MOTIVATION Quantitative studies of cellular morphodynamics rely on extracting leading-edge velocity time series based on accurate cell segmentation from live cell imaging. However, live cell imaging has numerous challenging issues regarding accurate edge localization. Fluorescence live cell imaging produces noisy and low-contrast images due to phototoxicity and photobleaching. While phase contrast microscopy is gentle to live cells, it suffers from the halo and shade-off artifacts that cannot be handled by conventional segmentation algorithms. Here, we present a deep learning-based pipeline, termed MARS-Net (Multiple-microscopy-type-based Accurate and Robust Segmentation Network), that utilizes transfer learning and data from multiple types of microscopy to localize cell edges with high accuracy, allowing quantitative profiling of cellular morphodynamics. SUMMARY To accurately segment cell edges and quantify cellular morphodynamics from live-cell imaging data, we developed a deep learning-based pipeline termed MARS-Net (multiple-microscopy-type-based accurate and robust segmentation network). MARS-Net utilizes transfer learning and data from multiple types of microscopy to localize cell edges with high accuracy. For effective training on distinct types of live-cell microscopy, MARS-Net comprises a pretrained VGG19 encoder with U-Net decoder and dropout layers. We trained MARS-Net on movies from phase-contrast, spinning-disk confocal, and total internal reflection fluorescence microscopes. MARS-Net produced more accurate edge localization than the neural network models trained with single-microscopy-type datasets. We expect that MARS-Net can accelerate the studies of cellular morphodynamics by providing accurate pixel-level segmentation of complex live-cell datasets.
Objectives: This study aimed to develop models that can automatically detect anterior disc displacement (ADD) of the temporomandibular joint (TMJ) on MRIs before orthodontic treatment to reduce the risk of developing serious complications after treatment. Methods: We used 9009 sagittal MRI of the TMJ as input and constructed three sets of deep learning models to detect ADD automatically. Deep learning models were developed using a convolutional neural network (CNN) based on the ResNet architecture and the “Imagenet” database. Five-fold cross-validation, oversampling, and data augmentation techniques were applied to reduce the risk of overfitting the model. The accuracy and area under the curve (AUC) of the three models were compared. Results: The performance of the maximum open mouth position model was excellent with accuracy and AUC of 0.970 (±0.007) and 0.990 (±0.005), respectively. For closed mouth position models, the accuracy and AUC of diagnostic Criteria 1 were 0.863 (±0.008) and 0.922 (±0.009), respectively significantly higher than that of diagnostic Criteria 2 with 0.839 (±0.013) (p = 0.009) and AUC of 0.885 (±0.018) (p = 0.003). The classification activation heat map also improved our understanding of the models and visually displayed the areas that play a key role in the model recognition process. Conclusion: Our CNN model resulted in high accuracy and AUC in detecting ADD and can therefore potentially be used by clinicians to assess ADD before orthodontic treatment, and hence improve treatment outcomes.
Time-lapse fluorescence live cell imaging has been widely used to study various dynamical processes in cell biology. However, fluorescence live cell images often have low contrast, noises, and uneven illumination, preventing accurate cell segmentation. The convolutional neural network has been successfully applied in natural image classification and segmentation by extracting hierarchical features, which could be transferred into other fields for image segmentation. Moreover, the temporal coherence in time-lapse images can allow us to extract sufficient features from a limited number of image frames to segment the entire time-lapse movies. In this paper, we propose a novel framework called vU-net, which integrates the VGG-16 1 pretrained model as an encoder and a U-net 2 derived simplified convolutional structure to reconstruct cell edge with a higher accuracy using limited training images. We evaluated our framework on the high-resolution images of paxillin, a canonical adhesion marker in migrating PtK1 cells acquired by a Total Internal Reflection Fluorescence (TIRF) microscope, and achieved higher accuracy of cell segmentation than conventional U-net. We also validated our framework on noisy confocal fluorescence live images of GFP-mDia1 in PtK1 cells. We demonstrated that vU-net could be practically applied to challenging live cell movies since it required limited training sets and achieved highly accurate segmentation.
Machine learning approaches have shown great promise in biology and medicine discovering hidden information to further understand complex biological and pathological processes. In this study, we developed a deep learning-based machine learning algorithm to meaningfully process image data and facilitate studies in vascular biology and pathology. Vascular injury and atherosclerosis are characterized by neointima formation caused by the aberrant accumulation and proliferation of vascular smooth muscle cells (VSMCs) within the vessel wall. Understanding how to control VSMC behaviors would promote the development of therapeutic targets to treat vascular diseases. However, the response to drug treatments among VSMCs with the same diseased vascular condition is often heterogeneous. Here, to identify the heterogeneous responses of drug treatments, we created an in vitro experimental model system using VSMC spheroids and developed a machine learning-based computational method called HETEROID (heterogeneous spheroid). First, we established a VSMC spheroid model that mimics neointima-like formation and the structure of arteries. Then, to identify the morphological subpopulations of drug-treated VSMC spheroids, we used a machine learning framework that combines deep learning-based spheroid segmentation and morphological clustering analysis. Our machine learning approach successfully showed that FAK, Rac, Rho, and Cdc42 inhibitors differentially affect spheroid morphology, suggesting that multiple drug responses of VSMC spheroid formation exist. Overall, our HETEROID pipeline enables detailed quantitative drug characterization of morphological changes in neointima formation, that occurs in vivo, by single-spheroid analysis.
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