Genotoxic damage of human adipose-tissue derived mesenchymal stem cells triggers their terminal differentiation.
ABSTRACT Human adipose tissue-derived mesenchymal (stromal) stem cells (AT-MSCs) and genetically modified to express cytosine deaminase:uracil phosphoribosyltransferase (CDy-AT-MSCs) were treated with hydrogen peroxide in order to induce DNA damage and subsequently evaluate their genetic stability by single cell gel electrophoresis. Both cells types (parental and transgene modified) did not differ in the sensitivity to DNA breaks induction. Potential tumorigenicity of AT-MSCs and CDy-AT-MSCs was tested by subcutaneous inoculation of cell suspension into flank of immunocompromised mice. Dose of 15x10(6) cells was not found to be tumorigenic in given experimental setup. AT-MSCs, CDy-AT-MSCs and MSCs isolated from human lipoma were treated with chemical carcinogen 4-nitroquinoline-1-oxide (4NQO) in attempts to transform them. Surviving cells after genotoxic stress were not transformed but underwent replicative senescence. Irreparable DNA damage caused triggered adipogenic terminal differentiation, rather than apoptosis induction in all kinds of cells tested.
- SourceAvailable from: Christopher D Porada[Show abstract] [Hide abstract]
ABSTRACT: Large animal models that accurately mimic human hemophilia A (HA) are in great demand for developing and testing novel therapies to treat HA. To re-establish a line of sheep exhibiting a spontaneous bleeding disorder closely mimicking severe human HA, fully characterize their clinical presentation, and define the molecular basis for disease. Sequential reproductive manipulations were performed with cryopreserved semen from a deceased affected ram. The resultant animals were examined for hematologic parameters, clinical symptoms, and responsiveness to human FVIII (hFVIII). The full coding region of sheep FVIII mRNA was sequenced to identify the genetic lesion. The combined reproductive technologies yielded 36 carriers and 8 affected animals. The latter had almost non-existent levels of FVIII:C and extremely prolonged aPTT, with otherwise normal hematologic parameters. These animals exhibited bleeding from the umbilical cord, prolonged tail and nail cuticle bleeding time, and multiple episodes of severe spontaneous bleeding, including hemarthroses, muscle hematomas and hematuria, all of which responded to hFVIII. Inhibitors of hFVIII were detected in four treated animals, further establishing the preclinical value of this model. Sequencing identified a premature stop codon and frame-shift in exon 14, providing a molecular explanation for HA. Given the decades of experience using sheep to study both normal physiology and a wide array of diseases and the high homology between human and sheep FVIII, this new model will enable a better understanding of HA and facilitate the development and testing of novel treatments that can directly translate to HA patients.Journal of Thrombosis and Haemostasis 11/2009; 8(2):276-85. · 6.08 Impact Factor
- Journal of the Saudi Heart Association 07/2011; 23(3):173-173.
- Cancer Letters - CANCER LETT. 01/2010; 1.
Genotoxic damage of human adipose-tissue derived mesenchymal stem cells
triggers their terminal differentiation
V. ALTANEROVA1, E. HORVATHOVA2, M. MATUSKOVA1, L. KUCEROVA1, C. ALTANER1*
1Laboratory of Molecular Oncology, 2Laboratory of Mutagenesis and Carcinogenesis, Cancer Research Institute SAS, Bratislava, Slovakia, e-mail:
Received April 26, 2009
Human adipose tissue-derived mesenchymal (stromal) stem cells (AT-MSCs) and genetically modified to express cyto-
sine deaminase:uracil phosphoribosyltransferase (CDy-AT-MSCs) were treated with hydrogen peroxide in order to induce
DNA damage and subsequently evaluate their genetic stability by single cell gel electrophoresis. Both cells types (parental
and transgene modified)didnotdifferinthesensitivitytoDNAbreaksinduction.PotentialtumorigenicityofAT-MSCsand
CDy-AT-MSCs was tested by subcutaneous inoculation of cell suspension into flank of immunocompromised mice. Dose
of 15x106 cells was not found to be tumorigenic in given experimental setup. AT-MSCs, CDy-AT-MSCs and MSCs isolated
from human lipoma were treated with chemical carcinogen 4-nitroquinoline-1-oxide (4NQO) in attempts to transform them.
Surviving cells after genotoxic stress were not transformed but underwent replicative senescence. Irreparable DNA damage
caused triggered adipogenic terminal differentiation, rather than apoptosis induction in all kinds of cells tested.
Key words: human adipose tissue-derived mesenchymal stem cells; genotoxic damage, single cell gel electrohoresis; chemical
carcinogen 4NQO, terminal differentiation
Human adipose tissue-derived mesenchymal (stromal)
stem cells (AT-MSCs) are multipotent adult stem cells derived
usually from lipoaspirate. Similarly as bone marrow derived
mesenchymal stem cells (MSC), AT-MSCs have the ability to
migrate to sites of injury to fulfill their role in repair of the
damage tissues . MSCs and AT-MSCs can be isolated by
their adherence to plastic tissue culture plates  Theyieldof
MSCs from adipose tissue is higher in comparison to the bone
marrow [1, 3, 4]. Theeasyandrepeatableaccesstosubcutane-
ous adipose tissue and the simple isolation procedures provides
a clear advantage over other sources . Therefore AT-MSCs
have potential therapeutic use as autologous and allogeneic
products for tissue engineering.
Mesenchymal stem cells both MSCs and AT-MSCs possess
unique ability to selectively migrate to tumors, metastases and
contribute to the formation of tumor-associated stroma. This
property predetermines them to become vehicles for stem cell
based targeted cancer gene therapy [5, 6].
Previously we have prepared yeast fusion cytosine deami-
nase:uracil phosphoribosyltransferase expressing human
adipose tissue derived mesenchymal stem cells (CDy-AT-
MSCs) by retrovirus transduction. CDy-AT-MSCs exerted
their anti-tumor potential on human colon cancer cells, human
malignant melanoma cell line, and human metastatic prostate
cells in the presence of prodrug 5-fluorocytosine (5-FC) [3,
7, 8]. CDy-AT-MSCs in combination with 5-FC augmented
tumor cells in co-culture in vitro. Moreover, strong inhibition
of subcutaneous tumor growth was achieved by systemically
administered CDy-AT-MSCs on xenotransplanted human
tumors grown on nude mice upon 5-FC treatment.
Therefore human mesenchymal stem cells derived from
adipose tissue are not only useful cells in regenerative medi-
cine, but when modifiedbyprodrugconvertinggenemayalso
serve as attractive delivery vehicles for stem cell-based targeted
cancer gene therapy. Genetic stability of AT-MSCs is an im-
portant attribute to asses, when they are intended to be used
in regenerative and cancer gene therapy clinical studies.
The aim of the present study was to evaluate genetic sta-
bility, of AT-MSCs and CDy-AT-MSCs, their sensitivity to
transformation induced by chemical carcinogen and potential
tumorigenicity. Here we demonstrate that irreparable DNA
damage caused by chemical carcinogen triggered adipose
* Corresponding author
NEOPLASMA 56, 6, 2009
GENOTOXIC DAMAGE OF HUMAN ADIPOSE TISSUE MESENCHYMAL CELLS
derived mesenchymal (stromal) cells to adipogenic, terminal
Materials and methods
Chemicals. Ethidium bromide, 4-nitroquinoline-1-oxide
(4NQO) were purchased from Sigma-Aldrich Co. (St. Louis,
MO). 4NQO was prepared as 5 mmol/L stock solutions in
100% dimethylsulfoxide (DMSO). Aliquots were diluted with
complete medium just before they were applied to culture
Mesenchymal stem cells isolation from human adipose
tissue, culture, and retrovirus transduction. AT-MSC cells
were isolated and cultivated as we described previously .
Lipoaspirates for their isolation were obtained from healthy
persons undergoing elective lipoaspiration, who provided an
informed consent. Cells were plated in low glucose (1,0 mg/L)
DMEM supplemented with 5% MSC-stimulatory supplement
(StemCell Technologies), 5% of human platelet extract and
antibiotic-antimycotic mixture. AT-MSCs were cultured at
370C in a humidified atmosphere containing 5% carbon di-
oxide with medium changes twice a week. Cells were plated
at a density of 3x103 to 5x103 nucleated cells/cm2. Adherent
cells were split upon reaching confluenceandAT-MSCswere
used for the experiments up to passage 5.
To prepare AT-MSCs expressing cytosine deaminase
(CDy-AT-MSCs), subconfluent cultures of AT-MSCs were
transduced thrice in 3 consecutive days with virus-contain-
ing medium from GP+envAM12/pST2 cells supplemented
with 100 μg/mL protamine sulphate. Transduced cells were
selected by cultivation in medium supplemented with 0.5
mg/ml of G418.
Experiments in vivo. Six- to 8-week-old athymic nude mice
(Balb/c-nu/nu) were used in accordance with institutional
guidelines under the approved protocols. The potential tu-
morigenicity of AT-MSCs and CDy-AT-MSCs was tested
by subcutaneous inoculation of cell suspension of 15x106
AT-MSCs or CDy-AT-MSCs in 0.1 ml PBS into flank of each
nude mouse (n = 2 in each treatment group). The animals
were inspected for tumor growth during 60 days and then the
tumor presence was control by autopsy.
Hydrogen peroxide treatment and DNA-damage testing.
The method of single cell gel electrophoresis (SCGE; comet
assay) according to  was performed with minor modifica-
tions suggested in [10, 11]. Briefly: 2.5 x 104 AT-MSCs or
CDy-AT-MSCs in 85μl of 0.75% low melting point agarose in
PBS were spread on a base layer of 100 μl 1% normal-melting
point agarose in PBS, placed on microscope slides and covered
with a cover slip. When the gel solidified, the cover slip was
removed. The cells were then exposed to different concentra-
tions of hydrogen peroxide (0, 100, 200, 300, and 400 μM).
Hydrogen peroxide treatment (5 min on ice) minimizes the
DNA repair process . The slides were washed with PBS
and placed in lysis solution (2.5 M NaCl, 0.1 M Na2EDTA, 10
mM Tris–HCl, pH 10.0, 1% Triton X-100) for 1 h at 4 0C. After
lysis the slides were transferred to an electrophoresis buffer
(0.3 M NaOH, 1 mM Na2EDTA, pH > 13.0) for unwinding
(40 min at 40C) and then subjected to electrophoresis at 25
V (current adjusted to 0.3 A) for 30 min at 40C. Theslideswere
neutralized with 0.4 M Tris–HCl (pH 7.5) and stained with
ethidium bromide (EtBr, 5μg/ml). For each sample, 100 EtBr-
stained nucleoids on triplicate slides were evaluated and scored
with an Olympus fluorescent microscope and computerized
image analysis (Komet 5.5, Kinetic Imaging, Liverpool, UK)
for determination of DNA in the tail, which is linearly related
to the frequency of single strand DNA breaks
Treatment of AT-MSCs, CDy-AT-MSCs, and MSCs from
lipoma with 4NQO.
Cells were exposed to single dose of 10 μM concentra-
tion of 4NQO. Survived fraction of cells (about ten percent)
was further cultivated in growth medium and when reached
confluence transferred by trypsin treatment. The cultivation
continued for several months. During this period of time signs
of senescence appeared, intervals of cell transfer was longer,
the number of passages never reach more than 10 before the
terminal differentiation appeared.
Statistics. Statistical comparisons of mean tail DNA values
in hydrogen peroxide treatment experiments were performed
using the Student’s t-test.
Induction of single stranded DNA breaks. Treatment of AT-
MSCs and CDy-AT-MSCs with hydrogen peroxide resulted
in an increase of DNA breaks in both parental AT-MSCs and
transgene-modified CDy-AT-MSCs when concentration of
hydrogen peroxide reached 100 μM. Higher hydrogen per-
oxide concentration did not further increase the number of
DNA breaks (Fig. 1.) Percentage of tail DNA was comparable
at each hydrogen peroxide concentration tested in both kinds
of cells. Thereforethegeneticallymodifiedmesenchymalcells
did not differ from parental cells in the sensitivity to DNA
Tumorigenicity of AT-MSCs and CDy-AT-MSCs testing.
Inoculums of cell suspension (15x106 of AT-MSCs or CDy-
AT-MSCs) in 0.1 ml PBS was injected subcutaneously into the
were inspected for tumor growth during period of time two
months. The palpable cell inoculum persisted on the site of
injection several days and slowly disappeared during firsttwo
weeks. By day 60 the experiments were ended and animals were
sacrificed and ispected for tumor presence. No tumors were
found either on the site of inoculation nor elsewhere during
the autopsy. ThereforetheAT-MSCsandCDy-AT-MSCswere
found not to be tumorigenic in given experimental setup.
Cytotoxicity of 4NQO to AT-MSC and CDy-AT-MSC.
Mesenchymal stem cells are known for their higher natural
resistance to toxic agents. In order to elucidate the sensitivity
544V. ALTANEROVA, E. HORVATHOVA2 M. MATUSKOVA, L. KUCEROVA, C. ALTANER
of AT-MSCs, CDy-AT-MSCs and MSCs isolated from lipoma,
potent chemical carcinogen and mutagen 4-nitroquinoline-1-
oxide (4NQO) was chosen for cell treatment. Concentrations
higher than 60 μM of 4NQO were found to be toxic to destroy
cells completely during 4 days of cultivation. At least 10 per-
cent of cells survived treatment with 10 μM concentration of
4NQO for 4 days. Recovered 4NQO-treated cells were further
cultivated and split upon reaching confluence. Cell prolifera-
tion declined with passages kept in culture. Cells became more
light-refractory, the cell morphology changed to cells more
prolonged (Fig. 2 B). Typical signs of aging-related phenotype
appeared (Fig. 2 C). Finally, the cells appeared to form drops of
fat, signs of mature differentiationtoadipocytesinallcells(Fig.
2 D). Accumulation of irreparable DNA damage in adipose
tissue derived mesenchymal stem cells apparently triggered
their terminal differentiation. Terminal adipogenic differen-
tiation after genotoxic stress caused by 4NGO was noticed in
AT-MSCs, in cytosine deaminase transduced CDy-ATMSCs
and also in MSCs isolated from human lipomas.
Several properties of human mesenchymal stem cells de-
rived from adipose tissue designate them attractive vehicles
for cell therapies. Theyareabletodifferentiatealonga variety
of lineage pathways, including bone, cartilage, adipose, neu-
ronal-like, and muscle in vitro [see 1 for a review]. They are
nonimmunogenic upon transplantation into allogeneic host [4,
13, 14]. Thecellsexhibita lowimmunogenicprofileasshown
by negative expression of MHC class II molecules and absence
of costimulatory molecule expression [15, 16, 17]. Furthermore
it was found that they are immunosuppressive [18, 19] as it
was demonstrated by their ability to control graft-versus-host
disease (GvHD) in humans .
Moreover AT-MSCs possess unique ability to selectively
migrate to tumors and metastases. When AT-MSCs are modi-
fied by prodrug converting gene they are attractive delivery
vehicles for anti-tumor agents such as IFNβ to tumors 
and for stem cell-based targeted cancer gene therapy [3, 7, 8,
22]. Taken in account nonimmunogenic properties of MSCs,
AT-MSCs may have potential therapeutic use as allogeneic
products for tissue engineering and for gene therapy upon
genetic modifications as well. For all these reasons, to assess
genetic stability of these cells is rather important, when they
are intended to be used in clinical studies.
There were several contradictive reports in literature with
regards to involvement of adult MSCs in tumor formation
. Increasing number of reports indicated that MSCs are
recruited in large numbers to the stroma of developing tumors
[21, 24], or they could act by enhancing the motility, invasion
and metastasis ability of adjacent cancer cells .
Experiments with human xenografts on nude mice have
shown that systemically administrated MSCs possess intrinsic
preferential migratory ability towards breast carcinoma cells,
lung metastasis of melanoma cells, intracranial glioma and co-
lon cancer cells [26, 27]. Moreover exogenously administered
MSCs could form a significantproportionoftumormass.
It was demonstrated that bone marrow derived MSCs may
also be involved in cancer metastasis forming pre-metastatic
niche . Thisfindingsupportstheideathattherapeutically
engineered MSCs may be used to treat metastases. Therefore
the introduction of a transgene into autologous stem cells pos-
sesses an attractive cell based delivery strategy [8, 29]. On the
contrary, other studies observed that MSCs may inhibit tumor
growth in animal models [30, 31] and posses antitumorigenic
effects on a model of Kaposi’s sarcoma .
Few contradictory reports appeared about spontaneous
transformation of cultured bone marrow derived murine
MSCs [32, 33, 34, 35] and human MSCs [36, 37]. It was
Figure 1. DNA-damaging effectofhydrogenperoxideonparentalandcytosinedeaminasetransducedhumanadiposetissuederivedmesenchymalstem
cells. Details of experimental procedure and evaluation is described in Material and Methods.
GENOTOXIC DAMAGE OF HUMAN ADIPOSE TISSUE MESENCHYMAL CELLS
reported that murine but not human mesenchymal stem
cells generated osteosarcoma-like lesions in the lung . If
murine MSCs were implanted subcutaneously together with
porous ceramic, host-derived sarcomas developed. Sarcomas
developed only in syngenic and immunodeficient recipients,
but not in allogenic hosts or when the cells were inoculated
as suspension .
In our experiments we did not observed any tumor forma-
tion when a high dose of AT-MSCs, or CDy-AT-MSCs was
inoculated subcutaneously as cell suspension in nude mice.
However, when these cells are co-injected with tumor cells,
they can support the growth of engrafts of human melanoma
xenografts, but not human glioma cells on nude mice, due
to their production of various growth factors in a paracrine
Mesenchymal stem cells as cells of vital importance for
organism are equipped with efficient DNA repair system as
well as high natural resistance to toxic agents . It was
therefore not surprising that DNA damage in CDy-AT-MSCs
was not influenced by the transduction with cytosine deami-
nase as documented by our results obtained with single-cell
gel electrophoresis, which represents a sensitive method for
measuring DNA damage.
In order to find whether adipose tissue derived mesen-
chymal stem cells can be transformed in vitro by chemical
carcinogen, we have chosen water soluble highly effective
carcinogen 4-nitroquinoline-1-oxide (4NQO). 4NQO is
metabolized into 4-acetoxyaminoquinoline-1-oxide (Ac-
4HAQO), which can form covalent adduct to deoxyguanine
and deoxyadenine in DNA. Thecarcinogenicactivityof4NQO
consist in DNA damage production and formation of irrevers-
ible topoisomerase I cleavage complexes (Top1cc) inducing
recombinations. Top1cc produced by 4NQO accumulate
progressively after 4NQO addition and persist to following
4NQO removal. . In addition 4NQO also produces oxida-
tive damage and DNA single-stranded breaks . Results of
our study indicate that replicative senescence of AT-MSCs as
a consequence of cumulating DNA damage is a continuous
process including far reaching alterations in phenotype like
the terminal differentiation. Similar observation with human
Figure 2. Adipogenic terminal differentiationofAT-MSCafteraccumulationofDNAdamagecausedby4NQO. A – untreatedcontrolAT-MSCs;B – AT-
MSCs exposed to 4NQO after serial passages became bigger; C- Senescence of damaged AT-MSCs progressed during serial passages, cells being more
light refractory and sparse; D – All 4NQO-treated AT-MSCs differentiated to adipocytes. Cells were stained with Oil Red-O. Magnification 100 fold.
546V. ALTANEROVA, E. HORVATHOVA2 M. MATUSKOVA, L. KUCEROVA, C. ALTANER
bone marrow derived MSC together with detail analysis on
molecular level was reported [44, 45]. The DNA damage,
a prime suspect in stem cell aging, causes graying and loss of
melanocyte stem cells by inducing premature differentiation,
without inducing apoptosis or senescence as it was reported
very recently . The irreparable DNA damage, as caused
by ionizing radiation, abrogated renewal of MSCs in mice.
Surprisingly, the DNA-damage response triggers MSC differ-
entiation into mature melanocytes in the niche, rather than
inducing their apoptosis or senescence .
Several other reports support the conclusion that accumula-
tion of DNA damage may cause somatic stem cell depletion [47,
48]. Typical sign of aging in mammals is hair graying caused
by the incomplete maintenance of melanocyte stem cells with
age . TheresultingMSCdepletionleadstoirreversiblehair
graying. Deficiency of Ataxia-telangiectasia mutated (ATM),
a central transducer kinase of the DNA-damage response,
sensitizes MSCs to ectopic differentiation,demonstratingthat
the kinase protects MSCs from their premature differentiation
by functioning as a “stemness checkpoint” to maintain the stem
cell quality and quantity .
We observed that genotoxic damage of human adipose
tissue derived mesenchymal stem cells caused by chemical
carcinogen 4NQO is not leading to cell transformation but
to process of ageing. The aged AT-MSCs are triggered to adi-
pogenic terminal differentiation.Thesedataareinagreement
with above mentioned observations of terminal differentiation
of melanocyte stem cells . Whether the elimination of
DNA damaged stem cells is generally provided by terminal
differentiation rather than apoptosis remains to be proven.
Acknowledgements. The study was supported by APVV grant
0260/07 (awarded to L. K.); VEGA grant 2/0072/09 (awarded to E. H.);
Capital Consult GmbH, Munich, Germany; and the League Against
Cancer. We thank D. Guba M.D., Institute of Medical Cosmetics, Bra-
tislava, Slovakia for providing us with material for AT-MSC isolation,
M. Dubrovcakova and V. Frivalska for technical assistance.
 SCHAFFLER A, BUCHLER C. Concise review: adipose tis-
sue-derived stromal cells – basic and clinical implications
for novel cell-based therapies. Stem Cells 2007; 25: 818–827.
ZUK PA, ZHU M, ASHJIAN P et al. Human adipose tissue
is a source of multipotent stem cells. Mol Biol Cell 2002; 13:
KUCEROVA L, ALTANEROVA V, MATUSKOVA M et
al. Adipose tissue derived human mesenchymal stem cells
mediated prodrug cancer gene therapy. Cancer Res 2007; 67:
KERN S, EICHLER H, STOEVE J et al. Comparative analysis
of mesenchymal stem cells from bone marrow, umbilical cord
blood or adipose tissue. Stem Cells 2006; 24: 1294–1301.
 ALTANER C. Prodrug cancer gene therapy. Cancer Letters
2008; 270: 191–201. doi:10.1016/j.canlet.2008.04.023
ALTANER C. Glioblastoma and stem cells Minireview. NEO-
PLASMA 2008; 55: 369–374.
KUCEROVA L, MATUSKOVA M, PASTORAKOVA A et al.
Cytosine deaminase expressing human mesenchymal stem
cells mediated tumour regression in melanoma bearing mice.
J Gene Med 2008; 10: 1071–1082. doi:10.1002/jgm.1239
CAVARRETTA I, ALTANEROVA V, MATUSKOVA M et al.
Adipose tissue-derived mesenchymal stem cells expressing
prodrug converting enzyme inhibit human prostate tumor
growth. Molecular Therapy (2009, in revision)
SINGH, NP, MCCOY MT, TICE RR et al. A simple technique
for quantitation of low levels of DNA damage in individual
cells. Exp. Cell Res.1988; 175: 184–191. doi:10.1016/0014-
SLAMENOVA D, GABELOVA A, RUZEKOVA L et al.
Detection of MNNG-induced DNA lesions inmamma-
lian cells: validation of comet assay against DNA unwinding
technique,alkaline elution of DNA and chromosomal aber-
rations. Mutat. Res. 1997; 383: 243–252.
SLAMENOVA D, HORVATHOVA E, MARSALKOVA L, et
al. Carvacrol given to rats in drinking water reduces the level
of DNA lesions induced in freshly isolated hepatocytes and
testicular cells by H2O2. Neoplasma. 2008; 55: 394–399.
GABELOVA A, SLAMENOVA D, RUZEKOVA L et al. Meas-
urement of DNA strand breakage and DNA repair induced
with hydrogen peroxide using single cell gel electrophoresis,
alkaline DNA unwinding and alkaline elution of DNA. Neo-
plasma 1997; 44: 380–388.
STREM BM, HICOK KC, ZHU M et al. Multipotential dif-
ferentiation of adipose tissue-derived stem cells. Keio J.Med.
2005; 54:132–141. doi:10.2302/kjm.54.132
KEATING A. Mesenchymal stromal cells. Curr. Opin. Hematol.
2006; 13: 419–425. doi:10.1097/01.moh.0000245697.54887.6f
MCINTOSH K, ZVONIC S, GARRETT S et al. The im-
munogenicity of human adipose derived cells: Temporal
changes in vitro. Stem Cells 2006; 24: 1246–1253. doi:10.1634/
NIEMEYER P, KORNACKER M, MEHLORN A et al.
Comparison of immunological properties of bone marrow
stromal cells and adipose tissue derived stem cells before and
after osteogenic differentiation in vitro. Tissue Eng 2007; 13:
CROP MJ, BAAN CC, KOREVAAR SS. et al. Donor-derived
mesenchymal stem cells suppress alloreactivity of kidney
transplant patients. Transplantation. 2009; 87: 896–906.
PUISSANT B., BARREAU C., BOURIN P et al. Immu-
nomodulatory effect of human adipose tissue derived adult
stem cells: comparison with bone marrow mesenchymal stem
cells. Br J Haematol 2005; 129: 118–129. doi:10.1111/j.1365-
CUI L, YIN S, LIU W et al. Expanded adipose derived
stem cells suppress mixed lymphocyte reaction by secre-
tion of prostaglandin E2. Tissue Eng 2007; 13: 1185–1195.
GENOTOXIC DAMAGE OF HUMAN ADIPOSE TISSUE MESENCHYMAL CELLS
 YAÑEZ R, LAMANA ML, GARCÍA-CASTRO J et al. Adipose
tissue-derived mesenchymal stem cells have in vivo immu-
nosuppressive properties applicable for the control of the
graft-versus host disease. Stem Cells 2006; 24: 2582–2591.
STUDENY FC, MARINI JL, DEMBINSKI C et al. Mesen-
chymal stem cells: potential precursors for tumor stroma and
targeted-delivery vehicles for anticancer agents, J. Natl. Cancer
Inst. 2004; 96: 1593–1603.
ABOODY KS, BUSH RA, GARCIA E, et al. Development of
a tumor selective approach to treat metastatic cancer. PLoS
ONE 2006;1: E23–31. doi:10.1371/journal.pone.0000023
LAZENNEC G, JORGENSEN C Concise review: adult multipo-
tent stromal cells and cancer: risk or benefit?StemCells.2008;
26: 1387–1394. doi:10.1634/stemcells.2007-1006
HUNG SC, DENG WP, YANG WK et al. Mesenchymal stem
cell targeting of microscopic tumors and tumor stroma devel-
opment monitored by noninvasive in vivo positron emission
tomography imaging. Clin. Cancer Res.2005; 11: 7749–7756.
KARNOUB AE, DASH AB, VO AP et al. Mesenchymal stem
cells within tumour stroma promote breast cancer metastasis.
Nature, 2007; 449: 557–563. doi:10.1038/nature06188
STUDENY M, MARINI FC, CHAMPLIN RE et al. Bone
marrow-derived mesenchymal stem cells as vehicles for
interferon-beta delivery into tumors, Cancer Res. 2002; 62:
NAKAMIZO A. MARINI F, AMANO T et al. Human bone
marrow-derived mesenchymal stem cells in the treatment of
gliomas. Cancer Res. 2005; 65: 3307–3318.
KAPLAN RN, PSAILA B, LYDEN D Bone marrow cells in
the‘pre-metastatic niche’: within bone and beyond, Cancer
Metastasis Rev 2006; 25: 521–529. doi:10.1007/s10555-006-
HAMADA H, KOBUNE M, NAKAMURA K et al. Mesen-
chymal stem cells (MSC) as therapeutic cytoreagents for gene
therapy, Cancer Sci. 2005; 96: 149–156. doi:10.1111/j.1349-
RAMASAMY R, LAM EW, SOEIRO I. et al. Mesenchymal
stem cells inhibit proliferation and apoptosis of tumor cells:
impact on in vivo tumor growth. Leukemia 2007; 21: 304–310.
OHLSSON LB, VARAS L, KJELLMAN C, et al. Mesenchy-
mal progenitor cell-mediated inhibition of tumor growth in
vivo and in vitro in gelatin matrix. Exp Mol Pathol. 2003; 75:
KHAKOO AY, PATI S, ANDERSON SA et al. Human mes-
enchymal stem cells exert potent antitumorigenic effects in a
model of Kaposi’s sarcoma, J. Exp. Med. 2006; 203: 1235–1247.
MIURA M, MIURA Y, PADILLA-NASH HM, et al. Accumulated
chromosomal instability in murine bone marrow mesenchymal
stem cells leads to malignant transformation. Stem Cells. 2006;
24: 1095–1103. doi:10.1634/stemcells.2005-0403
TOLAR J, NAUTA AJ, OSBORN MJ, et al. Sarcoma derived
from cultured mesenchymal stem cells. Stem Cells. 2007; 25:
 ZHOU YF, BOSCH-MARCE M, OKUYAMA H, et al.
Spontaneous transformation of cultured mouse bone mar-
row-derived stromal cells. Cancer Res. 2006; 66: 10849–10854.
RUBIO D, GARCIA-CASTRO J, MARTÍN MC, et al. Spon-
taneous human adult stem cell transformation. Cancer Res.
2005; 65: 3035–3039.
RØSLAND GV, SVENDSEN A, TORSVIK A, et al. Long-term
cultures of bone marrow-derived human mesenchymal stem
cells frequently undergo spontaneous malignant transforma-
tion. Cancer Res. 2009; 69: 5331–5339. doi:10.1158/0008-5472.
AGUILAR S, NYE E, CHAN J, et al. Murine but not human
mesenchymal stem cells generate osteosarcoma-like lesions
in the lung. Stem Cells. 2007; 25: 1586–1594. doi:10.1634/
TASSO R, AUGELLO A, CARIDA M et al. Development of
sarcomas in mice implanted with mesenchymal stem cells
seeded onto bioscaffolds. Carcinogenesis 2009; 30: 150–157.
KUCEROVA L, MATUSKOVA, M. HLUBINOVA, K et al.
Human adipose tissue-derived mesenchymal stromal cells as
modulators of tumor microenvironment. Molecular Cancer
HILDRESTRAND GA, DUGGAL S, BJØRÅS M, et al. Modu-
lation of DNA glycosylase activities in mesenchymal stem
cells. Exp Cell Res. 2009 May 25. [Epub ahead of print]
MIAO ZH, RAO VA, AGAMA K et al. 4-Nitroquinoline-
1-Oxide induces the formation of cellular topoisomerase
I-DNA cleavage complexes. Cancer Res. 2006; 66: 6540–6545.
NAGAO M, SUGIMURA T. Molecular biology of the car-
cinogen, 4-nitroquinoline 1-oxide. Adv Cancer Res 1976; 23:
WAGNER W, HORN P, CASTOLDI M, et al. Replicative
senescence of mesenchymal stem cells: a continuous and
organized process. PLoS One. 2008; 3: e2213. doi:10.1371/
WAGNER W, BORK S, HORN P, et al. Aging and replicative se-
nescence have related effectsonhumanstemandprogenitorcells.
PLoS One. 2009; 4: e5846. doi:10.1371/journal.pone.0005846
INOMATA K, AOTO T, BINH NT et al. Genotoxic stress
abrogates renewal of melanocyte stem cells by triggering
their differentiation.Cell.2009;137:1088–1099. doi:10.1016/
RUZANKINA Y, PINZON-GUZMAN C, Asare A, et al. Dele-
tion of the developmentally essential gene ATR in adult mice
leads to age-related phenotypes and stem cell loss. Cell Stem
Cell. 2007; 1: 113–126. doi:10.1016/j.stem.2007.03.002
INOUE-NARITA T, HAMADA K, SASAKI T, et al. Pten
and susceptibility to carcinogen-induced melanomagenesis.
Cancer Res. 2008; 68: 5760–5768. doi:10.1158/0008-5472.
NISHIMURA EK, GRANTER SR, FISHER DE Mechanisms of hair
graying: incomplete melanocyte stem cell maintenance in the niche.
Science. 2005; 307: 720–724. doi:10.1126/science.1099593