Date of Award

2009

Degree Name

Biological Sciences

College

College of Science

Type of Degree

M.S.

Document Type

Thesis

First Advisor

Jagan V. Valluri, Ph.D

Second Advisor

Pier Paolo Claudio, M.D., Ph.D.

Third Advisor

Vincent E. Sollars, Ph.D.

Abstract

The purpose of this study was to investigate the effects of simulated microgravity on cancer cell growth. Simulated microgravity is a condition in which cells are in constant free-fall and in which they are able to grow suspended without the shear and stress. Microgravity allows for important insights into the cellular mechanism and may be key in understanding why some cells thrive in this condition and others go into apoptosis. Cancer stem cells (CSCs) are considered a subset of the bulk tumor responsible for initiating and maintaining cancer. SAOS-2 is a human bone cancer cell line which consists of a heterogeneous population [10% of CD133(+) cells and 90% CD133(-)]. CD133 is a transmembrane pentameric glycoprotein and a surface marker for progenitor cells including neural and embryonic stem cells, and it is expressed in cancers, including some leukemias and brain tumors. We isolated CD133(+) cells from the SAOS-2 cell line by using a MACSorting system which magnetically sorts CD133(+) cells with the aid of an antibody against CD133 conjugated to a magnetic bead. SAOS-2 positivity to CD133 was assessed by flow cytometry. One million SAOS-2 CD133(+) MACSorted cells were cultured, in a Hydrodynamic Focusing Bioreactor (HFB) which was developed by NASA at the Johnson Space Center for modeling microgravity conditions. This experiment showed an increase of cell growth compared to the normal gravity control. We observed a 15-fold proliferation of the CD133(+) cellular fraction with cells that were cultured for 7-days at optimized conditions. Additionally, 100% of the harvested cells were found to be CD133(+), indicating that the HFB had selected for the SAOS-2 undifferentiated cellular fraction. CSCs are thought to be responsible for the formation and growth of neoplastic tissue and are naturally resistant to chemotherapy also in part due to their quiescent nature. We also tested the hypothesis that the microgravity environment could sensitize the CD133(+) resistant osteosarcoma cells to various chemotherapy agents at clinically relevant doses. SAOS-2 cells were cultured in three chemotherapy treatments: cisplatin, doxorubicin, and methotrexate. Toxicity and viability assays were performed on CD133(+) cells, CD133(-) cells, total SAOS-2 population, and on cells that were cultured in the HFB for 5-days with these different chemotherapies. Interestingly, we found, by FACS analysis and MTT assay, that the microgravity environment greatly sensitized the CSCs, which are normally resistant to chemotherapy treatments, to become susceptible to various chemotherapeutic agents. This experiment paves the way to less toxic and more effective chemotherapeutic treatments in patients.

Subject(s)

Cancer Cells -- Growth.

Cancer Cells -- Research.

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