Search Results (168)

Time-lapse microscopy of mesenchymal stem cell (MSC) cultures allows for the quantitative observation of their self-renewal, proliferation, and differentiation. However, the rigorous comparison of two conditions, baseline (A) versus perturbation (B) (the addition of molecular factors, environmental shifts, genetic modification, etc.), remains difficult because morphology, division timing, and migratory behavior are highly heterogeneous at the single-cell scale. MSCs can be used as an in vitro model to study cell morphology and kinetics in order to assess the effect of, for example, gene therapy and prime editing in the near future. By combining static, frame-wise morphology with dynamic descriptors, we can obtain weight profiles that highlight which morphological and behavioral dimensions drive divergence. In this study, we present A/B Cells Policy: a modular, open-source Python package implementing a robust cell tracking pipeline. It integrates a YOLO-based architecture as a two-stage assignment framework with fallback and recovery passes, re-identification of lost tracks, and lineage reconstruction. The framework links descriptive statistics to a transferable system, opening up avenues for regenerative medicine, pharmacology, and early translational pipelines. It does this by providing an interpretable, measurement-based bridge between in vitro imaging and in silico intervention strategy planning. Full article

Similarly to the short-lived messenger nitric oxide (NO), the more stable carbon monoxide (CO) molecule can also activate soluble guanylyl cyclase (sGC) to increase cGMP levels. However, CO-induced cGMP production is much less efficient. Using an accessible invertebrate model, we dissect a potential interaction between the canonical NO/sGC/cGMP and CO signalling pathways during development. The embryonic midgut of locusts is innervated by neurons that migrate in four discrete chains on its outer surface. Transcellular diffusing NO stimulates enteric neuron migration via cGMP signalling. The application of an NO donor results in virtually all enteric neurons being cGMP-immunoreactive while CO increases cGMP production only in approximately 33% of the migrating neurons. Cellular CO release appears to act as a slow down signal for motility. We quantify how CO specifically increases the interneuronal distance during chain migration. Moreover, time-lapse microscopy shows that CO reduces the directionality of the migrating neurons. These findings support the function of NO and CO as antagonistic signals for the coordination of collective cell migration during the development of the enteric nervous system. These experiments and the resulting insights into basic scientific questions prove once more that locust embryos are not only preparations for basic research, but also relevant models for screening of drugs targeting NO and CO signalling pathways as well as for isolating compounds affecting neuronal motility in general. Full article

Murine microglia exhibit rapid self-renewal upon removal from the postnatal brain. However, the signaling pathways that regulate microglial repopulation remain largely unclear. To address this knowledge gap, we depleted microglia from mixed glial cultures using anti-CD11b magnetic particles and cultured them for 4 weeks to monitor their repopulation ability in vitro. Flow cytometry and immunocytochemistry revealed that anti-CD11b bead treatment effectively eliminated >95% of microglia in mixed glial cultures. Following removal, the number of CX3CR1-positive microglia gradually increased; when a specific threshold was reached, repopulation ceased without any discernable rise in cell death. Cell cycle and 5-ethynyl-2′-deoxyuridine incorporation assays suggested the active proliferation of repopulating microglia at d7. Time-lapse imaging demonstrated post-removal division of microglia. Colony-stimulating factor 1 receptor-phosphoinositide 3-kinase-protein kinase B signaling was identified as crucial for microglial repopulation, as pharmacological inhibition or neutralization of the pathway significantly abrogated repopulation. Transwell cocultures revealed that resident microglia competitively inhibited microglial proliferation probably through contact inhibition. This in vitro microglial removal system provides valuable insights into the mechanisms underlying microglial proliferation. Full article

Objectives: With the development of artificial intelligence technology in medicine, an intelligent deep learning-based embryo scoring system (iDAScore) has been developed on full-time lapse sequences of embryos. It automatically ranks embryos according to the likelihood of achieving a fetal heartbeat with no manual input from embryologists. To ensure its performance, external validation studies should be performed at multiple clinics. Methods: A total of 6291 single vitrified–thawed blastocyst transfer cycles from 2018 to 2021 at the Reproductive Medicine Center, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology were retrospectively analyzed by the iDAScore model. Patients with two or more blastocysts transferred and blastocysts that were not cultured in a time-lapse incubator were excluded. Blastocysts were divided into four comparably sized groups by first sorting their iDAScore values in ascending order and then compared with the clinical, perinatal, and neonatal outcomes. Results: Our results showed that clinical pregnancy, miscarriage, and live birth significantly correlated with iDAScore (p < 0.001). For perinatal and neonatal outcomes, no significant difference was shown in four iDAScore groups, except sex ratio. Uni- and multivariable logistic regressions showed that iDAScore was significantly positively correlated with live birth rate (p < 0.05). Conclusions: In conclusion, the objective ranking can prioritize embryos reliably and rapidly for transfer, which could allow embryologists more time for processes requiring hands-on procedures. Full article

To address the challenges in identifying NAPL contamination within low-permeability clay sites, this study innovatively integrates high-density electrical resistivity tomography (ERT) with a UNet deep learning model to establish an intelligent contamination detection system. Taking an industrial site in Shanghai as the research object, we collected apparent resistivity data using the WGMD-9 system, obtained resistivity profiles through inversion imaging, and constructed training sets by generating contamination labels via K-means clustering. A semantic segmentation model with skip connections and multi-scale feature fusion was developed based on the UNet architecture to achieve automatic identification of contaminated areas. Experimental results demonstrate that the model achieves a mean Intersection over Union (mIoU) of 86.58%, an accuracy (Acc) of 99.42%, a precision (Pre) of 75.72%, a recall (Rec) of 76.80%, and an F1 score (f1) of 76.23%, effectively overcoming the noise interference in electrical anomaly interpretation through conventional geophysical methods in low-permeability clay, while outperforming DeepLabV3, DeepLabV3+, PSPNet, and LinkNet models. Time-lapse resistivity imaging verifies the feasibility of dynamic monitoring for contaminant migration, while the integration of the VGG-16 encoder and hyperparameter optimization (learning rate of 0.0001 and batch size of 8) significantly enhances model performance. Case visualization reveals high consistency between segmentation results and actual contamination distribution, enabling precise localization of spatial morphology for contamination plumes. This technological breakthrough overcomes the high-cost and low-efficiency limitations of traditional borehole sampling, providing a high-precision, non-destructive intelligent detection solution for contaminated site remediation. Full article

Constructing stable and flexible neuronal networks with multi-neurite wiring is essential for the in vitro modeling of brain function, connectivity, and neuroplasticity. However, most existing neuroengineering platforms rely on static microfabrication techniques, which limit the ability to dynamically control circuit architecture during cultivation. In this study, we developed a modifiable agarose gel-based platform that enables real-time microstructure fabrication using an infrared (IR) laser system under live-cell conditions. This approach allows for the stepwise construction of directional neurite paths, including sequential microchannel formation, cell chamber fabrication, and controlled neurite–neurite crossings. To support long-term neuronal health and network integrity in agarose microstructures, we incorporated direct glial co-culture into the system. A comparative analysis showed that co-culture significantly enhanced neuronal adhesion, neurite outgrowth, and survival over several weeks. The feeder layer configuration provided localized trophic support while maintaining a clear separation between glial and neuronal populations. Dynamic wiring experiments further confirmed the platform’s precision and compatibility. Neurites extended through newly fabricated channels and crossed pre-existing neurites without morphological damage, even when laser fabrication occurred after initial outgrowth. Time-lapse imaging showed a temporary growth cone stalling at crossing points, followed by successful elongation in all tested samples. Furthermore, the direct laser irradiation of extending neurites during microstructure modification did not visibly impair neurite elongation, suggesting minimal morphological damage under the applied conditions. However, potential effects on molecular signaling and electrophysiological function remain to be evaluated in future studies. Together, these findings establish a powerful, flexible system for constructive neuroengineering. The platform supports long-term culture, real-time modification, and multidirectional wiring, offering new opportunities for studying neural development, synaptic integration, and regeneration in vitro. Full article

Systemic chemotherapy is still the first-line treatment for cancer, and it’s associated with toxic side effects, chemoresistance, and ultimately cancer recurrence. Rapid gelling hydrogels can overcome this limitation by providing localized delivery of anti-cancer agents to solid tumors. Silk hydrogels are extremely biocompatible and suitable for anti-cancer drug delivery, but faster gelling formulations are needed. In this study, we introduce a rapid gelling hydrogel formulation (<3 min gelling time) due to chemical crosslinking between silk fibroin and curcumin, initiated by the addition of minute quantities of horseradish peroxidase (HRP) and hydrogen peroxide (H₂O₂). The novel observation in this study is that curcumin, while being a free-radical scavenger, also participates in accelerating silk di-tyrosine crosslinking in the presence of HRP and H₂O₂. Using UV-Vis, rheology, and time-lapse videos, we convincingly show that curcumin accelerates silk di-tyrosine crosslinking reaction in a concentration-dependent manner, and curcumin remains entrapped in the hydrogel post-crosslinking. FTIR results show an increase in secondary beta-sheet structures within hydrogels, with increasing concentrations of curcumin. Furthermore, we show that curcumin-silk di-tyrosine hydrogels are toxic to U2OS osteosarcoma cells, and most cancer cells are dead within short time scales of 4 h post-encapsulation. Full article

Time-lapse imaging has made possible the detailed observation of all stages of embryonic development, including also from the extrusion of the second polar body up to the first cleavage. By extensive observation, we achieved detection of a variety of behaviors of PBIIs such as (a) morphologically static behavior, (b) amoeboid movement, (c) shrinking, (d) fragmenting, and (e) ruffling. Retrospective analysis was performed on 282 ICSI zygotes derived from 69 ART treatment cycles from January to August 2019. Zygotes with morphologically static PBIIs (a) and PBIIs showing various behaviors (b)~(e) were classified into Group 1 (n = 70) and Group 2 (n = 212), respectively. Based on the rates of irregular division, good quality embryos, and the time from the PBII extrusion, pronuclear breakdown to the first cleavage was compared between groups (Study 1). Furthermore, the relationship between the type of PBII behaviors and ploidy in 94 biopsied blastocysts from 15 cycles was examined, in which one or more euploid embryos were obtained between August 2021 and July 2024 (Study 2). The results showed that good quality embryos tended to have morphologically static PBIIs, and that euploid embryos were absent in embryos with fragmenting and ruffling PBIIs. The behavior of PBIIs may be a new predictor of embryonic developmental potential, and, in the future, morphokinetic behaviors of PBIIs may be a useful parameter for AI-assisted embryo evaluation systems. Full article

Background: Phytocarriers are advanced drug delivery systems that use biocompatible and biodegradable materials to enhance the efficacy, stability, and bioavailability of natural products. The sea buckthorn (Hippophae rhamnoides L.) berry extract is rich in essential fatty acids and antioxidants, including vitamin C, vitamin E, and anthocyanins, which contribute to its wide-ranging health benefits. In this study, we assessed the morphology, intracellular delivery, and anti-inflammatory effect of sodium cholate (NaC) and sodium deoxycholate (NaDC)-based phytocarriers loaded with ethanolic extract from sea buckthorn berries (sea buckthorn carrier nanostructures, further defined as phytocarriers). Methods: Negative and electron cryo-microscopy were used to analyze hollow and loaded nanocarriers. The cyto-compatibility of nanocarriers was assessed by endpoint (LDH and MTS) and real-time cell assays, on both human fibroblasts (HS27) and human normal monocytes (SC). The anti-inflammatory effect of hollow and loaded nanocarriers was tested by multiplexing. Results: The negative and electron cryo-microscopy analyses showed that NaC-based phytocarriers were spherical, whilst NaDC-based phytocarriers were predominantly polymorphic. Moreover, the NaDC-based phytocarriers frequently formed large lipid networks or “plaques”. Although 24 h cytotoxicity testing showed both types of nanocarriers are biocompatible with human fibroblasts and monocytes, based on a long-term real-time assay, NaDC delayed fibroblast proliferation. NaC sea buckthorn phytocarriers did not impair fibroblast proliferation in the long term and they were uptaken by cells, as shown by hyperspectral microscopy. NaC nanocarriers and NaC sea buckthorn phytocarriers induced an anti-inflammatory effect, lowering IL-8 cytokine production in normal human monocytes as soon as 4 h of treatment lapsed. Conclusions: NaC-derived phytocarriers loaded with sea buckthorn alcoholic extract are a cell-compatible delivery system with anti-inflammatory properties. Full article

The main component of vacuum interrupters responsible for ensuring the correct flow of current is the contact system. In a vacuum environment, due to the higher values of the mean free path of electrons and particles in the contact gap, the material and condition of the contacts exert the greatest influence on the development of the arc discharge. To accurately analyze the phenomenon of discharge development in vacuum insulating systems, the authors conducted a time-lapse photographic analysis of a vacuum electric arc. For this purpose, they used a test setup comprising a discharge chamber, a vacuum pump set, a power and load assembly, an ultra-high-speed camera, and an oscilloscope with dedicated probes. The measurement process involved connecting the system, determining the power supply, load, and measurement parameters and subsequently performing contact opening operations while simultaneously recording the process using the oscilloscope and ultra-high-speed camera. An analysis of a low-current vacuum arc in a residual helium gas environment, with a pressure of p = 1.00 × 10¹ Pa was carried out. Different phases of vacuum arc burning between electrodes in the discharge chamber were identified. In the stable phase, the arc voltage remained constant, while in the unstable phase, the arc voltage increased. The results of the time-lapse analysis were compared with the characteristics recorded by the oscilloscope, revealing a correlation between the increase in vacuum arc voltage and the intensity of flashes in the interelectrode space. The movement of microparticles ejected from the surface of the contacts—either reflecting or adhering to one of the electrodes—was observed. This analysis provides a deeper understanding of the processes involved in discharge formation and development under reduced pressure conditions. Understanding these mechanisms can support the design of vacuum interrupters, particularly in the selection of suitable contact materials and shapes. Full article

The electrical resistivity tomography (ERT) method has been increasingly integrated with hydrogeological risk mitigation strategies to monitor the internal conditions and the stability of natural and artificial slopes. In this paper, we discuss a case study in which numerical simulations were essential to validate the interpretation of the resistivity images obtained from an ERT monitoring system installed on a critical slope in Italy. An initial analysis of the monitoring data after rainfall events in the study site showed that the resistivity values were decreased only in the central zone along the ERT line, but they were increased in the two sides of the profile. Opposite behaviors were observed during the drying processes following the rainfall events. Core samples show complex geology at the study site, which might justify uneven responses of the different subsurface bodies to meteorological events. However, we decided to investigate the possible inversion artifacts resulting from the individual inversion of the tomographic sections. Forward modeling simulations on simplified time-lapse models of the study site were performed to explore this problem and to compare the individual and time-lapse inversions. Synthetic tests confirmed the nature of these unexpected behaviors and assessed the absolute necessity of a time-lapse approach for a correct inversion of monitoring data in the presence of a complex geological model such as the one of this case study. By applying the time-lapse inversion approach to the real data, the inversion artifact problem was substantially solved, arriving after the proper calibration of the inversion parameters, mainly the time-lapse damping factor and the spatial and temporal roughness constraints, to a reduction in the inversion artifacts to less than 5%. Full article

A time-lapse live embryo monitoring system provides a powerful approach to recording dynamic developmental events of cultured embryos in detail. By obtaining continuous short-interval images, blastocyst formation can be predicted and embryos can be selected. The objective of this study was to investigate the morphokinetic parameters of fishing cat–domestic cat interspecies somatic cell nuclear transfer (iSCNT) embryos from one-cell to blastocyst stages, and in particular, the cleavage patterns of the first division in iSCNT and IVF embryos, as these play a central role in euploidy. Domestic cat in vitro fertilization (IVF) embryos were set up as controls. The results show that morula and blastocyst development rates were significantly lower in the iSCNT embryos compared to their IVF counterparts. All earlier time points of embryonic development before the onset of blastulation in the iSCNT embryos were significantly delayed when compared with their IVF counterparts. In iSCNT, normal embryos (defined as those that developed to the blastocyst stage) took a longer time to reach the morula stage, and these morulas were more likely to undergo compaction, compared to their arrested embryo counterparts. Direct cleavage in the first division is a morphological aberration, and was seen with greater prevalence in iSCNT embryos than control IVF embryos; these aberrant embryos displayed a significantly lower blastocyst development rate than embryos that had undergone normal cleavage. In conclusion, the morphokinetic parameters of fishing cat–domestic cat iSCNT embryos at early stages could be used to predict their potential for development to the blastocyst stage. A time-lapse imaging system is potentially a powerful tool for selecting early embryos with developmental potential for transfer, and hence, for improving feline iSCNT efficiency. Full article

We developed a portable microfluidic system that combines spontaneous lumen formation from human umbilical endothelial cells (HUVECs) in fibrin–collagen hydrogels with active perfusion controlled by a braille actuator. Adaptive interstitial flow and feedthrough perfusion switching enabled the successful culture of spontaneously formed naturally branched lumens for more than one month. We obtained many large-area (2 mm × 3 mm) long-term (more than 30 days per run) time-lapse image datasets of the in vitro luminal network using this microfluidic system. We also developed an automatic image analysis pipeline to extract the morphology of the lumen network and node–edge network structure weighted with segmentwise flow parameters. The automatic lumen area measurements revealed that almost all lumens were successfully cultured in this system for approximately 50 days, following the meshwork, sprouting, remodeling, stability, and erosion stages. We found that the optimization of the lumen network during the remodeling stage can be explained by the decrease in the betweenness centrality of the WSS-weighted network and the increase in the strength centrality of the flow-rate-weighted network. Full article

Background. Critical and progressive cachexia may be observed in numerous medical disciplines, but in patients with various diseases, several pathways overlap (endocrine, inflammatory and kidney diseases, heart failure, cancer). Methods. Unlike numerous cohort studies that examine thyroid cancer and risk factors, a different method was used to avoid bias and analyze the sequence of events, i.e., the pathway. A case-control analysis is presented on patients with initial immune-mediated thyroiditis complicated by cachexia, presenting pulmonary pathology coexisting with opportunistic infection, and ultimately diagnosed with cancer (TC—thyroid cancer, misdiagnosed as lung cancer). Results. Contrary to other patients with lung cancer, the presented patients were not active smokers and exclusively women who developed cachexia with existing autoimmune processes in the first phase. Furthermore, the coexistence of short overall survival without cancer progression in the most seriously ill patients, as well as correlation with sex (contrary to history of smoking) and predisposition to mycobacterial disease, are very suggestive. Although we describe three different autoimmune conditions (de Quervain’s, Graves’, and atrophic thyroiditis), disturbances in calcium and metabolic homeostasis, under the influence of hormonal and inflammatory changes, are crucial factors of cachexia and prognosis. Conclusions. The unique sequence sheds light on immune-mediated thyroid disease as a subclinical paraneoplastic process modified by various therapeutic regimens. However, it is also associated with cachexia, systemic consequences, and atypical sequelae, which require a holistic approach. The differential diagnosis of severe cachexia, adenocarcinoma with pulmonary localization, and tuberculosis reactivation requires an analysis of immunological and genetic backgrounds. Contrary to highly specialized teams (e.g., lung cancer units), immunotherapy and general medicine in aging populations require a multidisciplinary, holistic, and inquiring approach. The lack of differentiation, confusing biases, and discrepancies in the literature are the main obstacles to statistical research, limiting findings to correlations of common factors only. Time-lapse case studies such as this one may be among the first to build evidence of a pathway and an association between inflammatory and endocrine imbalances in cancer cachexia. Full article

The association between embryo morphokinetics and its developmental competence is well documented. For instance, early cleaved embryos are more competent in developing to blastocysts, whereas the proportion of abnormally cleaved embryos that further developed to blastocysts is low. Numerous factors, such as the parental age, lifestyle, health, and smoking habits have been reported to affect the embryo morphokinetics and, consequently, its development. However, less is known about the effect of environmental stressors on embryo morphokinetics. The current review discusses the effect of the most concerning environmental stressors on embryo morphokinetics. These stresses include heat stress and human-made chemicals such as phthalates (e.g., bis-(2-ethylhexyl phthalate, dibutyl phthalate, dimethyl phthalate, and their primary metabolites), herbicides (e.g., diaminochlorotriazine, the primary metabolite of atrazine), pharmaceutical compounds (e.g., carbamazepine, nocodazole) and pro-oxidant agents (cumene hydroperoxide, Triton X-100), as well as naturally occurring toxins such as mycotoxin (e.g., aflatoxin B1 and its metabolite, and ochratoxin A). In addition, this review discusses the effect of ionizing or non-ionizing radiation and viral infections (e.g., SARS-CoV-2, papillomavirus). Finally, it points out some potential mechanisms that underlie the impairment of embryo morphokinetics, and it suggests protective compounds, mainly the supplementation of antioxidants to improve the morphokinetics, and consequently, the embryo developmental competence. Full article