The Importance of the Stem Cell Marker Prominin-1/CD133 in the Uptake of Transferrin and in Iron Metabolism in Human Colon Cancer Caco-2 Cells

Abstract

As the pentaspan stem cell marker CD133 was shown to bind cholesterol and to localize in plasma membrane protrusions, we investigated a possible function for CD133 in endocytosis. Using the CD133 siRNA knockdown strategy and non-differentiated human colon cancer Caco-2 cells that constitutively over-expressed CD133, we provide for the first time direct evidence for a role of CD133 in the intracellular accumulation of fluorescently labeled extracellular compounds. Assessed using AC133 monoclonal antibody, CD133 knockdown was shown to improve Alexa488-transferrin (Tf) uptake in Caco-2 cells but had no impact on FITC-dextran or FITC-cholera-toxin. Absence of effect of the CD133 knockdown on Tf recycling established a role for CD133 in inhibiting Tf endocytosis rather than in stimulating Tf exocytosis. Use of previously identified inhibitors of known endocytic pathways and the positive impact of CD133 knockdown on cellular uptake of clathrin-endocytosed synthetic lipid nanocapsules supported that CD133 impact on endocytosis was primarily ascribed to the clathrin pathway. Also, cholesterol extraction with methyl-β-cyclodextrine up regulated Tf uptake at greater intensity in the CD133high situation than in the CD133low situation, thus suggesting a role for cholesterol in the inhibitory effect of CD133 on endocytosis. Interestingly, cell treatment with the AC133 antibody down regulated Tf uptake, thus demonstrating that direct extracellular binding to CD133 could affect endocytosis. Moreover, flow cytometry and confocal microscopy established that down regulation of CD133 improved the accessibility to the TfR from the extracellular space, providing a mechanism by which CD133 inhibited Tf uptake. As Tf is involved in supplying iron to the cell, effects of iron supplementation and deprivation on CD133/AC133 expression were investigated. Both demonstrated a dose-dependent down regulation here discussed to the light of transcriptional and post-transciptional effects. Taken together, these data extend our knowledge of the function of CD133 and underline the interest of further exploring the CD133-Tf-iron network.

Photochemical internalization (PCI) of immunotoxins targeting CD133 is specific and highly potent at femtomolar levels in cells with cancer stem cell properties

“…In addition, the intracellular ICC signal of the CD133/1 mAb, both in 2D and 3D was similar to the granular pattern of Dylight 549 labeled anti-CD133/1 mAb obtained by confocal microscopy. This indicates that in addition to plasma membrane expression, a substantial amount of the CD133-receptor is also expressed in the cytoplasm which is consistent with other reports [33,34].…”

“…Interestingly, the BxPC3 cells displayed reduced viability (~50 %) when exposed to the CD133/1 (AC133) mAb only, suggesting that blocking the CD133 receptor may affect the cell viability or growth in this cell line. The mechanism behind this is currently unclear; however, it was recently shown that the blockage of the CD133 receptor of un-differentiated human colorectal Caco-2 cells by using the AC133 antibody resulted in a significant reduction of the uptake of transferrin [33]. In addition, since transferrin is an essential factor for the proliferation of pancreatic cancer cells [35], this may explain the cytotoxic effects of AC133 in the BxPc3 cells.…”

Expression of CD133 in various premalignant and proliferative lesions

“…external factors such as the environment or habits that lead individuals exposed to substances that are carcinogenic, systemic disease, drugs, etc. 4 Whereas internal factors are including the factors, which are found in the body such as the genetic, hereditary carried by each individual, the cancer stem cell (CSC) and growth factors that exist in the surrounding environment such as cells and other hormone. 5 CSC defines as a small subpopulation of cancer cells that form a pool of autonomous cell with the exclusive ability to make the heterogeneous lineages of cancer cells that comprehend the tumor.…”

“…These lesions including: mucocel, papilloma, fibroepitelial polyp and epulis. 4,25 materials and methods A cross sectional observational research was carried out on patients who have suffered premalignant lesion (PML: leukoplakia, erythroplakia, oral lichen planus, lichenoid reaction, and submucous fibrosis.) and proliferative lesion (PL) such as papilloma, fibroma, epulis, and cysts that seek treatment to oral medicine clinic under informed consent.…”

Cd133/Epcam Cancer Stem Cell Markers of Tumour Stage in Colorectal Cancer Cells

“…CD133 is a pentaspan trans-membrane protein that is detected by an antibody binding to its two glycosylated extracellular moieties [9,10]. Whilst its function is not clearly understood, this glycoprotein is localised to cell membrane protrusions and microvilli, where it associates with cholesterol domains, suggesting a functional role in membrane transport processes [11,12].…”