BMEBT Master’s Thesis Proposal by Shrimathi Venugopalakrishnan
Title: Evaluation of extracellular vesicles released by 3D spheroids and 2D conventional cell culture of the SKOV3 ovarian cancer cell line
Abstract:
Cancer cells communicate with each other and with the surrounding tumour microenvironment by secreting extracellular vesicles (EVs), or exosomes. Exosomes play a critical role in cell-to-cell signalling and cancer progression by transferring bioactive cargo—such as proteins, lipids, mRNAs, miRNAs, and DNA fragments—from cancer cells to healthy cells, enabling distant organ colonization and metastasis. SKOV3 is a widely used human ovarian adenocarcinoma cell line traditionally cultured under anchorage-dependent (2D monolayer) conditions. However, suspension culture more closely mimics the three-dimensional, non-adherent environment cancer cells experience in vivo, leading to altered cell signalling, spheroid formation, and potentially distinct exosome release profiles. In preliminary experiments, SKOV3 cells successfully adapted to suspension culture in ultra-low attachment plates and formed compact spheroids, confirming their ability to grow under non-adherent conditions. This research project compares exosomes secreted by SKOV3 cells cultured in (1) conventional anchorage-dependent conditions (T-flasks) and (2) suspension conditions using a stirred-tank bioreactor with impeller agitation, to maintain cells in suspension while ensuring adequate nutrient and oxygen transfer. Exosomes will be isolated from conditioned culture supernatant using a sequential process: low-speed centrifugation to remove cells and debris, ultrafiltration, and final purification via size-exclusion chromatography (SEC) using Sepharose columns. This protocol yields high-purity exosomes free of contaminating proteins and other EV subtypes. Isolated exosomes will be characterized using techniques such as Dynamic Light Scattering (DLS), Transmission Electron Microscopy (TEM), Flow cytometry for surface marker expression and Raman spectroscopy for the molecular fingerprints (lipid, protein, and nucleic acid vibrational signatures) to generate condition-specific spectral libraries.
Advisor: Dr. Milana Vasudev, Department of Bioengineering
Committee: Dr. Tracie Ferreira, and Dr. Lamya Karim, Department of Bioengineering.
All BMEBT students are encouraged to attend, and all interested parties are invited.
LIB 426
Milana C. Vasudev
508-999-9284
mvasudev@umassd.edu