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Understanding Dynamic Behavior of Embryonic Stem Cells. Shubham Debnath University of Minnesota-Twin Cities [email protected]. Advisor: Dr. Bir Bhanu BRITE REU 2009 University of California-Riverside. Overview. Introduction to embryonic stem cells and importance - PowerPoint PPT Presentation
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Understanding Dynamic Behavior of Embryonic Stem Cells
Shubham DebnathUniversity of Minnesota-Twin Cities
Advisor: Dr. Bir BhanuBRITE REU 2009
University of California-Riverside
Overview
• Introduction to embryonic stem cells and importance• Description of video processing and image segmentation
methods used for study• Stem cell videos used for data and examples of use of
segmentation and image analysis methods• Results and Analysis• Conclusions• Future pursuits
Stem Cells and Importance
• Derived from the inner cell mass of early stage embryos, known as blastocysts
• Known to be pluripotent and can differentiate into a variety of cell types
• Very important towards study in the future of medicine and healthcare
Stem Cells and Importance• Attach to substrate to
differentiate based on the environment they are placed in
• For mitosis to proceed, cells must unattach themselves, divide, then reattach.
• Behavior of embryonic stem cells is not fully understood
• Past and continued research at Stem Cell Center at UCR• Effects of smoke and alcohol
on stem cell behavior
Mitosis• Process by which eukaryotic
cells divide into two identical daughter cells
• Consists of various phases in which the nucleus and cytoplasm divide ending with cytokinesis and cleavage into two cells
• Importance for maintenance of genome set
• Rate of mitosis depends on tissue renewal for stem cells; varies for different cell types
Introduction to Methods• Otsu’s Algorithm for Binary
Thresholding:
• Inputs a grey-scale image, automatically finds a threshold value, splits the image accordingly
• Threshold value is found by histogram analysis
• Outputs a binary image showing regions of interest
• Connected Components Analysis:
• Only done on binary images• For each pixel, checks
neighboring pixels and labels each region accordingly
• Labeled with random pseudo colors for visual identification of each connected component
Otsu’s Algorithm
Threshold Value: 129
Otsu’s Algorithm
Original Image Segmented Image
Connected Components
1 01 1 1
1 1
1
1 11
1
1 1
0
0 0 0 0
0 0 0 0
1
0
00
0
0
0
0
00 0
0
0
1 0
0
0
0 1
01
0 0
1 1
0
0 1
0 0
11
1
0
0
0 0 0001
1 01 1 2
1 2
2
3 33
3
4 5
0
0 0 0 0
0 0 0 0
5
0
00
0
0
0
0
00 0
0
0
2 0
0
0
0 2
06
0 0
6 6
0
0 2
0 0
55
5
0
0
0 0 0
00
7
Binary Image Result of Connected Components Algorithm
Examples
Video of Stem Cells
Hypotheses• Number of mitosis occurrences: approximately 6-10• Cell count should be similar and beginning and end of video
– Cells going through mitosis, attaching, apoptotic events– If recorded over a longer period of time, cell count remains
essentially the same• Large jumps, spikes in data represent colonies of cells unattaching
together, multiplying• Sharp drop shows colony reattaching to substrate• Low number of unattached cells should correspond to low number of
pixels in “white” membranes– More pixels in darker mass of cell showing how surface area
increases with attachment to substrate
Plate 9 – 20X
Results
Results
Plate 6 – 20X
Results
Results
Analysis
• Graphs complement each other – directly related
• Spikes in graphs accurately show points of mitosis
• Shows how surface area of cell changes with attachment and cell division
• Problems with counting come with colonies of cells
• Background noise, light
Conclusions
• Video processing can be used for the segmentation of stem cell videos for their characterization.
• Mitosis is important cell differentiation in stem cells and for regulation of processes in the human body
• Mitosis count between 6 and 10 divisions based on resulting graphs– Watching videos agrees with these estimations
• Time: 4 to 6 frames for process of mitosis to start and complete– 8 to 12 minutes
• Colonies tend to unattach and multiply together
Future Research
• Use of the relaxation gradient algorithm• Choosing of different thresholds• Use of a new Nikon Biostation can be used to record videos for
longer times and with higher magnification and resolution.• More biomedical engineering based objective: behavior can be
simulated computationally with macromolecular interactions• The free energy of cells in various states can be calculated to
find a minimum at which the cell responds to diverse changes.
Acknowledgements• Special thanks to my research
advisor, Professor Bir Bhanu of UC-Riverside
• Students at Center for Research in Intelligent System (CRIS)
• Thanks to Dr. Prue Talbot and students at the Stem Cell Center at UCR for providing the data
• Thanks to the BRITE REU program funded by the National Science Foundation (NSF)
Questions?