Presented By: Biomedical Engineering
BME 500 Seminar Series: Alon Greenbaum, Ph.D.
Alon Greenbaum, Ph.D.
BME Faculty Candidate and Guest Speaker
California Institute of Technology
Abstract:
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. Towards this end, 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, for instance, our recent demonstration of single molecule RNA transcript detection in thick mouse brain tissue (2).
Although tissue clearing methods work well on soft tissue, vital organs with complex composition (e.g. bones and cartilage) are challenging to clear. Given that bone tissue harbors unique and essential physiological processes, such as hematopoiesis, bone growth, and 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+ osteoprogenitor cells in the tibia, femur, and vertebral column of adult transgenic mice. To obtain a complete distribution map of these osteoprogenitor cells, we developed a computational pipeline that semiautomatically detects individual Sox9-tdTomato+ cells in their native three-dimensional environment. Our computational method counted all labeled osteoprogenitor cells without relying on sampling techniques and displayed increased precision when compared with traditional stereology techniques for estimating the total number of these rare cells. 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.
From the brain to bone, I believe that with continued development, methods that measure cellular and molecular phenotyping at the whole organ level will be the de facto approach to studying biological questions and diseases.
BME Faculty Candidate and Guest Speaker
California Institute of Technology
Abstract:
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. Towards this end, 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, for instance, our recent demonstration of single molecule RNA transcript detection in thick mouse brain tissue (2).
Although tissue clearing methods work well on soft tissue, vital organs with complex composition (e.g. bones and cartilage) are challenging to clear. Given that bone tissue harbors unique and essential physiological processes, such as hematopoiesis, bone growth, and 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+ osteoprogenitor cells in the tibia, femur, and vertebral column of adult transgenic mice. To obtain a complete distribution map of these osteoprogenitor cells, we developed a computational pipeline that semiautomatically detects individual Sox9-tdTomato+ cells in their native three-dimensional environment. Our computational method counted all labeled osteoprogenitor cells without relying on sampling techniques and displayed increased precision when compared with traditional stereology techniques for estimating the total number of these rare cells. 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.
From the brain to bone, I believe that with continued development, methods that measure cellular and molecular phenotyping at the whole organ level will be the de facto approach to studying biological questions and diseases.
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