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3D Computerized Reconstruction of The Human Embryonic Lens Scholar(s): Luis Noboa , Jih-Shu Ruey. Mentor(s): Dr. Richard Hendrix, Dr Joel Hernandez Borough of Manhattan Community College Abstract Discussion We are interested in these reconstructions because they make excellent model systems at the Tissue, Molecular, and Cellular level. Our interest is in the geometrical transformation the tissue goes through in the course of production. We are developing an efficient way to measure the geometric properties of human embryonic ocular lenses by means of computer aided design. Specifically, a new software program called WinSURF is being tested for its ability to provide us with more accurate numerical measurements at greater speed. We are working with serially sectioned human eye specimens from the Carnegie Collection kindly donated by the National Museum of Health and Medicine. The Materials come to us as digitized serial micrographs of entire human embryos between stages 10 (circa 20 days post conception) and 17 (circa 50 days post conception). These serial sections are aligned and subjected to 3D computerized reconstruction using the WinSURF modeling system, which produces 3D virtual images of staged human ocular lenses. Linear, surface and volume measurements will constitute the geometric data. Analysis of this data will allow us to compare the geometrodynamics of human ocular lens development with previous research on lamprey and chickens. These new analyses will aid in the understanding of the forces contributing to the construction of normal human ocular lenses. Ultimately, it should also give us insight into congenital eye abnormalities such as Cyclopia and Micropthalmia. Flat plates are seen during early development, by stage 12 they get larger. We suspect the major driving force is the growth and division of cells. During the placode stage (13) there is an increase of cells, that leads to a cup formation in stages (14, 15). A deep cup forms in stage 16 called the lens pit. In stage 17 the ectoderm pinches off to form the hollow lens vesicle. These stages are not unlike what we see in the development of the chick and mouse lens. However they differ in size of the tissue, cellular size, and developmental time. We are asking whether the succession of geometries seen at different stages represents a conserved mechanism in development. That is to say, how do the driving forces establish the structures in the early lens, that are similar across the vertebrate lineage? To establish this we took advantage of digitized serial sections provided by the Carnegie Foundation Materials & Methods Introduction Our human embryological specimens of study were originally gathered by the Carnegie Foundation and have since become property of the National Museum of Health and Medicine, and the Walter Reed Army Medical Center in Washington, DC. Human Embryos were placed in a fixative over 100 years ago. At a later time the fixed embryos were processed in an alcohol and xylene series. Finally, they were embedded in paraffin and serial sectioned in their entirely at 10-30 microns on a paraffin microtome. Digital serial photographs were produced from the sections, and samples were obtained from the National Museum of Health and Medicine. Data was extracted from targeted internal structures (like the lens) of a human embryonic fetus for the study of the formation of the visual system - in particular the development of the embryonic lens at various fetal stages. Studying the shape and location of these structures and how they are connected to each other is essential for understanding human development. It is also the basis for knowing how and when errors in development occur and if a possibility exists for a corrective intervention. Data Stage Time (days) Tissue Height (μm) Tissue Thickness (μm) 13 32 35.79 10 46040.45 46040.45 920808.90 921 17 45 42.81 15 63641.78 45775.49 1569686.17 1570 Section Stage 17 Perimete Perimete Area Section r Base r Apex Base 1083 1084 1085 1086 1087 1088 1089 1090 352.328 563.771 661.151 723.998 709.644 643.905 433.57 154.418 228.442 433.362 580.965 644.75 649.382 514.798 - 7910.43 17802.70 24323.60 28076.60 29796.50 28287.30 13930.10 1748.38 Section Area Apex Surface Area Tissue Height 2636.81 8622.31 14146.5 14385.7 17694.5 15427.9 - 5273.62 9180.39 10177.10 13690.90 12102.00 12859.40 13930.10 1748.38 41.58192 48.08064 38.38262 48.00982 40.40596 40.40596 Basal Area Apical Area (μm2) (μm2) Tissue Volume (μm3) Cell Number (Vt/C#) Tissue Thicknes s Section Volume (A*t) Pb*Tt Surface Base Surface Apex Cell Volume 19.19131 24.00491 19.19131 24.00491 20.20298 24.20329 10 10 101207.68 220374.44 195311.88 328648.82 244496.46 311239.79 139301.00 17483.80 6761.64 13533.27 12688.35 17379.51 14336.92 15584.62 4335.70 1544.18 4384.101 10402.82 11149.48 15477.17 13119.45 12459.81 - 1000 1000 1000 1000 1000 1000 1000 1000 Cell Tissue Number Volume (Vt/C#) ***** 321582.1 516894 845542.8 1090039 1401279 1540580 1558064 ***** 321.5821 516.894 845.5428 1090.039 1401.279 1540.58 1558.064 G t to Vt 3.3223log10 ( ) Vo • We began by obtaining digital photographs of sections of the specimen from the Carnegie Collection. • In stage 13 of the developing human embryonic lens there were 18 sections dedicated to our targeted region, each of which is 15 microns thick. • A 3D modeling program called WinSURF will be used to model our lenses. • In order to obtain accurate data from the software we had to first create a relative scale with our images by using a magnification grid provided with the digital sections. • After the scaling was set. We then manually traced the regions of interest. • This procedure was repeated throughout each section. • The software then takes the tracing of the image, and stacks them in it’s respective order to create a 3D model. • We can then obtain ‘volumetrics’ from the software, which provides us with; perimeter length, surface area, and volume measurements. • Using the data we can then derive addition geometric properties of our model such as: Tissue Height, Basel Area, and Apex Area. Using the derived data with reference data (Cell Volume), we can find the Cell number for each lens. Bibliography Dolye, M., Ang, C., Raju, R. Willisams, B., DeFanti, T., Goshtasby, A., Grzesczuk, R., Noe, A. 1993 “Processing of Cross-Sectional Image Data for Reconstruction of Human Developmental Anatomy from Museum Specimens.” ACM SIG BIO Newsletter, 13:9-14. Bozanic, D., Saraga-Babie, M. 2004 “Cell proliferation during the early stages of human eye development.” ANAT. EMBRYOL 208:381-388. Cohen, J. 2002 “Embryo Development At the Click of a Mouse.” SCIENCE, 297: 1629 Zwann, J., Hendrix, R. 1973 “Changes in Cell and Organ Shape during Early Development of the Ocular Lens.” AMERICAN ZOOL. 13:1039-1049. POSTER TEMPLATE BY: www.PosterPresentations.com