The Study of Establishment of a Label Free Cancer Stem Cell Selection System

Cancer is believed to arise from a series of sequential mutations. Stem cell biology could provide new insights into cancer biology, thus leading us to the notion that tumors might contain cancer stem cells (CSCs) with the self renewal ability that drive the formation and growth of tumors. Hepatocullular carcinoma (HCC) is typically comprised of morphologically diverse cells that express a variety of hepatic lineage markers and have been reported that subpopulations with CSCs function exist in some HCC cell lines. In the proposed research, we will construct a series of biomaterials to fabricate a simple, rapid and label-free isolated system for extensive selection, maintenance and enrichment of HCC CSCs and program the effect of cancer stem cell microenviroments by using three kinds of cancer cell line. During the first year, we will construct a series of layer by layer polyelectrolyte multilayer (PEM) films to study the surface properties variation. Besides, a series of methylcellulose hydrogel with different concentration will also be prepared. We will combine sequential measurements by Quartz Crystal Microbalance with Dissipation (QCM-D), atomic force microscopy (AFM) and ellipsometry, to characterize the surface properties variation. PEM could provide a series variation of surface properties such as charge, thickness, roughness, hydration for investigation of the CSCs behaviors under different microenviroments. Different concentration of hydrogel also could provide a series of surface properties variation for sphere formation. Finally, sphere and colony morphology, sphere or colony number and size distribution of CSCs on a series substrate will be counted and analyzed. During the second year, three kinds of cancer cells will be used. The cell viability, proliferation rate, and cytotoxicity of colonies selected from different substrates and normal population cancer cells will be analysis and compared. The constitutive expression of cancer stem cell markers including the CD133, CD90, CD44, EpCAM etc, will be determined by flow cytometry and immunocytochemistry and reverse transcription-polymerase chain reaction (RT-PCR) analysis. In addition, the colonies or sphere isolated from different substrates will be measured with drug sensitivity, such as the doxorubicin, and in comparison with normal population. During the third year, in order to make sure the colony selected from PEM will express the CSC properties in vivo, the in vivo tumorigenic assays will also be proceeded to investigate the ability of tumor formation. Besides, we also will use the western blot to determine the signal pathway of the self renewal ability and drug resistance of CSCs. The successful demonstration of the proposed goals will lead to a new category of cancer biology and cancer therapy.

Project ID:PB10207-0370
External Project ID:NSC102-2221-E182-007