KEYWORDS: Image segmentation, 3D modeling, Education and training, 3D image processing, Prostate, Data modeling, Biopsy, Pathology, Prostate cancer, Performance modeling
SignificanceIn recent years, we and others have developed non-destructive methods to obtain three-dimensional (3D) pathology datasets of clinical biopsies and surgical specimens. For prostate cancer risk stratification (prognostication), standard-of-care Gleason grading is based on examining the morphology of prostate glands in thin 2D sections. This motivates us to perform 3D segmentation of prostate glands in our 3D pathology datasets for the purposes of computational analysis of 3D glandular features that could offer improved prognostic performance.AimTo facilitate prostate cancer risk assessment, we developed a computationally efficient and accurate deep learning model for 3D gland segmentation based on open-top light-sheet microscopy datasets of human prostate biopsies stained with a fluorescent analog of hematoxylin and eosin (H&E).ApproachFor 3D gland segmentation based on our H&E-analog 3D pathology datasets, we previously developed a hybrid deep learning and computer vision-based pipeline, called image translation-assisted segmentation in 3D (ITAS3D), which required a complex two-stage procedure and tedious manual optimization of parameters. To simplify this procedure, we use the 3D gland-segmentation masks previously generated by ITAS3D as training datasets for a direct end-to-end deep learning-based segmentation model, nnU-Net. The inputs to this model are 3D pathology datasets of prostate biopsies rapidly stained with an inexpensive fluorescent analog of H&E and the outputs are 3D semantic segmentation masks of the gland epithelium, gland lumen, and surrounding stromal compartments within the tissue.ResultsnnU-Net demonstrates remarkable accuracy in 3D gland segmentations even with limited training data. Moreover, compared with the previous ITAS3D pipeline, nnU-Net operation is simpler and faster, and it can maintain good accuracy even with lower-resolution inputs.ConclusionsOur trained DL-based 3D segmentation model will facilitate future studies to demonstrate the value of computational 3D pathology for guiding critical treatment decisions for patients with prostate cancer.
Non-destructive 3D microscopy enables the accurate characterization of diagnostically and prognostically significant microstructures in clinical specimens with significantly increased volumetric coverage than traditional 2D histology. We are using open-top light-sheet microscopy to image prostate cancer biopsies and investigating the prognostic significance of 3D spatial features of nuclei within prostate cancer microstructures. Using a previously published 3D nuclear segmentation workflow, we identify a preliminary set of 3D graph-based nuclear features to quantify the 3D spatial arrangement of nuclei in prostate cancer biopsies. Using a machine classifier, we identify the features which prognosticate prostate cancer risk and demonstrate agreement with patient outcomes.
Glandular architecture is currently the basis for the Gleason grading of prostate biopsies. To visualize and computationally analyze glandular features in large 3D pathology datasets, we developed an annotation-free segmentation method for 3D prostate glands that relies upon synthetic 3D immunofluorescence (IF) enabled by generative adversarial networks. By using a fluorescent analog of H and E (cheap and fast stain) as an input, our strategy allows for accurate glandular segmentation that does not rely upon subjective and tedious human annotations or slow and expensive 3D immunolabeling. We aim to demonstrate that this 3D segmentation will enable improved prostate cancer prognostication.
Fluorescence-based slide-free digital pathology techniques, including 3D microscopy, are gaining interest as alternatives to traditional slide-based histology. Since pathologists are accustomed to the appearance of standard histology stains, the ability to render grayscale fluorescent images with color palettes that mimic traditional histology is valuable. We present FalseColor-Python, an open-source rapid digital-staining package that renders two-channel fluorescence images to mimic standard histology. Our package offers consistent color-space representations that are robust to both intra-specimen and inter-specimen variations in staining and imaging conditions, along with GPU-accelerated methods to process large datasets efficiently.
A current challenge is providing an accurate diagnosis in a timely manner for patients at risk of having prostate cancer. We developed and demonstrated a non-destructive procedure in which 12 biopsies can be cleared, fluorescently labeled, imaged with an open-top light-sheet (OTLS) microscope, and then diagnosed by a pathologist within an hour of biopsy. Using conventional histology as the gold standard, the accuracy, sensitivity, and specificity of 1Hr2Dx were all >90%. Such a method could potentially provide patients with a preliminary on-site diagnosis after a biopsy procedure, thereby alleviating anxiety and potentially expediting treatments.
Significance: Processing and diagnosing a set of 12 prostate biopsies using conventional histology methods typically take at least one day. A rapid and accurate process performed while the patient is still on-site could significantly improve the patient’s quality of life.
Aim: We develop and assess the feasibility of a one-hour-to-diagnosis (1Hr2Dx) method for processing and providing a preliminary diagnosis of a set of 12 prostate biopsies.
Approach: We developed a fluorescence staining, optical clearing, and 3D open-top light-sheet microscopy workflow to enable 12 prostate needle core biopsies to be processed and diagnosed within an hour of receipt. We analyzed 44 biopsies by the 1Hr2Dx method, which does not consume tissue. The biopsies were then processed for routine, slide-based 2D histology. Three pathologists independently evaluated the 3D 1Hr2Dx and 2D slide-based datasets in a blinded, randomized fashion. Turnaround times were recorded, and the accuracy of our method was compared with gold-standard slide-based histology.
Results: The average turnaround time for tissue processing, imaging, and diagnosis was 44.5 min. The sensitivity and specificity of 1Hr2Dx in diagnosing cancer were both >90 % .
Conclusions: The 1Hr2Dx method has the potential to improve patient care by providing an accurate preliminary diagnosis within an hour of biopsy.
Intraoperative assessment of breast surgical margins will be of value for reducing the rate of re-excision surgeries for lumpectomy patients. While frozen-section histology is used for intraoperative guidance of certain cancers, it provides limited sampling of the margin surface (typically <1 % of the margin) and is inferior to gold-standard histology, especially for fatty tissues that do not freeze well, such as breast specimens. Microscopy with ultraviolet surface excitation (MUSE) is a nondestructive superficial optical-sectioning technique that has the potential to enable rapid, high-resolution examination of excised margin surfaces. Here, a MUSE system is developed with fully automated sample translation to image fresh tissue surfaces over large areas and at multiple levels of defocus, at a rate of ∼5 min / cm2. Surface extraction is used to improve the comprehensiveness of surface imaging, and 3-D deconvolution is used to improve resolution and contrast. In addition, an improved fluorescent analog of conventional H&E staining is developed to label fresh tissues within ∼5 min for MUSE imaging. We compare the image quality of our MUSE system with both frozen-section and conventional H&E histology, demonstrating the feasibility to provide microscopic visualization of breast margin surfaces at speeds that are relevant for intraoperative use.
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