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A high-performance machine learning-assisted gas sensor strategy based on the integration of supervised and unsupervised learning with a gas-sensitive semiconductor metal oxide (SMO) gas sensor array is introduced. A 4-SMO sensor array was chosen as a test sensor system for detecting carbon monoxide (CO) and ethyl alcohol (C2H5OH) mixtures using 15 different combinations. Gas sensing detection/classification was performed with different numbers of gas sensor and machine learning algorithms. K-Means clustering was successfully employed to rationally identify the similarity features of targeted gases among 4 different groups, i.e., matrix gas, two single-component gases, and one two-gas mixture, based on only unlabeled voltage-based gas sensing information. Detailed classification was performed through a multitude of supervised algorithms, i.e., 2-layer artificial neural networks (ANNs), 4-layer deep neural networks (DNNs), 1-dimensional convolutional neural networks (1D CNNs), and 2-dimensional CNNs (2D CNNs). The numerical-based DNNs and image-based CNNs are shown to be excellent approaches for gas detection and classification, as indicated by the highest accuracy and lowest loss indicators. Through the analysis of the influence of the number of sensors on the arrayed gas sensor system, the application of machine learning methodology to an arrayed gas sensor system demonstrates four unique features, i.e., a data augmentation methodology, machine learning approach of combining K-means clustering and neural networks, and a systematic approach to optimized sensor combinations, potentially leading to the practical sensor networks based on chemical sensors. Even two SMO sensor combinations are shown to be highly effective in gas discrimination against diverse gas environments assisted through numeric-based DNNs and image-based 1D CNNs, overcoming the simple clustering proposed through the unsupervised K-means clustering.

SUMMARY Spermatogonial stem cells (SSCs) are essential for male fertility. Here, we report that mouse SSC generation is driven by a transcription factor (TF) cascade controlled by the homeobox protein, RHOX10, which acts by driving the differentiation of SSC precursors called pro-spermatogonia (ProSG). We identify genes regulated by RHOX10 in ProSG in vivo and define direct RHOX10-target genes using several approaches, including a rapid temporal induction assay: iSLAMseq. Together, these approaches identify temporal waves of RHOX10 direct targets, as well as RHOX10 secondary-target genes. Many of the RHOX10-regulated genes encode proteins with known roles in SSCs. Using an in vitro ProSG differentiation assay, we find that RHOX10 promotes mouse ProSG differentiation through a conserved transcriptional cascade involving the key germ-cell TFs DMRT1 and ZBTB16. Our study gives important insights into germ cell development and provides a blueprint for how to define TF cascades., Graphical abstract, In brief Using a battery of genome-wide approaches, Tan et al. identify genes regulated by the RHOX10 transcription factor in mouse pro-spermatogonia. Temporal waves of RHOX10 targets, including both direct and indirect targets, are defined. Among these genes are those that participate in a conserved transcription factor signaling cascade driving pro-spermatogonia differentiation.

Transposable elements (TEs) are mobile sequences that engender widespread mutations and thus are a major hazard that must be silenced. The most abundant active class of TEs in mammalian genomes is long interspersed element class 1 (LINE1). Here, we report that LINE1 transposition is suppressed in the male germline by transcription factors encoded by a rapidly evolving X-linked homeobox gene cluster. LINE1 transposition is repressed by many members of this RHOX transcription factor family, including those with different patterns of expression during spermatogenesis. One family member—RHOX10—suppresses LINE1 transposition during fetal development in vivo when the germline would otherwise be susceptible to LINE1 activation because of epigenetic reprogramming. We provide evidence that RHOX10 suppresses LINE transposition by inducing Piwil2, which encodes a key component in the Piwi-interacting RNA pathway that protects against TEs. The ability of RHOX transcription factors to suppress LINE1 is conserved in humans but is lost in RHOXF2 mutants from several infertile human patients, raising the possibility that loss of RHOXF2 causes human infertility by allowing uncontrolled LINE1 expression in the germline. Together, our results support a model in which the Rhox gene cluster is in an evolutionary arms race with TEs, resulting in expansion of the Rhox gene cluster to suppress TEs in different biological contexts.

Transposable elements (TEs) are mobile sequences that engender widespread mutations and thus are a major hazard that must be silenced. The most abundant active class of TEs in mammalian genomes is long interspersed element class 1 (LINE1). Here, we report that LINE1 transposition is suppressed in the male germline by transcription factors encoded by a rapidly evolving X-linked homeobox gene cluster. LINE1 transposition is repressed by many members of this RHOX transcription factor family, including those with different patterns of expression during spermatogenesis. One family member-RHOX10-suppresses LINE1 transposition during fetal development in vivo when the germline would otherwise be susceptible to LINE1 activation because of epigenetic reprogramming. We provide evidence that RHOX10 suppresses LINE transposition by inducing Piwil2, which encodes a key component in the Piwi-interacting RNA pathway that protects against TEs. The ability of RHOX transcription factors to suppress LINE1 is conserved in humans but is lost in RHOXF2 mutants from several infertile human patients, raising the possibility that loss of RHOXF2 causes human infertility by allowing uncontrolled LINE1 expression in the germline. Together, our results support a model in which the Rhox gene cluster is in an evolutionary arms race with TEs, resulting in expansion of the Rhox gene cluster to suppress TEs in different biological contexts.

DNA-barcoded antibodies enable single-cell multiomics approaches that examine protein alongside mRNA in a single cell. Although it is a powerful approach, leading methods are currently limited to surface proteins, which limits understanding of signal-transduction and transcriptional pathways that are often governed by expression changes and posttranslational modifications of intracellular proteins. The ability to analyze intracellular proteins alongside the transcriptome is complicated by the fix and perm processes necessary to access intracellular epitopes, which can negatively impact RNA assay sensitivity. Furthermore, nonspecific binding of antibody-oligos to nucleic acids can generate high background signals. To address these issues, we generated a protocol that can recover high-quality RNA alongside specific intracellular and surface protein information. Using this protocol in conjunction with the BD Rhapsody™ Single-Cell Analysis System, we simultaneously profiled over 50 protein markers including 14 intracellular proteins with the whole transcriptome. We identified predicted patterns, including increased phosphorylation of AKT protein correlated with the proliferation marker Ki67 upon immune stimulation, and transcription factor (T-bet) expression in a specific cell subset. Together these preliminary results suggest that analysis of intracellular proteins together with the transcriptome may be possible using single-cell sequencing technologies and can enable a deeper understanding of cellular states. For Research Use Only. Not for use in diagnostic or therapeutic procedures.

Spermatogonial stem cells (SSCs) are essential for male fertility. How they initially form is poorly understood. Here, we report that SSC generation is driven by a transcription factor (TF) cascade controlled by the homeobox protein RHOX10, which acts by stimulating the differentiation of SSC precursors called pro-spermatogonia (ProSG). We identified genes regulated by RHOX10 in ProSG in vivo using genome-wide transcriptome profiling. We defined RHOX10-direct target genes using the CUT&Tag protocol coupled with a rapid temporal induction assay we designed: inducible thiol(SH)-linked alkylation for the metabolic sequencing of RNA (iSLAMseq). Together, these genome-wide assays identified temporal waves of RHOX10-direct targets, as well as secondary-target genes. Many of these RHOX10-regulated genes encode proteins with known roles in SSCs. Using an in vitro ProSG differentiation assay that we developed, we found that RHOX10 promotes ProSG differentiation through a conserved transcriptional cascade involving the key germ-cell TFs DMRT1 and ZBTB16. Our study gives important insights into germ cell development and provides a blueprint for how to define TF cascades.

Spermatogonial stem cells (SSCs) are essential for the generation of sperm and have potential therapeutic value for treating male infertility, which afflicts >100 million men world-wide. While much has been learned about rodent SSCs, human SSCs remain poorly understood. Here, we molecularly characterize human SSCs and define conditions favoring their culture. To achieve this, we first identified a cell-surface protein, PLPPR3, that allowed purification of human primitive undifferentiated spermatogonia (uSPG) highly enriched for SSCs. Comparative RNA-sequencing analysis of these enriched SSCs with differentiating SPG (KIT+ cells) revealed the full complement of genes that shift expression during this developmental transition, including genes encoding key components in the TGF-β, GDNF, AKT, and JAK-STAT signaling pathways. We examined the effect of manipulating these signaling pathways on cultured human SPG using both conventional approaches and single-cell RNA-sequencing analysis. This revealed that GDNF and BMP8B broadly support human SPG culture, while activin A selectively supports more advanced human SPG. One condition-AKT pathway inhibition-had the unique ability to selectively support the culture of primitive human uSPG. This raises the possibility that supplementation with an AKT inhibitor could be used to culture human SSCs in vitro for therapeutic applications.

Pro-spermatogonia (SG) serve as the gateway to spermatogenesis. Using single-cell RNA sequencing (RNAseq), we studied the development of ProSG, their SG descendants, and testicular somatic cells, during the perinatal period in mice. We identified both gene and protein markers for 3 temporally distinct ProSG cell subsets, including a migratory cell population with a distinct transcriptome from the previously defined T1- and T2-ProSG stages. This intermediate (I)-ProSG subset translocates from the center of seminiferous tubules to the spermatogonial stem cell (SSC) “niche” in its periphery soon after birth. We identified 3 undifferentiated SG subsets at postnatal day 7, each of which express distinct genes, including transcription factor and signaling genes. Two of these subsets have the characteristics of newly emergent SSCs. We also molecularly defined the development of Sertoli, Leydig, and peritubular myoid cells during the perinatal period, allowing us to identify candidate signaling pathways acting between somatic and germ cells in a stage-specific manner during the perinatal period. Our study provides a rich resource for those investigating testicular germ and somatic cell developmental during the perinatal period.

Comment on “A microRNA cluster in the Fragile‐X region expressed during spermatogenesis targets FMR1” by Ramaiah et al.[Image: see text]

As digital devices which are capable of viewing contents easily such as mobile phones and tablet PCs have become widespread, the number of digital crimes using these digital contents also increases. Usually, the data which can be the evidence of crimes is compressed and the header of data is damaged to conceal the contents. Therefore, it is necessary to identify the characteristics of the entire bits of the compressed data to discriminate the compression type not using the header of the data. In this paper, we propose a method for distinguishing 16 dictionary-based compression types. We utilize 5-layered Convolutional Neural Network (CNN) for classification of compression type using Spatial Pyramid Pooling (SPP) layer. We evaluate our proposed method on the Wikileaks Dataset, which is a text file database. The average accuracy of 16 dictionary-based compression algorithms is 99%. We expect that our proposed method will be useful for providing evidence for Digital Forensics.

High-throughput single cell RNA sequencing (scRNA-seq) has recently emerged as a powerful tool for profiling complex cell populations. While traditional scRNA-seq captures information at the transcript level, recent technology using DNA-barcoded antibodies enable multiomic analysis, examining protein alongside mRNA. Although this approach allows analyzing a large number of protein markers, the sequencing cost can escalate quickly for large panels, particularly ones with high antigen-density markers, and workflows can be long. Therefore, panels should be optimized to gain meaningful results efficiently and cost-effectively. Here, we introduce the BD® AbSeq Immune Discovery Panel (IDP), a lyophilized panel consisting of 30 antibodies against major human immune cell-types and immune response markers for analyzing multiple immune cell subsets. The BD® AbSeq IDP can be used to uncover phenotypic changes in cell subsets such as unique cell input differences or changes in the immune cell status. To test this capability, we used the IDP alongside a mRNA analysis using targeted mRNA panel and whole transcriptome assay to compare the immune phenotypes of cells from disease and in vitro stimulated samples at the single-cell level using the BD Rhapsody™ System. We propose that the BD® AbSeq IDP can be used to identify the cell types and molecular pathways impacted by the disease at the single cell level. For Research Use Only. Not for use in diagnostic or therapeutic procedures. BD, the BD Logo, and Rhapsody are trademarks of Becton, Dickinson and Company or its affiliates. © 2019 BD. All rights reserved.

INTRODUCTION AND OBJECTIVES:Over 100 million men worldwide suffer from infertility. In most cases, the underlying cause is unknown. Treating human infertility requires detailed knowledge of human s...

Medical imaging devices such as X-ray, CT and MRI provide information such as disease, abnormality and anatomic land-mark of inside of human body which cannot be seen directly. This information can be used for diagnosis or surgical planning. Since there are hundreds of slices to examine for CT and MRI data, it is time-consuming to examine all slices. Also, some noises inherent in these devices disturb the diagnosis process. For an efficient and clear Computer Aided Diagnosis, it is im-portant to process the images in order to remove the degrada-tion and to extract meaningful information from slices. This is the reason why the computer vision algorithms are essential for the Computer Aided Diagnosis.In the Computer Aided Diagnosis, extracting surgical infor-mation from image is processed through pattern recognition algorithms. In the pattern recognition procedure, image pro-cessing method is applied to eliminate possible noises from the imaging device and to enhance the image quality such as image contrast and sharpness. After that, the structural information of vessel, bone, organs is extracted via a segmentation method. Segmentation is the process which separates an image into seg -ments based on its pixel information. Each segment should rep-resent a region of human body on the image. Finally, the pattern

The objective of the present study was to compare the contents of antioxidants (sesamol and sesamolin) and benzo(a)pyrene (BaP), as well as the volatile profiles in sesame oil obtained from sesame seeds cultivated in four different areas in Korea, and those cultivated in other countries, including Ethiopia, India, and China. The sesamol content was lower in Korean sesame oils than in Ethiopian, Indian, and Chinese sesame oils, whereas sesamolin content was higher in sesame oils derived from Korea than those in sesame oils derived from other countries. There was also a difference in the contents of BaP in the sesame oils obtained from seeds in different cultivated areas. The volatiles that allowed Korean samples to be discriminated from those of other countries were mainly pyrroles and sulfur-containing compounds such as 1-ethyl-1H-pyrrole, 2-ethyl-4-methyl-1H-pyrrole, 2,4-dimethylthiazole, and 5-ethyl-4-methylthiazole.

The Augmented Reality (AR) and Virtual Reality (VR) tech -nologies provide many contents that we can barely experience in reality. Further, the developments in performance of comput-er and GPU are based on putting virtual environment into our reality efficiently. The core technologies in developing contents of AR and VR are 3D character modeling and 3D character ani -mation. These are used in various areas such as 3D film, game industry, Physics, Medical simulation and Educational contents. In this review, we discuss the method of animating contents in the virtual reality. There are two topics of 3D character animation, namely, Fa-cial animation and Body animation. Facial animation implies the representation of natural facial expressions. There are two methods of producing this type of animation. One is the Geom-etry-based approach which is based on control point in 3D char-acter face mesh and the other is Blend shape model-based ap-proach which utilizes 3D facial expression template blending. The tree-structured skeleton is usually used for animating body motion. In addition, to achieve more realistic character move-ments, not only physics but also non-rigid deformation of skin or clothes are adopted. In this paper, we discuss Geometry- and Blend shape model-based of facial animation as well as Skeleton- and Physics-based of body animation.

Effects of adsorbent treatments used for reduction of benzo(a)pyrene (BaP) levels in volatile profiles and sesamol and sesamolin antioxidant contents in sesame oil were evaluated. Charcoal-based activated carbons with different mesh sizes and celite were used as adsorbents. Treatment with activated carbons led to reductions in BaP contents of sesame oil. A small charcoal-based activated carbon particle size was more effective for removal of BaP. Celite; however, did not show an effect on the BaP content of sesame oil under the experimental conditions. Volatile profiles of sesame oil were not influenced by addition of any adsorbent, and the sesamol and sesamolin contents were reduced by all adsorbents.

High-throughput single cell RNA sequencing (RNA-seq) has recently emerged as a powerful tool for profiling complex cell populations. While traditional single cell RNA-seq captures information about transcript expression, recent technological advances using oligo-conjugated antibodies enable simultaneous detection of proteins alongside mRNA in high-throughput sequencing. This multiomic technology enables high parameter protein analysis that can simultaneously discriminate 100 protein markers. However, experiment workflow time and cost can escalate quickly with high plexy assays. To address these issues we investigated a number of methods to decrease cost and assay time while maintaining data quality using the BD® AbSeq on the BD Rhapsody™ Single-Cell Analysis sequencing system. To begin, we investigated the performance of 20 core proteins when used in a 20plex, 40plex, 60plex, 80plex, or 100plex in model cells. Results from 3 experiments showed that performance was maintained as plexy increased, however rigorous experimental approaches must be followed to minimize impact on specificity or sensitivity. After establishing performance of this 20plex we investigated pre-cocktailing of antibodies as a method to shorten the day-of-experiment workflow, and mixing of oligo-labeled and unlabeled antibodies to high-expressing proteins (“signal muting”) as a way to decrease sequencing costs. Results showed a decrease in specificity as soon as 24 hours after the cocktailing of antibodies and increasing non-specific binding with longer cocktail storage times. On the other hand, signal muting was effective at decreasing sequencing requirements by up to 70%, without impacting data for high expressors. Together these results can be used to aid in optimizing experimental design when working with high-plex AbSeq protein panels, to better harness the information gained from multiomic single cell sequencing data. For Research Use Only. Not for use in diagnostic or therapeutic procedures.BD, the BD Logo, and Rhapsody are trademarks of Becton, Dickinson and Company or its affiliates. © 2019 BD. All rights reserved. Citation Format: Punya Narayan, Hye-Won Song, Margaret Nakamoto, Elisabeth Walczak, Katherine Lazaruk, Stefanie Mortimer. Optimizing experimental design for high-parameter protein analysis using BD AbSeq technology [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 1348.

Advances in high-throughput single cell whole transcriptome analysis (WTA) have enabled the discovery of biomarkers and cellular pathways critical to resolving diversity in diseases. Despite these advances, challenges still remain in comprehensively profiling single cells. Here we address two challenges, limited use of frozen tissues, and inability to resolve post-transcriptional differences from cell to cell. Single-cell RNAseq often requires intact single cells to get good resolution of signal, preventing the use of archival frozen samples. To overcome this limitation, we have tested the possibility to use dissociated nuclei that can be isolated from frozen samples in the BD RhapsodyTM Single-Cell Analysis system. As a proof of concept, we compared high throughput single-nucleus WTA data to that produced from single-cell RNAseq from the same cell type. Our data showed that mRNAs from single nuclei can be captured, and amplified using the BD RhapsodyTM system. Furthermore although the number of molecules detected is lower in nucleus, probably due to lower mRNA content, their expression profiles are well correlated with Pearson's correlation coefficient over 0.9. This opens up the possibility to conduct single cell transcriptomic analysis on the large pool of archived frozen specimens, expanding the variety and number of disease samples that can be examined at the single cell level in the BD RhapsodyTM system. A second limitation of WTA analysis is that it only provides information about gene regulation at the transcript level. Given that regulation critical for cellular pathways is often found not only at the transcriptional level but also at the post-transcriptional level, quantitative analysis of proteins and mRNAs at the single cell level can provide deeper understanding of the disease cells. Here, we used DNA-barcoded antibodies BD® AbSeq to enable multiomic analysis, examining protein alongside mRNA expression and enabling simultaneous transcriptional and post-transcriptional gene profiling in single cells. Our data show that upon activation human peripheral blood mononuclear cells and isolated T cells undergo regulation of markers at the protein level that cannot be resolved at the mRNA level. Several key markers used to define activation states including CD69 and L-Selectin were found to be regulated at the protein level confirming that addition of protein analysis to WTA can generate more comprehensive cell profiling. These approaches offer flexibility and choice in experimental design and allow users to utilize archived frozen samples or to obtain mRNA and protein profiles from single cells. For Research Use Only. Not for use in diagnostic or therapeutic procedures. BD, the BD Logo, and Rhapsody are trademarks of Becton, Dickinson and Company or its affiliates. © 2019 BD. All rights reserved. Citation Format: Hye-Won Song, Gretchen Y. Lam, Margaret Nakamoto, Punya Narayan, Ian Taylor, Stefanie Mortimer. Advances in single cell whole transcriptome analysis: Single nucleus RNAseq and simultaneous protein and mRNA profiling using the BD RhapsodyTM Single-Cell Analysis system and BD® AbSeq [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 1345.

Obesity can weaken the body's immune system and reduce its ability to fight off infections. Current analysis on immune cells from obese mouse models suggests that the immune cells suffer from chronic inflammation. To understand the root cause of this problem, we used single cell sequencing to examine thousands of immune cells isolated from primary and secondary lymphoid organs as well as adipose tissue and compared a diet-induced obesity mouse model with control mice. Cells were stained with 30 BD® AbSeq DNA-barcoded antibodies to enable multiomic analysis, i.e. examining protein alongside mRNA expression in tandem. We also utilized DNA-barcoded universal antibodies from the BD® Single-Cell Multiplexing Kit, which allowed us to combine 8 samples from different mice and tissue types into a single pooled sample, significantly reducing experimental scale and cost while eliminating potential batch effects. The pooled samples were loaded on the BD Rhapsody™ system to perform cell lysis and individual mRNA and cell barcoding, allowing measurement of ~400 immune-related mRNAs and 30 proteins at the single cell level. We were able to efficiently de-multiplex the pooled samples after sequencing. The targeted mRNA and AbSeq panel provided robust clustering of immune cell types and showed that genes related to critical immune responses, including inflammation and lymphocyte activation, are differentially regulated in specific immune-cell subsets in the obese mouse model. Using this multiomic analysis of genes differentially regulated in immune cells from different tissues, we propose a model to explain the immuno-phenotype we observed in obese mouse. For Research Use Only. Not for use in diagnostic or therapeutic procedures. BD, the BD Logo, and Rhapsody are trademarks of Becton, Dickinson and Company or its affiliates. © 2019 BD. All rights reserved. Citation Format: Hye-Won Song, Gisele Baracho, Ian Taylor, Stephanie Widmann, Stefanie Mortimer. An efficient multiplexing approach to avoid batch effects in single cell RNA- and Ab-seq studies demonstrated in a mouse model of chronic inflammation and obesity [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 2750.

High-throughput single cell RNA sequencing (scRNA-seq) has recently emerged as a powerful tool for profiling complex cell populations. While traditional scRNA-seq captures information at transcript level, recent technology using DNA-barcoded antibodies enable multiomic analysis, examining protein alongside mRNA. Although this approach allows high parameter antibody panels, the sequencing cost can escalate quickly for high plexy panel with high antigen density markers. Therefore, thought should be put on panel design in order to gain meaningful results efficiently. Here, we have designed a panel for analyzing multiple immune responses, BD® AbSeq pan immune-response panel. Our immune response panel excludes abundant cell lineage markers which take up a huge amount of reads but provide only cell lineage information that can be easily resolved by RNA profiles only. This panel is focused on analyzing the various immune responses in multiple cell lineages. Therefore, it has a potential to be used to uncover phenotypic changes upon disease status with various immune stimuli. To test the capability of this AbSeq panel in discovery of phenotypic changes in multiple immune response, we have tested it in the various scenarios. We propose that the BD® AbSeq pan immune-response panel can be used to evaluate unpredictable disease phenotypes to identify the cell types affected from the disease as well as the responsible molecular pathways of the disease at a single cell level. For Research Use Only. Not for use in diagnostic or therapeutic procedures. BD, the BD Logo, and Rhapsody are trademarks of Becton, Dickinson and Company or its affiliates. © 2019 BD. All rights reserved.

Blendshape is one of the most common facial representation used for 3D animation, 3D game and virtual reality. In this paper, four representative blendshape approaches are benchmarked: global, delta, mean-delta, and SVD-based blend-shapes. When fitting the blendshape models to sparse facial points, the obtained facial shape highly depends on fitting approach due to the lack of the fitted points. Therefore, it is important to set up appropriate criteria for comparing and verifying the performance of the approaches. In this paper, we use four kinds of metrics that are utilized to measure the performance of the approaches: fitting, landmark, and vertex errors and coefficient sparsity. Through the experimental results, it is verified that the benchmarks are very effective to measure the subjective quality of blendshape.

Facial landmark is one of the most basic elements for obtaining facial information such as facial expression and emotion. However, detecting dense landmarks on an image is challenging due to various facial poses. In this paper, a deep architecture for dense facial landmark detection, called ConcatNet, is proposed. In our architecture, we propose a CNN-based dense landmark detector on part regions of a face, which extends a given set of sparse landmarks to more accurate and dense landmarks. By introducing interface layers for coordinate normalization and part region localization, we concatenate a network for sparse landmark detection to ConcatNet in a global-to-local manner and the whole network to operate in an end-to-end manner. The experimental results on LFW and 300W datasets show that ConcatNet not only expands the number of the sparse landmarks but also increases the accuracy of the landmark positions remarkably. Also, ConcatNet shows high accuracy in detecting the dense landmarks with a smaller dataset and without additional data on an image such as 3D position annotations when compared to 3D model-based detection method.

In this paper, we present a robust framework for facial pose estimation from binocular stereoscopic vision. Unlike prior work on the facial pose estimation that employs the whole landmarks even located in the wrong position, we propose a landmark selection method to remove the erroneous landmarks for better performance, especially in the large facial pose case. For this purpose, we train a convolutional neural network (CNN) in order to measure the confidence of each facial landmark detected by using a well-known landmark detection algorithm. Also, by fitting selected landmarks to 3D space, our framework becomes more robust even when a small number of landmarks are selected. Due to the absence of public dataset for the binocular stereo facial pose, we construct facial pose data sets using a motion sensor for performance validation. In our experiments, our method achieves the higher accuracy of the pose estimation than the previous method, especially for large facial pose cases.

Testis‐expressed X‐linked genes typically evolve rapidly. Here, we report on a testis‐expressed X‐linked microRNA (miRNA) cluster that despite rapid alterations in sequence has retained its position in the Fragile‐X region of the X chromosome in placental mammals. Surprisingly, the miRNAs encoded by this cluster (Fx‐mir) have a predilection for targeting the immediately adjacent gene, Fmr1, an unexpected finding given that miRNAs usually act in trans, not in cis. Robust repression of Fmr1 is conferred by combinations of Fx‐mir miRNAs induced in Sertoli cells (SCs) during postnatal development when they terminate proliferation. Physiological significance is suggested by the finding that FMRP, the protein product of Fmr1, is downregulated when Fx‐mir miRNAs are induced, and that FMRP loss causes SC hyperproliferation and spermatogenic defects. Fx‐mir miRNAs not only regulate the expression of FMRP, but also regulate the expression of eIF4E and CYFIP1, which together with FMRP form a translational regulatory complex. Our results support a model in which Fx‐mir family members act cooperatively to regulate the translation of batteries of mRNAs in a developmentally regulated manner in SCs.

As digital devices which are capable of viewing contents easily such as mobile phones and tablet PCs have become widespread, publications are being digitized rapidly and the market for digital publications is growing up. However, digital publications are illegally distributed in the form of digital images. It is necessary to identify each digital image for protecting the copyrights of digital publications. We can detect the copyright infringement using publication identification algorithm after applying publication classification. In this paper, we suggest the publication classification method for mainly 4 types such as text, cartoon, webtoon, and regular picture. We use 2-layered CNN for publication classification using histogram images, which are extracted by NSS(Natural Scene Statistics), which usually is used for figuring out distortion in a natural image. We expect that our proposed method will be useful for protecting copyrights of publications.

Single cell RNA-sequencing (scRNA-seq) is a powerful tool for understanding the sample heterogeneity of individual cells. Many methods rely on whole transcriptome analysis (WTA) to get a snapshot of the entire cellular landscape. Although WTA analysis can be used to discover novel biomarkers, this technique can be expensive. To enable scaling of experiments, WTA data can be mined to design targeted gene panels. To showcase this, we used single cell sequencing to examine thousands of B cells isolated from the bone marrow and peripheral blood of chronic lymphocytic leukemia (CLL) and healthy donors. B cells that had been sorted using the BD FACSMelody™ cell sorter were multiplexed using the BD™ Human Single-Cell Multiplexing Kit and pooled before being processed on the BD Rhapsody™ system, thereby minimizing batch effects. The resulting data was mined to design a panel of differentially expressed genes between CLL and healthy B cells that could be used for subsequent CLL phenotyping. By combining this panel with the BD Rhapsody™ Immune Response Panel (together comprising ~500 mRNAs), along with 36 DNA-barcoded BD™ AbSeq antibodies, we were able to simultaneously analyze mRNA and protein targets from a new subset of CLL and healthy B cells for additional high-resolution analysis. This study showcases the power of using WTA data to design specific gene panels that can be used alone or in combination with existing targeted panels for routine and cost-effective transcriptional profiling at a single cell level. For Research Use Only. Not for use in diagnostic or therapeutic procedures. BD, the BD Logo, FACSMelody, and Rhapsody are trademarks of Becton, Dickinson and Company. © 2019 BD and its subsidiaries. All rights reserved.

Obesity can weaken the body’s immune system and trigger chronic inflammation. To understand the root cause of this problem, we used single cell sequencing to examine thousands of immune cells isolated from primary and secondary lymphoid organs as well as adipose tissue and compared a diet-induced obesity mouse model with control mice. Cells were stained with 30 DNA-barcoded antibodies from BD™ AbSeq reagents to enable multiomic analysis, i.e. examining protein alongside mRNA expression in tandem. We also utilized DNA-barcoded universal antibodies from the BD™ Single-Cell Multiplexing Kit, which allowed us to combine 8 samples from different mice and tissue types into a single pooled sample, significantly reducing experimental scale and cost while eliminating potential batch effects. The pooled samples were loaded on the BD Rhapsody™ system to perform cell lysis and individual mRNA and cell barcoding, allowing measurement of ~400 immune-related mRNAs and 30 proteins at the single cell level. We were able to de-multiplex the pooled samples with high specificity after sequencing. The targeted mRNA and AbSeq panel provided robust clustering of immune cell types and showed that genes related to critical immune responses, including inflammation and lymphocyte activation, are differentially regulated in specific immune-cell subsets in obese mouse. Using this multiomic analysis of genes differentially regulated in immune cells from different tissues, we propose a model to explain the immuno-phenotype we observed in obese mouse. For Research Use Only. Not for use in diagnostic or therapeutic procedures. BD, the BD Logo, and Rhapsody are trademarks of Becton, Dickinson and Company. © 2019 BD and its subsidiaries. All rights reserved.

study question: What human tissues and cell types express the X-linked reproductive homeobox (RHOX) gene cluster? summary answer: The RHOX homeobox genes and proteins are selectively expressed in germ cells in both the ovary and testis. what is known already: The RHOX homeobox transcription factors are encoded by an X-linked gene cluster whose members are selectively expressed in the male and female reproductive tract of mice and rats. The Rhox genes have undergone strong selection pressure to rapidly evolve, making it uncertain whether they maintain their reproductive tissue-centric expression pattern in humans, an issue we address in this report. study design, size, duration: We examined the expression of all members of the human RHOX gene cluster in 11 fetal and 8 adult tissues. The focus of our analysis was on fetal testes, where we evaluated 16 different samples from 8 to 20 weeks gestation. We also analyzed fixed sections from fetal testes, adult testes and adult ovaries to determine the cell type-specific expression pattern of the proteins encoded by RHOX genes. participants/materials, setting, methods: We used quantitative reverse transcription –polymerase chain reaction analysis to assay human RHOX gene expression. We generated antisera against RHOX proteins and used them for western blotting, immunohistochemical and immunofluorescence analyses of RHOXF1 and RHOXF2/2B protein expression. main results and the role of chance: We found that the RHOXF1 and RHOXF2/2B genes are highly expressed in the testis and exhibit low or undetectable expression in most other organs. Using RHOXF1- and RHOXF2/2B-specific antiserum, we found that both RHOXF1 and RHOXF2/2B are primarily expressed in germ cells in the adult testis. Early stage germ cells (spermatogonia and early spermatocytes) express RHOXF2/2B, while later stage germ cells (pachytene spermatocytes and round spermatids) express RHOXF1. Both RHOXF1 and RHOXF2/2B are expressed in prespermatogonia in human fetal testes. Consistent with this, RHOXF1 and RHOXF2/2B mRNA expression increases in the second trimester during fetal testes development when gonocytes differentiate into prespermatogonia. In the human adult ovary, we found that RHOXF1 and RHOXF2/2B are primarily expressed in oocytes. limitations, reasons for caution: While the average level of expression of RHOX genes was low or undetectable in all 19 human tissues other than testes, it is still possible that RHOX genes are highly expressed in a small subset of cells in some of these nontesticular tissues. As a case in point, we found that RHOX proteins are highly expressed in oocytes within the human ovary, despite low levels of RHOX mRNA in the whole ovary. wider implications of the findings: The cell type-specific and developmentally regulated expression pattern of the RHOX transcription factors suggests that they perform regulatory functions during human fetal germ cell development, spermatogenesis and

The X-linked reproductive homeobox (RHOX) gene cluster encodes transcription factors preferentially expressed in reproductive tissues. This gene cluster has important roles in male fertility based on phenotypic defects of Rhox-mutant mice and the finding that aberrant RHOX promoter methylation is strongly associated with abnormal human sperm parameters. However, little is known about the molecular mechanism of RHOX function in humans. Using gene expression profiling, we identified genes regulated by members of the human RHOX gene cluster. Some genes were uniquely regulated by RHOXF1 or RHOXF2/2B, while others were regulated by both of these transcription factors. Several of these regulated genes encode proteins involved in processes relevant to spermatogenesis; e.g. stress protection and cell survival. One of the target genes of RHOXF2/2B is RHOXF1, suggesting cross-regulation to enhance transcriptional responses. The potential role of RHOX in human infertility was addressed by sequencing all RHOX exons in a group of 250 patients with severe oligozoospermia. This revealed two mutations in RHOXF1 (c.515G > A and c.522C > T) and four in RHOXF2/2B (-73C > G, c.202G > A, c.411C > T and c.679G > A), of which only one (c.202G > A) was found in a control group of men with normal sperm concentration. Functional analysis demonstrated that c.202G > A and c.679G > A significantly impaired the ability of RHOXF2/2B to regulate downstream genes. Molecular modelling suggested that these mutations alter RHOXF2/F2B protein conformation. By combining clinical data with in vitro functional analysis, we demonstrate how the X-linked RHOX gene cluster may function in normal human spermatogenesis and we provide evidence that it is impaired in human male fertility.

Gene duplication is a major evolutionary force driving adaptation and speciation, as it allows for the acquisition of new functions and can augment or diversify existing functions. Here, we report a gene duplication event that yielded another outcome--the generation of antagonistic functions. One product of this duplication event--UPF3B--is critical for the nonsense-mediated RNA decay (NMD) pathway, while its autosomal counterpart--UPF3A--encodes an enigmatic protein previously shown to have trace NMD activity. Using loss-of-function approaches in vitro and in vivo, we discovered that UPF3A acts primarily as a potent NMD inhibitor that stabilizes hundreds of transcripts. Evidence suggests that UPF3A acquired repressor activity through simple impairment of a critical domain, a rapid mechanism that may have been widely used in evolution. Mice conditionally lacking UPF3A exhibit "hyper" NMD and display defects in embryogenesis and gametogenesis. Our results support a model in which UPF3A serves as a molecular rheostat that directs developmental events.

The generation of functional sperm in vitro has been a goal for almost a century. Until recently, researchers have only succeeded in reproducing the early steps of spermatogenesis. This is not surprising given that spermatogenesis is a complicated process that requires the coordinated efforts of germ cells and several somatic cells within the tubular structure of the testis. Finally—last year—Sato et al. reported the successful in vitro production of functional sperm, thereby potentially opening up a new era of reproductive biology. Here, we summarize the history of research directed toward reproducing steps of spermatogenesis in vitro, detail the seminal findings of Sato et al., and suggest ways that their approach can be applied toward clinical applications and addressing fundamental questions about the underlying mechanism of spermatogenesis.

Homeobox genes encode transcription factors that regulate diverse developmental events. The largest known homeobox gene cluster – the X-linked mouse reproductive homeobox (Rhox) cluster – harbors genes whose expression patterns and functions are largely unknown. Here, we report that a member of this cluster,Rhox10, is expressed in male germ cells.Rhox10is highly transcribed in spermatogoniain vivoand is upregulated in response to the differentiation-inducing agent retinoic acidin vitro. Using a specific RHOX10 antiserum that we generated, we found that RHOX10 protein is selectively expressed in fetal gonocytes, germline stem cells, spermatogonia, and early spermatocytes. RHOX10 protein undergoes a dramatic shift in subcellular localization as germ cells progress from mitotically arrested gonocytes to mitotic spermatogonia and from mitotic spermatogonia to early meiotic spermatocytes, consistent with RHOX10 performing different functions in these stages.

Porphyrias are a group of diseases characterized by an enzyme deficiency in the heme biosynthesis pathway, resulting in accumulation of precursor molecules in the tissue. Some porphyric patients develop progressive liver disease that requires liver transplantation. This case report describes special anesthetic challenges, including careful selection of drugs and the use of special filters that can exclude harmful wavelengths of ultraviolet, in a patient with porphyria who underwent living donor liver transplantation. Understanding the patient's status and disease process, and avoiding triggering factors of porphyria attacks, are important for successful liver transplantation anesthesia in patients with porphyria.

SummaryThe developmental origins of most adult stem cells are poorly understood. Here, we report the identification of a transcription factor—RHOX10—critical for the initial establishment of spermatogonial stem cells (SSCs). Conditional loss of the entire 33-gene X-linked homeobox gene cluster that includes Rhox10 causes progressive spermatogenic decline, a phenotype indistinguishable from that caused by loss of only Rhox10. We demonstrate that this phenotype results from dramatically reduced SSC generation. By using a battery of approaches, including single-cell-RNA sequencing (scRNA-seq) analysis, we show that Rhox10 drives SSC generation by promoting pro-spermatogonia differentiation. Rhox10 also regulates batteries of migration genes and promotes the migration of pro-spermatogonia into the SSC niche. The identification of an X-linked homeobox gene that drives the initial generation of SSCs has implications for the evolution of X-linked gene clusters and sheds light on regulatory mechanisms influencing adult stem cell generation in general.

Summary Nonsense-mediated mRNA decay (NMD) is a conserved RNA decay pathway that degrades aberrant mRNAs and directly regulates many normal mRNAs. This dual role for NMD raises the possibility that its magnitude is buffered to prevent the potentially catastrophic alterations in gene expression that would otherwise occur if NMD were perturbed by environmental or genetic insults. In support of this, here we report the existence of a negative feedback regulatory network that directly acts on seven NMD factors. Feedback regulation is conferred by different branches of the NMD pathway in a cell type-specific and developmentally regulated manner. We identify feedback-regulated NMD factors that are rate limiting for NMD and demonstrate that reversal of feedback regulation in response to NMD perturbation is crucial for maintaining NMD. Together, our results suggest the existence of an intricate feedback network that maintains both RNA surveillance and the homeostasis of normal gene expression in mammalian cells.

The early development of metazoans is mainly regulated by differential translation and localization of maternal mRNAs in the embryo. In general, these processes are orchestrated by RNA-binding proteins interacting with specific sequence motifs in the 3'-untranslated region (UTR) of their target RNAs. Hermes is an RNA-binding protein, which contains a single RNA recognition motif (RRM) and is found in various vertebrate species from fish to human. In Xenopus laevis, Hermes mRNA and protein are localized in the vegetal region of oocytes. A subpopulation of Hermes protein is concentrated in a specific structure in the vegetal cortex, called the germ plasm (believed to contain determinants of the germ cell fate) where Hermes protein co-localizes with Xcat2 and RINGO/Spy mRNAs. The level of total Hermes protein decreases during maturation. The precocious depletion of Hermes protein by injection of Hermes antisense morpholino oligonucleotide (HE-MO) accelerates the process of maturation and results in cleavage defects in vegetal blastomeres of the embryo. It is known that several maternal mRNAs including RINGO/Spy and Mos are regulated at the translational level during meiotic maturation and early cleavage in Xenopus. The ectopic expression of RINGO/Spy or Mos causes resumption of meiotic maturation and cleavage arrests, which resemble the loss of Hermes phenotypes. We found that the injection of HE-MO enhances the acceleration of maturation caused by the injection of RINGO/Spy mRNA, and that Hermes protein is present as mRNP complex containing RINGO/Spy, Mos, and Xcat2 mRNAs in vivo. We propose that as an RNA-binding protein, Hermes may be involved in maturation, cleavage events at the vegetal pole and germ cell development by negatively regulating the expression of RINGO/Spy, Mos, and Xcat2 mRNAs.

Alginate based polymeric matrices were designed for controlled release and stabilization of cefaclor in gastrointestinal fluid. Cefaclor is known to be acid stable and subjected to be degraded at neutral and alkaline pHs. In order to achieve an effective release profile of cefaclor in gastrointestinal tract, a particular strategy in dosage form design should be required from the view point of maintaining its activity. The amphiphilic nature of cefaclor allowed its controlled release using ionic polymers based on ionic interaction between the drug and polymers. The thrust of this study was to develop a technique that delivers cefaclor keeping effective release rate in the intestinal tract. Considering the fast degradation of cefaclor in the intestinal fluid, the matrices were designed to release surplus amount of cefaclor. The alginate based matrices demonstrated increase in release rate in the simulated intestinal fluid, which was favorable to compensate the degraded portion of cefaclor. In addition, stabilization of cefaclor in the intestinal fluid was obtained by employing citric acid that provides an local acidic environment. The matrices might be valuably used for the development of an oral cefaclor dosage form.