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Whole-organ transcriptomic analyses have emerged as a common method for characterizing developmental transitions in mammalian organs. However, it is unclear if all cell types in an organ follow the whole-organ defined developmental trajectory. Recently, a postnatal two-stage developmental process was described for the mouse stomach. Here, using laser capture microdissection to obtain in situ transcriptomic data, we show that mouse gastric pit cells exhibit four postnatal developmental stages. Interestingly, early stages are characterized by the up-regulation of genes associated with metabolism, a functionality not typically associated with pit cells. Hence, beyond revealing that not all constituent cells develop according to the whole-organ determined pathway, these results broaden our understanding of the pit cell phenotypic landscape during stomach development.

With improvements in the survival rate of patients with cancer, fertility maintenance has become a major concern in terms of cancer treatment for women of reproductive age. Thus, it is important to examine the impact on fertility of anticancer drugs that are used clinically or are undergoing trials. The HuR small-molecule inhibitor MS-444 has been used in many cancer treatment studies, but its reproductive toxicity in females is unknown. Here, we reported that MS-444 blocked the nucleocytoplasmic transport of Agbl2 mRNA by inhibiting HuR dimerization, resulting in the developmental arrest of 2-cell stage embryos in mouse. Combining analysis of low-input RNA-seq for MS-444-treated 2-cell embryos and mapping binding sites of RNA-binding protein, Agbl2 was predicted to be the target gene of MS-444. For further confirmation, RNAi experiment in wild-type zygotes showed that Agbl2 knockdown reduced the proportion of embryos successfully developed to the blastocyst stage: from 71% in controls to 23%. Furthermore, RNA-FISH and luciferase reporter analyses showed that MS-444 blocked the nucleocytoplasmic transport of Agbl2 mRNA and reduced its stability by inhibiting HuR dimerization. In addition, optimized stochastic optical reconstruction microscopy (STORM) imaging showed that MS-444 significantly reduced the HuR dimerization, and HuR mainly existed in cluster form in 2-cell stage embryos. In conclusion, this study provides clinical guidance for maintaining fertility during the treatment of cancer with MS-444 in women of reproductive age. And also, our research provides guidance for the application of STORM in nanometer scale studies of embryonic cells. HuR inhibitor MS-444 arrested embryonic development at 2-cell stage. Low-input RNA-seq revealed that Agbl2 was the target gene of MS-444. MS-444 blocked the nucleocytoplasmic transport of Agbl2 mRNA by inhibiting HuR dimerization and reduced the stability of Agbl2 mRNA. STORM with our optimized protocol showed that HuR tended to form elliptical and dense clusters in 2-cell stage embryos.

There is presently intense interest in the development of synthetic nanopores that recapitulate the functional properties of biological water channels for a wide range of applications. To date, all known synthetic water channels have a hydrophobic lumen, and while many exhibit a comparable rate of water transport as biological water channels, there is presently no rationally designed system with the ability to regulate water transport, a critical property of many natural water channels. Here, we describe a self-assembling nanopore consisting of stacked macrocyclic molecules with a hybrid hydrophilic/hydrophobic lumen exhibiting water transport that can be regulated by alkali metal ions. Stopped-flow kinetic assays reveal a non-monotonic-dependence of transport on cation size as well as a strikingly broad range of water flow, from essentially none in the presence of the sodium ion to as high a flow as that of the biological water channel, aquaporin 1, in the absence of the cations. All-atom molecular dynamics simulations show that the mechanism underlying the observed sensitivity is the binding of cations to defined sites within this hybrid pore, which perturbs water flow through the channel. Thus, beyond revealing insights into factors that can modulate a high-flux water transport through sub-nm pores, the obtained results provide a proof-of-concept for the rational design of next-generation, controllable synthetic water channels., A nanopore of macrocycles with a hydrophilic/hydrophobic lumen exhibits water transport that is regulated by cations, providing a proof-of-concept for next-generation, controllable synthetic channels.

Objective : To alleviate the severe limitation of the prohibitively long process of immune-fluorescence labeling on the routine applications of revolutionary intact tissue clearing techniques in diverse biomedical arenas. Methods : We proposed an easily adaptable approach, electro-enhanced rapid staining (EERS), for highly efficient and fast immuno-labeling of thick clarified tissues. In EERS, an optimized and precisely controlled weak external electric field is engineered into a compact device to enable efficient and uniform transport of antibodies into clarified tissues while minimizing the detrimental effect of macromolecular crowding at the tissue-solution interface. Results and Conclusions : The experimental results show that, with EERS, a current density of only ∼0.2 mA mm−2 is sufficient to achieve uniform labeling of clarified tissues of several millimeters thick in a few hours without detectable tissue damage. In addition, the amount of antibodies required is also several-fold lower than conventional immuno-labeling assays under comparable conditions. Significance : It is expected that the implementation of EERS in most laboratories should significantly expedite the application of tissue clearing in a broad range of research explorations, both basic and clinical.

Chromatin immunoprecipitation coupled with high-throughput sequencing (ChIP-seq) can profile genome-wide epigenetic marks associated with regulatory genomic elements. However, conventional ChIP-seq is challenging when examining limited numbers of cells. Here, we developed a new technique by supplementing carrier materials of both chemically modified mimics with epigenetic marks and dUTP-containing DNA fragments during conventional ChIP procedures (hereafter referred to as 2cChIP-seq), thus dramatically improving immunoprecipitation efficiency and reducing DNA loss of low-input ChIP-seq samples. Using this strategy, we generated high-quality epigenomic profiles of histone modifications or DNA methylation in 10–1000 cells. By introducing Tn5 transposase-assisted fragmentation, 2cChIP-seq reliably captured genomic regions with histone modification at the single-cell level in about 100 cells. Moreover, we characterized the methylome of 100 differentiated female germline stem cells (FGSCs) and observed a particular DNA methylation signature potentially involved in the differentiation of mouse germline stem cells. Hence, we provided a reliable and robust epigenomic profiling approach for small cell numbers and single cells.

The folding of interphase chromatin into highly compact mitotic chromosomes is one of the most recognizable changes during the cell cycle. However, the structural organization underlying this drastic compaction remains elusive. Here, we combine several super resolution methods, including structured illumination microscopy (SIM), binding-activated localization microscopy (BALM), and atomic force microscopy (AFM), to examine the structural details of the DNA within the mitotic chromosome, both in the native state and after up to 30-fold extension using single-molecule micromanipulation. Images of native chromosomes reveal widespread ∼125 nm compact granules (CGs) throughout the metaphase chromosome. However, at maximal extensions, we find exclusively ∼90 nm domains (mitotic nanodomains, MNDs) that are unexpectedly resistant to extensive forces of tens of nanonewtons. The DNA content of the MNDs is estimated to be predominantly ∼80 kb, which is comparable to the size of the inner loops predicted by a recent nested loop model of the mitotic chromosome. With this DNA content, the total volume expected of the human genome assuming closely packed MNDs is nearly identical to what is observed. Thus, altogether, these results suggest that these mechanically stable MNDs, and their higher-order assembly into CGs, are the dominant higher-level structures that underlie the compaction of chromatin from interphase to metaphase.

The precise spatial positioning of functional groups in biomacromolecules leads to astonishing binding, catalytic, and transport capabilities. In contrast, synthetic frameworks capable of convergently locking functional groups with minimized conformational uncertainty are highly desirable but rare. Here we report C5-symmetric aromatic pentaamide macrocycles c5a-c synthesized in one pot from the corresponding monomers. The crystal structure of c5c reveals a fully constrained backbone that enforces ten alternating NH/CH hydrogen-bond donors and five large amide dipoles to point to the center of the macrocycle. With a highly electropositive cavity in a high-energy unbound state, macrocycles c5 bind anions in a 1:1 stoichiometry in solution, with Ka values up to 106 M-1 for halides and 108 M-1 for oxoanions. Macrocycle c5a was able to selectively transport chloride ions across lipid bilayers. The extraordinary binding of phosphate-related ions by c5 also enabled catalytic ester amidation owing to the stabilization of the corresponding transition states.

BackgroundWith the improvement of the survival rate of cancer patients, fertility maintenance has become a major concern of cancer treatment for women of reproductive age. Thus, it is important to examine the impact on fertility of anticancer drugs that have been clinically used or are undergoing trials. The HuR small-molecule inhibitor MS-444 has been used in many cancer treatment studies, but its reproductive toxicity in females is unknown. ResultsCombining analysis of low-input RNA-seq for MS-444-treated 2-cell embryos and mapping binding sites of RNA binding protein, Agbl2 was predicted to be the target gene of MS-444. For further confirmation, RNAi experiment in wild-type zygotes showed that Agbl2 knockdown reduced the proportion of embryos successfully develop to the blastocyst stage: from 71% in controls to 23%. Furthermore, RNA-FISH and luciferase reporter analysis showed that MS-444 blocked the nucleocytoplasmic transport of Agbl2 mRNA and reduced its stability by inhibiting HuR dimerization. In addition, optimized stochastic optical reconstruction microscopy (STORM) imaging showed that MS-444 significantly reduced the HuR dimerization, and HuR mainly existed in the form of clusters in 2-cell stage embryos.ConclusionThose results showed that MS-444 blocked the nucleocytoplasmic transport of Agbl2 mRNA by inhibiting HuR dimerization, resulting in the developmental arrest of 2-cell stage embryos in mouse. This study provides clinical guidance for maintaining fertility during the treatment of cancer with MS-444 in women of reproductive age. And also, our research provides guidance for the application of STORM in the nanometer scale study of embryonic cells.

Accurate reconstruction of the 3D morphology and spatial distribution of myelinated axons in mouse brains is very important for understanding the mechanism and dynamic behavior of long-distance information transmission between brain regions. However, it is difficult to segment and reconstruct myelinated axons automatically due to two reasons: the amount of it is huge and the morphology of it is different between brain regions. Traditional artificial labeling methods usually require a large amount of manpower to label each myelinated axon slice by slice, which greatly hinders the development of the mouse brain connectome. In order to solve this problem and improve the reconstruction efficiency, this paper proposes an annotation generation method that takes the myelinated axon as prior knowledge, which can greatly reduce the manual labeling time while reaching the level of manual labeling. This method consists of three steps. Firstly, the 3D axis equation of myelinated axons is established by sparse axon artificial center point labels on slices, and the region to be segmented is pre-extracted according to the 3D axis. Subsequently, the U-Net network was trained by a small number of artificially labeled myelinated axons and was used for precise segmentation of output by the last step, so as to obtain accurate axon 2D morphology. Finally, based on the segmentation results, the high-precision 3D reconstruction of axons was performed by imaris software, and the spatial distribution of myelinated axons in the mouse brain was reconstructed. In this paper, the effectiveness of this method was verified on the dataset of high-resolution X-ray microtomography of the mouse cortex. Experiments show that this method can achieve an average MIoU 81.57, and the efficiency can be improved by more than 1400x compared with the manual labeling method.

The activation of monocytes and their trans-differentiation into macrophages are critical processes of the immune response. Prior work has characterized the differences in the expression between monocytes and macrophages, but the transitional process between these cells is poorly detailed. Here, we analyzed the temporal changes of the transcriptome during trans-differentiation of primary human monocytes into M0 macrophages. We find changes with many transcription factors throughout the process, the vast majority of which exhibit a maximally different expression at the intermediate stages. A few factors, including AP-1, were previously known to play a role in immunological transitions, but most were not. Thus, these findings indicate that this trans-differentiation requires the dynamic expression of many transcription factors not previously discussed in immunology, and provide a foundation for the delineation of the molecular mechanisms associated with healthy or pathological responses that involve this transition.

Immortalized cell lines have long been used as model systems to systematically investigate biological processes under controlled and reproducible conditions, providing insights that have greatly advanced cellular biology and medical sciences. Recently, the widely used monocytic leukemia cell line, THP-1, was comprehensively examined to understand mechanistic relationships between the 3D chromatin structure and transcription during the trans-differentiation of monocytes to macrophages. To corroborate these observations in primary cells, we analyze in situ Hi-C and RNA-seq data of human primary monocytes and their differentiated macrophages in comparison to that obtained from the monocytic/macrophagic THP-1 cells. Surprisingly, we find significant differences between the primary cells and the THP-1 cells at all levels of chromatin structure, from loops to topologically associated domains to compartments. Importantly, the compartment-level differences correlate significantly with transcription: those genes that are in A-compartments in the primary cells but are in B-compartments in the THP-1 cells exhibit a higher level of expression in the primary cells than in the THP-1 cells, and vice versa. Overall, the genes in these different compartments are enriched for a wide range of pathways, and, at least in the case of the monocytic cells, their altered expression in certain pathways in the THP-1 cells argues for a less immune cell-like phenotype, suggesting that immortalization or prolonged culturing of THP-1 caused a divergence of these cells from their primary counterparts. It is thus essential to reexamine phenotypic details observed in cell lines with their primary counterparts so as to ensure a proper understanding of functional cell states in vivo. Supplementary Information The online version contains supplementary material available at 10.1186/s41065-021-00205-w.

The functioning of tissues is fundamentally dependent upon not only the phenotypes of the constituent cells but also their spatial organization in the tissue. However, acquisition of comprehensive transcriptomes of spatially- and phenotypically-defined cells in situ remains challenging. Here we present a general and robust method based on immunofluorescence-guided laser capture microdissection (immuno-LCM-RNAseq) to acquire finely resolved spatial transcriptomes including isoforms with as few as tens of cells from snap-frozen or RNAlater-treated clinical tissues, circumventing the problem of significant RNA degradation during this time-consuming process. The efficacy of this approach is exemplified by the characterization of differences at the transcript isoform level between the mouse small intestine lacteal cells at the tip versus the main capillary body. With the extensive repertoire of specific antibodies that are presently available, our method provides a powerful means by which spatially resolved cellular states can now be delineated in situ with preserved tissues. Moreover, such high quality spatial transcriptomes defined by immunomarkers can be used to compare with the phenotypes derived from single-cell RNAseq of dissociated cells as well as applied to bead-based spatial transcriptomic approaches that require such information a priori for cell identification.

Atomic force microscopy-based single-molecule-force spectroscopy is limited by low throughput. We introduce addressable DNA origami to study multiple target molecules. Six target DNAs that differed by only a single base-pair mismatch were clearly differentiated a rupture force of only 4 pN.

Immortalized cell lines have long been used as model systems to systematically investigate biological processes under controlled and reproducible conditions, providing insights that have greatly advanced cellular biology and medical sciences. Recently, the widely used monocytic leukemia cell line, THP-1, was comprehensively examined to understand mechanistic relationships between the 3D chromatin structure and transcription during the trans-differentiation of monocytes to macrophages. To corroborate these observations in primary cells, we analyze in situ Hi-C and RNA-seq data of human primary monocytes and their differentiated macrophages in comparison to that obtained from the monocytic/macrophagic THP-1 cells. Surprisingly, we find significant differences between the primary cells and the THP-1 cells at all levels of chromatin structure, from loops to topologically associated domains to compartments. Importantly, the compartment-level differences correlate significantly with transcription: those genes that are in A-compartments in the primary cells but are in B-compartments in the THP-1 cells exhibit a higher level of expression in the primary cells than in the THP-1 cells, and vice versa. Overall, the genes in these different compartments are enriched for a wide range of pathways, and, at least in the case of the monocytic cells, their altered expression in certain pathways in the THP-1 cells argues for a less immune cell-like phenotype, suggesting that immortalization or prolonged culturing of THP-1 caused a divergence of these cells from their native counterparts. It is thus essential to reexamine phenotypic details observed in cell lines with their native counterparts so as to ensure a proper understanding of functional cell states in vivo.

Single-cell chromatin studies provide insights into how chromatin structure relates to functions of individual cells. However, balancing high-resolution and genome wide-coverage remains challenging. We describe a computational method for the reconstruction of large 3D-ensembles of single-cell (sc) chromatin conformations from population Hi-C that we apply to study embryogenesis in Drosophila. With minimal assumptions of physical properties and without adjustable parameters, our method generates large ensembles of chromatin conformations via deep-sampling. Our method identifies specific interactions, which constitute 5–6% of Hi-C frequencies, but surprisingly are sufficient to drive chromatin folding, giving rise to the observed Hi-C patterns. Modeled sc-chromatins quantify chromatin heterogeneity, revealing significant changes during embryogenesis. Furthermore, >50% of modeled sc-chromatin maintain topologically associating domains (TADs) in early embryos, when no population TADs are perceptible. Domain boundaries become fixated during development, with strong preference at binding-sites of insulator-complexes upon the midblastula transition. Overall, high-resolution 3D-ensembles of sc-chromatin conformations enable further in-depth interpretation of population Hi-C, improving understanding of the structure-function relationship of genome organization., Balancing high resolution and broad genome coverage in single-cell Hi-C approaches remains challenging. Here, the authors describe a computational method for the reconstruction of a large 3D-ensemble of single-cell chromatin conformations from population Hi-C measurements and apply this model to study embryogenesis in Drosophila.

Chromatin interactions are important for gene regulation and cellular specialization. Emerging evidence suggests many-body spatial interactions play important roles in condensing super-enhancer regions into a cohesive transcriptional apparatus. Chromosome conformation studies using Hi-C are limited to pairwise, population-averaged interactions; therefore unsuitable for direct assessment of many-body interactions. We describe a computational model, CHROMATIX, which reconstructs ensembles of single-cell chromatin structures by deconvolving Hi-C data and identifies significant many-body interactions. For a diverse set of highly active transcriptional loci with at least 2 super-enhancers, we detail the many-body functional landscape and show DNase accessibility, POLR2A binding, and decreased H3K27me3 are predictive of interaction-enriched regions., Genome Biology, 21, ISSN:1474-760X

Additional file 1 Supplementary information. Detailed methodology for fractal Monte Carlo sampling and Bayesian deconvolution of Hi-C specific contacts. Also, additional figures referenced for this study.

Monocyte-to-macrophage trans-differentiation has long been studied to better understand this immunological response and aspects of developmental processes more generally. A key question is the nature of the corresponding changes in chromatin conformation and its relationship to the transcriptome during this process. This question is especially intriguing since this trans-differentiation is not associated with progression through mitosis, often considered a necessary step for gross changes in chromosomal structure. Here, we characterized the transcriptional and genomic structural changes during macrophage development of primary human monocytes using RNA-seq and in situ Hi-C. We found that, during this transition, the genome architecture undergoes a massive remodeling to a degree not observed before between structured genomes, with changes in ~90% of the topologically associating domains (TADs). These changes in the TADs are associated with changed expression of immunological genes. These structural changes, however, differ extensively from those described recently in a study of the leukemia cell line, THP-1. Furthermore, up-regulation of the AP-1 family of genes that effected functionally important changes in the genomic structure during the differentiation of the THP-1 cells was not corroborated with the primary cells. Taken together, our results provide a comprehensive characterization of the changes in genomic structure during the monocyte-to-macrophage transition, establish a framework for the elucidation of processes underlying differentiation without proliferation, and demonstrate the importance of verifying with primary cells the mechanisms discovered with cultured cells.

Topologically associating domains (TADs) are fundamental elements of the eukaryotic genomic structure. However, recent studies suggest that the insulating complexes, CTCF/cohesin, present at TAD borders in mammals are absent from those in Drosophila melanogaster, raising the possibility that border elements are not conserved among metazoans. Using in situ Hi-C with sub-kb resolution, here we show that the D. melanogaster genome is almost completely partitioned into >4000 TADs, nearly sevenfold more than previously identified. The overwhelming majority of these TADs are demarcated by the insulator complexes, BEAF-32/CP190, or BEAF-32/Chromator, indicating that these proteins may play an analogous role in flies as that of CTCF/cohesin in mammals. Moreover, extended regions previously thought to be unstructured are shown to consist of small contiguous TADs, a property also observed in mammals upon re-examination. Altogether, our work demonstrates that fundamental features associated with the higher-order folding of the genome are conserved from insects to mammals., Topologically associating domain (TAD) boundaries in flies seem to be different from those in mammals. Here, the authors use Hi-C with sub-kb resolution to identify about 4000 TADs in flies, most demarcated by the insulator complexes BEAF-32/CP190 or BEAF-32/Chromator like CTCF/cohesin in mammals.

Mechanistic understanding of how living systems sense, transduce, and respond to mechanical cues has important implications in development, physiology, and therapy. Here, the authors use an integrated atomic force microscope (AFM) and brightfield/epifluorescent microscope platform to precisely simulate living single cells or groups of cells under physiological conditions, in real time, concomitantly measuring the single‐cell autophagic response and its transmission to neighboring cells. Dual‐color fluorescence monitoring of the cellular autophagic response reveals the dynamics of autophagosome formation, degradation, and induction in neighboring contacting and noncontacting cells. Autophagosome formation is dependent on both the applied force and contact area of the AFM tip. More importantly, the enhancement of the autophagic responses in neighboring cells via a gap junction‐dependent mechanism is observed. This AFM‐based nanoacupuncture platform can serve as a tool for elucidating the primary mechanism underlying mechanical stimulation of living systems and other biomechanical therapeutics., An integrated atomic force microscope and brightfield/epifluorescent microscope platform is employed to quantify real‐time force application at the single cell level, which reveals the dynamics of autophagosome formation, degradation, and induction in neighboring contacting and noncontacting cells.

Sentinel lymph node (SLN) imaging and biopsy has been advocated as an important technique to evaluate the metastatic status of regional lymph nodes and determine subsequent surgical procedure for many cancers, yet there is no reliable means to provide accurate and rapid diagnosis of metastatic SLN during surgery. Here we develop a new approach, named “Ratiometric Raman dual-nanotag strategy”, that using folic acid functionalized targeted and nontargeted gap-enhanced Raman tags (FA-GERTs and Nt-GERTs) to detect metastatic SLN based on Raman imaging combined with classical least square data processing methods. By using this strategy, with built-in self-calibration for signal correction, rather than absolute intensity-dependent signal readout, we realize the visualization and prompt intraoperative diagnosis of metastatic SLN with a high accuracy of 87.5 % when the cut-off value of ratio (FA-GERTs/Nt-GERTs) set at 1.255. This approach may outperform the existing histopathological assessment in diagnosing SLN metastasis and is promising for guiding surgical procedure in the future.

Hexakis(m-phenylene ethynylene) (m-PE) macrocycles 1–4, sharing the same hydrogen-bonding side chains but having backbones of different electronic properties, are designed to probe the effectiveness of multiple H-bonding interactions in enforcing columnar assemblies. 1H NMR, absorption, fluorescence, and circular dichroism (CD) spectroscopy indicate that, compared with analogous macrocycles that self-associate based on aromatic stacking which is highly sensitive to the electronic nature of the macrocyclic backbones, macrocycles 1–4 all exhibit strong aggregation down to the micromolar (μM) concentrations in nonpolar solvents. Increasing solvent polarity quickly weakens aggregation. In THF and DMF, the macrocycles exist as free molecules. The observed solvent effects, along with the behavior of 5-F6 that cannot self-associate via H-bonding, confirm that H-bonding plays the dominating role in driving the self-association of 1–4. The backbone electronic nature does not change the self-assembling pattern comm...

As a naturally-abundant biopolymer, chitosan (CS) exhibit pH-sensitive structural transformation within a narrow pH range. Integrating hydrophobic groups to CS molecules gives modified CS polymers with more adjustable pH responsiveness. In this paper, near-infrared (NIR) photoluminescent Ag2S QDs capped by long-chain carboxylic acid were synthesized and then conjugated with CS via esterification reaction. The anticancer drug doxorubicin (DOX) has an affinity for the hydrophobic oleoyl groups and was entrapped by them to produce Ag2S(DOX)@CS nanospheres. A variety of experiments were performed to characterize the nanospheres. In vitro and in vivo experiments showed that the nanospheres can release DOX at lowered pH in tumor cells and have high antitumor efficacy. In addition, the strong NIR signal derived from the encapsulated Ag2S QDs makes real-time monitoring of the nanosphere distribution in a body possible. This study provides a new CS-based nanocomposite drug carrier for efficient cancer therapy.

Chromatin interactions are important for gene regulation and cellular specialization. Emerging evidence suggests many-body spatial interactions can play important roles in condensing super-enhancer regions into a cohesive transcriptional apparatus. Chromosome conformation studies using Hi-C are limited to pairwise, population-averaged interactions; therefore, not suitable for direct assessment of many-body interactions. We describe a computational model, CHROMATIX, that reconstructs structural ensembles based on Hi-C data and identifies significant many-body interactions. For a diverse set of highly-active transcriptional loci with at least 2 super-enhancers, we detail the many-body functional landscape and show DNase-accessibility, POLR2A binding, and decreased H3K27me3 are predictive of interaction-enriched regions.

Accurate segmentation of the acquired 3D stack fluorescence images of mice gland is a prerequisite for qualitative and quantitative analysis. However, the traditional methods cannot fully satisfy the need of accurate segmentation for 3D reconstruction result of the generally low signal-to-noise ratio of the 3D fluorescence images and the unique structure of mice gland. In this article, the task of mice gland segmentation is regarded as a multi-label classification problem and design to bind the widely-used method U-Net up with an ensemble learning method for 3D segmentation of mice gland. The proposed method is tested on our 3D confocal images of mice gastric gland. The experiment results show that our method can implement effective and fast segmentation of mice gastric gland. We would expect that this method is applicable for similar structural segmentation in other glands and organs.

Summary: Neurons require proper polarization for precise positioning and axon-dendrite formation. Their intrinsic regulators and underlying mechanisms are poorly understood. Here, we show that Wdr47 is a key polarization regulator. Wdr47-deficient newborn mice die of suffocation due to central nervous system defects including axonal tracts agenesis and slowed radial migration. Wdr47 deficiency represses the multipolar-bipolar transition of cortical neurons, reduces neurite tip-directed microtubule dynamics, and causes multi-axon formation. Overexpression of Wdr47 in wild-type neurons inhibits axon specification and neutralizes Taxol-induced neurite overgrowth and axon overproduction. Wdr47 interacts with the Camsap family of microtubule minus-end-binding proteins; associates with microtubules through Camsap3, whose gene disruption also causes multi-axons; and promotes Camsap3 accumulation in neurites of unpolarized neurons. Furthermore, Camsap overexpression rescues the polarization defects of Wdr47-deficient neurons. Our results indicate that Wdr47 facilitates proper neurite remodeling through Camsaps to fine-tune regional microtubule dynamics and organization during early stages of neuronal polarization. : Proper axon-dendrite polarity ensures unidirectional information processing of neurons. Here, Chen et al. uncover that Wdr47 is critical for neuronal intrinsic polarity control machinery. It functions through Camsap proteins to fine-tune regional microtubule dynamics and organization in unpolarized neurons. Wdr47-deficient neonatal mice die of developmental central nervous system defects. Keywords: Wdr47, Camsap, microtubule dynamics, multi-axon formation, multipolar-bipolar transition, neonatal death, radial migration