AI applications in renal pathology.

The explosive growth of artificial intelligence (AI) technologies, especially deep learning methods, has been translated at revolutionary speed to efforts in AI-assisted healthcare. New applications of AI to renal pathology have recently become available, driven by the successful AI deployments in digital pathology. However, synergetic developments of renal pathology and AI require close interdisciplinary collaborations between computer scientists and renal pathologists. Computer scientists should understand that not every AI innovation is translatable to renal pathology, while renal pathologists should capture high-level principles of the relevant AI technologies. Herein, we provide an integrated review on current and possible future applications in AI-assisted renal pathology, by including perspectives from computer scientists and renal pathologists. First, the standard stages, from data collection to analysis, in full-stack AI-assisted renal pathology studies are reviewed. Second, representative renal pathology-optimized AI techniques are introduced. Last, we review current clinical AI applications, as well as promising future applications with the recent advances in AI.

The two-component system, BasSR, is involved in the regulation of biofilm and virulence in avian pathogenic Escherichia coli.

Avian pathogenic Escherichia coli (APEC) is a subgroup of extra-intestinal pathogenic E. coli (ExPEC) strains that cause avian colibacillosis, resulting in significant economic losses to the poultry industry worldwide. It has been reported that a few two-component signal transduction systems (TCS) participate in the regulation of the virulence factors of APEC infection. In this study, a basSR-deficient mutant strain was constructed from its parent strain APECX40 (WT), and high-throughput sequencing (RNA-seq) was performed to analyse the transcriptional profile of WT and its mutant strain XY1. Results showed that the deletion of basSR down-regulated the transcript levels of a series of biofilm- and virulence-related genes. Results of biofilm formation assays and bird model experiments indicated that the deletion of basSR inhibited biofilm formation in vitro and decreased bacterial virulence and colonization in vivo. In addition, electrophoretic mobility shift assays confirmed that the BasR protein could bind to the promoter regions of several biofilm- and virulence-related genes, including ais, opgC and fepA. This study suggests that the BasSR TCS might be a global regulator in the pathogenesis of APEC infection. RESEARCH HIGHLIGHTS Transcriptional profiling showed that BasSR might be a global regulator in APEC. BasSR increases APEC pathogenicity in vivo. BasSR positively regulates biofilm- and the virulence-associated genes. BasSR can bind to the promoter regions of virulence-associated genes ais, opgC and fepA.

FNPC-SAM: Uncertainty-Guided False Negative/Positive Control for SAM on Noisy Medical Images.

The Segment Anything Model (SAM) is a recently developed all-range foundation model for image segmentation. It can use sparse manual prompts such as bounding boxes to generate pixel-level segmentation in natural images but struggles in medical images such as low-contrast, noisy ultrasound images. We propose a refined test-phase prompt augmentation technique designed to improve SAM's performance in medical image segmentation. The method couples multi-box prompt augmentation and an aleatoric uncertainty-based false-negative (FN) and false-positive (FP) correction (FNPC) strategy. We evaluate the method on two ultrasound datasets and show improvement in SAM's performance and robustness to inaccurate prompts, without the necessity for further training or tuning. Moreover, we present the Single-Slice-to-Volume (SS2V) method, enabling 3D pixel-level segmentation using only the bounding box annotation from a single 2D slice. Our results allow efficient use of SAM in even noisy, low-contrast medical images. The source code has been released at: https://github.com/MedICL-VU/FNPC-SAM.

Robust fiber orientation distribution function estimation using deep constrained spherical deconvolution for diffusion-weighted magnetic resonance imaging.

Purpose: Diffusion-weighted magnetic resonance imaging (DW-MRI) is a critical imaging method for capturing and modeling tissue microarchitecture at a millimeter scale. A common practice to model the measured DW-MRI signal is via fiber orientation distribution function (fODF). This function is the essential first step for the downstream tractography and connectivity analyses. With recent advantages in data sharing, large-scale multisite DW-MRI datasets are being made available for multisite studies. However, measurement variabilities (e.g., inter- and intrasite variability, hardware performance, and sequence design) are inevitable during the acquisition of DW-MRI. Most existing model-based methods [e.g., constrained spherical deconvolution (CSD)] and learning-based methods (e.g., deep learning) do not explicitly consider such variabilities in fODF modeling, which consequently leads to inferior performance on multisite and/or longitudinal diffusion studies. Approach: In this paper, we propose a data-driven deep CSD method to explicitly constrain the scan-rescan variabilities for a more reproducible and robust estimation of brain microstructure from repeated DW-MRI scans. Specifically, the proposed method introduces a three-dimensional volumetric scanner-invariant regularization scheme during the fODF estimation. We study the Human Connectome Project (HCP) young adults test-retest group as well as the MASiVar dataset (with inter- and intrasite scan/rescan data). The Baltimore Longitudinal Study of Aging dataset is employed for external validation. Results: From the experimental results, the proposed data-driven framework outperforms the existing benchmarks in repeated fODF estimation. By introducing the contrastive loss with scan/rescan data, the proposed method achieved a higher consistency while maintaining higher angular correlation coefficients with the CSD modeling. The proposed method is assessing the downstream connectivity analysis and shows increased performance in distinguishing subjects with different biomarkers. Conclusion: We propose a deep CSD method to explicitly reduce the scan-rescan variabilities, so as to model a more reproducible and robust brain microstructure from repeated DW-MRI scans. The plug-and-play design of the proposed approach is potentially applicable to a wider range of data harmonization problems in neuroimaging.

Cross-scale multi-instance learning for pathological image diagnosis.

Analyzing high resolution whole slide images (WSIs) with regard to information across multiple scales poses a significant challenge in digital pathology. Multi-instance learning (MIL) is a common solution for working with high resolution images by classifying bags of objects (i.e. sets of smaller image patches). However, such processing is typically performed at a single scale (e.g., 20× magnification) of WSIs, disregarding the vital inter-scale information that is key to diagnoses by human pathologists. In this study, we propose a novel cross-scale MIL algorithm to explicitly aggregate inter-scale relationships into a single MIL network for pathological image diagnosis. The contribution of this paper is three-fold: (1) A novel cross-scale MIL (CS-MIL) algorithm that integrates the multi-scale information and the inter-scale relationships is proposed; (2) A toy dataset with scale-specific morphological features is created and released to examine and visualize differential cross-scale attention; (3) Superior performance on both in-house and public datasets is demonstrated by our simple cross-scale MIL strategy. The official implementation is publicly available at https://github.com/hrlblab/CS-MIL.

Cell Spatial Analysis in Crohn's Disease: Unveiling Local Cell Arrangement Pattern with Graph-based Signatures.

Crohn's disease (CD) is a chronic and relapsing inflammatory condition that affects segments of the gastrointestinal tract. CD activity is determined by histological findings, particularly the density of neutrophils observed on Hematoxylin and Eosin stains (H&E) imaging. However, understanding the broader morphometry and local cell arrangement beyond cell counting and tissue morphology remains challenging. To address this, we characterize six distinct cell types from H&E images and develop a novel approach for the local spatial signature of each cell. Specifically, we create a 10-cell neighborhood matrix, representing neighboring cell arrangements for each individual cell. Utilizing t-SNE for non-linear spatial projection in scatter-plot and Kernel Density Estimation contour-plot formats, our study examines patterns of differences in the cellular environment associated with the odds ratio of spatial patterns between active CD and control groups. This analysis is based on data collected at the two research institutes. The findings reveal heterogeneous nearest-neighbor patterns, signifying distinct tendencies of cell clustering, with a particular focus on the rectum region. These variations underscore the impact of data heterogeneity on cell spatial arrangements in CD patients. Moreover, the spatial distribution disparities between the two research sites highlight the significance of collaborative efforts among healthcare organizations. All research analysis pipeline tools are available at https://github.com/MASILab/cellNN.

Alleviating tiling effect by random walk sliding window in high-resolution histological whole slide image synthesis.

Multiplex immunofluorescence (MxIF) is an advanced molecular imaging technique that can simultaneously provide biologists with multiple (i.e., more than 20) molecular markers on a single histological tissue section. Unfortunately, due to imaging restrictions, the more routinely used hematoxylin and eosin (H&E) stain is typically unavailable with MxIF on the same tissue section. As biological H&E staining is not feasible, previous efforts have been made to obtain H&E whole slide image (WSI) from MxIF via deep learning empowered virtual staining. However, the tiling effect is a long-lasting problem in high-resolution WSI-wise synthesis. The MxIF to H&E synthesis is no exception. Limited by computational resources, the cross-stain image synthesis is typically performed at the patch-level. Thus, discontinuous intensities might be visually identified along with the patch boundaries assembling all individual patches back to a WSI. In this work, we propose a deep learning based unpaired high-resolution image synthesis method to obtain virtual H&E WSIs from MxIF WSIs (each with 27 markers/stains) with reduced tiling effects. Briefly, we first extend the CycleGAN framework by adding simultaneous nuclei and mucin segmentation supervision as spatial constraints. Then, we introduce a random walk sliding window shifting strategy during the optimized inference stage, to alleviate the tiling effects. The validation results show that our spatially constrained synthesis method achieves a 56% performance gain for the downstream cell segmentation task. The proposed inference method reduces the tiling effects by using 50% fewer computation resources without compromising performance. The proposed random sliding window inference method is a plug-and-play module, which can be generalized for other high-resolution WSI image synthesis applications. The source code with our proposed model are available at https://github.com/MASILab/RandomWalkSlidingWindow.git.

Nucleus subtype classification using inter-modality learning.

Understanding the way cells communicate, co-locate, and interrelate is essential to understanding human physiology. Hematoxylin and eosin (H&E) staining is ubiquitously available both for clinical studies and research. The Colon Nucleus Identification and Classification (CoNIC) Challenge has recently innovated on robust artificial intelligence labeling of six cell types on H&E stains of the colon. However, this is a very small fraction of the number of potential cell classification types. Specifically, the CoNIC Challenge is unable to classify epithelial subtypes (progenitor, endocrine, goblet), lymphocyte subtypes (B, helper T, cytotoxic T), or connective subtypes (fibroblasts, stromal). In this paper, we propose to use inter-modality learning to label previously un-labelable cell types on virtual H&E. We leveraged multiplexed immunofluorescence (MxIF) histology imaging to identify 14 subclasses of cell types. We performed style transfer to synthesize virtual H&E from MxIF and transferred the higher density labels from MxIF to these virtual H&E images. We then evaluated the efficacy of learning in this approach. We identified helper T and progenitor nuclei with positive predictive values of 0.34 ± 0.15 (prevalence 0.03 ± 0.01) and 0.47 ± 0.1 (prevalence 0.07 ± 0.02) respectively on virtual H&E. This approach represents a promising step towards automating annotation in digital pathology.

An End-to-end Pipeline for 3D Slide-wise Multi-stain Renal Pathology Registration.

Tissue examination and quantification in a 3D context on serial section whole slide images (WSIs) were labor-intensive and time-consuming tasks. Our previous study proposed a novel registration-based method (Map3D) to automatically align WSIs to the same physical space, reducing the human efforts of screening serial sections from WSIs. However, the registration performance of our Map3D method was only evaluated on single-stain WSIs with large-scale kidney tissue samples. In this paper, we provide a Docker for an end-to-end 3D slide-wise registration pipeline on needle biopsy serial sections in a multi-stain paradigm. The contribution of this study is three-fold: (1) We release a containerized Docker for an end-to-end multi-stain WSI registration. (2) We prove that the Map3D pipeline is capable of sectional registration from multi-stain WSI. (3) We verify that the Map3D pipeline can also be applied to needle biopsy tissue samples. The source code and the Docker have been made publicly available at https://github.com/hrlblab/Map3D.

An Accelerated Pipeline for Multi-label Renal Pathology Image Segmentation at the Whole Slide Image Level.

Deep-learning techniques have been used widely to alleviate the labour-intensive and time-consuming manual annotation required for pixel-level tissue characterization. Our previous study introduced an efficient single dynamic network - Omni-Seg - that achieved multi-class multi-scale pathological segmentation with less computational complexity. However, the patch-wise segmentation paradigm still applies to Omni-Seg, and the pipeline is time-consuming when providing segmentation for Whole Slide Images (WSIs). In this paper, we propose an enhanced version of the Omni-Seg pipeline in order to reduce the repetitive computing processes and utilize a GPU to accelerate the model's prediction for both better model performance and faster speed. Our proposed method's innovative contribution is two-fold: (1) a Docker is released for an end-to-end slide-wise multi-tissue segmentation for WSIs; and (2) the pipeline is deployed on a GPU to accelerate the prediction, achieving better segmentation quality in less time. The proposed accelerated implementation reduced the average processing time (at the testing stage) on a standard needle biopsy WSI from 2.3 hours to 22 minutes, using 35 WSIs from the Kidney Tissue Atlas (KPMP) Datasets. The source code and the Docker have been made publicly available at https://github.com/ddrrnn123/Omni-Seg.

Omni-Seg: A Scale-Aware Dynamic Network for Renal Pathological Image Segmentation.

Comprehensive semantic segmentation on renal pathological images is challenging due to the heterogeneous scales of the objects. For example, on a whole slide image (WSI), the cross-sectional areas of glomeruli can be 64 times larger than that of the peritubular capillaries, making it impractical to segment both objects on the same patch, at the same scale. To handle this scaling issue, prior studies have typically trained multiple segmentation networks in order to match the optimal pixel resolution of heterogeneous tissue types. This multi-network solution is resource-intensive and fails to model the spatial relationship between tissue types. In this article, we propose the Omni-Seg network, a scale-aware dynamic neural network that achieves multi-object (six tissue types) and multi-scale (5× to 40× scale) pathological image segmentation via a single neural network. The contribution of this article is three-fold: (1) a novel scale-aware controller is proposed to generalize the dynamic neural network from single-scale to multi-scale; (2) semi-supervised consistency regularization of pseudo-labels is introduced to model the inter-scale correlation of unannotated tissue types into a single end-to-end learning paradigm; and (3) superior scale-aware generalization is evidenced by directly applying a model trained on human kidney images to mouse kidney images, without retraining. By learning from 150,000 human pathological image patches from six tissue types at three different resolutions, our approach achieved superior segmentation performance according to human visual assessment and evaluation of image-omics (i.e., spatial transcriptomics).

Influence of Cell-type Ratio on Spatially Resolved Single-cell Transcriptomes using the Tangram Algorithm: Based on Implementation on Single-Cell and MxIF Data.

The Tangram algorithm is a benchmarking method of aligning single-cell (sc/snRNA-seq) data to various forms of spatial data collected from the same region. With this data alignment, the annotation of the single-cell data can be projected to spatial data. However, the cell composition (cell-type ratio) of the single-cell data and spatial data might be different because of heterogeneous cell distribution. Whether the Tangram algorithm can be adapted when the two data have different cell-type ratios has not been discussed in previous works. In our practical application that maps the cell-type classification results of single-cell data to the Multiplex immunofluorescence (MxIF) spatial data, cell-type ratios were different, though they were sampled from adjacent areas. In this work, both simulation and empirical validation were conducted to quantitatively explore the impact of the mismatched cell-type ratio on the Tangram mapping in different situations. Results show that the cell-type difference has a negative influence on classification accuracy.

Deep Constrained Spherical Deconvolution for Robust Harmonization.

Diffusion weighted magnetic resonance imaging (DW-MRI) captures tissue microarchitecture at millimeter scale. With recent advantages in data sharing, large-scale multi-site DW-MRI datasets are being made available for multi-site studies. However, DW-MRI suffers from measurement variability (e.g., inter- and intra-site variability, hardware performance, and sequence design), which consequently yields inferior performance on multi-site and/or longitudinal diffusion studies. In this study, we propose a novel, deep learning-based method to harmonize DW-MRI signals for a more reproducible and robust estimation of microstructure. Our method introduces a data-driven scanner-invariant regularization scheme to model a more robust fiber orientation distribution function (FODF) estimation. We study the Human Connectome Project (HCP) young adults test-retest group as well as the MASiVar dataset (with inter- and intra-site scan/rescan data). The 8th order spherical harmonics coefficients are employed as data representation. The results show that the proposed harmonization approach maintains higher angular correlation coefficients (ACC) with the ground truth signals (0.954 versus 0.942), while achieves higher consistency of FODF signals for intra-scanner data (0.891 versus 0.826), as compared with the baseline supervised deep learning scheme. Furthermore, the proposed data-driven framework is flexible and potentially applicable to a wider range of data harmonization problems in neuroimaging.

Feasibility of Universal Anomaly Detection without Knowing the Abnormality in Medical Images.

Many anomaly detection approaches, especially deep learning methods, have been recently developed to identify abnormal image morphology by only employing normal images during training. Unfortunately, many prior anomaly detection methods were optimized for a specific "known" abnormality (e.g., brain tumor, bone fraction, cell types). Moreover, even though only the normal images were used in the training process, the abnormal images were often employed during the validation process (e.g., epoch selection, hyper-parameter tuning), which might leak the supposed "unknown" abnormality unintentionally. In this study, we investigated these two essential aspects regarding universal anomaly detection in medical images by (1) comparing various anomaly detection methods across four medical datasets, (2) investigating the inevitable but often neglected issues on how to unbiasedly select the optimal anomaly detection model during the validation phase using only normal images, and (3) proposing a simple decision-level ensemble method to leverage the advantage of different kinds of anomaly detection without knowing the abnormality. The results of our experiments indicate that none of the evaluated methods consistently achieved the best performance across all datasets. Our proposed method enhanced the robustness of performance in general (average AUC 0.956).

Democratizing Pathological Image Segmentation with Lay Annotators via Molecular-empowered Learning.

Multi-class cell segmentation in high-resolution Giga-pixel whole slide images (WSI) is critical for various clinical applications. Training such an AI model typically requires labor-intensive pixel-wise manual annotation from experienced domain experts (e.g., pathologists). Moreover, such annotation is error-prone when differentiating fine-grained cell types (e.g., podocyte and mesangial cells) via the naked human eye. In this study, we assess the feasibility of democratizing pathological AI deployment by only using lay annotators (annotators without medical domain knowledge). The contribution of this paper is threefold: (1) We proposed a molecular-empowered learning scheme for multi-class cell segmentation using partial labels from lay annotators; (2) The proposed method integrated Giga-pixel level molecular-morphology cross-modality registration, molecular-informed annotation, and molecular-oriented segmentation model, so as to achieve significantly superior performance via 3 lay annotators as compared with 2 experienced pathologists; (3) A deep corrective learning (learning with imperfect label) method is proposed to further improve the segmentation performance using partially annotated noisy data. From the experimental results, our learning method achieved F1 = 0.8496 using molecular-informed annotations from lay annotators, which is better than conventional morphology-based annotations (F1 = 0.7015) from experienced pathologists. Our method democratizes the development of a pathological segmentation deep model to the lay annotator level, which consequently scales up the learning process similar to a non-medical computer vision task. The official implementation and cell annotations are publicly available at https://github.com/hrlblab/MolecularEL.

Predicting Crohn's disease severity in the colon using mixed cell nucleus density from pseudo labels.

Crohn's disease (CD) is a debilitating inflammatory bowel disease with no known cure. Computational analysis of hematoxylin and eosin (H&E) stained colon biopsy whole slide images (WSIs) from CD patients provides the opportunity to discover unknown and complex relationships between tissue cellular features and disease severity. While there have been works using cell nuclei-derived features for predicting slide-level traits, this has not been performed on CD H&E WSIs for classifying normal tissue from CD patients vs active CD and assessing slide label-predictive performance while using both separate and combined information from pseudo-segmentation labels of nuclei from neutrophils, eosinophils, epithelial cells, lymphocytes, plasma cells, and connective cells. We used 413 WSIs of CD patient biopsies and calculated normalized histograms of nucleus density for the six cell classes for each WSI. We used a support vector machine to classify the truncated singular value decomposition representations of the normalized histograms as normal or active CD with four-fold cross-validation in rounds where nucleus types were first compared individually, the best was selected, and further types were added each round. We found that neutrophils were the most predictive individual nucleus type, with an AUC of 0.92 ± 0.0003 on the withheld test set. Adding information improved cross-validation performance for the first two rounds and on the withheld test set for the first three rounds, though performance metrics did not increase substantially beyond when neutrophils were used alone.

Topological-Preserving Membrane Skeleton Segmentation in Multiplex Immunofluorescence Imaging.

Multiplex immunofluorescence (MxIF) is an emerging imaging technology whose downstream molecular analytics highly rely upon the effectiveness of cell segmentation. In practice, multiple membrane markers (e.g., NaKATPase, PanCK and ß-catenin) are employed to stain membranes for different cell types, so as to achieve a more comprehensive cell segmentation since no single marker fits all cell types. However, prevalent watershed-based image processing might yield inferior capability for modeling complicated relationships between markers. For example, some markers can be misleading due to questionable stain quality. In this paper, we propose a deep learning based membrane segmentation method to aggregate complementary information that is uniquely provided by large scale MxIF markers. We aim to segment tubular membrane structure in MxIF data using global (membrane markers z-stack projection image) and local (separate individual markers) information to maximize topology preservation with deep learning. Specifically, we investigate the feasibility of four SOTA 2D deep networks and four volumetric-based loss functions. We conducted a comprehensive ablation study to assess the sensitivity of the proposed method with various combinations of input channels. Beyond using adjusted rand index (ARI) as the evaluation metric, which was inspired by the clDice, we propose a novel volumetric metric that is specific for skeletal structure, denoted as clDiceSKEL. In total, 80 membrane MxIF images were manually traced for 5-fold cross-validation. Our model outperforms the baseline with a 20.2% and 41.3% increase in clDiceSKEL and ARI performance, which is significant (p<0.05) using the Wilcoxon signed rank test. Our work explores a promising direction for advancing MxIF imaging cell segmentation with deep learning membrane segmentation. Tools are available at https://github.com/MASILab/MxIF_Membrane_Segmentation.

Dense Multi-Object 3D Glomerular Reconstruction and Quantification on 2D Serial Section Whole Slide Images.

There has been a long pursuit for precise and reproducible glomerular quantification in the field of renal pathology in both research and clinical practice. Currently, 3D glomerular identification and reconstruction of large-scale glomeruli are labor-intensive tasks, and time-consuming by manual analysis on whole slide imaging (WSI) in 2D serial sectioning representation. The accuracy of serial section analysis is also limited in the 2D serial context. Moreover, there are no approaches to present 3D glomerular visualization for human examination (volume calculation, 3D phenotype analysis, etc.). In this paper, we introduce an end-to-end holistic deep-learning-based method that achieves automatic detection, segmentation and multi-object tracking (MOT) of individual glomeruli with large-scale glomerular-registered assessment in a 3D context on WSIs. The high-resolution WSIs are the inputs, while the outputs are the 3D glomerular reconstruction and volume estimation. This pipeline achieves 81.8 in IDF1 and 69.1 in MOTA as MOT performance, while the proposed volume estimation achieves 0.84 Spearman correlation coefficient with manual annotation. The end-to-end MAP3D+ pipeline provides an approach for extensive 3D glomerular reconstruction and volume quantification from 2D serial section WSIs.

Compound Figure Separation of Biomedical Images: Mining Large Datasets for Self-supervised Learning.

With the rapid development of self-supervised learning (e.g., contrastive learning), the importance of having large-scale images (even without annotations) for training a more generalizable AI model has been widely recognized in medical image analysis. However, collecting large-scale task-specific unannotated data at scale can be challenging for individual labs. Existing online resources, such as digital books, publications, and search engines, provide a new resource for obtaining large-scale images. However, published images in healthcare (e.g., radiology and pathology) consist of a considerable amount of compound figures with subplots. In order to extract and separate compound figures into usable individual images for downstream learning, we propose a simple compound figure separation (SimCFS) framework without using the traditionally required detection bounding box annotations, with a new loss function and a hard case simulation. Our technical contribution is four-fold: (1) we introduce a simulation-based training framework that minimizes the need for resource extensive bounding box annotations; (2) we propose a new side loss that is optimized for compound figure separation; (3) we propose an intra-class image augmentation method to simulate hard cases; and (4) to the best of our knowledge, this is the first study that evaluates the efficacy of leveraging self-supervised learning with compound image separation. From the results, the proposed SimCFS achieved state-of-the-art performance on the ImageCLEF 2016 Compound Figure Separation Database. The pretrained self-supervised learning model using large-scale mined figures improved the accuracy of downstream image classification tasks with a contrastive learning algorithm. The source code of SimCFS is made publicly available at https://github.com/hrlblab/ImageSeperation.

Circle Representation for Medical Object Detection.

Box representation has been extensively used for object detection in computer vision. Such representation is efficacious but not necessarily optimized for biomedical objects (e.g., glomeruli), which play an essential role in renal pathology. In this paper, we propose a simple circle representation for medical object detection and introduce CircleNet, an anchor-free detection framework. Compared with the conventional bounding box representation, the proposed bounding circle representation innovates in three-fold: (1) it is optimized for ball-shaped biomedical objects; (2) The circle representation reduced the degree of freedom compared with box representation; (3) It is naturally more rotation invariant. When detecting glomeruli and nuclei on pathological images, the proposed circle representation achieved superior detection performance and be more rotation-invariant, compared with the bounding box. The code has been made publicly available: https://github.com/hrlblab/CircleNet.