The Effects of Vitamin C on MG-63 Cancer Cell Survivorship

Slides:



Advertisements
Similar presentations
Smokeless Tobacco’s Influence on Microbial Life
Advertisements

By Sonali Dadoo School Pine-Richland High School Grade 11 Category
JACK LEECH GRADE 12 PITTSBURGH CENTRAL CATHOLIC HIGH SCHOOL PJAS 2011 Estradiol Effects on a Mammalian Stem Cell Line.
Creatine Effects on Oxidative- Stressed Stem Cells Anthony DeRenzo Pittsburgh Central Catholic HS Grade 11 February 6, 2010.
The Effects of Ibuprofen On Stem Cell Development Kevin Pfau Central Catholic High School Grade 11.
Magnetic Stress Remediation on Mammalian Cell Lines J.R. Stragar Grade 11 Pittsburgh Central Catholic High School.
Brandon Perrotte Grade 11 Pittsburgh Central Catholic High School Effects of UV Radiation on C2C12 Cells.
DAVID MCFALL GRADE 11 CENTRAL CATHOLIC HIGH SCHOOL PJAS 2012 Sweetener Effects on Cancer Cell Proliferation.
The Effects of UVC Light on C2C12 Stem Cells Cory Soltys Pittsburgh Central Catholic High School Grade 12 PJAS 2015.
Cranberry Juice Antimicrobial Properties Zane Stiles Central Catholic High School Grade 9.
EFFECTS OF FRACKING FLUID ON STAPH. EPIDERMIDIS AND E. COLI LUKE WEARDEN GRADE 11 CENTRAL CATHOLIC HIGH SCHOOL.
ALOE EFFECTS ON 3T3 CELL PROLIFERATION AND SURVIVORSHIP Raashmi Krishnasamy Peters Township High School Grade 12 6th Year in PJAS.
The Effects of Chlorinated Water on Microbial Life Jeff Van Kooten 9 th Grade Central Catholic High School.
Vitamin D Effects on Microbial Flora
Effect of Microwave Radiation on C2C12 Stem Cells
James Constantin Central Catholic High School Grade 11.
The Effect of Electromagnetic Radiation on Yeast Jamison Beiriger Grade 11 Central Catholic High School.
The Impact of Red Food Coloring on C2C12 Stem Cells Christian Ford Grade 12 Central Catholic High School PJAS 2015.
The Effect of Potassium Nitrate on Microbes By Liam O'Malley 9th Grade Central Catholic High School.
Magnetic Stress Remediation on Mammalian Cell Lines
1 Neil Carleton Grade 11 Pittsburgh Central Catholic High School PJAS 2012.
The Effects of Isotretinoin on C2C12 Stem Cells Colm Parrish Pittsburgh Central Catholic HS Grade 11.
Grade 11 Central Catholic High School Effects of Whey Protein on Microbial Survivorship William McCarthy.
L-Arginine Remediation of Stressed C2C12 Cells Warren Austin Pittsburgh Central Catholic High School Grade 12 3 rd Year in PJAS.
Vitamin C Attenuation of Smoke Stress Will Maher Central Catholic High School.
Frank Wolf Pittsburgh Central Catholic High School Grade 11 PJAS 2010.
Steroid Influence on Myoblast Stem Cell Line Nathan Lampenfeld CCHS Grade 11 PJAS 2015.
The Effects of Vitamins B6 and C on Yeast Growth By: Gianmarco Frezza Pittsburgh Central Catholic HS Grade 9.
Clarke Bacharach 12 th Grade Central Catholic High School Light Wavelength Influence on Stem Cell Behavior.
Antioxidant Attenuation of Yeast Mutagenesis Peter Chekan Central Catholic High School Pittsburgh.
David McFall Grade 9 Central Catholic High School.
Frank Wolf Pittsburgh Central Catholic High School Grade 11 PJAS 2010.
By : Phillip DeRenzo 9th Grade Central Catholic High School.
The Effects of Stress on Post-Natal Stem Cell Proliferation and Differentiation Robert McKinstry Pittsburgh Central Catholic High School.
JACK LEECH PITTSBURGH CENTRAL CATHOLIC HIGH SCHOOL GRADE Pesticide Effects on Yeast Survivorship.
By: Luke Beck Pittsburgh Central Catholic HS PJAS 2012 Grade 11 Synergistic Drug Effects on Microbial Flora.
Effects of Citrus Aurantium on Stem Cell Differentiation and Proliferation By Kelly Hyland Oakland Catholic High School Grade 11 PJAS 2011.
EFFECTS OF METHYLHEXANAMINE (DMAA) ON C2C12 AND 3T3 STEM CELLS Cameron Franz Pittsburgh Central Catholic High School Grade 11.
Fracking Chemical Effect on Yeast Mutagenesis Cam Coco Pittsburgh Central Catholic 11 th Grade.
XENOESTROGEN EFFECTS ON STEM CELL BEHAVIOR Maria DeRenzo Oakland Catholic High School Grade 10.
By : Phillip DeRenzo 9th Grade Central Catholic High School.
The Effects of Aspartame on Mammalian Stem Cells
UV Light Effects on Vitamin D Stressed Staph Cells
Creatine Effects on Oxidative-Stressed Stem Cells
Grape Seed Extract Effects on C2C12 Cell Behavior
Xenoestrogen Effects on Stem Cell Behavior
Microbial Survivorship in River Water
Hydrogen Peroxide Anti-Microbial Effects
L-Theanine Effects on C2C12 Stem Cells
Yohimbine Influence on a Mammalian Stem Cell Line
Electromagnetic Field Effects on MG-63 Cell Lines
Adrenaline Effects on Adult Stem Cells
UV Light Effects on Vitamin D Stressed Staph Cells
The Effects of Capsaicin on Microbes
Magnetic Stress Remediation on Mammalian Cell Lines
Theraflu Effects on Microbial Flora
Effects of BMP-2 and Estradiol on 3T3 Fibroblast Cell Line
Effects Of Fertilizer on Yeast Cell and E. Coli Survivorship
Effects of Microwave Radiation on Bacteria
The EFFECTS OF OILS ON MG63 CANCER CELLS
Acid Rain Effects on Microbial Survivorship
Effects of Bisphenol A on Cancer Cell Proliferation
Ascorbic Acid Remediation of UV Stressed E. Coli
The Effects of antibacterial hand soap on bacteria survivorship
Ascorbic Acid Remediation of UV Stressed E. Coli
Joe Stern Pittsburgh Central Catholic Grade 9
The Effect of FGF on Mammalian Stem Cells
Narcan Effects on Cancer Cell Behavior
DECELLULARIZE D SCAFFOLDS
The Effects of Atrazine on MG-63 Cancer Cells
Presentation transcript:

The Effects of Vitamin C on MG-63 Cancer Cell Survivorship Joe Ziccarelli Grade 12 Central Catholic High School PJAS 2015

An Overview of Cancer Cells Cancer cells are cells that grow and divide at an irregular, unregulated pace. Apoptosis does not occur in cancerous cells; their mutations are passed on to the second generation, eventually clustering and forming tumors. Tumors can be malignant (aggressive) or benign.

MG63 Cancer Cell Line Human cancer cell line Osteosarcoma cells, an aggressive form of bone cancer Useful model to test the effects of variables on cancer cell proliferation

Antioxidants Molecules capable of slowing or preventing the oxidation of other molecules May be able to prevent cancer and coronary heart disease Body produces antioxidants Can obtain through Diet Past studies have shown that the antioxidant nature of Vitamin C has anti-tumoregenic effects

Variable: Ascorbic Acid (Vitamin C) Antioxidant Enzyme cofactor In Oranges, Strawberries, and Grapefruit Recommended daily intake: 60 mg The disease scurvy occurs from lack of Vitamin C

Purpose To determine the effects of ascorbic acid on MG-63 cell survivorship.

Hypotheses Null Hypothesis: Ascorbic will not have a significant effect on MG63 cell survivorship. Alternative Hypothesis: Ascorbic Acid will significantly alter the survivorship of MG63 cells.

Materials Cryotank Four 75mm2 tissue culture treated flasks, Sixteen 25 mm2 tissue culture treated flasks MG63 Osteosarcoma Cancer Cell Line Trypsin-EDTA DMEM Media 1 Molar Ascorbic Acid Solution Incubator Evo Imaging System Macropipette + sterile macropipette tips (1 mL, 5 mL, 10, mL, 20 mL) Micropipettes + sterile tips Labeling Tape Hemacytometer Sterile PBS Ethanol (70% and 100%) Laminar Flow Hood UV and Sterilizing Lamp Purple Nitrile gloves Safety goggles

Procedure 1: Cell Culturing A 1 mL aliquot of MG63 cells from a Cryotank was used to inoculate 30 mL of 10% serum DMEM media in one 75mm2 culture flask yielding a cell density of approximately 106 to 2 x 106 cells. The media was replaced with 15 mL of fresh media to remove cryo-freezing fluid and incubated (37° C, 5% CO2) for 2 days until a cell density of approximately 4 x 106 to 5 x 106 cells/mL was reached. The culture was passed into two sets of 3 flasks in preparation for experiment and incubated for 2 days at 37° C, 5% CO2.

Procedure 1: Proliferation Experiment- Day 0 (Addition of Variable) After trypsinization, cells from all of the flasks were pooled into 1 common 75mm2 flask (cell density of approximately 1 million cells/mL). 0.1 mL of the cell suspension was added to eight 25 mm2 tissue culture treated flasks containing 5 mL of DMEM (com) media, creating a cell density of approximately 105 cells per flask. The stock solution of ascorbic acid (one molar) was created using 50 mL of ethanol and 10 g of ascorbic acid. This was then used in a serial dilution to create a 10-2, 10-4, 10-6 M concentration. The cells were incubated at 37°C, 5% CO2 for the remainder of the study.

Procedure 2: Cell Culturing A 1 mL aliquot of MG63 cells from a Cryotank was used to inoculate 30 mL of 10% serum DMEM media in one 75mm2 culture flask yielding a cell density of approximately 106 to 2 x 106 cells. The media was replaced with 15 mL of fresh media to remove cryo-freezing fluid and incubated (37° C, 5% CO2) for 2 days until a cell density of approximately 4 x 106 to 5 x 106 cells/flask was reached. The culture was passed into two sets of 3 flasks in preparation for experiment and incubated for 2 days at 37° C, 5% CO2.

Procedure 2: Proliferation Experiment- Day 0 (Addition of Variable) After trypsinization, cells from all of the flasks were pooled into 1 common 75mm2 flask (cell density of approximately 1 million cells/mL). 0.1 mL of the cell suspension was added to eight 25 mm2 tissue culture treated flasks containing 5 mL of DMEM (com) media, creating a cell density of approximately 105 cells per flask. The stock solution of ascorbic acid (one molar) was created using 50 mL of ethanol and 10 g of ascorbic acid. This was then used in a serial dilution to create a 10-2, 10-4, 10-6 M concentration. The cells were incubated at 37°C, 5% CO2 for the remainder of the study.

Procedure: Proliferation Experiment- Days 1 and 3 Using one flask from each group, cell densities were determined as follows: The cells were trypsinized and collected into cell suspension. 25 µl aliquots were transferred to a Hemocytometer for quantification (eight counts per flask). Day 1 and Day 3 The previous procedure for determining densities was used again, and a Evo Imaging System was used to take images of representative areas of each flask.

Results of Proliferation Analysis (MG63) P-value 4.15E-15 P-value 7.25E-22

Dunnett’s Test (Vitamin C Exposure After Two Days) Concentration T-Value T-Critical (0.05) Variation MG63 - 10^-6 M 2.057 2.88 Insignificant 10^-4 M 18.322 Significant 10^-2 M 26.357

Results of Proliferation Analysis (MG63) P-value 2.09E-13 P-value 2.15E-13

Dunnett’s Test (Variable Added Without Two Day Proliferation) Concentration T-Value T-Critical (0.05) Variation MG63 - 10^-6 M 0.095 2.88 Insignificant 10^-4 M 19.205 Significant 10^-2 M 26.551

Conclusions MG63 (variable added after two day proliferation) Based upon the results gathered from the ANOVA and Dunnett’s statistical analyses, it appears that the addition of ascorbic acid at higher concentrations significantly affects cancer cell proliferation. Therefore, the null hypothesis can be rejected. MG63 (variable added without two day proliferation)

Future Changes Limitations Extensions Hemacytometer counts in proliferation experiment were subject to clumping error Limited range of concentrations Limited days of exposure Only accessed proliferation/survivorship Include further trials in order to maximize data accuracy Use a wider range of concentrations Test synergistic effects of other vitamins

Acknowledgements Mark Krotec, PTEI Merisant Company

Works Cited http://cancerres.aacrjournals.org/content/ 40/3/734.full.pdf http://www.cancer.gov/cancertopics/pdq/c am/highdosevitaminc/patient/page2 http://www.mayoclinic.org/diseases- conditions/cancer/expert- answers/alternative-cancer-treatment/faq- 20057968

Works Cited (cont.) http://lpi.oregonstate.edu/s- s00/vitaminc.html http://www.webmd.com/cancer/news/201 40205/intravenous-vitamin-c-may-boost- chemos-cancer-fighting-power

Statistical Analyses of the Proliferation Results ANOVA Compares variation within groups to variation between groups. Using the ANOVA, a p-value less than the alpha of .05 was gathered (significant variation). Reject the null hypothesis. Dunnett’s test Compares each experimental group to control individually. 0.05 alpha was used, and the t-value compared to the t- critical value of 2.88

Additional Information DMEM Serum - 1% and Complete Media (4 mM L-glutamine, 4500 mg/L glucose, 1 mM sodium pyruvate, and 1500 mg/L sodium bicarbonate + [ 10% fetal bovine serum for complete])