Tumor initiating cells: development and critical characterization of a model derived from the A431 carcinoma cell line forming spheres in suspension.
To investigate the tumor fraction with cancer stem/tumor initiating cell (CSC/TIC) characteristics, we tested the human cervical carcinoma cell lines A431, Caski and SiHa, by growth as non-adherent spheres in specific media and aldehyde dehydrogenase (ALDH) enzymatic activity. A good correlation between the two parameters was observed and the highest levels were observed in A431 cell line that was selected for characterization of the CSC/TIC fraction. A431 parental cells already displayed characteristics common to CSC/TIC, such as sphere forming efficiency, adherent holoclone formation and high ALDH activity. Non-adherent spheres maintained or increased these properties, and, in particular, ALDH-positive fraction increased from 46 to 65% and a transient induction of stem cell markers such as Nanog, Nestin and Oct4 was observed. Furthermore, a significant increase of paraclone forming cells was observed, suggesting that differentiation took place inside sphere cell populations. As compared to parental cells, spheres were characterized by: (1) a ten-fold higher verapamil-sensitive side population fraction; (2) the appearance of a podoplanin-positive subpopulation characterized by a small cell size; (3) the ability to propagate tumors in nude mice at a lower cell dose. The global gene expression analysis demonstrated a strong and reversible modulation of 'sphere' phenotype in comparison to parental and sphere cells re-induced to adherent conditions. All together our results indicated that the growth of A431 cells as a non-adherent sphere was not sufficient by itself to define a stem-like population, but it was essential for the emergence of a small population of tumor cells with CSC properties.
Unit of Molecular Therapies, Experimental Oncology and Molecular Medicine Department, Fondazione IRCCS Istituto Nazionale Tumori, Milan, Italy.
Ileana
I
Bortolomai
Silvana
S
Canevari
Ileana
I
Facetti
Loris
L
De Cecco
Giancarlo
G
Castellano
Alberto
A
Zacchetti
Malcolm R
MR
Alison
Silvia
S
Miotti
D000444:Aldehyde Dehydrogenase / Q000378:metabolism; D000818:Animals; D014408:Biomarkers, Tumor / Q000378:metabolism; D002294:Carcinoma, Squamous Cell / Q000201:enzymology / Q000473:pathology; D049109:Cell Proliferation; D002999:Clone Cells; D005260:Female; D020869:Gene Expression Profiling; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D051379:Mice; D008819:Mice, Nude; D008954:Models, Biological; D014411:Neoplastic Stem Cells / Q000378:metabolism / Q000473:pathology*; D058985:Side-Population Cells / Q000473:pathology; D018874:Spheroids, Cellular / Q000378:metabolism / Q000473:pathology*; D014407:Tumor Cells, Cultured; D002583:Uterine Cervical Neoplasms / Q000201:enzymology / Q000473:pathology
D014408:Biomarkers, Tumor; D000444:Aldehyde Dehydrogenase