Outer-Boundary Assisted Segmentation and Quantification of Trabecular Bones by an Imagej Plugin

Summary

We present a workflow for segmenting and quantifying trabecular bones for 2D and 3D images based on the bone's outer boundary using an ImageJ plugin. This approach is more efficient and accurate than the current manual hand-contouring approach, and provides layer-by-layer quantifications, which are not available in current commercial software.

Abstract

Micro-computed tomography (micro-CT) is routinely used to assess bone quantity and trabecular microstructural properties in small animals under different bone loss conditions. However, the standard approach for trabecular analysis of micro-CT images is slice-by-slice semi-automatic hand-contouring, which is labor intensive and error prone. Described here is an efficient method for automatic segmentation of trabecular bones according to the bone's outer boundaries, where trabecular bones can be identified and segmented automatically with accuracy with less operator bias when appropriate segmentation parameters are set. To profile satisfactory segmentation parameters, an image stack of segmentation results is displayed, where all possible combinations of the segmentation parameters are changed one by one in sequence, and segmentation results with associated parameters can easily be visually checked. As a quality-control feature of the plugin, simulated standard objects are quantified where the measured quantities can be compared with theoretical values. Layer-by-layer quantification of trabecular properties and trabecular thicknesses are reported by such a plugin, and the distributions of such properties within the selected regions can be profiled easily. Although layer-by-layer quantification retains more information about trabecular bones and facilitates further statistical analysis of structural changes, such measures are unavailable from the output of current commercial software, where only a single quantified value for each parameter is reported for each sample. Therefore, the described workflows are better approaches for analyzing trabecular bones with accuracy and efficiency.

Introduction

Micro-CT analysis of trabecular bones is the standard approach for tracking morphological changes of the bones in small animals under different bone loss conditions^1,2,3, where several variables related to the structures of the bones are reported⁴. However, such parameters are not evenly distributed in the metaphysis of long bones⁵, and only a summarized or averaged value is reported for each structural variable of each sample by current commercial micro-CT machines^6,7, ....

Protocol

Procedures involving animal subjects were conducted in accordance with the Guide for the Care and Use of Laboratory Animals (NIH publication, 8th edition, 2011), and have been reviewed and approved by the Institutional Animal Care and Use Committee of Wuhan University.

1. Software Installation

1. Install ImageJ software. Download the Windows version of the ImageJ (version 1.51p) software bundled with 64-bit Java from https://imagej.nih.gov/ij/. Extract the downloaded software into a folder, which will be subsequently referred to as the "ImageJ Directory".
   NOTE: Trabecular analysis plugins require 64-bit java run time....

Results

The trabecular analysis plugin is designed to automatically segment and quantify trabecular bones with accuracy. Initially, bone outer boundary is detected and delineated followed by a hole-filling operation where any holes within bone outer cortical shells are filled. Then an erosion operation is performed to exclude the outer cortical bones and get the segmented trabecular bones. Finally, measures of trabecular bones in the segmented region are quantified.

Discussion

This study describes an ImageJ plugin for analyzing trabecular bones, which is automatic, efficient, and user friendly. The plugin can also be used to quantify any 2D or 3D object for layer-by-layer measures of areas, volumes, and thicknesses. Currently, only a single measured value for each trabecular parameter is reported for each sample by standard micro-CT analysis, which cannot fully represent the characteristics of the measured entity in the selected analyzing region. The described plugin reports layer-by-layer qua.......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
ImageJ	NIH	imagej	Any version with a java 1.8 run time
trabecular analysis plugin	Bomomics	bomomics	free or commercial version
Micro CT scanner	Scanco	μ-50	micro CT from any vendor
Computer System	Lenovo		any brand
Windows Operating System	Microsoft	Windows 7 x64	any 64-bit Windows operating system
Office Software	Microsoft	Office 2010	any speadsheet software that has xy chart function