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Mismatch Repair Responses to Carcinogenic Polycyclic Aromatic Hydrocarbons

Presented by: Sarah Ferrer Under the m entorship of Dr. Andrew Buermeyer of the OSU Environmental and Molecular Toxicology Department. Mismatch Repair Responses to Carcinogenic Polycyclic Aromatic Hydrocarbons. Relevance of Research.

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Mismatch Repair Responses to Carcinogenic Polycyclic Aromatic Hydrocarbons

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  1. Presented by: Sarah Ferrer Under the mentorship of Dr. Andrew Buermeyer of the OSU Environmental and Molecular Toxicology Department Mismatch Repair Responses to Carcinogenic Polycyclic Aromatic Hydrocarbons

  2. Relevance of Research • MMR deficiency linked to colorectal (and other) cancer predisposition. • Lynch Syndrome causes premature cancer occurrence and greater reoccurrence.

  3. Background: Mismatch Repair • MMR protects against DNA mutations.

  4. Background: Carcinogenic Polycyclic Aromatic Hydrocarbons • PAHs are carcinogens found in the environment. • Metabolized by the liver and colon into diolepoxides. • PAHs used in my research project: • Benzo-[a]-pyrene • Benzo-[c]-phenanthrene • MMR proteins can bind to and recognize PAH adducted DNA

  5. Hypothesis MMR proficient cells are more effective at maintaining DNA integrity in human lymphoblastic cell lines than MMR deficient cells when exposed to PAHs benzo-[a]-pyrene and benzo-[c]-phenanthrene.

  6. Method: Cell Culture • Human lymphoblastoid cell lines TK6 and MT1 • Maintained under the following conditions: • 10% complete RPMI media • Incubated with 5% carbon dioxide and at 38°C • Cell density between 5 x 104 cells/mL and 1 x 106 cells/mL

  7. Method: Growth Curve Characterization • Growth curves created over several dilutions and compared to the characterized doubling times.

  8. Method: HPRT Gene Spontaneous Mutant Frequency Characterization • Hypoxanthine guanine phosphoribosyltransferase (HPRT) reporter gene. Culture with 12 million cells RPMI w/ HAT media Normal RPMI media 6-TG exposed plate Plating efficiency plate

  9. Method: PAH Exposed HPRT Mutant Frequency • Re-run previous experiment to determine MF. • Pick 6-TG resistant colonies and analyze for types of mutations. Cell with HPRT- DNA, but HPRT+ proteins Normal HPRT+ cell Cell with HPRT- DNA and proteins

  10. Prediction When exposed to benzo-[a]-pyrene and benzo-[c]-phenanthrene, MMR deficient cells lines exhibit a higher mutant frequency in the HPRT gene than MMR proficient cells.

  11. Acknowledgements • Dr. Andrew Buermeyer • VidyaSchalk • Kevin Ahern • Howard Hughes Medical Institute

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