouse retina.

Light-induced degeneration in rodent retinas is an established model for of retinal degeneration, including the roles of oxidative stress and neuroinflammatory activity. In these models, photoreceptor death is elicited via photo-oxidative stress, and is exacerbated by recruitment of subretinal macrophages and activation of immune pathways including complement propagation. Existing light damage models have relied heavily on albino rodents, and mostly using acute light stimuli. These albino models have proven valuable in uncovering the pathogenic mechanisms of such pathways in the context of retinal disease. However, their inherent albinism hinders comparability to normal retinal physiology, and also makes gene technology analysis time-consuming due to the predominance of the pigmented mouse strains in these applications. In this study, we characterise a new light damage model utilising C57BL/6J mice over a 7 day period of chronic light exposure. We use high-efficiency LED technology to deliver a sustained intensity of 100 k lux with negligible modulation of ambient temperature. We show that in the C57BL/6J mouse, chronic light exposure elicits the cardinal features of light damage including photoreceptor degeneration, atrophy of the choriocapillaris, decreased retinal function and increases in oxidative stress markers 4-HNE and 8-OHG, which emerge progressively over the 7 day period of exposure. These changes are accompanied by robust recruitment of IBA1+ and F4/80 + microglia/macrophages to the ONL and subretinal space, followed the strong up-regulation of monocyte-chemoattractants Ccl2, Ccl3, and Ccl12, as well as increases in expression of complement component C3. These findings are in agreement with prior damage models conducted in albino rodents such as Balb/c mice, and support the use of this new model in further investigating the causative features of oxidative stress and inflammation in retinal disease.

Near-Infrared Photobiomodulation in Retinal Injury and Disease.

Evidence is growing that exposure of tissue to low energy photon irradiation in the far-red (FR) to near-infrared (NIR) range of the spectrum, collectively termed "photobiomodulation" (PBM) can restore the function of damaged mitochondria, upregulate the production of cytoprotective factors and prevent apoptotic cell death. PBM has been applied clinically in the treatment of soft tissue injuries and acceleration of wound healing for more than 40 years. Recent studies have demonstrated that FR/NIR photons penetrate diseased tissues including the retina. The therapeutic effects of PBM have been hypothesized to result from intracellular signaling pathways triggered when FR/NIR photons are absorbed by the mitochondrial photoacceptor molecule, cytochrome c oxidase, culminating in improved mitochondrial energy metabolism, increased cytoprotective factor production and cell survival. Investigations in rodent models of methanol-induced ocular toxicity, light damage, retinitis pigmentosa and age-related macular degeneration have demonstrated the PBM attenuates photoreceptor cell death, protects retinal function and exerts anti-inflammatory actions.

Chemokine-mediated inflammation in the degenerating retina is coordinated by Müller cells, activated microglia, and retinal pigment epithelium.

BACKGROUND: Monocyte infiltration is involved in the pathogenesis of many retinal degenerative conditions. This process traditionally depends on local expression of chemokines, though the roles of many of these in the degenerating retina are unclear. Here, we investigate expression and in situ localization of the broad chemokine response in a light-induced model of retinal degeneration. METHODS: Sprague-Dawley (SD) rats were exposed to 1,000 lux light damage (LD) for up to 24 hrs. At time points during (1 to 24 hrs) and following (3 and 7 days) exposure, animals were euthanized and retinas processed. Microarray analysis assessed differential expression of chemokines. Some genes were further investigated using polymerase chain reaction (PCR) and in situ hybridization and contrasted with photoreceptor apoptosis using terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL). Recruitment of retinal CD45 (+) leukocytes was determined via fluorescence activated cell sorting (FACS), and expression of chemokine receptors determined using PCR. RESULTS: Exposure to 24 hrs of LD resulted in differential expression of chemokines including Ccl3, Ccl4, Ccl7, Cxcl1, and Cxcl10. Their upregulation correlated strongly with peak photoreceptor death, at 24 hrs exposure. In situ hybridization revealed that the modulated chemokines were expressed by a combination of Müller cells, activated microglia, and retinal pigment epithelium (RPE). This preceded large increases in the number of CD45(+) cells at 3- and 7-days post exposure, which expressed a corresponding repertoire of chemokine receptors. CONCLUSIONS: Our data indicate that retinal degeneration induces upregulation of a broad chemokine response whose expression is coordinated by Müller cells, microglia, and RPE. The findings inform our understanding of the processes govern the trafficking of leukocytes, which are contributors in the pathology of retinal degenerations.

Red/near-infrared irradiation therapy for treatment of central nervous system injuries and disorders.

Irradiation in the red/near-infrared spectrum (R/NIR, 630-1000 nm) has been used to treat a wide range of clinical conditions, including disorders of the central nervous system (CNS), with several clinical trials currently underway for stroke and macular degeneration. However, R/NIR irradiation therapy (R/NIR-IT) has not been widely adopted in clinical practice for CNS injury or disease for a number of reasons, which include the following. The mechanism/s of action and implications of penetration have not been thoroughly addressed. The large range of treatment intensities, wavelengths and devices that have been assessed make comparisons difficult, and a consensus paradigm for treatment has not yet emerged. Furthermore, the lack of consistent positive outcomes in randomised controlled trials, perhaps due to sub-optimal treatment regimens, has contributed to scepticism. This review provides a balanced précis of outcomes described in the literature regarding treatment modalities and efficacy of R/NIR-IT for injury and disease in the CNS. We have addressed the important issues of specification of treatment parameters, penetration of R/NIR irradiation to CNS tissues and mechanism/s, and provided the necessary detail to demonstrate the potential of R/NIR-IT for the treatment of retinal degeneration, damage to white matter tracts of the CNS, stroke and Parkinson's disease.

670 nm light mitigates oxygen-induced degeneration in C57BL/6J mouse retina.

BACKGROUND: Irradiation with light wavelengths from the far red (FR) to the near infrared (NIR) spectrum (600 nm -1000 nm) has been shown to have beneficial effects in several disease models. In this study, we aim to examine whether 670 nm red light pretreatment can provide protection against hyperoxia-induced damage in the C57BL/6J mouse retina. Adult mice (90-110 days) were pretreated with 9 J/cm2 of 670 nm light once daily for 5 consecutive days prior to being placed in hyperoxic environment (75% oxygen). Control groups were exposed to hyperoxia, but received no 670 nm light pretreatment. Retinas were collected after 0, 3, 7, 10 or 14 days of hyperoxia exposure (n = 12/group) and prepared either for histological analysis, or RNA extraction and quantitative polymerase chain reaction (qPCR). Photoreceptor damage and loss were quantified by counting photoreceptors undergoing cell death and measuring photoreceptor layer thickness. Localization of acrolein, and cytochrome c oxidase subunit Va (Cox Va) were identified through immunohistochemistry. Expression of heme oxygenase-1 (Hmox-1), complement component 3 (C3) and fibroblast growth factor 2 (Fgf-2) genes were quantified using qPCR. RESULTS: The hyperoxia-induced photoreceptor loss was accompanied by reduction of metabolic marker, Cox Va, and increased expression of oxidative stress indicator, acrolein and Hmox-1. Pretreatment with 670 nm red light reduced expression of markers of oxidative stress and C3, and slowed, but did not prevent, photoreceptor loss over the time course of hyperoxia exposure. CONCLUSION: The damaging effects of hyperoxia on photoreceptors were ameliorated following pretreatment with 670 nm light in hyperoxic mouse retinas. These results suggest that pretreatment with 670 nm light may provide stability to photoreceptors in conditions of oxidative stress.

670-nm light treatment reduces complement propagation following retinal degeneration.

AIM: Complement activation is associated with the pathogenesis of age-related macular degeneration (AMD). We aimed to investigate whether 670-nm light treatment reduces the propagation of complement in a light-induced model of atrophic AMD. METHODS: Sprague-Dawley (SD) rats were pretreated with 9 J/cm(2) 670-nm light for 3 minutes daily over 5 days; other animals were sham treated. Animals were exposed to white light (1,000 lux) for 24 h, after which animals were kept in dim light (5 lux) for 7 days. Expression of complement genes was assessed by quantitative polymerase chain reaction (qPCR), and immunohistochemistry. Counts were made of C3-expressing monocytes/microglia using in situ hybridization. Photoreceptor death was also assessed using outer nuclear layer (ONL) thickness measurements, and oxidative stress using immunohistochemistry for 4-hydroxynonenal (4-HNE). RESULTS: Following light damage, retinas pretreated with 670-nm light had reduced immunoreactivity for the oxidative damage maker 4-HNE in the ONL and outer segments, compared to controls. In conjunction, there was significant reduction in retinal expression of complement genes C1s, C2, C3, C4b, C3aR1, and C5r1 following 670 nm treatment. In situ hybridization, coupled with immunoreactivity for the marker ionized calcium binding adaptor molecule 1 (IBA1), revealed that C3 is expressed by infiltrating microglia/monocytes in subretinal space following light damage, which were significantly reduced in number after 670 nm treatment. Additionally, immunohistochemistry for C3 revealed a decrease in C3 deposition in the ONL following 670 nm treatment. CONCLUSIONS: Our data indicate that 670-nm light pretreatment reduces lipid peroxidation and complement propagation in the degenerating retina. These findings have relevance to the cellular events of complement activation underling the pathogenesis of AMD, and highlight the potential of 670-nm light as a non-invasive anti-inflammatory therapy.

The question of dissection in medical training: Not just "if," but "when"? A student perspective.

While debate about the use of-and alternatives to-human cadaveric dissection in medical training is robust, little attention has been paid to questions about timing. This study explores the perspectives of medical students and recent graduates with regard to two key questions: when in the degree program do students prefer dissection opportunities and what are the students getting out of participating in dissection? Self-report survey data from students in preclinical years (n = 105), clinical years (n = 57), and graduates (n = 13) were analyzed. Most (89%) preferred dissection during the preclinical years, with no effect by training year (χ2  = 1.98, p = 0.16), previous anatomy (χ2  = 3.64, p = 0.31), or dissection (χ2  = 3.84, p = 0.26) experience. Three key findings emerged. First, the majority of students prefer to dissect in the preclinical years because they view dissection as important for developing foundation knowledge and delivering an opportunity for consolidation prior to transitioning to primarily clinical studies. In addition, students recognize that it is a time-consuming activity requiring specialized facilities. Second, three main understandings of the purpose of dissection were reported: depth of learning, learning experience, and real-world equivalence. Third, these student perspectives of the purpose of dissection are associated with timing preferences for dissection opportunities. The results identify the preclinical phase as the optimal time to strategically integrate dissection into medical training in order to maximize the benefits of this unique learning opportunity for students and minimize its impact upon curricular time.

Comparison of 2 open-sourced 3-dimensional modeling techniques for orthopaedic application.

Objectives: Although 3-dimensional (3D) printing is becoming more widely adopted for clinical applications, it is yet to be accepted as part of standard practice. One of the key applications of this technology is orthopaedic surgical planning for urgent trauma cases. Anatomically accurate replicas of patients' fracture models can be produced to guide intervention. These high-quality models facilitate the design and printing of patient-specific implants and surgical devices. Therefore, a fast and accurate workflow will help orthopaedic surgeons to generate high-quality 3D printable models of complex fractures. Currently, there is a lack of access to an uncomplicated and inexpensive workflow. Methods: Using patient DICOM data sets (n = 13), we devised a novel, simple, open-source, and rapid modeling process using Drishti software and compared its efficacy and data storage with the 3D Slicer image computing platform. We imported the computed tomography image directory acquired from patients into the software to isolate the model of bone surface from surrounding soft tissue using the minimum functions. One pelvic fracture case was further integrated into the customized implant design practice to demonstrate the compatibility of the 3D models generated from Drishti. Results: The data sizes of the generated 3D models and the processing files that represent the original DICOM of Drishti are on average 27% and 12% smaller than that of 3D Slicer, respectively (both P < 0.05). The time frame needed to reach the stage of viewing the 3D bone model and the exporting of the data of Drishti is 39% and 38% faster than that of 3D Slicer, respectively (both P < 0.05). We also constructed a virtual model using third-party software to trial the implant design. Conclusions: Drishti is more suitable for urgent trauma cases that require fast and efficient 3D bone reconstruction with less hardware requirement. 3D Slicer performs better at quantitative preoperative planning and multilayer segmentation. Both software platforms are compatible with third-party programs used to produce customized implants that could be useful for surgical training. Level of Evidence: Level V.

Morphological, functional and gene expression analysis of the hyperoxic mouse retina.

This study examined the impact of prolonged (up to 35 day) exposure to hyperoxia on the morphology and function of the retina, in the C57BL/6J mouse, as a basis for interpretation of gene expression changes. Mice of the C57BL/6J strain were raised from birth in dim cyclic illumination (12 h 5 lux, 12 h dark). Adult animals (90-110 days) were exposed to continuous hyperoxia (75% oxygen) for up to 35 d. Retinas were examined after 0 d (controls), 3 d, 7 d, 14 d and 35 d. Spatial and temporal patterns of photoreceptor death were mapped, using the TUNEL technique. Immunohistochemistry and a specific assay were used to assess the expression of a stress-related protein (GFAP) and the activity of key antioxidant enzymes (SOD). The dark-adapted flash electroretinogram was used to assess the function of rods and cones. RNA hybridized to Affymetrix Genechips was used to assess gene expression during the first 3 d of exposure. Photoreceptors were stable during the first 7 d exposure to hyperoxia, but thereafter showed progressive damage and degeneration, which began in a 'hot-spot' 0.5 mm inferior to the optic disc, then spread into surrounding retina. SOD activity was upregulated at 14 d, but not at earlier time points. GFAP expression was upregulated in Müller cells from 3 d. Rod and cone components of the ERG were supernormal at 3 d and 7 d, but then fell below control levels. Gene expression changes suggested possible mechanisms for this early supernormality of function. At 14 d exposure, damage to and death of photoreceptors were prominent and spreading, and function was correspondingly degraded. However at 3 d exposure, hyperoxia-induced supernormal functional responses in rods, while leaving their structure apparently undamaged. Variations in early (3 days) gene expression provide a partial insight into the mechanisms involved in this.

Examining the Short-, Medium-, and Long-Term Success of an Embodied Learning Activity in the Study of Hand Anatomy for Clinical Application.

A student's own body provides an often disregarded site of knowledge production and corporeal wisdom. Learning via cognitive processes anchored in physical movement and body awareness, known as embodied learning, may aid students to visualize structures and understand their functions and clinical relevance. Working from an embodied learning perspective, the current article evaluates the use of an offline physical learning tool (Anatomical Glove Learning System; AGLS) for teaching hand anatomy for clinical application in medical students. Two student samples (N1  = 105; N2  = 94) used the AGLS in two different ways. In the first sample, the AGLS was compared to a traditional approach using hand bones, models and prosected specimens. Secondly, the AGLS and traditional approach were combined. The evaluation consisted of three outcomes: short-term learning (post-test), medium-term applications (mock-objective structured clinical examination, MOSCE), and longer-term assessment (objective structured clinical examination, OSCE). Findings from the first sample indicated no significant differences between the AGLS and traditional laboratory groups on short- (F(1,78) = 0.036, P = 0.849), medium- (F(1,50) = 0.743, P = 0.393), or longer-term (F(1,82) = 0.997, P = 0.321) outcomes. In the second sample using the AGLS in combination with a traditional approach was associated with significantly better short-term post-test scores (F(2,174) = 5.98, P = 0.003) than using the AGLS alone, but demonstrated no effect for long-term OSCE scores. These results suggest an embodied learning experience alone does not appear to be advantageous to student learning, but when combined with other methods for studying anatomy there are learning gains.

Differential gene expression in mouse retina related to regional differences in vulnerability to hyperoxia.

PURPOSE: In the C57BL/6J mouse retina, hyperoxia-induced degeneration of photoreceptors shows strong regional variation, beginning at a locus ~0.5 mm inferior to the optic disc. To identify gene expression differences that might underlie this variability in vulnerability, we have used microarray techniques to describe regional (superior-inferior) variations in gene expression in the retina. METHODS: Young adult C57BL/6J mice raised in dim cyclic illumination (12 h at 5 lx and 12 h in darkness) were exposed to hyperoxia (75% oxygen for two weeks). Retinas were collected from hyperoxia-exposed and control animals without fixation and divided into superior and inferior halves. RNA was extracted from each sample, purified, and hybridized to Mouse Gene 1.0 ST arrays (Affymetrix). The consistency of the microarray results was assessed using quantitative PCR for selected genes. Expression data were analyzed to identify genes and ncRNAs whose differential expression between the superior and inferior retina could be associated with relative vulnerability to hyperoxia. RESULTS: In control retinas, only two genes showed a fold difference in expression >2 between the superior and inferior retina; another 25 showed a fold difference of 1.5-2.0. Of these 27, the functions of six genes, including ventral anterior homeobox containing gene 2 (Vax2) and T-box 5 (Tbox5), are related to parameters of anatomic development and the functions of five are related to sensory perception. Among the latter, short-wave-sensitive cone opsin (Opn1sw) was more strongly expressed in the inferior retina and medium-wave-sensitive cone opsin (Opn1mw) in the superior retina. This is consistent with known differences in S- and M-cone distribution, confirming our separation of retinal regions. The highest fold difference was reported for membrane metalloendopeptidase (Mme), a member from the metallothionein group of cytoprotective proteins. To identify genes whose regulation by hyperoxia was significantly different between the inferior and superior retina, we calculated the "fold margin" (FM, the difference between hyperoxia-induced regulation in the inferior and superior retina) for each gene, and identified genes for which abs(FM) > 0.5. Genes thus identified numbered 112, and included many immune-, cell defense-, and inflammation- related genes. CONCLUSIONS: Gene expression analysis revealed relatively subtle differences between inferior and superior regions of control C57BL/6J retinas, with only 27 genes showing an expression difference >1.5 fold. Among these, genes related to cytoprotection and apoptosis were included, along with genes related to central projections and cone-type differences. After hyperoxia-induced photoreceptor degeneration had begun, the number of genes that showed significant expression differences between the inferior and superior retina more than quadrupled, with genes related to immune processes, defense processes, and inflammation being numerically dominant.

Gene and noncoding RNA regulation underlying photoreceptor protection: microarray study of dietary antioxidant saffron and photobiomodulation in rat retina.

PURPOSE: To identify the genes and noncoding RNAs (ncRNAs) involved in the neuroprotective actions of a dietary antioxidant (saffron) and of photobiomodulation (PBM). METHODS: We used a previously published assay of photoreceptor damage, in which albino Sprague Dawley rats raised in dim cyclic illumination (12 h 5 lux, 12 h darkness) were challenged by 24 h exposure to bright (1,000 lux) light. Experimental groups were protected against light damage by pretreatment with dietary saffron (1 mg/kg/day for 21 days) or PBM (9 J/cm(2) at the eye, daily for 5 days). RNA from one eye of four animals in each of the six experimental groups (control, light damage [LD], saffron, PBM, saffronLD, and PBMLD) was hybridized to Affymetrix rat genome ST arrays. Quantitative real-time PCR analysis of 14 selected genes was used to validate the microarray results. RESULTS: LD caused the regulation of 175 entities (genes and ncRNAs) beyond criterion levels (p<0.05 in comparison with controls, fold-change >2). PBM pretreatment reduced the expression of 126 of these 175 LD-regulated entities below criterion; saffron pretreatment reduced the expression of 53 entities (50 in common with PBM). In addition, PBM pretreatment regulated the expression of 67 entities not regulated by LD, while saffron pretreatment regulated 122 entities not regulated by LD (48 in common with PBM). PBM and saffron, given without LD, regulated genes and ncRNAs beyond criterion levels, but in lesser numbers than during their protective action. A high proportion of the entities regulated by LD (>90%) were known genes. By contrast, ncRNAs were prominent among the entities regulated by PBM and saffron in their neuroprotective roles (73% and 62%, respectively). CONCLUSIONS: Given alone, saffron and (more prominently) PBM both regulated significant numbers of genes and ncRNAs. Given before retinal exposure to damaging light, thus while exerting their neuroprotective action, they regulated much larger numbers of entities, among which ncRNAs were prominent. Further, the downregulation of known genes and of ncRNAs was prominent in the protective actions of both neuroprotectants. These comparisons provide an overview of gene expression induced by two neuroprotectants and provide a basis for the more focused study of their mechanisms.