UvA-DARE (Digital Academic Repository) 3D atlas of human embryology New insights in human development de Bakker, B.S. Publication date 2018 Document Version Other version License Other Link to publication Citation for published version (APA): de Bakker, B. S. (2018). 3D atlas of human embryology: New insights in human development. General rights It is not permitted to download or to forward/distribute the text or part of it without the consent of the author(s) and/or copyright holder(s), other than for strictly personal, individual use, unless the work is under an open content license (like Creative Commons). Disclaimer/Complaints regulations If you believe that digital publication of certain material infringes any of your rights or (privacy) interests, please let the Library know, stating your reasons. In case of a legitimate complaint, the Library will make the material inaccessible and/or remove it from the website. Please Ask the Library: https://uba.uva.nl/en/contact, or a letter to: Library of the University of Amsterdam, Secretariat, Singel 425, 1012 WP Amsterdam, The Netherlands. You will be contacted as soon as possible. UvA-DARE is a service provided by the library of the University of Amsterdam (https://dare.uva.nl) Download date:05 Oct 2021 CHAPTER 6.1 - HITHERTO UNKNOWN DETAILED MUSCLE ANATOMY IN AN 8 WEEKS OLD EMBRYO Moritz V Warmbrunn* Bernadette S de Bakker* Jaco Hagoort Pauline B Alefs-de Bakker Roelof-Jan Oostra * = both authors contributed equally to the manuscript. Journal of Anatomy, minor revisions Supplementary material is available online at http://www.3datlasofhumanembryology.com "It is not the strongest of the species that survives, nor the most intelligent that survives. It is the one that is the most adaptable to change." Charles Darwin (1809-1882) 3D ATLAS OF HUMAN EMBRYOLOGY Abstract Congenital muscle diseases, such as myopathies or dystrophies, occur relatively frequent with estimated incidences of up to 4.7 per 100.000 newborns. To diagnose congenital diseases in the early stages of pregnancy, and to interpret the results of increasingly advanced in utero imaging techniques, profound knowledge of normal human morphological development of the locomotor system and the nervous system is necessary. Muscular development however, is an often neglected or only generally described topic in embryology textbooks and papers. To provide the required detailed and updated comprehensive picture of embryologic muscular anatomy, three-dimensional (3D) reconstructions were created based on serial histological sections of a human embryo at Carnegie stage 23 (eight weeks of development, crown-rump length of 23.8 mm), using Amira reconstruction software. Reconstructed muscles, tendons, bones and nerves were exported in a 3D-PDF file to permit interactive viewing. Almost all adult skeletal muscles of the trunk and limbs could be individually identified in their relative adult position. The pectoralis major muscle was divided in three separate muscle heads. The reconstructions showed remarkable highly developed extraocular, infrahyoid and suprahyoid muscles at this age but surprisingly also absence of the facial muscles that were described to be present at this stage of development. The overall stage of muscle development suggests heterochrony of skeletal muscle development. Several individual muscle groups were found to be developed earlier and in more detail than was described in current literature. Introduction Congenital muscle diseases, such as myopathies or dystrophies, occur relatively frequent and form a heavy burden for society but firstly for the affected child and its family. Global incidences of congenital muscular dystrophies are unknown, but described incidences of 4.7 live born per 100.000 inhabitants in north-eastern Italy between 1979-1993 (Mostacciuolo et al. 1996) and a prevalence of 3.95 cases per 100.000 inhabitants in northern England (Norwood et al. 2009) provide an indication. An epidemiological study of neuromuscular disorders in children under the age of 16 in western Sweden showed a prevalence of 63.1 per 100.000 inhabitants (Darin, Tulinius 2000). Comprehensive understanding of congenital muscle diseases requires knowledge of muscle anatomy and physiology. In adults, imaging techniques such as ultrasound and magnetic resonance imaging (MRI) are already contributing to diagnose the type of muscle disease (Wattjes, Kley & Fischer 2010) and could therefore also be used for in-utero imaging. However, little is known about late embryonic morphological development. Since fetal medicine is developing rapidly (Danzer, Johnson 2014, Moldenhauer 2014) and prenatal diagnostics are regularly performed in modern medical practice, the need for detailed anatomical knowledge in clinical settings becomes more pressing. Because skeletal muscles contribute significantly to the total volume of the human body, they can often be used as anatomical landmarks (De Battista et al. 2011). Accurate knowledge of late embryonic and fetal spatial anatomic relations is therefore crucial for effectively interpreting high resolution prenatal imaging techniques. Procedures such as sonography or fetoscopy during prenatal 234 Hitherto unknown detailed muscle anatomy in an 8 weeks old embryo diagnostics and fetal surgery can even predict embryonic health, as it is already in use for fetal health (Hutchinson, Kieser & Kramer 2014, D'Addario et al. 2005). Morphological muscle development is an often neglected or vaguely described topic in embryology textbooks and articles (Deries, Thorsteinsdottir 2016, Pu, Huang & Brand-Saberi 2016, Schoenwolf et al. 2009, Sadler 2012). Based on our experience in creating the 3D Atlas of Human Embryology (de Bakker et al. 2016), we hypothesized that human embryonic muscle anatomy is more advanced than would be concluded from descriptions in literature. To provide a more detailed and comprehensive picture of embryonic muscular anatomy, we used Amira reconstruction software to create three-dimensional (3D) reconstructions based on serial histological sections of a human embryo at Carnegie stage 23 (CS23, eight weeks of development), the final stage of embryonic development, with a crown-rump length of 23.8 mm. Reconstructed muscles, tendons, bones and nerves were exported in a 3D-PDF file to permit interactive investigation of the morphology of skeletal muscles at CS23. Comprehensive muscle reconstructions are novel to our knowledge and were not included in previous publications from our research group. Materials and Methods 3D Reconstructions Image acquisition and alignment was done as previously described by de Bakker et al. (2016, 2012). Digital images were captured from the series of sections of a CS23 specimen number 950 of the Carnegie collection in Silver Spring, MD, USA. This male specimen was collected after a miscarriage in 1914 and has been graded by specialists of the Carnegie collection as ‘good’ (O'Rahilly, Müller 1987). This specimen was fixed in formalin, transversally sectioned in 42.71 µm and then stained with an aluminum cochineal staining (de Bakker et al. 2016, O'Rahilly, Müller 1987). Based on the section 4.2 thickness and number of slices we calculated an approximate crown-rump length of 23.8 mm. Labeling of the 557 serial sections was accomplished by trained medical students, under the supervision of a physical therapist and four experienced embryologists. Skeletal muscles and tendons were manually segmented, with a Bamboo tablet and pen (http://www.wacom.com) based on digital images of the serial sections. Segmentation of structures on the serial sections was performed on grey scale images in Amira® software (version 5.3 - 5.6, http://www.amira.com), while the high-resolution full color dataset was displayed on a second computer screen. This way, every single muscle, including its tendons, was traced from origin to insertion. The nerves and bones of this specimen were already reconstructed in previous studies (de Bakker et al. 2016, de Bakker et al. 2012) and could therefore give 6.1 information about the topographic position of muscles relative to bones or nerves. The use of bones and nerves as anatomical landmarks allowed distinction between different types of tissues and provided topographical orientation throughout the body. If demarcation between individual muscles was not possible, muscles were labeled as muscle groups. Structure identification After reconstructing, muscles and tendons were identified using adult human anatomy atlases (Gilroy et al. 2009, Netter 2014, Drake et al. 2014) by establishing their origin and insertion and topographic relation to bones, nerves and surrounding organs. English names of the muscles were used, based on ‘Terminologia Anatomica’ (Baud, Neumann & Sprumont 1998). 235 3D ATLAS OF HUMAN EMBRYOLOGY Interactive 3D-PDF Amira surface files of the 3D reconstructions were converted to a .u3d file with Fiji (ImageJ, https://imagej.net). Deep Exploration (version 6.5 CSE, part of Corel DESIGNER Technical Suite X5 http://www.corel.com) was used for grouping and labeling of structures. Finally, Adobe Acrobat XI Pro (http://www.adobe.com) allowed to create an interactive 3D-PDF file containing all reconstructed structures and a custom user interface. A 3D-PDF can be viewed with a recent version of Adobe Reader® (X or higher, freeware, http://www.adobe.com) on MS Windows or MacOS systems, with javascript and playing of 3D content enabled. The Adobe Reader program allows users to not only view the 3D-PDF file, but also to zoom in or out on a structure, to hide it or to show it transparently. It provides the user with a clear view of the embryo in a 3D interactive fashion and thus to obtain a better understanding of the spatial relations between segmented organs and structures, in this case the musculoskeletal system in the embryo. This technique has proved its usefulness in earlier publications (de Bakker et al. 2016, de Bakker et al. 2012, Sizarov et al. 2011, Sizarov et al. 2012, de Boer et al. 2011, van den Berg et al. 2009, van den Berg, Moorman 2011). Fig. 1. Transversospinal muscle reconstructions. A: Left lateral view of the head and vertebral column of a stage 23 human embryo (56-60 days of development), specimen number 950.