Abstract
We examined CD133 distribution in a human hepatoblastoma cell line (HuH-6 clone 5). We directly observed the cultured cells on a pressure-resistant thin film (silicon nitride thin film) in a buffer solution by using the newly developed atmospheric scanning electron microscope (ASEM), which features an open sample dish with a silicon nitride thin film window at its base, through which the scanning electron microscope beam scans samples in solution, from below. The ASEM enabled observation of the ventral cell surface, which could not be observed using standard SEM. However, observation of the dorsal cell surface was difficult with the ASEM. Therefore, we developed a new method to observe the dorsal side of cells by using Aclar® plastic film. In this method, cells are cultured on Aclar plastic film and the dorsal side of cells is in contact with the thin silicon nitride film of the ASEM dish. A preliminary study using the ASEM showed that CD133 was mainly localized in membrane ruffles in the peripheral regions of the cell. Standard transmission electron microscopy and scanning electron microscopy revealed that CD133 was preferentially concentrated in a complex structure comprising filopodia and the leading edge of lamellipodia. We also observed co-localization of CD133 with F-actin. An antibody against CD133 decreased cell migration. Methyl-β-cyclodextrin treatment decreased cell adhesion as well as lamellipodium and filopodium formation. A decrease in the cholesterol level may perturb CD133 membrane localization. The results suggest that CD133 membrane localization plays a role in tumor cell adhesion and migration.
Highlights
The hypothesis of cancer stem cells (CSCs) was proposed to explain the origin of cancer cells
The cells were cultured on a 100-nm thick silicon nitride film and AclarVR plastic film (Nisshin EM, Tokyo, Japan) in Dulbecco’s modified Eagle’s medium (DMEM) supplemented with 10% fetal bovine serum (FBS) and 50 mg/mL gentamicin, according to a method previously reported by Hayashi et al (2011)
Strong positive results were seen for CD133 at the cell periphery (Fig. 1), where it co-localized with F-actin (Figs. 2a–2c)
Summary
The hypothesis of cancer stem cells (CSCs) was proposed to explain the origin of cancer cells. CSCs are a small fraction of tumor cells with the capacity of both self-renewal and unlimited slow proliferation. They are often resistant to chemotherapy and radiation and are responsible for continuously supplying new cancer cells (Zhao et al, 2008). Quintana et al (2008) and Shackleton et al (2009) showed that tumors that arose from both CD1332 and CD1331 cells sorted from an original melanoma re-established the original ratios of CD1332 and CD1331 cells This experiment indicates that individual cancer cells can recapitulate the marker heterogeneity of the tumors from which they are derived
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
