Towards Anatomical and Transcriptional Profiling of Intact Organisms
Monday, March 26, 2018
12:00 PM-1:30 PM
BIOMED Special Seminar
Towards Anatomical and Transcriptional Profiling of Intact Organisms with Tissue Clearing, Custom Microscopy and Algorithms
Alon Greenbaum, PhD
Life Sciences Research Foundation Postdoctoral Fellow
Division of Biology and Biological Engineering
California Institute of Technology (Caltech)
Organs are dynamic, complex and coordinated networks of cells. To understand organ function in normal and diseased states, their anatomy, including transcriptional profile and intracellular pathways, needs to be captured as a whole. Tissue clearing and labeling methods were devised to render entire organs transparent by removing lipids, allowing light penetration throughout (1). Cleared tissue facilitates the 3D structural mapping and connectivity of whole organs with unprecedented resolution and detail (2).
Although tissue clearing methods work well on soft tissue, vital organs with complex composition (e. g. bones) are challenging to clear. Given that bone tissue harbors essential physiological processes, such as bone remodeling, we decided to enable visualization of these processes at the cellular level in an intact environment. Therefore, we developed “Bone CLARITY,” a bone tissue clearing method (3). We used Bone CLARITY and a custom-built light-sheet fluorescence microscope to detect the endogenous fluorescence of Sox9-tdTomato+ osteoprogenitors cells in the tibia, femur, and vertebral column of adult mice. We demonstrate the value of the clearing-imaging pipeline by quantifying changes in the population of Sox9-tdTomato–labeled osteoprogenitor cells after sclerostin antibody treatment. Bone tissue clearing is able to provide fast and comprehensive visualization of biological processes in intact bone tissue.
(1) J. B. Treweek, K. Y. Chan, N. C. Flytzanis, B. Yang, B. E. Deverman, A. Greenbaum, A. Lignell, C. Xiao, L. Cai, M. S. Ladinsky, P. J. Bjorkman, C. C. Fowlkes, V. Gradinaru, Whole-body tissue stabilization and selective extractions via tissue-hydrogel hybrids for high-resolution intact circuit mapping and phenotyping, Nat. Protoc. 10, 1860–1896 (2015).
(2) S. Shah, E. Lubeck, M. Schwarzkopf, T.-F. He, A. Greenbaum, C. H. Sohn, A. Lignell, H. M. T. Choi, V. Gradinaru, N. A. Pierce, L. Cai, Single-molecule RNA detection at depth by hybridization chain reaction and tissue hydrogel embedding and clearing, Development 143, 2862–2867 (2016).
(3) A. Greenbaum, K. Y. Chan, T. Dobreva, D. Brown, D. H. Balani, R. Boyce, H. M. Kronenberg, H. J. McBride, V. Gradinaru, Bone CLARITY: Clearing, imaging, and computational analysis of osteoprogenitors within intact bone marrow, Sci. Transl. Med. 9, eaah6518 (2017).
Alon Greenbaum, PhD, holds BS and MS degrees in Electrical Engineering from Tel Aviv University, and a PhD in Electrical Engineering from University of California, Los Angeles. In his doctorate degree, he conducted biophotonics research under Prof. Aydogan Ozcan, focusing on high-resolution and high-throughput microscopy for biomedical applications. In Prof. Viviana Gradinaru's lab (California Institute of Technology), he is exploring innovative imaging techniques for various bone and neuroscience applications.
Dr. Greenbaum has been awarded the Good Ventures Postdoctoral Fellowship of the Life Sciences Research Foundation, as well as the Howard Hughes Medical Institute International Student Research Fellowship. He has published over 25 peer reviewed journal articles and his work has been presented in over 30 conferences.