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Abstract This study introduced a novel dual fixation method for the pulmonary vasculature and lung tissue in pulmonary hypertension (PH) rats, addressing the limitations of traditional fixation methods that failed to accurately preserve the in vivo status of pulmonary vascular morphology. The modified method involved a dual fixation process, combining individualized ventilation support and vascular perfusion to simulate the respiratory motion, pulmonary artery pressure and right ventricular output of the rat under in vivo conditions. Utilizing a monocrotaline-induced PH rat model, this study compared the dual fixation with the traditional immersion fixation, focusing on the quantitative assessment of alveolar expansion degree, capillary patency, endothelial cell quantity and wall thickness of pulmonary vein and artery. The results demonstrated that the dual fixation is superior in maintaining the authenticity and integrity of lung tissue and more sensitive in the evaluation of pulmonary artery hypertrophy, providing a more reliable representation of pulmonary vascular remodeling associated with PH.

Abstract Multicolor microscopy and super-resolution optical microscopy are two widely used techniques that greatly enhance the ability to distinguish and resolve structures in cellular imaging. These methods have individually transformed cellular imaging by allowing detailed visualization of cellular and subcellular structures, as well as organelle interactions. However, integrating multicolor and super-resolution microscopy into a single method remains challenging due to issues like spectral overlap, crosstalk, photobleaching, phototoxicity, and technical complexity. These challenges arise from the conflicting requirements of using different fluorophores for multicolor labeling and fluorophores with specific properties for super-resolution imaging. We propose a novel multicolor super-resolution imaging method called phasor-based fluorescence spatiotemporal modulation (Phasor-FSTM). This method uses time-resolved detection to acquire spatiotemporal data from encoded photons, employs phasor analysis to simultaneously separate multiple components, and applies fluorescence modulation to create super-resolution images. Phasor-FSTM enables the identification of multiple structural components with greater spatial accuracy on an enhanced laser scanning confocal microscope using a single-wavelength laser. To demonstrate the capabilities of Phasor-FSTM, we performed two-color to four-color super-resolution imaging at a resolution of ~λ/5 and observed the interactions of organelles in live cells during continuous imaging for a duration of over 20 min. Our method stands out for its simplicity and adaptability, seamlessly fitting into existing laser scanning microscopes without requiring multiple laser lines for excitation, which also provides a new avenue for other super-resolution imaging technologies based on different principles to build multi-color imaging systems with the requirement of a lower budget.

This paper investigates the multi-Unmanned Aerial Vehicle (UAV)-assisted wireless-powered Mobile Edge Computing (MEC) system, where UAVs provide computation and powering services to mobile terminals. We aim to maximize the number of completed computation tasks by jointly optimizing the offloading decisions of all terminals and the trajectory planning of all UAVs. The action space of the system is extremely large and grows exponentially with the number of UAVs. In this case, single-agent learning will require an overlarge neural network, resulting in insufficient exploration. However, the offloading decisions and trajectory planning are two subproblems performed by different executants, providing an opportunity for problem-solving. We thus adopt the idea of decomposition and propose a 2-Tiered Multi-agent Soft Actor-Critic (2T-MSAC) algorithm, decomposing a single neural network into multiple small-scale networks. In the first tier, a single agent is used for offloading decisions, and an online pretrained model based on imitation learning is specially designed to accelerate the training process of this agent. In the second tier, UAVs utilize multiple agents to plan their trajectories. Each agent exerts its influence on the parameter update of other agents through actions and rewards, thereby achieving joint optimization. Simulation results demonstrate that the proposed algorithm can be applied to scenarios with various location distributions of terminals, outperforming existing benchmarks that perform well only in specific scenarios. In particular, 2T-MSAC increases the number of completed tasks by 45.5% in the scenario with uneven terminal distributions. Moreover, the pretrained model based on imitation learning reduces the convergence time of 2T-MSAC by 58.2%.

Abstract Background The Bucephalidae is a large family of digenean trematodes but most previous analyses of its phylogenetic position have relied on a single mitochondrial gene or morphological features. Mitochondrial genomes (mitogenomes) remain unavailable for the entire family. To address this, we sequenced the complete mitogenome of Dollfustrema vaneyi and analyzed the phylogenetic relationships with other trematodes. Results The circular genome of Dollfustrema vaneyi spanned 14,959 bp and contained 12 protein-coding genes, 22 transfer RNA genes, 2 ribosomal RNA genes, and a major non-coding region. We used concatenated amino acid and nucleotide sequences of all 36 genes for phylogenetic analyses, conducted using MrBayes, IQ-TREE and PhyloBayes. We identified pronounced topological instability across different analyses. The addition of recently sequenced two mitogenomes for the Aspidogastrea subclass along with the use of a site-heterogeneous model stabilized the topology, particularly the positions of Azygiidae and Bucephalidae. The stabilized results indicated that Azygiidae was the closest lineage to Bucephalidae in the available dataset, and together, they clustered at the base of the Plagiorchiida. Conclusions Our study provides the first comprehensive description and annotation of the mitochondrial genome for the Bucephalidae family. The results indicate a close phylogenetic relationship between Azygiidae and Bucephalidae, and reveal their basal placement within the order Plagiorchiida. Furthermore, the inclusion of Aspidogastrea mitogenomes and the site-heterogeneous model significantly improved the topological stability. These data will provide key molecular resources for future taxonomic and phylogenetic studies of the family Bucephalidae.

Abstract Background Cigarette smoking is a recognized risk factor for orthopedic disorders, particularly intervertebral disc (IVD) degenerative disease. However, the IVD pathophysiology, especially the spatial–temporal remodeling progression in the context of cigarette smoking, remains unclear. This study aimed to address this knowledge gap through a quantitative assessment of IVD structural composition and diffusion properties using a Sprague–Dawley rat model. Methods Twenty‐four rats were divided into control and smoke exposure cohorts, each with two sub‐groups of six rats. One smoke exposure sub‐group was sacrificed after 2 months of daily cigarette smoke exposure in a custom smoking apparatus, while the other was sacrificed after an additional 5 months of smoke cessation. The control groups were age‐matched to the smoke exposure groups. A fluorescent recovery after photobleaching (FRAP) technique was used to determine solute diffusivities and multi‐photon excitation (MPE) imaging was performed to characterize structural changes in the annulus fibrosus (AF), nucleus pulposus (NP), and cartilage endplate (CEP). Results A decrease in diffusivity was observed in the CEP and the AF (radial direction only) after 2 months of smoke exposure. MPE imaging showed aberrant CEP calcification and reduced AF radial collagen fiber bundle diameter, suggesting that the IVD exhibits regionally dependent structural remodeling due to smoke exposure. Furthermore, the smoke cessation group showed deteriorating alterations of structure and diffusivities in all three‐disc regions, including the NP, indicating that five‐month smoke cessation alone didn't reverse the progression of IVD degenerative remodeling during aging. Conclusion This study advances the understanding of IVD pathophysiology in the context of cigarette smoke exposure and cessation, laying the groundwork for potential earlier diagnosis and optimized interventions.

IntroductionProtein acetylation is an extensively investigated post-translational modification (PTM). In addition to lysine acetylation, three new types of lysine acylations characterized by the presence of an acidic carboxylic group have been recently identified and validated. These included lysine malonylation (Kmal), lysine succinylation (Ksucc) and lysine glutarylation (Kglu). Pathogens belonging to the genus Mycobacterium elicit severe diseases in mammalian hosts through the modulation of energy metabolism pathways. Throughout this process, malonyl-CoA, succinyl-CoA and glutaryl-CoA are important intermediates in metabolic pathways, including the tricarboxylic acid (TCA) cycle, amino acid and lipid metabolism. These short-chain acyl-CoAs serve as substrates for corresponding acidic lysine acylation reactions. However, the landscape of these acyl-CoAs dependent acidic lysine acylomes remains unclear.MethodsWe used the high-affinity antibody enrichment combined with high-resolution LC-MS/MS analysis to systematically investigate the global proteomic characteristics of the three acidic lysine acylations in Mycobacterium smegmatis. Subsequently, we employed in vitro enzymatic assays to validate the functional impact of acylated substrates, adenylate kinase and proteasome-associated ATPase. Furthermore, we investigated the effects of overexpressing these two substrates on the in vitro growth of Mycobacterium smegmatis, its invasion of THP-1 cells, and the influence on inflammatory cytokines.ResultsWe systematically investigated the global substrate characterization of 1,703 lysine malonylated sites, 5,320 lysine succinylated sites and 269 lysine glutarylated sites in the non-pathogenic model strain Mycobacterium smegmatis. Bioinformatics analysis demonstrated a correlation between these acidic lysine acylations and the functional roles of ribosomes, in addition to their roles in various metabolic pathways. Furthermore, we investigated the impact of lysine acylations on the functional activity of adenylate kinase and proteasome-associated ATPase, as well as their roles in mycobacterial infection process.DiscussionCollectively, our study provided an important resource on substrate characterization and functional regulation of acidic lysine acylations in Mycobacterium smegmatis, giving valuable insights into their interrelation with the biology of infectious process.

Abstract Decreased myocardial capillary density has been reported as an important histopathological feature associated with various heart disorders. Quantitative assessment of cardiac capillarization typically involves double immunostaining of cardiomyocytes (CMs) and capillaries in myocardial slices. In contrast, single immunostaining of basement membrane protein is a straightforward approach to simultaneously label CMs and capillaries, presenting fewer challenges in background staining. However, subsequent image analysis always requires expertise and laborious manual work to identify and segment CMs/capillaries. Here, we developed an image analysis tool, AutoQC, for automatic identification and segmentation of CMs and capillaries in immunofluorescence images of basement membrane. Commonly used capillarization-related measurements can be derived from segmentation results. By leveraging the power of a pre-trained segmentation model (Segment Anything Model, SAM) via prompt engineering, the training of AutoQC required only a small dataset with bounding box annotations instead of pixel-wise annotations. AutoQC outperformed SAM (without prompt engineering) and YOLOv8-Seg, a state-of-the-art instance segmentation model, in both instance segmentation and capillarization assessment. Thus, AutoQC, featuring a weakly supervised algorithm, enables automatic segmentation and high-throughput, high-accuracy capillarization assessment in basement-membrane-immunostained myocardial slices. This approach reduces the training workload and eliminates the need for manual image analysis once AutoQC is trained.

Abstract Background Mesenchymal stem cells (MSCs) hold a great promise for cell-based therapy in the field of regenerative medicine. In this study, we aimed to evaluate the safety and efficacy of intravenous infusion of human umbilical cord-derived MSCs (HUC-MSCs) in patients with aging frailty. Methods In this randomized, double-blind, placebo-controlled trial, participants diagnosed with aging frailty were randomly assigned to receive intravenous administrations of HUC-MSCs or placebo. All of serious adverse events and AEs were monitored to evaluate the safety of treatment during the 6-month follow-up. The primary efficacy endpoint was alteration of physical component scores (PCS) of SF-36 qualities of life at 6 months. The secondary outcomes including physical performance tests and pro-inflammatory cytokines, were also observed and compared at each follow-up visits. All evaluations were performed at 1 week, 1, 2, 3 and 6 months following the first intravenous infusion of HUC-MSCs. Results In the MSCs group, significant improvements in PCS of SF-36 were observed from first post-treatment visit and sustained throughout the follow-up period, with greater changes compared to the placebo group (p = 0.042). EQ-VAS scores of MSCs group improved significantly at 2 month (p = 0.023) and continued until the end of the 6-month visit (p = 0.002) in comparison to the placebo group. The timed up and go (TUG) physical performance test revealed significant group difference and showed continual enhancements over 6 months (p

Background: In N-way model merging, model matching plays an important role. However, the N-way model matching problem has been recognized as NP-hard. Aim: To search the optimal or near-optimal matching solution efficiently, this paper proposes an N-way model matching algorithm based on the Artificial Bee Colony (ABC) algorithm. Method: This algorithm combines global heuristic search and local search to deal with the complexity of N-way model matching. We evaluated the proposed N-way model merging approach through case studies and we evaluated the proposed ABCMatch algorithm by comparing it with Genetic Algorithm (GA) and Elephant Herding Optimization (EHO). Results: The experimental results show that ABCMatch can obtain more accurate model matching solutions in a shorter time, and the average model matching accuracy of ABCMatch is 2.7725% higher than GA and 1.8804% higher than EHO. Conclusion: Results demonstrate that our method provides an effective way for software engineers to merge UML models in collaborative modeling scenarios.

Abstract The evidence that parasitic animals exhibit elevated mitogenomic evolutionary rates is inconsistent and limited to Arthropoda. Similarly, the evidence that mitogenomic evolution is faster in species with low locomotory capacity is limited to a handful of animal lineages. We hypothesised that these two variables are associated and that locomotory capacity is a major underlying factor driving the elevated rates in parasites. Here, we study the evolutionary rates of mitogenomes of 10,906 bilaterian species classified according to their locomotory capacity and parasitic/free-living life history. In Bilateria, evolutionary rates were by far the highest in endoparasites, much lower in ectoparasites with reduced locomotory capacity and free-living lineages with low locomotory capacity, followed by parasitoids, ectoparasites with high locomotory capacity, and finally micropredatory and free-living lineages. The life history categorisation (parasitism) explained ≈45%, locomotory capacity categorisation explained ≈39%, and together they explained ≈56% of the total variability in evolutionary rates of mitochondrial protein-coding genes in Bilateria. Our findings suggest that these two variables play major roles in calibrating the mitogenomic molecular clock in bilaterian animals.

Within the myocardium, cardiomyocytes reside in a complex and dynamic extracellular matrix (ECM) consisting of a basement membrane (BM) and interstitial matrix. The interactions between cardiomyocytes and the myocardial ECM play a critical role in maintaining cardiac geometry and function throughout cardiac development and in adult hearts. Understanding how the structural changes of the myocardial ECM affect cardiomyocyte function requires knowledge of pericellular structures. These structures are of a size beyond the resolution of conventional optical microscopy. Here, we demonstrated multi-scale and multi-aspect characterization of the cardiomyocyte microenvironment in myocardial tissue sections using multimodal stimulated emission depletion (STED) microscopy. Second harmonic generation and autofluorescence facilitated multiplexed imaging, enabling the interpretation of protein distribution in 3D. STED imaging modality revealed BM structures of cardiomyocytes and myocardial capillaries at the subdiffractional level. Moreover, meaningful measurements retrieved from acquired images, such as sarcomere length and capillary density, enabled quantitative assessment of myocardial structures.

Objective: Chondrocyte viability (CV) can be measured with the label-free method using second harmonic generation (SHG) and two-photon excitation autofluorescence (TPAF) imaging. To automate the image processing for the label-free CV measurement, we previously demonstrated a two-step deep-learning method: Step 1 used a U-Net to segment the lacuna area on SHG images; Step 2 used dual CNN networks to count live cells and the total number of cells in extracted cell clusters from TPAF images. This study aims to develop one-step deep learning methods to improve the efficiency of CV measurement. Method: TPAF/SHG images were acquired simultaneously on cartilage samples from rats and pigs using two-photon microscopes and were merged to form RGB color images with red, green, and blue channels assigned to emission bands of oxidized flavoproteins, reduced forms of nicotinamide adenine dinucleotide, and SHG signals, respectively. Based on the Mask R-CNN, we designed a deep learning network and its denoising version using Wiener deconvolution for CV measurement. Results: Using training and test datasets from rat and porcine cartilage, we have demonstrated that Mask R-CNN-based networks can segment and classify individual cells with a single-step processing flow. The absolute error (difference between the measured and the ground-truth CV) of the CV measurement using the Mask R-CNN with or without Wiener deconvolution denoising reaches 0.01 or 0.08, respectively; the error of the previous CV networks is 0.18, significantly larger than that of the Mask R-CNN methods. Conclusions: Mask R-CNN-based deep-learning networks improve efficiency and accuracy of the label-free CV measurement.

ABSTRACT In response to the problems of high impurity content in ears and poor stalk crushing efficiency during the harvesting of fresh corn, a double fan high-pressure impurity removal and cleaning device was designed based on the operating conditions and mechanical analysis of harvester. Field trials were conducted using fan speed (1200 rpm, 1300 rpm and 1400 rpm) and harvester operating speed (6 km/h, 8 km/h and 10 km/h) as test conditions, with ear impurity content rate and stalk crushing length qualification rate as indicators. Ear impurity content rate ranged from 0.25% to 0.50% and stalk crushing length qualification rate ranged from 96.1% to 98.6%, with test results meeting operation standard under all test conditions. The designed device was able to meet the operational requirements.

Autofluorescence (AF) poses challenges for detecting proteins of interest in situ when employing immunofluorescence (IF) microscopy. This interference is particularly pronounced in strongly autofluorescent tissues such as myocardium, where tissue AF can be comparable to IF. Although various histochemical methods have been developed to achieve effective AF suppression in different types of tissue, their applications on myocardial samples have not been well validated. Due to inconsistency across different autofluorescent structures in sometypes of tissue, it is unclear if these methods can effectively suppress AF across all autofluorescent structures within the myocardium. Here, we quantitatively evaluated the performance of several commonly used quenching treatments on formaldehyde-fixed myocardial samples, including 0.3 M glycine, 0.3% Sudan Black B (SBB), 0.1% and 1% sodium borohydride (NaBH4), TrueVIEW® and TrueBlack®. We further assessed their quenching performance by employing the pre-treatment and post-treatment protocols, designed to cover two common IF staining scenarios where buffers contained detergents or not. The results suggest that SBB and TrueBlack® outperform other reagents in AF suppression on formaldehyde-fixed myocardial samples in both protocols. Furthermore, we inspected the quenching performance of SBB and TrueBlack® on major autofluorescent myocardial structures and evaluated their influence on IF imaging. The results suggest that SBB outperforms TrueBlack® in quenching major autofluorescent structures, while TrueBlack® excels in preserving IF labeling signal. Surprisingly, we found the treatment of NaBH4 increased AF signal and enhanced the AF contrast of major autofluorescent structures. This finding suggests that NaBH4 has the potential to act as an AF enhancer and may facilitate the interpretation of myocardial structures without the need for counterstaining.

Laser speckle contrast imaging or laser speckle imaging (LSI) is a noninvasive imaging technology that can detect areas of dynamic perfusion or vascular flow. Thus, LSI has shown increasing diagnostic utility in various pathologies and has been employed for intraoperative, postoperative, and long-term monitoring in many medical specialties. Recently, LSI has gained traction in clinical dermatology because it can be effective in the assessment of pathologies that are associated with increased perfusion and hypervascularity compared with that of normal tissue. To date, LSI has been found to be highly accurate in monitoring skin graft reperfusion, determining the severity of burns, evaluating neurosurgical revascularization, assessing persistent perfusion in capillary malformations after laser therapy, and differentiating malignant and benign skin lesions. LSI affords the advantage of noninvasively assessing lesions before more invasive methods of diagnosis, such as tissue biopsy, while remaining inexpensive and exhibiting no adverse events to date. However, potential obstacles to its clinical use include tissue movement artifact, primarily qualitative data, and unclear impact on clinical practice given the lack of superiority data compared with the current standard-of-care diagnostic methods. In this review, we discuss the clinical applications of LSI in dermatology for use in the diagnosis and monitoring of vascular, neoplastic, and inflammatory skin conditions.

Arabidopsis thaliana heme oxygenase‐1 (AtHO‐1), a metabolic enzyme in the heme degradation pathway, serves as a prototype for study of the bilin‐related functions in plants. Past biological analyses revealed that AtHO‐1 requires ferredoxin‐NADP+ reductase (FNR) and ferredoxin for its enzymatic activity. Here, we characterized the binding and degradation of heme by AtHO‐1, and found that ferredoxin is a dispensable component of the reducing system that provides electrons for heme oxidation. Furthermore, we reported the crystal structure of heme‐bound AtHO‐1, which demonstrates both conserved and previously undescribed features of plant heme oxygenases. Finally, the electron transfer pathway from FNR to AtHO‐1 is suggested based on the known structural information.

Cul3 encodes Cullin-3, a core component of the ubiquitin E3 ligase that is involved in protein ubiquitination. Recent studies have identified Cul3 as a high-confidence risk gene in neurodevelopmental disorders (NDDs), especially autism spectrum disorder (ASD). Different strategies have been used to generate animal models with Cul3 deficiency in the central nervous system, including whole-brain knockout (KO), cell-type specific conditional KO (cKO), and brain region-specific knockdown. In this review, we revisited the basic properties of CUL3 and its function under physiological and pathological conditions. Recent clinical studies including case reports and large cohort sequencing studies related to CUl3 in NDDs have been summarized. Moreover, we characterized the behavioral, electrophysiological, and molecular changes in newly developed Cul3 deficiency models. This would guide further studies related to Cul3 in CNS and provide potential therapeutic targets for Cul3-deficiency-induced NDDs, including ASD.

CTNNB1 is the gene that encodes β-catenin which acts as a key player in the Wnt signaling pathway and regulates cellular homeostasis. Most CTNNB1-related studies have been mainly focused on its role in cancer. Recently, CTNNB1 has also been found involved in neurodevelopmental disorders (NDDs), such as intellectual disability, autism, and schizophrenia. Mutations of CTNNB1 lead to the dysfunction of the Wnt signaling pathway that regulates gene transcription and further disturbs synaptic plasticity, neuronal apoptosis, and neurogenesis. In this review, we discuss a wide range of aspects of CTNNB1 and its physiological and pathological functions in the brain. We also provide an overview of the most recent research regarding CTNNB1 expression and its function in NDDs. We propose that CTNNB1 would be one of the top high-risk genes for NDDs. It could also be a potential therapeutic target for the treatment of NDDs.

Abstract Long-lived interlayer excitons (IXs) in van der Waals heterostructures (HSs) stacked by monolayer transition metal dichalcogenides (TMDs) carry valley-polarized information and thus could find promising applications in valleytronic devices. Current manipulation approaches for valley polarization of IXs are mainly limited in electrical field/doping, magnetic field or twist-angle engineering. Here, we demonstrate an electrochemical-doping method, which is efficient, in-situ and nonvolatile. We find the emission characteristics of IXs in WS2/WSe2 HSs exhibit a large excitonic/valley-polarized hysteresis upon cyclic-voltage sweeping, which is ascribed to the chemical-doping of O2/H2O redox couple trapped between WSe2 and substrate. Taking advantage of the large hysteresis, a nonvolatile valley-addressable memory is successfully demonstrated. The valley-polarized information can be non-volatilely switched by electrical gating with retention time exceeding 60 min. These findings open up an avenue for nonvolatile valley-addressable memory and could stimulate more investigations on valleytronic devices.

Collagen has been ubiquitously applied in the food, cosmetic, biomedical, and pharmaceutical industries. However, the existing collagen in the market cannot meet the demand of consumers. In this study, double-spotted pufferfish collagen (DPC) was extracted from the skin of the pufferfish and characterized as type I collagen, which maintains a native triple helical structure. Liquid chromatography-tandem mass spectrometry analysis revealed that α1 and α2 subunits of DPC comprised 378 and 372 uninterrupted Gly-X-Y triplets, respectively, which were 78.3 % and 78.4 % of the total amino acid content. Secondary structure analysis revealed that DPC consisted of 50.23 % α-helices, 26.51 % β-sheets, 23.26 % β-turns, and 0 % random coils. Using XRD, the distance between collagen molecular chains was found to be 11.45 Å. The yield of DPC was 49.83 % ± 1.85 % (dry weight basis), which was higher than that of pigskin collagen. Dynamic sweep and steady-state shear of DPC were significantly affected by temperature and concentration. DPC solutions exhibited shear-thinning rheological behavior. DPC exhibited water holding and oil absorption capacities superior to terrestrial collagen, such as that of chicken feet collagen. The foaming properties and emulsifying properties of DPC were superior to commercial collagen, such as that of black ruff collagen. DPC was nontoxic to MC3T3-E1 cells and had good cytocompatibility. These results suggest that DPC is a new alternative for collagen with potential for further applications in food, cosmetic, and biomedical fields.

ABSTRACT Chroococcidiopsis spp. can withstand extremely harsh environments, including a Mars-like environment. However, studies are lacking on the molecular mechanisms of Chroococcidiopsis sp. surviving in Mars-like environments. In the HH-21-5 mission, the desert cyanobacterium Chroococcidiopsis sp. was exposed to a Mars-like environment (near space; 35 km altitude) for 4 h, and a single-factor environment of near space was simulated on the ground. We investigated the survival and endurance mechanisms of Chroococcidiopsis sp. ASB-02 after exposing it to near space by studying its physiological and transcriptional properties. After the exposure, Chroococcidiopsis sp. ASB-02 exhibited high cell viability, although photosystem II activity decreased and the levels of reactive oxygen species increased. The single-factor simulation experiments revealed that for the survival of Chroococcidiopsis sp. ASB-02 in near space, UV radiation was the most important limiting factor, and it was followed by temperature. The near space environment triggered multiple metabolic pathway responses in Chroococcidiopsis sp. ASB-02. The upregulation of extracellular polysaccharides as well as carotenoid and scytonemin biosynthesis genes in response to UV radiation attenuated the extent of radiation reaching the cells. At the same time, genes related to protein synthesis were upregulated in response to the low temperature, overcoming the decrease in metabolic activity that was caused by the low temperature. In near space and after rehydration, the genes involved in various DNA and photosystem II repair pathways were upregulated. This reflected the damage to the DNA and photosystem II protein subunits in cells during the flight and suggested that repair mechanisms play an important role in the recovery of Chroococcidiopsis sp. ASB-02. IMPORTANCE This study reported that the protective and repair mechanisms of Chroococcidiopsis sp. ASB-02 contributed to its endurance ability in a Mars-like near space environment. In Chroococcidiopsis sp. ASB-02, a Mars-like near space environment activated the expression of genes involved in extracellular polysaccharides (EPS), carotenoid, scytonemin, and protein syntheses, which provided additional protection. Additionally, the cell damage repair process enhanced the recovery rate of Chroococcidiopsis sp. ASB-02 after the flight. This study will help to enhance the understanding of the tolerance mechanism of Chroococcidiopsis sp. and to provide important guidance as to the survival requirements for microbial life in a Mars-like environment.

In the space environment, cosmic rays and high-energy particles may cause a single-event upset (SEU) during program execution, and further cause silent data corruption (SDC) errors in program outputs. After extensive research on SEU and SDC errors, it has been found that SDC errors in the routing program in satellite networks may lead to the emergence of Sinkhole (SH) and Grayhole (GH) nodes in the network, which may cause damage to satellite networks. To find and solve the problems in time, a digital-twin-based detection and protection framework for SDC-induced SH and GH nodes in satellite networks is proposed. First, the satellite network fault model under SEU and the generation mechanism of SH and GH nodes induced by SDC errors are described. Then, the data structure based on digital twins required by the proposed detection and protection framework is designed, and the detection methods of SH and GH nodes induced by SDC errors are proposed. SKT and LLFI simulation tools are used to build a simulated Iridium satellite network and carry out fault injection experiments. Experiment results show that the accuracy of the proposed detection method is 98–100%, and the additional time cost of routing convergence caused by the proposed framework is 3.1–28.2%. Compared with existing SH and GH detection methods, the proposed methods can timely and accurately detect faults during the routing update stage.

The protective ozone layer is continually depleting owing to an increase in the levels of solar UV-B radiation, which has harmful effects on organisms. Algae in desert soil can resist UV-B radiation, but most research on the radiation resistance of desert algae has focused on cyanobacteria. In this study, we found that desert green algae, Chlorella sp., could maintain high photosynthetic activity under UV-B stress. To examine the tolerance mechanism of the desert green algae photosystem, we observed the physiological and transcriptome-level responses of Chlorella sp. to high doses of UV-B radiation. The results showed that the reactive oxygen species (ROS) content first increased and then decreased, while the malondialdehyde (MDA) content revealed no notable lipid peroxidation during the UV-B exposure period. These results suggested that Chlorella sp. may have strong system characteristics for scavenging ROS. The antioxidant enzyme system showed efficient alternate coordination, which exhibited a protective effect against enhanced UV-B radiation. DNA damage and the chlorophyll and soluble protein contents had no significant changes in the early irradiation stage; UV-B radiation did not induce extracellular polysaccharides (EPS) synthesis. Transcriptomic data revealed that a strong photosynthetic system, efficient DNA repair, and changes in the expression of genes encoding ribosomal protein (which aid in protein synthesis and improve resistance) are responsible for the high UV-B tolerance characteristics of Chlorella sp. In contrast, EPS synthesis was not the main pathway for UV-B resistance. Our results revealed the potential cell damage repair mechanisms within Chlorella sp. that were associated with high intensity UV-B stress, thereby providing insights into the underlying regulatory adaptations of desert green algae.

Compared with single-point laser-induced plasma, multi-point laser-induced plasma shows some advantages in ignition and flameholding, but with little research. Therefore, numerical studies of multi-point laser-induced plasma in supersonic flow are conducted using an instantaneous energy deposition model. In the studies, the single-pulse laser energy of every laser focus is 50 mJ, the laser focus configuration is linear, the inflow velocities are from Ma 1.5 to Ma 2.5, and the distances between adjacent focal spots are from 2 to 4 mm. The evolution process of shock waves and plasma kernels in supersonic flow has been described in detail when the inflow velocity is Ma 2 and when the distance between adjacent focal spots is 2 mm. Besides, the evolution law of the velocity field, the streamline, and the plasma kernel position in the flow field have also been analyzed. The results reveal that the volume of the plasma kernel increases rapidly within 15 µs and that the initial shock wave has a strong deflection effect on the supersonic flow. Moreover, the effects of inflow velocities and distances between adjacent focal spots on multi-point laser-induced plasma are compared using the plasma kernel’s average temperature, volume, and specific surface area. Under the conditions of the studies, the results show that the distance between adjacent focal spots is the main reason affecting the evolution. Thus, a distance longer than 4 mm between adjacent focal spots should be avoided for multi-point laser-induced plasma in supersonic flow because the plasma kernel could not be fused in a timely manner.

Objective: To develop an UPLC-MS/MS method for the accurate quantification of Estriol (E3), a new radioprotective agent, and observe the variation in pharmacokinetic characteristics of E3 after irradiation. As a hormone drug, the gender differences of E3 metabolism were also concerned here. Methods: Various chromatographic and mass spectrometric conditions of E3 were optimized. The specificity, linearity, precision and accuracy of UPLC-MS/MS method were validated. Twenty SD rats, half male and half female, were administered with intramuscular (im) injection of 2.7 ​mg/kg E3, and then divided randomly into two groups, sham-irradiated group (Con) and irradiated group (IR). IR group was irradiated to 7 ​Gy of γ-rays. Blood samples were collected at different times post-irradiation and E3 concentrations were detected. The changes of concentration-time variation and pharmacokinetic parameters for E3 after irradiation were investigated, together with metabolic differences between male and female rats. Results: The range of the calibration curve of UPLC-MS/MS for E3 was 1.00–200.0 ​ng/mL. Con group reached maximum concentration (Cmax) (77.57 ​± ​18.71) ng/mL at (0.68 ​± ​0.29) h (Tmax) after im injection. The drug concentration-time profiles and pharmacokinetic parameters of E3 were consistent before and after irradiation. The areas under time curve (AUC0-t) of E3 were (353 ​± ​74) h·ng/mL for Con group, and (299 ​± ​74) h·ng/mL for IR group (P ​> ​0.05). There were also no statistical differences in pharmacokinetic parameters between female and male rats. The elimination half-lifes (T1/2) of males and females were (3.02 ​± ​0.68) h and (3.01 ​± ​0.42) h in Con group, and (3.64 ​± ​0.51) h and (3.38 ​± ​0.60) h in IR group, respectively (P ​> ​0.05). Conclusion: A rapid and sensitive UPLC-MS/MS method for determination of E3 was established. The pharmacokinetic characteristics of E3 in rat were not affected by 7 ​Gy irradiation and gender differences. This study provided a theoretical basis for the development and application of new radiation injury treatment drug.

During long-distance flight, unmanned aerial vehicles (UAVs) need to perform cross-domain authentication to prove their identity and receive information from the ground control station (GCS). However, the GCS needs to verify all drones arriving at the area it is responsible for, which leads to the GCS being unable to complete authentication in time when facing cross-domain requests from a large number of drones. Additionally, due to potential threats from attackers, drones and GCSs are likely to be deceived. To improve the efficiency and security of cross-domain authentication, we propose an efficient blockchain-based cross-domain authentication scheme for the Internet of Drones (BCDAIoD). By using a consortium chain with a multi-chain architecture, the proposed method can query and update different types of data efficiently. By mutual authentication before cross-domain authentication, drones can compose drone groups to lighten the authentication workload of domain management nodes. BCDAIoD uses the notification mechanism between domains to enable path planning for drones in advance, which can further improve the efficiency of cross-domain authentication. The performance of BCDAIoD was evaluated through experiments. The results show that the cross-domain authentication time cost and computational overhead of BCDAIoD are significantly lower those of than existing methods when the number of drones is large.

It is challenging to quantify anisotropic diffusion in biological systems. Here the authors report light-sheet imaging-based Fourier transform fluorescence recovery after photobleaching (LiFT-FRAP) to noninvasively determine 3D diffusion tensors of various biomolecules at physiological diffusivity.

4-Hydroxyphenylpyruvate dioxygenase (HPPD) is not only an important target enzyme for the treatment of type I tyrosinemia, but also a new target for design bleaching herbicides, and it plays key role in the biosynthesis of tocopherol and plastoquinone. Thirty-six known active pyridine derivatives were collected, and comparative molecular field analysis (CoMFA) and comparative molecular similarity index analysis (CoMSIA) models based on common skeleton were constructed to obtain novel HPPD herbicides with higher activity. Two new HPPD inhibitors were rationally designed and synthesized according to the CoMFA and CoMSIA models and verified by enzyme activity, biological assays, and molecular docking. The promising compound W1 ((E)-5-(3-(4-bromophenyl)acryloyl)-6-hydroxy-2,3-dihydropyridin-4(1H)-one) showed better AtHPPD inhibitory activity, and the bioassay results revealed that some weeds showed bleaching symptoms. The good binding stability of W1 and protein was confirmed by molecular dynamics simulation in 100 ns. These results would be highly useful in the progress of new HPPD inhibitors discovery.

Due to the high global warming potential and rapidly increasing atmospheric concentrations of fluorinated greenhouse gases (hereinafter referred to as F-gases), the emission estimation has been widely studied. Additionally, the atmospheric observation supports the emission estimation. However, there are few F-gas studies with respect to chemical plants. In this study, we firstly carried out atmospheric observation studies of F-gases around chemical plants in China. Four typical chemical plants were selected, and the concentration levels, their concentration differences (hereinafter referred to as the up–down difference) and enhanced ratios (hereinafter referred to as the up–down enhanced ratio) of the F-gases at upwind and downwind sites in the four plants were observed. Both the upwind and downwind observation results of each plant showed that the average concentrations of HFCs were the highest (57%–85%), followed by PFCs, while the total average concentrations of SF6 and NF3 only accounted for less than 5%. The F-gases, with both the larger up–down enhanced ratios and the larger up–down differences in each plant, may suggest larger emission strengths during fluorinated production, by-products and usage processes. Additionally, the numbers of F-gas species emitted from Plants A, B and C were all significantly higher than those from Plant D. This may be because the F-gases in Plants A, B and C were emitted not only during fluorinated production and by-product processes, but also during usage processes, with respect to refrigeration and air-conditioning or foam insulation materials. The F-gases emitted by each plant did not cause toxicity to humans at sensitive sites; however, they can cause nonnegligible climate impacts. In the future, it is necessary to select more typical chemical plants to carry out the atmospheric observation of F-gases, with a higher observation frequency and larger sample size, to further estimate F-gas emissions from chemical plants in China.

In recent years, with the intricacy of international politics and economic situation and the anti-globalization trend, China’s trade with world is facing many serious challenges. There are more factors that effects China export and import. Because of that, high-precision forecasting for international trade is beneficial for nation’s government, guild and export and import enterprise that need a judgement or decision for future. To better promote future research, the paper reviews the paper written by experts from world in trade forecasting field, classifies and summarizes their opinion according in their adopting machine learning method.

Element geochemistry, Sr and Nd isotope, and LA-ICP-MS zircon U-Pb isotope data have been obtained for the granitoids of Dazesan pluton in the Jiaodong Peninsula, East China, and their intermediate microgranular dark enclaves so as to reveal their petrogenesis and tectonic implications. These granitoids have high SiO2 (68.25–71.56 wt.%), K2O (3.44–5.50 wt.%), total alkalis (K2O + Na2O = 7.29–9.00 wt.%), Sr (451–638 ppm), Ba (1157–2842 ppm) and light rare earth elements (LREEs) (131.57–210.08), with strong depletion both in heavy rare earth element (HREE) and high field strength element (HFSE) concentrations as well as unclearly Eu anomalies, showing typical signatures of high Ba-Sr granitoids. They possess high (La/Yb)N (32–50) and Sr/Y (50–79) values and low MgO (0.76–1.11 wt.%), Cr (9.9–19.6 ppm) and Ni (4.51–7.04 ppm) concentrations. All the above geochemical compositions are similar to those of late Early Cretaceous granitoids, in combination with zircon LA-ICP-MS U-Pb ages of 119.6 ± 1.3 to 120 ± 1 Ma for these granitoids obtained in this study, indicating c. 120 Ma probably represents the lower limit of ages when late Early Cretaceous granitoids emplaced in the Jiaodong Peninsula. The microgranular dark enclaves, forming a linear trend with their host granitoids on the oxide against SiO2 plots, display higher MgO contents of 3.05–4.39 wt.% at lower SiO2 concentrations of 54.25–56.84 wt.% and possess a zircon LA-ICP-MS U-Pb age of 119 ± 2 Ma, identical to those of these granitoids, indicating the acid magma and intermediate magma were coeval. Furthermore, dark enclaves and their host granitoids have indistinguishable (87Sr/86Sr)i values of 0.709523–0.70972 and 0.709361–0.709858, respectively, and plot within a two-liquid immiscible field on the Greig pseudoternary phase diagram. In addition, they have markedly parallel REE patterns, with the dark enclaves having much greater REE and HFSE abundances than those of their host granitoids. Therefore, it is suggested that liquid immiscibility is a viable model to explain the chemical compositional variations between the Dazeshan granitoids and their dark enclaves. Based on the element geochemistry, geochronology and Sr- and Nd-isotope of the Dazeshan granitoids and their dark enclaves, it is envisaged the crust-derived acid melts due to partial melting of ancient continental lower crust in the Jiaodong Peninsula (mainly Neoarchean-Palaeoproterozoic basement in the Jiaobei terrane) containing a subduction-related material, resulting from the addition of the enriched subcontinental lithospheric mantle-derived melts, assimilated the lithospheric mantle-derived basic melts and formed the homogeneous magma chamber at the crust base, then split into two immiscible liquids, with one granitic liquid producing the Dazeshan granitoids and the other intermediate one forming the dark enclave during its ascent. Combined with previous studies, the identification of a lithospheric mantle-derived material in the Dazeshan granitoids suggests a catastrophic lithospheric thinning at c. 120 Ma, reflecting an abrupt change in the direction of Palaeo-Pacific plate subducting and the corresponding regional tectonic transition from E–W extension to NW–SE extension.

N6-methyladenosine (m6A) modification remains the most pivotal epigenetic modification on RNA. As we know, m6A not only affects physiological processes but is also involved in carcinoma. Noncoding RNAs play an indispensable role in the occurrence and development of carcinoma. However, a large amount of research is focused on mRNA currently. Insufficient research has been done on the relationship between noncoding RNA (ncRNA) methylation and cancer. Therefore, this review aims to introduce the theoretical knowledge of m6A modification in noncoding RNA, discuss its function in tumorigenesis and progression, and ultimately summarize its potential clinical applications.

Photoacoustic imaging (PAI) is a fast evolving imaging technology enabling in vivo imaging with high specificity and spatial resolution. However, due to strong background signals from various intrinsic chromospheres such as melanin, photoacoustic imaging of targeting objects labeled by contrast agents remain a challenge. The transient triplet differential (TTD) method has shown a significant potential for background-free photoacoustic imaging. Here, we develop a photoacoustic system using an ultrasonic semicircular ring array for transient triplet differential imaging. Pt(II) Octaethylporphine (PtOEP) and black ink are used as the contrast agent and the phantom of melanoma, respectively. Using the TTD method, we could remove the strong background signal from black ink. The ratio between contrast agent signal and background signal is increased to about 10 times the previous one. Our finding demonstrates the potential of the TTD method on molecular imaging for strong background removal.

This paper investigates the dual-mode power-saving strategy designed for the 100G Energy Efficient Ethernet. The process of this strategy is a sequence of cycles, where each cycle is an interval elapsed between two consecutive instants when the buffer of the Ethernet interface becomes empty. In each cycle, the interface first enters the fast-wake mode to perform the conditional sleep-mode selection of the strategy. If the number of arrivals during the fast-wake mode reaches a threshold, the interface directly wakes up; otherwise, it proceeds to the deep-sleep mode. The sequence of cycles switches between two types: deep-sleep cycles with deep-sleep mode and light-sleep cycles without deep-sleep mode. We analyze the dual-mode strategy based on the condition of the number of arrivals during the fast-wake mode. We first derive the weights of conditions according to the feature of the conditional sleep-mode operation, and then calculate the unconditioned performance measure of the dual-mode strategy based on the weighted average of that of these two kinds of cycles. Finally, we obtain the close-form expressions of the power efficiency and the mean delay, based on which we provide a set of parameter selection rules. We show that the dual-mode strategy with these rules can select suitable sleep modes according to the instantaneous traffic rate, and thus perform well under bursty input traffic.

Cytidine is an antiviral and anticancer drug intermediate, its primary method of manufacture being fermentation. Uridine-cytidine kinase (UCK) catalyzes the reverse process of phosphorylation of cytidine to produce cytidylic acid, which influences cytidine accumulation in the Escherichia coli cytidine biosynthesis pathway. The cytidine-producing strain E. coli NXBG-11 was used as the starting strain in this work; the udk gene coding UCK was knocked out of the chromosomal genome using clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 technology. The mutant strain E. coli NXBG-12 was obtained; its transcriptomics were studied to see how udk gene deletion affected cytidine synthesis and cell-wide transcription. The mutant strain E. coli NXBG-12 generated 1.28 times more cytidine than the original strain E. coli NXBG-11 after 40 h of shake-flask fermentation at 37 °C. The udk gene was knocked out, and transcriptome analysis showed that there were 1168 differentially expressed genes between the mutant and original strains, 559 upregulated genes and 609 downregulated genes. It was primarily shown that udk gene knockout has a positive impact on the cytidine synthesis network because genes involved in cytidine synthesis were significantly upregulated (p < 0.05) and genes related to the cytidine precursor PRPP and cofactor NADPH were upregulated in the PPP and TCA pathways. These results principally demonstrate that udk gene deletion has a favorable impact on the cytidine synthesis network. The continual improvement of cytidine synthesis and metasynthesis is made possible by this information, which is also useful for further converting microorganisms that produce cytidine.

Abstract Nanovaccines have attracted booming interests in vaccinology studies, but the profound impacts of their delivery mode on immune response remain unrealized. Herein, immunostimulatory CpG‐modified tumor‐derived nanovesicles (CNVs) are used as a nanovaccine testbed to initially evaluate the impacts of three distinct delivery modes, including mono‐pulse CNVs, staggered‐pulse CNVs, and gel‐confined CNVs. Fundamentally, delivery mode has enormous impacts on the immunomodulatory effects, altering the spatiotemporal distribution of nanovaccine residence and dendritic cell–T cell interaction in lymph nodes, and finally affecting subsequent T cell‐mediated immune performance. As a result, the gel‐confined delivery mode offers the best therapeutic performance in multiple tumor models. When extending evaluation to examine how the various delivery modes impact the performance of liposome‐based nanovaccines, similar trends in intralymph node distribution and antitumor effect are observed. This work provides a strong empirical foundation that nanovaccine researchers should position delivery mode near the top of their considerations for the experimental design, which should speed up nanovaccine development and facilitate efficient selection of appropriate delivery modes in the clinic.

Laser cell patterning is a distinctly effective technique for creating cell arrangements in culture that replicate in vivo tissue structure for studying contact-mediated cell-cell interactions. Conventional laser-based single-cell-manipulation techniques are limited by their inability to pattern irregularly shaped cells, such as rod-shaped cardiomyocytes. We report use of a spatial light modulator loaded with a computer-generated phase map to shape a single laser source into multiple laser-guidance beams distributed around the outer contour of an irregularly shaped cell to achieve accurate cell patterning. In addition to describing the principle and practice of the system design, we present what is to our knowledge the first achievement of patterning large, irregularly (rod) shaped adult rat cardiomyocytes in an end-to-end connected alignment to replicate the in vivo heart muscle cell connection without use of substrate surface modifications, which can interfere with in vivo-like cell-cell and cell-extracellular matrix interactions. Our research demonstrates that two-stage multiple beam laser guidance is effective: 1) A dual-beam configuration horizontally translates a cell from the outlet of the microfluidic cell delivery channel to a position above the cell deposition site and 2) A quad-beam configuration rapidly propels the cell axially through the suspension medium to the culture substrate with vertical movement of the cell patterning chamber. Our study reveals that 90% of the patterned cells maintained end-to-end connection 30 min after patterning, and mechanical junctions could be reinstalled between laser connected cells after overnight incubation. This demonstrates that multiple beam laser patterning is an outstanding tool for in vitro studying contact-mediated cell-cell interactions among irregularly shaped cells.

This paper studies the Ethernet passive optical network (EPON) with limited service. The transmission window (TW) is limited in this system to guarantee a bounded delay experienced by disciplined users and to constrain malicious users from monopolizing the transmission channel. Thus, selecting an appropriate TW size is critical to the performance of EPON with limited service discipline. In this paper, we develop an M/G/1 queuing model with vacation times and limited-service discipline to investigate the impact of TW size on packet delay. A distinguishing feature of this model is that there are two queues in the buffer of each optical network unit (ONU): one queue is inside the gate and the other one is outside the gate, from which we derive the generalized formula of mean waiting time. Furthermore, based on the service-level agreements of users and the Chernoff bound of queue length, we provide a simple rule to determine an optimum TW size for limited-service EPONs. The analytic results reported in this paper are all verified by simulations.

Uncontrolled hemorrhage from trauma or surgery can lead to death. In this study, chitosan/kaolin (CSK) and chitosan/montmorillonite (CSMMT) composites were prepared from chitosan (CS), kaolin (K), and montmorillonite (MMT) as raw materials to control bleeding. The physiochemical properties and surface morphology of CSK and CSMMT composites were analyzed by Fourier transform infrared spectrometry (FT-IR), X-ray diffraction (XRD), scanning electron microscopy (SEM), zeta potentials, and X-ray fluorescence (XRF). The hemostatic mechanism was measured in vitro by activated partial thromboplastin time (APTT), prothrombin time (PT), in vitro clotting time, erythrocyte aggregation, and thromboelastogram (TEG). The hemostasis ability was further verified by using tail amputation and arteriovenous injury models in rats. The biocompatibility of CSK and CSMMT was evaluated by in vitro hemolysis, cytotoxicity assays, as well as acute toxicity test and skin irritation tests. The results show that CSK and CSMMT are promising composite materials with excellent biocompatibility and hemostatic properties that can effectively control bleeding.

Pacemaker cardiomyocytes of the sinoatrial node (SAN) beat more rapidly than cells of the working myocardium. Beating in SAN cells responds to β-adrenergic and cholinergic signaling by speeding up or slowing, respectively. Beat rate has traditionally been assessed using voltage or calcium sensitive dyes, however these may not reflect the true rate of beating because they sequester calcium. Finally, in vitro differentiated cardiomyocytes sometimes briefly pause during imaging giving inaccurate beat rates. We have developed a MATLAB automation to calculate cardiac beat rates directly from video clips based on changes in pixel density at the edges of beating areas. These data are normalized to minimize the effects of secondary movement and are converted to frequency data using a fast Fourier transform (FFT). We find that this gives accurate beat rates even when there are brief pauses in beating. This technique can be used to rapidly assess beating of cardiomyocytes in organoid culture. This technique could also be combined with field scanning techniques to automatically and accurately assess beating within a complex cardiac organoid.

Bessel beams have been successfully used in two-photon laser scanning fluorescence microscopy to extend the depth of field (EDF), which makes it possible to observe fast events volumetrically. However, the depth information is lost due to integration of fluorescence signals along the propagation direction. We describe the design and implementation of two-photon lasers scanning stereomicroscopy, which allows viewing dynamic processes in three-dimensional (3D) space stereoscopically in real-time with shutter glasses at the speed of 1.4 volumes per second. The depth information can be appreciated by human visual system or be recovered with correspondence algorithms for some cases.

Structured illumination microscopy (SIM) with axially optical sectioning capability has found widespread applications in three-dimensional live cell imaging in recent years, since it combines high sensitivity, short image acquisition time, and high spatial resolution. To obtain one sectioned slice, three raw images with a fixed phase-shift, normally 2π/3, are generally required. In this paper, we report a data processing algorithm based on the one-dimensional Hilbert transform, which needs only two raw images with arbitrary phase-shift for each single slice. The proposed algorithm is different from the previous two-dimensional Hilbert spiral transform algorithm in theory. The presented algorithm has the advantages of simpler data processing procedure, faster computation speed and better reconstructed image quality. The validity of the scheme is verified by imaging biological samples in our developed DMD-based LED-illumination SIM system.

Two-photon microscopy has enabled the study of individual cell behavior in live animals. Many organs and tissues cannot be studied, especially longitudinally, because they are located too deep, behind bony structures or too close to the lung and heart. Here we report a novel mouse model that allows long-term single cell imaging of many organs. A wide variety of live tissues were successfully engrafted in the pinna of the mouse ear. Many of these engrafted tissues maintained the normal tissue histology. Using the heart and thymus as models, we further demonstrated that the engrafted tissues functioned as would be expected. Combining two-photon microscopy with fluorescent tracers, we successfully visualized the engrafted tissues at the single cell level in live mice over several months. Four dimensional (three-dimensional (3D) plus time) information of individual cells was obtained from this imaging. This model makes long-term high resolution 4D imaging of multiple organs possible.

Focusing an annular laser beam can improve the axial trapping efficiency due to the reduction of the scattering force, which enables the use of a lower numerical aperture (NA) objective lens with a long working distance to trap particles in deeper aqueous medium. In this paper, we present an axicon-to-axicon scheme for producing parallel annular beams with the advantages of higher efficiency compared with the obstructed beam approach. The validity of the scheme is verified by the observation of a stable trapping of silica microspheres with relatively low NA microscope objective lenses (NA = 0.6 and 0.45), and the axial trapping depth of 5 mm is demonstrated in experiment.