tumors derive from a single cell clone but consist of heterogeneous cell subpopulations with diverse features and functions . sequencing analysis has revealed that tumors accumulate mutations , the majority of which do not provide selective benefits , while a small subset of mutations drive cancer development by conferring a selective advantage . a limited number of subclones with a selective advantage survive treatment with anticancer drugs or radiation . such subclones can initiate tumors when inoculated into immunocompromised mice and are known as cancer stem or cancer - initiating cells ( cics ) . cics can lead to tumor recurrence because they escape apoptosis by effluxing antitumor drugs and degrading reactive oxygen species ( ros ) , which are related to radiation - induced apoptosis . cics are generally defined on the basis of their ability to initiate a tumor in a transplantation assay in immunocompromised mice . in leukemia , breast , lung , and colon cancers , however , recent reports reveal that in some tumor types , such as melanomas , the majority of tumor cells can initiate tumors . asymmetrical division of stem cells yields a stem cell and a more highly differentiated cell . as in the case of physiological stem cells , cics are derived from cics but not from non - cics under normal conditions . however , under some conditions , such as severe hypoxia , non - cics can yield cics ; this phenomenon is known as  plasticity  . the dynamic exchange between cics and non - cics may enable tumors to survive under unfavorable conditions . therefore , cics may not be a distinct cell type but may be a mode or phenotype of tumor cells . this review briefly describes isolation of cics from tumor cell populations and the microenvironmental factors that regulate cic phenotypes in uterine cancer and lymphoma . cics express high levels of cell membrane - associated abc transporters . when stained with hoechst 33342 dye and incubated for approximately 1 hour in contrast , a small population with high efflux ability eliminates hoechst 33342 dye ( hoechst 33342-low ) . the upper part shows an image of hoechst 33342 staining , in which hoechst 33342 dye is designated by stars . the lower part shows the dot - blot pattern of flow cytometers , in which hoechst 33342-high cells are contained in the mp , whereas hoechst 33342-low cells in the sp . upper part shows the image of cm - h2dcfda staining , in which ros is designated as flowers . when treated with hydrogen peroxide , most tumor cells are stained with cm - h2dcfda , but a small population remains unstained . the upper part shows an aldefluor assay , in which aldh - high population is figured out in the box ( red part ) . these boxed cells are diminished when treated with n , n - diethylaminobenzaldehyde , the inhibitor of aldh . the lower part shows the results of immunohistochemical analysis using an anti - aldh antibody , in which a portion of uterine endometrioid carcinoma cells is aldh - positive . ) . cics express high levels of cell membrane - associated atp - binding cassette ( abc ) transporters , which play important roles in efflux of antitumor drugs . when stained with hoechst 33342 dye , most tumor cells retain the dye and are termed hoechst 33342-high . however , a small population with high efflux ability eliminates hoechst 33342 dye and are termed hoechst 33342-low . most tumor cells are hoechst 33342-high , and are termed the  main population ( mp )  , whereas hoechst 33342-low cells are known as the  side population ( sp )  . flow cytometers equipped with ultraviolet lasers can be used to distinguish the mp and sp , which enable isolation of sp cells . because sp cells can efflux compounds efficiently , cics are considered to be involved in the sp . patrawala et al . reported that the sp cells in several tumor cell lines are more tumorigenic than mp cells . this finding is consistent with the concept that cics are enriched in the sp . cics express high levels of cell membrane - associated abc transporters . when stained with hoechst 33342 dye and incubated for approximately 1 hour in contrast , a small population with high efflux ability eliminates hoechst 33342 dye ( hoechst 33342-low ) . the upper part shows an image of hoechst 33342 staining , in which hoechst 33342 dye is designated by stars . the lower part shows the dot - blot pattern of flow cytometers , in which hoechst 33342-high cells are contained in the mp , whereas hoechst 33342-low cells in the sp . upper part shows the image of cm - h2dcfda staining , in which ros is designated as flowers . when treated with hydrogen peroxide , most tumor cells are stained with cm - h2dcfda , but a small population remains unstained . the lower part shows the staining of endometrioid carcinoma cells . the upper part shows an aldefluor assay , in which aldh - high population is figured out in the box ( red part ) . these boxed cells are diminished when treated with n , n - diethylaminobenzaldehyde , the inhibitor of aldh . the lower part shows the results of immunohistochemical analysis using an anti - aldh antibody , in which a portion of uterine endometrioid carcinoma cells is aldh - positive . the second method of isolating cics is based on their ability to degrade ros ( fig . upon x - ray irradiation , tumor cells produce a large quantity of ros and undergo apoptosis . cics express high levels of ros - degrading enzymes , such as superoxide dismutase , which facilitate their escape from apoptosis by degrading intracellular ros . intracellular ros can be stained with 5-(and 6)-chloromethyl-2,7-dichlorohydrofluorescein diacetate ( cm - h2dcfda ) . when treated with hydrogen peroxide , most tumor cells are stained with cm - h2dcfda , but a small population remains unstained . these cm - h2dcfda - low cells are more tumorigenic than cm - h2dcfda - high cells , and cics are enriched in the cm - h2dcfda - low population . s lymphoma cell lines , most foxo3a - expressing cells are contained in the cm - h2dcfda - low population . a small subpopulation of hodgkin s lymphoma cells expressing foxo3a can degrade ros efficiently and is resistant to apoptosis . sternberg ( rs ) cells ; however , foxo3a - expressing cells are small mononucleated cells . the l1236 and l428 hodgkin s lymphoma cell lines include both small mononucleated and giant multinucleated cells . in these cell lines , multinucleated cells are derived from mononucleated cells , but mononucleated cells are not derived from multinucleated cells . these findings suggest that some mononucleated cells , but not rs cells , are the cics of hodgkin s lymphoma . several such markers , such as cd133 and cd44 ( cd44v isoform ) , have been reported ; aldehyde dehydrogenase ( aldh ) is one of the most commonly used . high aldh activity is present in not only normal stem cells , such as hematopoietic and neural stem cells , but also various types of cics . indeed , immunohistochemical analysis using an anti - aldh antibody revealed that a portion of uterine endometrioid carcinoma cells are aldh - positive , and cases with high aldh activity have an unfavorable prognosis ( fig . aldh - high endometrioid carcinoma cells are resistant to apoptosis , mobile , and highly invasive . sp and ros - degrading cells can only be detected while alive . by contrast , aldh - high cells can be detected immunohistochemically in formalin - fixed paraffin - embedded tissues . uterine endometrioid carcinoma cells usually express steroid hormone receptors , such as estrogen and progesterone receptors . however , double staining of clinical samples with anti - aldh and anti - hormone receptor antibodies revealed that aldh - high cells are negative for these hormone receptors . aldh - high endometrioid carcinoma cells appear to be immature and negative for  differentiation  markers , such as estrogen and progesterone receptors . because aldh - high cells are readily isolated by flow cytometry , their proteome can be investigated using mass spectroscopy ( table 1table 1 . list of proteins preferentially expressed in the aldh - high population of uterine endometrioid adenocarcinoma cells ) . these cells also exhibit impaired akt phosphorylation and matrix metalloproteinase-2 activation , which account for their impaired proliferation and invasion , respectively . elevated expression of s100a4 is related to myometrial and lymphatic invasion in well- to moderately - differentiated endometrioid carcinoma . notably , strong and diffuse expression of s100a4 is observed in tumor tissues with a microcystic , elongated , and fragmented (  melf  ) pattern , which is associated with a highly invasive phenotype . however , aldh is not an almighty marker of cics , because , for example , aldh is highly expressed in stromal cells , such as dendritic cells and macrophages , in lymphoma . this suggests the presence of microenvironmental factors that induce or inhibit aldh expression and suggests that intercellular signaling between tumor cells might regulate aldh expression . indeed , phosphorylated smad-2 , an indicator of tgf- signaling activation , is present at higher levels in aldh - low than -high endometrioid carcinoma cells . tgf- family members regulate cell fates in developing embryos ; for example , lefty determines the left similarly , the development or differentiation of tumor cells might be controlled by tgf- family members . among the tgf- receptor ligands immunohistochemical analysis of clinical samples has revealed nodal - high tumor cells to be aldh - low cells , and vice versa . these findings suggest that nodal inhibits aldh expression via stimulation of tgf- signaling in uterine endometrioid carcinoma cells . lymphomas can be categorized using the surface markers of differentiation status expressed by lymphocytes . for example , peripheral t - cell lymphoma cells express the t - cell marker cd3 , and diffuse large b - cell lymphoma cells express the b - cell markers cd79a and cd20 . lymphoplasmacytic lymphoma ( lpl ) is a rare indolent disease that affects the bone marrow , and is associated with expression of the b- and plasma - cell markers cd20 and cd138 . mwcl-1 cells are derived from lpl , and express cd20 and cd138 as surface markers . mwcl-1 cells include three subpopulations : cd20 cd138 , cd20 cd138 , and cd20 cd138 cells . the first subpopulation expresses neither b- nor plasma - cell markers , the second subpopulation expresses only b - cell markers , and the third subpopulation expresses both b- and plasma - cell markers . when cultured , cd20 cd138 cells yield all three subpopulations , but cd20 cells do not yield cd20 cd138 cells ( fig . mwcl-1 cells derived from lpl include three subpopulations : cd20 cd138 , cd20 cd138 , and cd20 cd138 cells . when cultured , cd20 cd138 cells yield all three subpopulations , but cd20 cells do not yield cd20 cd138 cells . the cd20 cd138 subpopulation expresses the chemokine receptor cxcr7 , the ligand of which is cxcl12 . the number of cd20 cd138 cells increases in a time- and dose - dependent manner when mwcl-1 cells are cultured in the presence of cxcl12 . ) . cd20 cd138 cells have higher ros degradation and in vitro colony formation activities than cd20 cd138 and cd20 cd138 cells . when cultured in the absence of serum or the presence of an anticancer drug , cd20 cd138 cells immunohistochemical analysis of clinical samples has indicated that lpl tumor cells undergoing apoptosis are cd138 . the production of all three subpopulations , efficient ros degradation and in vitro colony formation activities , and resistance to apoptosis suggest that cd20 cd138 cells are candidate cics in lpl . mwcl-1 cells derived from lpl include three subpopulations : cd20 cd138 , cd20 cd138 , and cd20 cd138 cells . when cultured , cd20 cd138 cells yield all three subpopulations , but cd20 cells do not yield cd20 cd138 cells . the cd20 cd138 subpopulation expresses the chemokine receptor cxcr7 , the ligand of which is cxcl12 . the number of cd20 cd138 cells increases in a time- and dose - dependent manner when mwcl-1 cells are cultured in the presence of cxcl12 . as described above , cics are plastic under stress conditions , in which non - cics are transformed to cics . indeed , hypoxia induces conversion of cd20 cd138 cells to cd20 cd138 cells , whereas normoxic conditions do not . among the proteins preferentially expressed by the cd20 cd138 subpopulation , expression of the chemokine receptor cxcr7 it is possible that the conversion from cd20 cd138 to cd20 cd138 cells is mediated by signaling via cxcr7 ( fig . this is plausible because the number of cd20 cd138 cells increases in a time- and dose - dependent manner when mwcl-1 cells are cultured in the presence of cxcl12 , a ligand of cxcr7 . recently , a constitutively active mutation of cxcr4 , a receptor related to cxcr7 , has been reported in lpl . the fact that cics are resistant to apoptosis and related to aggressive behavior is consistent with the notion that the cics of lpl utilize the cxcl12-cxcr4/cxcr7 axis . cics , which are resistant to anticancer drugs and escape from apoptosis , may be a good therapeutic target for cancers . however , they are , under some circumstances , derived from non - cics due to their plasticity . therefore , the regulatory mechanism of cics should be determined . reagents that block the signals controlling cics may interfere with their plasticity , leading to their complete elimination . the tgf- family member nodal regulates aldh activity in uterine endometrioid carcinoma , whereas signaling via the chemokine receptor cxcr7 regulates cics in non - hodgkin s lymphoma . therefore , investigation of the cics of each tumor type is necessary to facilitate development of novel therapeutics . aldh - high tumor cells express a large number of factors , some of which play roles in invasion and some of which play roles in mobility . although aldh - high cells are invasive and mobile , these characteristics are likely mediated by different factors . genome - editing techniques will enable determination of the function of such factors in cics , which will facilitate development of novel therapeutics .
tumors derive from a single cell clone but consist of heterogeneous cell subpopulations with diverse features and functions . a limited number of subclones with a selective advantage can initiate tumors when inoculated into immunocompromised mice , and are called cancer - initiating cells ( cics ) . 
 cics can be isolated from the bulk of tumors on the basis of their characteristics , such as high reagent efflux , degradation of reactive oxygen species , and aldehyde dehydrogenase ( aldh ) activity . under normal conditions , 
 new cics are produced by existing cics rather than non - cics . however , 
 under stress conditions , non - cics can occasionally produce cics , a phenomenon known as plasticity . 
 the dynamic exchange between cics and non - cics may enable tumors to survive under unfavorable conditions . 
 cics are located in a small portion of tumors . 
 this suggests that microenvironmental factors induce or inhibit the cic phenotype , which might be regulated by intercellular signaling between tumor cells . 
 this review describes isolation of cics from tumor cell populations and the microenvironmental factors that regulate cic phenotypes in uterine cancer and lymphoma .