Hypoxia Enhances Tumor Stemness by Increasing the Invasive and Tumorigenic Side Population Fraction

Abstract

Although advances have been made in understanding the role of hypoxia in the stem cell niche, almost nothing is known about a potentially similar role of hypoxia in maintaining the tumor stem cell (TSC) niche. Here we show that a highly tumorigenic fraction of side population (SP) cells is localized in the hypoxic zones of solid tumors in vivo. We first identified a highly migratory, invasive, and tumorigenic fraction of posthypoxic side population cells (SPm [hox] fraction) in a diverse group of solid tumor cell lines, including neuroblastoma, rhabdomyosarcoma, and small-cell lung carcinoma. To identify the SPm (hox) fraction, we used an "injured conditioned medium" derived from bone marrow stromal cells treated with hypoxia and oxidative stress. We found that a highly tumorigenic SP fraction migrates to the injured conditioned medium in a Boyden chamber. We show that as few as 100 SPm (hox) cells form rapidly growing tumors in vivo. In vitro exposure to hypoxia increases the SPm (hox) fraction significantly. Quantitative realtime polymerase chain reaction and immunofluorescence studies showed that SPm (hox) cells expressed Oct-4, a "stemness" gene having a potential role in TSC maintenance. In nude mice xenografts, SPm (hox) cells were localized to the hypoxic zones, as demonstrated after quantum dot labeling. These results suggest that a highly tumorigenic SP fraction migrates to the area of hypoxia; this migration is similar to the migration of normal bone marrow SP fraction to the area of injury/hypoxia. Furthermore, the hypoxic microenvironment may serve as a niche for the highly tumorigenic fraction of SP cells. STEM

Oct-4+/Tenascin C+ neuroblastoma cells serve as progenitors of tumor-derived endothelial cells

“…Previous studies [20,22] have demonstrated that sidepopulation cells of the SK-NB-BE(2) NB cell line and tumor-initiating human NB cells express the embryonic stem cell [12][13][14] and cancer stem cell marker Oct-4 [15][16][17][18][19][20]. The human Oct-4 gene can generate at least three transcripts (Oct-4A, Oct-44B and Oct-4B1) and four protein isoforms (OCT4A, OCT4B-190, OCT4B-265 and OCT4B-164) by alternative splicing and alternative translation initiation [31].…”

“…All isotypic controls used for immunofluorescence tested negative ( Figure 2B Figure S1, 4,8,12,16,20).…”

“…The Octamer-binding transcription factor 4 (Oct-4), together with SOX-2 and NANOG, maintains self-renewal and blocks cell differentiation in embryonic stem cells [12][13][14] and is expressed by some cancer stem cells [15][16][17][18][19][20]. Among these factors, Oct-4 and SOX-2 appear to be indispensable for stemness maintenance [21].…”

“…Among these factors, Oct-4 and SOX-2 appear to be indispensable for stemness maintenance [21]. Previous studies have demonstrated that Oct-4 is expressed by side-population cells and tumor-initiating cells from human NB, indicating that Oct-4 + NB cells include tumor stem/progenitor cells [20,22].…”

Dual SP/ALDH Functionalities Refine the Human Hematopoietic Lin−CD34+CD38− Stem/Progenitor Cell Compartment

“…Given the role of SP and ALDH functionalities in HSC quiescence/self-renewal [14,[38][39][40], reflecting two processes involved in early stem cell regulation, the isolation of cells expressing both activities is of great interest to better understand these processes. Furthermore, because SP and ALDH are involved in stem cell multidrug resistance [41], this coupled technology is of clinical significance, allowing the comparison, quantification, and manipulation of normal and leukemic stem cells (LSCs) [39]. Actually, it is hypothesized that LSCs are quiescent and closely associated with a specific niche environment, from which they can be forced to escape to expose them to antileukemic drugs [42].…”