1. Project Description
This project investigates the effects of different doses of gamma radiation on
cellular senescence of lymphocytes derived from human blood samples.
Lymphocytes are a type of white blood cell which recognize and respond to
particular pathogens. Their long-living nature makes them useful subjects in
investigating the effects of ionizing radiation on human cells.
By studying the percentage of lymphocytes which senesce following different
doses of radiation, the cellular consequences of radiation can be better
understood.
When exposed to gamma (γ) rays (high-energy electromagnetic
waves that have the ability to cause ionization and break
chemical bonds), the affected lymphocytes may become
unstable. This instability can create mutations in the DNA that
lead to cancerous cells. However, cellular senescence can
suppress the potential of these cancer cells to grow into lethal
tumors by stopping cell division.
Experimental Methods
1. Irradiate blood samples
2. Dilute blood using PBS
3. Carefully layer diluted blood samples on
Ficoll
4. Centrifuge to separate blood
into different layers
5. Wash lymphocytes
and transfer into RPMI media
6. Incubate for 48 hours
7. Centrifuge tube and discard supernatant
8. Vortex and add ethanol to cell pellet
9. Incubate at -20°C for at least 2 hours
10. Add p16 antibody into cell suspension
11. Incubate for 20-30 minutes in the dark
12. Proceed to flow cytometric analysis
Stephen Liu, Mihai Dumbrava, Adrienne Wan, Laura Paterson
Future Work
The results of this project can be applied in
the medical field of cancer research.
Radiation therapy is currently a common
procedure used to eliminate malignant
tumors, but unfortunately this method is not
always effective. This research will look to
enhance cancer treatment by investigating
the level of radiation which maximizes the
occurrence of cellular senescence and thus
preventing cancerous cells from replicating.
The main purpose of this project is to observe
how radiation can affect cellular processes and
lead to senescence by analyzing the p16
protein using fluorescent probes.
The gathered data will also contribute to a
larger project that examines how radiation
exposure leads to various other forms of cell
death, including apoptosis (programmed cell
death) and necrosis (premature death of cells
resulting in inflammation).
Purpose and Objectives
Cellular senescence has already been
recognized for its various purposes in the
human body. Senescence can serve as a
tumor suppressor. As demonstrated by
experiments on model organisms, senescent
cells have a strong influence on the
proliferation of neighbouring cells, both
healthy and cancerous. Furthermore,
senescence can also help in repairing
damaged tissues.
Existing Work
• Blood samples
• Gamma cell
• 15 ml test-tubes
• Centrifuge
• Phosphate
Buffered Saline
(PBS)
• P16 antibody
• Vortexer
• Ethanol
• Pipettes
• RPMI 1640
• Flow cytometer
• Ficoll
Materials and Equipment
Cell death is a fundamental cellular response
that has a crucial role in shaping our bodies by
eliminating unwanted cells.
Cellular senescence refers to a natural process
in which cell growth and development are
irreversibly halted. Exposure to certain stimuli,
such as radiation, can lead to senescence in
cells. Although they no longer replicate,
senescent cells remain metabolically active.
Background Information
Bibliography
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European Molecular Biology Organization. "Workshop." Programmed Cell Death in Model Organisms. European Molecular Biology Organization, Feb. 2012. Web. 10 July 2014.
Jefferson Lab. "Radiation Biological Effects." Radiation Biological Effects. Jefferson Lab, n.d. Web. 10 July 2014.
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Radiation Effects Research Foundation. "How Radiation Affects Cells." Radiation Effects Research Foundation. Radiation Effects Research Foundation, 2007. Web. 8 July 2014.
Ris, Katie. "Senescence comes of age." Nature.com. Nature Publishing Group, n.d. Web. 11 July 2014.
Rodier, F., and J. Campisi. "Four Faces of Cellular Senescence." The Journal of Cell Biology 192.4 (2011): 547-56. JCB. The Rockefeller University Press, 14 Feb. 2011. Web. 8 July 2014.
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Sõti, Csaba, Amere Sreedhar, and Péter Csermely. "Apoptosis, necrosis and cellular senescence: chaperone." linkgroup. Anatomical Society of Great Britain and Ireland, n.d. Web. 8 July 2014.
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