A novel technique of three-dimensional reconstruction segmentation and analysis for sliced images of biological tissues

A novel technique of three-dimensional reconstruction segmentation and analysis for sliced images of biological tissues 1 2 first-author †E-mail:li_jingkm@126.com †E-mail:li_jingkm@126.com li_jingkm@126.com 3 1 4 4 2 2 Department of Equipments, Kunming General Hospital, Kunming 650032, China Department of Equipments, Kunming General Hospital, Kunming 650032, China State Key Laboratory for Biocontrol, School of Life Science, Sun Yat-sen University, Guangzhou 510275, China State Key Laboratory for Biocontrol, School of Life Science, Sun Yat-sen University, Guangzhou 510275, China Department of Ophthalmology, Yunnan Province People’s Hospital, Kunming 650032, China Department of Ophthalmology, Yunnan Province People’s Hospital, Kunming 650032, China Department of Orthopaedics, Kunming General Hospital, Kunming 650032, China Department of Orthopaedics, Kunming General Hospital, Kunming 650032, China A novel technique of three-dimensional (3D) reconstruction, segmentation, display and analysis of series slices of images including microscopic wide field optical sectioning by deconvolution method, cryo-electron microscope slices by Fourier-Bessel synthesis and electron tomography (ET), and a series of computed tomography (CT) was developed to perform simultaneous measurement on the structure and function of biomedical samples. The paper presents the 3D reconstruction segmentation display and analysis results of pollen spore, chaperonin, virus, head, cervical bone, tibia and carpus. At the same time, it also puts forward some potential applications of the new technique in the biomedical realm. Sliced images 3D reconstruction and analysis 3D segmentation Chaperonin Virus INTRODUCTION 1 The past decades witnessed unprecedented advances in the imaging science fields (Yelbuz et al., 2002 2002 ; Campagnola et al., 2002 2002 ; Sharpe et al., 2002 2002 ). The technique of 3D image reconstruction and analysis plays a significant role in the basic research of life science, which developed from 2D image processing but provides more information than 2D image, and has rapidly expanded to molecular and cellular field (Guo et al., 1995 1995 ; Trehan et al., 2003 2003 ). Despite the rapid development of the new field, many challenges are still waiting for resolution (Paddock, 2002 2002 ). Rare 3D image analysis techniques can be used for macroscopic and microcosmic fields. This paper presents a new method of 3D segmentation and analysis, by which the tomograms of the cellular and sub-cellular inner structure of tissue can be displayed, segmented and analyzed in 3D mode. METHODS 1 3D display 2 The 3D display method we used is volume display in real space based on light projection algorithm. Firstly, each point of the volume data is evaluated with the color and transparency values, and then the viewed plane and observation direction are determined. Then the reference frame is transferred into the 3D image coordinates, and the illumination of each point can be calculated by the method of light projection model. Finally, the 3D image can be synthesized by summing up the contributions of all points to each pixel of the image. 3D segmentation and analysis 2 We developed a new method called “virtual dissection” (Li et al., 2001 2001 ), which controls the volume projection in 3D space by Open GL interface. The function of the segmented line and the angle of segmented volume are properly resolved by applying circular cylindrical coordinates which are unified by differential and light tracing methods. The segmentation of any part and in any way can be realized in 3D volume while complicated calculation was avoided. Even more important, accurate measurement of area, volume, distance and angle in 3D mode can be done by analyzing the characteristics of the 3D volume and summing up the pixels. RESULTS AND DISCUSSION 1 3D analysis of optical section 2 The devices for taking pictures are: Nikon TE300 microscope, CCD camera (IMAC-CCD30 768×527), and computer for image displaying, storing, processing and analysis. After 2D image acquisition, the optical section can be obtained by removing the defocused background by deconvolution methods. The 3D segmentation of pollen spore is shown in Fig. 1 1 where the thickness of each section is 1.0 μm. The segmented part can be moved and quantified. The measurement results of the segmented parts A1 A2 are given in Table 1 1 . Item A1 A2 V1 V2 Surface area (μm 3 ) 18.1 23.3 Volume (μm 3 ) 97.3 127.7 3D analysis of tomogram by ET 2 The results of 3D reconstruction and arbitrary segmentation of Bombyx mori Cytoplasmic Polyhedrosis virus (CPV) are shown in Fig. 2a 2a whose tomograms were collected by Fourier-Bessel synthesis (Crowther, 1971 1971 ). The result of 3D reconstruction of chaperonin is shown in Fig. 2b 2b whose tomograms were collected by electron tomography (Nitsch et al., 1998 1998 ). Arbitrary parts can be segmented from the whole. Important information about super macromolecular organization and interactions can thus be obtained by analyzing the sub-part’s spatial relationships inside a whole. The technique can also be useful for determining the 3D conformation of macromolecular and molecular modelling in the future. 3D analysis of X-ray CT 2 The application of 3D segmentation in orthopaedics is very useful for computer-aided surgery plan and biomechanical analysis (Li et al., 2003 2003 ).The results of using the technique in Computer Aided Cranio-Maxillofacial Surgery System are shown in Fig. 3a 3a , the 3D segmentation of cervical bone is shown in Fig. 3b 3b , the 3D segmentation of tibia is shown in Fig. 3c 3c and the results on carpus are shown in Fig. 3d 3d . The technique can be applied to relatively bigger tissues such as those of the head or tibia and smaller tissues such as those of the carpus. CONCLUDING REMARK 1 Software has been developed based on the new technique of 3D segmentation and analysis presented in this paper. The 3D inner structure can be clearly displayed and measured accurately by this method. The new technique can be used as a common method for 3D reconstruction, display, and analysis of both macroscopic and microscopic sectioning of images, and can also be widely used for medical applications such as computer-aided surgery, virtual reality and minimally invasive medical procedures and so on. References PJ Campagnola , AC Millard , M Terasaki , PE Hoppe , CJ Malone , WA Mohler . Three dimensional high resolution second harmonic generation imaging of endogenous structural proteins in biological tissues . Biophys J , 2002 . 82 493 - 508 . . 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