NUT Expression Is of Diagnostic Utility in the Distinction of Digital Papillary Carcinoma From Poroid Hidradenoma.

ABSTRACT: The distinction between digital papillary adenocarcinoma (DPAC) and benign cutaneous adnexal tumors is clinically important and can be challenging. Poroid hidradenoma frequently occurs at acral sites and can show a number of histological features, which overlap with digital papillary adenocarcinoma. Recent work has shown that YAP1-NUTM1 fusions are frequent in poroid hidradenoma and are associated with nuclear protein in testis (NUT) expression by immunohistochemistry. We evaluated the expression of NUT-1 by immunohistochemistry in 4 cases of DPAC and 4 cases of poroid hidradenoma. Three of 4 cases of poroid hidradenoma showed strong NUT-1 expression, with no staining in any of the cases of DPAC. These results suggest that NUT-1 immunohistochemistry may be a useful additional tool in evaluating this differential diagnosis.

A simulated comparison of lung tumor target verification using stereoscopic tomosynthesis or radiography.

PURPOSE: Mobile lung tumors are increasingly being treated with ablative radiotherapy, for which precise motion management is essential. In-room stereoscopic radiography systems are able to guide ablative radiotherapy for stationary cranial lesions but not optimally for lung tumors unless fiducial markers are inserted. We propose augmenting stereoscopic radiographic systems with multiple small x-ray sources to provide the capability of imaging with stereoscopic, single frame tomosynthesis. METHODS: In single frame tomosynthesis, nine x-ray sources are placed in a 3 × 3 configuration and energized simultaneously. The beams from these sources are collimated so that they converge on the tumor and then diverge to illuminate nine non-overlapping sectors on the detector. These nine sector images are averaged together and filtered to create the tomosynthesis effect. Single frame tomosynthesis is intended to be an alternative imaging mode for existing stereoscopic systems with a field of view that is three times smaller and a temporal resolution equal to the frame rate of the detector. We simulated stereoscopic tomosynthesis and radiography using Monte Carlo techniques on 60 patients with early-stage lung cancer from the NSCLC-Radiomics dataset. Two board-certified radiation oncologists reviewed these simulated images and rated them on a 4-point scale (1: tumor not visible; 2: tumor visible but inadequate for motion management; 3: tumor visible and adequate for motion management; 4: tumor visibility excellent). Each tumor was independently presented four times (two viewing angles from radiography and two viewing angles from tomosynthesis) in a blinded fashion over two reading sessions. RESULTS: The fraction of tumors that were rated as adequate or excellent for motion management (scores 3 or 4) from at least one viewing angle was 53% using radiography and 90% using tomosynthesis. From both viewing angles, the corresponding fractions were 7% for radiography and 48% for tomosynthesis. Readers agreed exactly on 62% of images and within 1 point on 98% of images. The acquisition technique was estimated to be 75 mAs at 120 kVp per treatment fraction assuming one verification image per breath, approximately one order of magnitude less than a standard dose cone beam CT. CONCLUSIONS: Stereoscopic tomosynthesis may provide a noninvasive, low dose, intrafraction motion verification technique for lung tumors treated by ablative radiotherapy. The system architecture is compatible with real-time video capture at 30 frames per second. Simulations suggest that most, but not all, lung tumors can be adequately visualized from at least one viewing angle.

Dose Sculpting Intensity Modulated Radiation Therapy for Vertebral Body Sparing in Children With Neuroblastoma.

PURPOSE: To assess the effect of dose sculpting intensity modulated radiation therapy on vertebral body growth in children with neuroblastoma. METHODS AND MATERIALS: From 2000 to 2011, 88 children with neuroblastoma underwent radiation at the authors' institution. Children with paravertebral tumors with at least 3 years of evaluable posttreatment imaging were included, and children who underwent spine reirradiation before follow-up were excluded. If vertebral bodies could not be spared, these "target" vertebral bodies were treated to at least 18 Gy. Thoracic and lumbar vertebral bodies were assessed separately. Dose data for target, spared, and internal control vertebral bodies were extracted. Multivariate generalized estimating equation modeling was used to assess the effect of dose and other clinical factors on vertebral body growth. RESULTS: A total of 34 patients (20 boys, 14 girls) met study criteria. Median age at start of radiation was 4.3 years; all but 1 had prior high-dose chemotherapy with stem cell rescue. Mean growth rates of target, spared, and control vertebral bodies (cm/body/y) were, respectively, 0.027, 0.032, and 0.044 in thoracic spine and 0.033, 0.055, and 0.083 in lumbar spine. On multivariate generalized estimating equation analysis, higher dose, older treatment age, male gender, and thoracic spine location were significantly associated with decreased vertebral body growth (P<.0001, P<.0001, P=.007, and P<.0001, respectively). Dose and spine location were significant in a 3-way interaction model (P<.0001). CONCLUSIONS: Vertebral bodies spared by intensity modulated radiation therapy grew faster than target vertebrae. Regardless of intent to spare or target, multivariate analysis confirms that lower dose results in significantly increased growth rate. This technique should be investigated prospectively.

Real-time tomosynthesis for radiation therapy guidance.

PURPOSE: Fluoroscopy has been a tool of choice for monitoring treatments or interventions because of its extremely fast imaging times. However, the contrast obtained in fluoroscopy may be insufficient for certain clinical applications. In stereotactic ablative radiation therapy of the lung, fluoroscopy often lacks sufficient contrast for gating treatment. The purpose of this work is to describe and assess a real-time tomosynthesis design that can produce sufficient contrast for guidance of lung tumor treatment within a small field of view. METHODS: Previous tomosynthesis designs in radiation oncology have temporal resolution on the order of seconds. The proposed system design uses parallel acquisition of multiple frames by simultaneously illuminating the field of view with multiple sources, enabling a temporal resolution of up to 30 frames per second. For a small field of view, a single flat-panel detector could be used if different sectors of the detector are assigned to specific sources. Simulated images were generated by forward projection of existing clinical datasets. The authors varied the number of tubes and the power of each tube in order to determine the impact on tumor visualization. RESULTS: Visualization of the tumor was much clearer in tomosynthesis than in fluoroscopy. Contrast generally improved with the number of sources used, and a minimum of four sources should be used. The high contrast of the lung allows very low system power, and in most cases, less than 1 mA was needed. More power is required in the lateral direction than the AP direction. CONCLUSIONS: The proposed system produces images adequate for real-time guidance of radiation therapy. The additional hardware requirements are modest, and the system is capable of imaging at high frame rates and low dose. Further development, including a prototype system and a dosimetry study, is needed to further evaluate the feasibility of this device for radiation therapy guidance.

Organization of the connections between claustrum and cortex in the mouse.

The connections between the claustrum and the cortex in mouse are systematically investigated with adeno-associated virus (AAV), an anterograde viral tracer. We first define the boundary and the three-dimensional structure of the claustrum based on a variety of molecular and anatomical data. From AAV injections into 42 neocortical and allocortical areas, we conclude that most cortical areas send bilateral projections to the claustrum, the majority being denser on the ipsilateral side. This includes prelimbic, infralimbic, medial, ventrolateral and lateral orbital, ventral retrosplenial, dorsal and posterior agranular insular, visceral, temporal association, dorsal and ventral auditory, ectorhinal, perirhinal, lateral entorhinal, and anteromedial, posteromedial, lateroposterior, laterointermediate, and postrhinal visual areas. In contrast, the cingulate and the secondary motor areas send denser projections to the contralateral claustrum than to the ipsilateral one. The gustatory, primary auditory, primary visual, rostrolateral visual, and medial entorhinal cortices send projections only to the ipsilateral claustrum. Primary motor, primary somatosensory and subicular areas barely send projections to either ipsi- or contralateral claustrum. Corticoclaustral projections are organized in a rough topographic manner, with variable projection strengths. We find that the claustrum, in turn, sends widespread projections preferentially to ipsilateral cortical areas with different projection strengths and laminar distribution patterns and to certain contralateral cortical areas. Our quantitative results show that the claustrum has strong reciprocal and bilateral connections with prefrontal and cingulate areas as well as strong reciprocal connections with the ipsilateral temporal and retrohippocampal areas, suggesting that it may play a crucial role in a variety of cognitive processes. J. Comp. Neurol. 525:1317-1346, 2017. © 2016 Wiley Periodicals, Inc.

Prognostication of serial post-intensity-modulated radiation therapy undetectable plasma EBV DNA for nasopharyngeal carcinoma.

Plasma Epstein-Barr virus (EBV) DNA titers have been used to monitor treatment response and provide prognostic information on survival for nasopharyngeal carcinoma (NPC). However, the long-term prognostic role of pretreatment and posttreatment titers after radical contemporaneous radiation therapy remains uncertain. We recruited 260 evaluable patients with non-metastatic NPC treated with radical intensity-modulated radiation therapy (IMRT) with or without adjunct chemotherapy. Plasma EBV DNA titers at baseline and then 8 weeks and 6 months after IMRT were measured. Cox regression models were employed to identify interaction between post-IMRT 8th week and 6th month undetectable titers and 3-year survival endpoints. Concordance indices (Ct) from time-dependent receiver-operating characteristics (TDROC) were compared between patients with post-IMRT undetectable and those with detectable titers. After a median follow-up duration of 3.4 years (range 1.4-4.6 years), patients with post-IMRT 8th week and 6th month undetectable plasma EBV DNA titers enjoyed longer 3-year survival endpoints than those who had detectable titers at the same time points. Post-IMRT 8th week, and more significantly, post-IMRT 6th month undetectable plasma EBV DNA were the only significant prognostic factors of 3-year survival endpoints. Ct values for all 3-year survival endpoints for both post-IMRT 8th week and 6th month undetectable plasma EBV DNA were significantly higher in those with stage IVA-IVB diseases compared to stage I-III counterparts. Early post-IMRT undetectable plasma EBV DNA titers were prognostic of 3-year survival endpoints in patients with non-metastatic NPC. Intensified treatment should be further explored for patients with persistently detectable titers after IMRT.

Effects of Chronic Sleep Restriction during Early Adolescence on the Adult Pattern of Connectivity of Mouse Secondary Motor Cortex.

Cortical circuits mature in stages, from early synaptogenesis and synaptic pruning to late synaptic refinement, resulting in the adult anatomical connection matrix. Because the mature matrix is largely fixed, genetic or environmental factors interfering with its establishment can have irreversible effects. Sleep disruption is rarely considered among those factors, and previous studies have focused on very young animals and the acute effects of sleep deprivation on neuronal morphology and cortical plasticity. Adolescence is a sensitive time for brain remodeling, yet whether chronic sleep restriction (CSR) during adolescence has long-term effects on brain connectivity remains unclear. We used viral-mediated axonal labeling and serial two-photon tomography to measure brain-wide projections from secondary motor cortex (MOs), a high-order area with diffuse projections. For each MOs target, we calculated the projection fraction, a combined measure of passing fibers and axonal terminals normalized for the size of each target. We found no homogeneous differences in MOs projection fraction between mice subjected to 5 days of CSR during early adolescence (P25-P30, ≥ 50% decrease in daily sleep, n=14) and siblings that slept undisturbed (n=14). Machine learning algorithms, however, classified animals at significantly above chance levels, indicating that differences between the two groups exist, but are subtle and heterogeneous. Thus, sleep disruption in early adolescence may affect adult brain connectivity. However, because our method relies on a global measure of projection density and was not previously used to measure connectivity changes due to behavioral manipulations, definitive conclusions on the long-term structural effects of early CSR require additional experiments.

Mapping social behavior-induced brain activation at cellular resolution in the mouse.

Understanding how brain activation mediates behaviors is a central goal of systems neuroscience. Here, we apply an automated method for mapping brain activation in the mouse in order to probe how sex-specific social behaviors are represented in the male brain. Our method uses the immediate-early-gene c-fos, a marker of neuronal activation, visualized by serial two-photon tomography: the c-fos-GFP+ neurons are computationally detected, their distribution is registered to a reference brain and a brain atlas, and their numbers are analyzed by statistical tests. Our results reveal distinct and shared female and male interaction-evoked patterns of male brain activation representing sex discrimination and social recognition. We also identify brain regions whose degree of activity correlates to specific features of social behaviors and estimate the total numbers and the densities of activated neurons per brain areas. Our study opens the door to automated screening of behavior-evoked brain activation in the mouse.

Efferent pathways of the mouse lateral habenula.

The lateral habenula (LHb) is part of the habenula complex of the dorsal thalamus. Recent studies of the LHb have focused on its projections to the ventral tegmental area (VTA) and rostromedial tegmental nucleus (RMTg), which contain γ-aminobutyric acid (GABA)ergic neurons that mediate reward prediction error via inhibition of dopaminergic activity. However, older studies in the rat have also identified LHb outputs to the lateral and posterior hypothalamus, median raphe, dorsal raphe, and dorsal tegmentum. Although these studies have shown that the medial and lateral divisions of the LHb have somewhat distinct projections, the topographic specificity of LHb efferents is not completely understood, and the relative extent of these projections to brainstem targets is unknown. Here we have used anterograde tracing with adeno-associated virus-mediated expression of green fluorescent protein, combined with serial two-photon tomography, to map the efferents of the LHb on a standard coordinate system for the entire mouse brain, and reconstruct the efferent pathways of the LHb in three dimensions. Using automated quantitation of fiber density, we show that in addition to the RMTg, the median raphe, caudal dorsal raphe, and pontine central gray are major recipients of LHb efferents. By using retrograde tract tracing with cholera toxin subunit B, we show that LHb neurons projecting to the hypothalamus, VTA, median raphe, caudal dorsal raphe, and pontine central gray reside in characteristic, but sometimes overlapping regions of the LHb. Together these results provide the anatomical basis for systematic studies of LHb function in neural circuits and behavior in mice. J. Comp. Neurol. 523:32-60, 2015. © 2014 Wiley Periodicals, Inc.

A mesoscale connectome of the mouse brain.

Comprehensive knowledge of the brain's wiring diagram is fundamental for understanding how the nervous system processes information at both local and global scales. However, with the singular exception of the C. elegans microscale connectome, there are no complete connectivity data sets in other species. Here we report a brain-wide, cellular-level, mesoscale connectome for the mouse. The Allen Mouse Brain Connectivity Atlas uses enhanced green fluorescent protein (EGFP)-expressing adeno-associated viral vectors to trace axonal projections from defined regions and cell types, and high-throughput serial two-photon tomography to image the EGFP-labelled axons throughout the brain. This systematic and standardized approach allows spatial registration of individual experiments into a common three dimensional (3D) reference space, resulting in a whole-brain connectivity matrix. A computational model yields insights into connectional strength distribution, symmetry and other network properties. Virtual tractography illustrates 3D topography among interconnected regions. Cortico-thalamic pathway analysis demonstrates segregation and integration of parallel pathways. The Allen Mouse Brain Connectivity Atlas is a freely available, foundational resource for structural and functional investigations into the neural circuits that support behavioural and cognitive processes in health and disease.

Preoperative embolization of aberrant systemic artery in sequestration of lung.

A 40-year-old man presented with left lower lobe pneumonia that failed to resolve on antibiotic therapy. Computed tomography revealed intralobar sequestration of the left lower lobe supplied by a large artery from the descending aorta. The aberrant artery was embolized using polyvinyl alcohol particles. The sequestered tissue was resected 3 weeks later. Identification and control of the aberrant artery is essential to avoid inadvertent injury and massive hemorrhage.

A genome-wide RNAi screen identifies multiple RSK-dependent regulators of cell migration.

To define the functional pathways regulating epithelial cell migration, we performed a genome-wide RNAi screen using 55,000 pooled lentiviral shRNAs targeting ∼11,000 genes, selecting for transduced cells with increased motility. A stringent validation protocol generated a set of 31 genes representing diverse pathways whose knockdown dramatically enhances cellular migration. Some of these pathways share features of epithelial-to-mesenchymal transition (EMT), and together they implicate key regulators of transcription, cellular signaling, and metabolism, as well as novel modulators of cellular trafficking, such as DLG5. In delineating downstream pathways mediating these migration phenotypes, we observed universal activation of ERKs and a profound dependence on their RSK effectors. Pharmacological inhibition of RSK dramatically suppresses epithelial cell migration induced by knockdown of all 31 genes, suggesting that convergence of diverse migratory pathways on this kinase may provide a therapeutic opportunity in disorders of cell migration, including cancer metastasis.

Vascular density has prognostic value in Merkel cell carcinoma.

Merkel cell carcinomas are aggressive tumours for which histological prognostic factors need to be established. This study examines the prognostic role of vascular density, based on CD34 immunohistochemical staining in Merkel cell carcinoma. Thirty-six cases of Merkel cell carcinoma were immunohistochemically stained for the endothelial marker CD34. Vascular density was assessed in the tumor and stroma with a Chalkley eyepiece graticule. The scores of vascular density were correlated with other clinical and histological parameters to determine the prognostic significance of tumor vascularity. Increased vascular density was shown to be significantly associated with a worse prognosis (P = 0.005). A 1-unit increase in total vessel score was associated with a 3.9 times increase in the risk of death (95% hazard ratio confidence limits 1.50-10.32). Other factors associated with a worse outcome included tumor size (P = 0.05), the presence of lymphovascular invasion (P = 0.03), and tumor mast cell count (P < 0.002). Increased vascular density is associated with a worse prognosis in Merkel cell carcinomas. Assessment of vascular density may assist in predicting clinical behavior in these tumors and in evaluating the effects of adjuvant therapy.

Mast cells have prognostic value in Merkel cell carcinoma.

We examined the role of mast cell infiltrates and other clinical and histological factors in the prognosis of Merkel cell carcinoma. Mast cells were stained immunohistochemically in 36 Merkel cell carcinomas with an antibody to tryptase. The number of stainable cells was quantified within the tumors and surrounding stroma. Other clinical and histological parameters were examined, statistically analyzed, and compared to subsequent clinical course and prognosis. Patient prognosis was worse with higher tumor mast cell numbers (P < 0.002). Prognosis was also found to be adversely affected by the presence of lymphovascular invasion (P = 0.03) and increased tumor size (P = 0.05). Increased mast cells counts, tumor size, and lymphovascular invasion are associated with an adverse prognosis in Merkel cell carcinomas. Evaluation of mast cell infiltrates may provide useful prognostic data and ultimately could assist in selecting patients that require adjuvant treatment in this aggressive form of skin cancer.

Quantitative image analysis: software systems in drug development trials.

Multi-dimensional image analysis is being used increasingly to arrive at surrogate end-points for drug development trials. Various imaging modalities such as computed tomography (CT), magnetic resonance imaging (MRI), positron emission tomography (PET) and ultrasound are used to analyze treatments for diseases such as cancer, multiple sclerosis, osteoarthritis, and Alzheimer's disease. However, extracting information from images can be tedious and is prone to high user variability. The medical image analysis community is moving towards advanced software systems specifically designed for drug development trials. These systems can automatically identify the anatomy of interest in medical images (segmentation methods), can compare the anatomy over time or between patients (registration methods) and allow the quantitative extraction of anatomical features and the integration of the data and results into a database management system, automatically tracking the changes made to the data (audit trail generation). In this article, we present a case study using a prototype system that is used for quantifying multiple sclerosis lesions from multivariate MRI.