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Breast Cancer Program, Karmanos Cancer Institute [M. P. V. S., J. W., S. S., R. J. P., L. T.], and Departments of Pathology [M. P. V. S., L. T.] and Internal Medicine [R. J. P.], Wayne State University School of Medicine, Detroit, Michigan 48201
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Introduction |
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Because commitment to the morphogenetic and differentiation programs requires the establishment of intercellular communication between breast stromal and epithelial cells, we have established a novel three-dimensional cell-cell interaction model to study the molecular and cellular basis of epithelial-fibroblast-endothelial cell interactions. In this experimental system, we have compared the abilities of specific mesenchymal cell types and HUVECs to induce three-dimensional morphogenesis and growth of normal-behaving MCF10A and preneoplastic MCF10AT1-EIII8 human breast epithelial cells. Our results not only demonstrate a requirement for breast-specific fibroblasts but also show the dominant manner by which normal (reduction mammoplasty) and tumor-derived breast fibroblasts suppress or induce, respectively, growth and ductal-alveolar morphogenesis of MCF10A and MCF10AT1-EIII8 breast epithelial cells.
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Materials and Methods |
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HUVECs purchased from ATCC at passage 13 were maintained in SFM (Life Technologies, Inc., Grand Island, NY) supplemented with 10 ng/ml EGF, 20 ng/ml bFGF, and 10 µg/ml fibronectin. Cultures were maintained at 37°C in a humidified atmosphere containing 5% CO2 in air and cultured up to 15 passages.
Primary cultures of human breast fibroblasts were established from reduction mammoplasty tissues, tumors, and benign tissues ipsilateral but distal to the tumors acquired after protocol review and approval by the Wayne State University human investigation committee (16) . Tumors, confirmed by histology, were finely minced and digested overnight with collagenase (150 IU/ml) in DMEM/Ham’s F-12 medium (1:1) containing 20% calf serum. Larger pieces were allowed to settle, and cells in the supernatant, predominantly fibroblasts, were recovered by centrifugation and plated in Waymouth’s MB752 supplemented with 10 mmol/liter HEPES and 15% FCS (FIB; Ref. 16 ). Reduction mammoplasty and benign tissues were digested as above with the addition of hyaluronidase (100 IU/ml) and were further processed as described for tumors. Portions of reduction mammoplasty and benign tissues used for culture were examined histologically to confirm the absence of neoplasia. Fibroblasts were routinely cultured up to 15 passages and were used at passages 4 to 6. Fibroblasts were characterized immunocytochemically with monoclonal antibodies to pan cytokeratin (Dako Corp., Carpintaria, CA), cytokeratin 14 (Novacastra Laboratories, Ltd., Newcastle upon Tyne, United Kingdom), and vimentin (V9; Dako Corp.) as described in Ref. 16 . Normal human lung fibroblasts (IMR-90) and the mesenchymally derived malignant human fibrosarcoma cell line HT1080 were obtained from ATCC and maintained in Eagle’s MEM supplemented with 2 mM L-glutamine, 0.1 mM nonessential amino acids, 1.0 mM sodium pyruvate, and 10% fetal bovine serum. The NIH 3T3 mouse embryo fibroblast line was purchased from ATCC and maintained in DMEM supplemented with 4 mM glutamine, 4.5 g/liter glucose, 1.0 mM sodium pyruvate, and 10% FCS.
Homotypic and Heterotypic Three-Dimensional Basement Membrane
Culture of Epithelial, Fibroblast, and Endothelial Cells.
Homotypic three-dimensional cultures were set up as described in Ref.
14
. Briefly, 10 x 104 cells were seeded as a single-cell suspension
in eight-well chamber slides coated with Matrigel (Collaborative
Biomedical Products, Bedford, MA) in DMEM-F12-supplemented medium for
MCF10A/EIII8 cells, in SFM-supplemented medium for HUVECs, in FIB
medium for human breast fibroblasts, or in the appropriate medium for
IMR-90, HT1080, or NIH 3T3 cells as described above. For heterotypic
cocultures, 50 x 103
MCF10A or
EIII8 cells were mixed with an equal number of human breast fibroblasts
(reduction mammoplasty, benign or tumor-derived), IMR-90, HT1080, or
NIH 3T3 cells. For heterotypic tricultures, MCF10A or EIII8 cells were
mixed with equivalent numbers of human breast fibroblasts (reduction
mammoplasty, benign or tumor-derived) and HUVECs and seeded onto
chamber slides coated with Matrigel as described for homotypic
cultures. Typically, heterotypic cultures were performed in SFM
supplemented with EGF and bFGF because it allows optimal viability,
growth, and three-dimensional organization of HUVECs, fibroblasts, and
MCF10A cells. Heterotypic cocultures were routinely maintained for up
to 3 weeks, and morphological development was analyzed by
phase-contrast microscopy.
Quantitation of Effects of Interaction of Preneoplastic Breast
Epithelial Cells with Reduction Mammoplasty/Tumor-derived Breast
Fibroblasts and Endothelial Cells on Three-Dimensional Growth.
Three-dimensional cultures were set up by seeding 50 x 103
EIII8, HUVECs, 31R or 14R (reduction
mammoplasty fibroblasts), and 38T or 12T (tumor-derived breast
fibroblasts) either alone or by mixing 50 x 103
EIII8 cells with an equivalent number of 31R
(or 14R) or 38T (or 12T) in the absence or presence of an equivalent
number of HUVEC cells onto chamber slides coated with Matrigel as
described above. Cultures were incubated at 37°C for 6 days,
after which the cell viability was measured in the three-dimensional
cultures. The medium was removed, wells were rinsed with PBS, and the
Matrigel was digested with dispase for 2 h at 37°C. The digested
material was centrifuged at 4000 x g for 10
min, and the pellet was treated with trypsin to recover single cells
from the three-dimensional structures. The number of viable cells was
determined by trypan blue exclusion in a hemocytometer
(14)
, and results were expressed as the average ± SE from three independent experiments. The effects of
E2 and the pure antiestrogen ICI 182,780 (a gift
from Dr. A. Wakeling, Zeneca Pharmaceuticals, Cheshire, United Kingdom)
on three-dimensional growth of heterotypic cultures of EIII8–31R-HUVEC
and EIII8–38T-HUVEC cells were tested as described above, except that
after seeding of cells, slides were incubated overnight to allow cells
to attach to the surface and were treated with vehicle (0.01% ethanol,
v/v), 100 nM ICI 182,780, or 1 or 10
nM E2 (Sigma Chemical Co.,
St. Louis, MO) or with a combination of 1 nM
E2 and 100-fold molar excess of ICI
182,780.
Morphological Evaluation of Three-Dimensional Cultures.
Whole mounts of EIII8–12T cocultures were prepared by staining
formalin-fixed cultures with toluidine blue and were mounted on slides
with Permount (17)
. For histological evaluation,
three-dimensional cocultures (EIII8–12T or EIII8–12T-HUVEC) were
fixed in buffered formalin and embedded in paraffin, and 4-µm
sections were stained in H&E. For immunohistochemical evaluation of
breast epithelial, fibroblast, and endothelial cell functional markers,
sections were incubated with monoclonal antibodies directed to the
following human proteins: pan cytokeratin 5/6/8/18, a cocktail of
monoclonal antibodies designed to recognize epithelial cells and their
tumors (Novacastra Laboratories, Ltd.), vimentin (V9; Dako Corp.),
muc-1 glycoprotein (a mammary-type apomucin also known as milk
fat globule membrane antigen; Novacastra Laboratories, Ltd.), cd31 or
PECAM-1 (Dako Corp.), Factor VIII-related antigen (Dako Corp.), Ki-67
(Dako Corp.), VEGF receptor-2 (Chemicon International, Inc., Tamecula,
CA), 
vß3 integrin
(Chemicon International, Inc.), MMP-9 (Oncogene Science, Cambridge,
MA), and rabbit polyclonal antibody to VEGF (Oncogene Science). In each
case, negative controls were overlaid with appropriate mouse or rabbit
IgG isotype. The slides were overlaid with avidin-biotin-conjugated
goat antimouse or antirabbit IgG (Vectastain ABC kit; Vector
Laboratories, Burlingame, CA), developed with peroxidase substrate, and
counterstained by Mayer’s hematoxylin (Sigma Chemical Co.).
Statistical Analysis.
Specific differences in growth between cocultures of EIII8 cells with
reduction mammoplasty or tumor-derived breast fibroblasts in the
absence and presence of endothelial cells, and those affected by
hormonal treatments were examined using Student’s t test.
Statistical significance was determined using Student’s t
test with P < 0.02 considered as
statistically significant.
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Results |
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TopABSTRACTIntroductionMaterials and MethodsResultsDiscussionREFERENCES |
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, panels A and E; Ref.
14
). In contrast, MCF10A or EIII8 cells, seeded on
Matrigel in contact-dependent cocultures with fibroblasts derived from
reduction mammoplasty or breast tumor tissues, produce sealed
structures consisting of centrally located fibroblasts with epithelial
buds emerging from the central fibroblast core (Fig. 1, A and B)
. When cocultured with MCF10A or EIII8 cells,
reduction mammoplasty fibroblasts 31R or 14R retain their capacity for
chemotaxis but are unable, or only weakly able, to support growth and
differentiation of MCF10A and EIII8 cells, respectively (Fig. 1A
, panels B and F). In contrast,
cocultures of MCF10A or EIII8 cells with 12T tumor-derived (or 12B
benign), or 38T tumor-derived (or 38B benign) breast fibroblasts,
besides concentrating cells by chemotaxis, induce growth and
ductal-alveolar morphogenesis of both MCF10A and EIII8 cells (Fig.
1B, panels A, C, and E).
However, a principal difference between the MCF10A and EIII8 cultures
is the formation of a larger number of highly proliferative,
ductal-alveolar units with EIII8 cells as compared with those formed by
MCF10A cells (Fig. 1B, compare panel A with
C and E). These results indicate major
differences in the ability of fibroblasts derived from reduction
mammoplasty versus benign (data not shown) or tumor breast
tissues to support the growth and morphogenesis of normal and
preneoplastic breast epithelial cells.
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,
panel C, and Fig. 1B
, panel B), or
EIII8 cocultures with reduction mammoplasty fibroblasts (Fig. 1A
, panel G) failed to alter the growth
characteristics induced by fibroblasts. In contrast, the inclusion of
HUVECs into EIII8 cocultures with fibroblasts from benign/tumor tissues
(12B, 12T, 38B, or 38T) resulted in a dramatic (Fig. 1B
,
panels D and F) induction of branching
ductal-alveolar morphogenesis. These results are consistent with our
previous data that showed that, unlike normal MCF10A cells,
preneoplastic EIII8 cells have the capacity to support endothelial
survival and growth (14)
. Similar cocultures set up with
HT1080 fibrosarcoma cells, normal IMR-90 lung cells, or
embryonal NIH 3T3 fibroblasts failed to elicit similar effects on
breast epithelial growth and morphogenesis as those induced by human
breast fibroblasts (data not shown).
Quantitative Evaluation of the Effects of Fibroblasts and
Endothelial Cells on Three-Dimensional Growth of EIII8 Cells, and EIII8
Responsiveness to Estrogen.
Because the results from Fig. 1, A and B
,
revealed remarkable differences in the abilities of fibroblasts from
reduction mammoplasty and breast tumor tissues to mediate
three-dimensional organization and morphogenesis of EIII8 cells and
further induction by endothelial cells, we measured cell growth in
homotypic (EIII8, 31R, 38T, HUVEC) and heterotypic (EIII8–31R,
EIII8–38T) cocultures, both in the presence and absence of endothelial
cells, by direct cell counting of viable cells from dispase-digested
Matrigel. Consistent with the visual effects on growth and morphology
of the cocultures observed in Fig. 1, A and B
,
contact-dependent coculture of EIII8 with 38T tumor-derived breast
fibroblasts elicited a 3-fold increase in growth
(P < 0.001) over corresponding cultures with
31R reduction mammoplasty fibroblasts (Fig. 2A)
. Inclusion of HUVECs into EIII8–38T cocultures caused
further enhancement of cell proliferation that was 
1.7-fold higher
(P < 0.01) than EIII8–38T cocultures and
3.5-fold higher (P < 0.005) than
EIII8–31R-HUVEC tricultures. These data are not unique to 38T or 31R
fibroblasts, because similar effects on growth were observed with 12T
tumor-derived fibroblasts or 14R reduction mammoplasty fibroblasts
(data not shown).
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. Similar treatment of EIII8
tricultures containing 38T tumor-derived breast fibroblasts invoked
only a marginal increase in growth (P < 0.03) that was apparent only with 10 nM
E2 (Fig. 2B)
. However, the inductory
effects of exogenous E2 became more obvious on
addition of the pure estrogen antagonist ICI 182,780 because it
significantly inhibited (P < 0.001) growth
in both vehicle- and E2-treated cultures (Fig. 2B)
. Because no E2-mediated
stimulatory effects were observed in EIII8 cocultures with reduction
mammoplasty fibroblasts, these results suggest that the
E2-induced growth effects observed in
vehicle-treated EIII8–38T-HUVEC tricultures were not caused by
contaminating estrogen or estrogenic compounds in the phenol red-free
culture media but probably originated from the tumor-derived
fibroblasts. From these data, it can be inferred that breast
fibroblasts play a dominant role in the maintenance and induction of
epithelial growth and morphogenesis: on the one hand, fibroblasts from
normal breast tissues exert growth inhibitory effects that can override
genetic constraints imposed by preneoplastic breast epithelial cells
whereas, on the other hand, tumor-derived breast fibroblasts exercise
growth stimulatory effects that can overcome or augment the genetic
constraints imposed by normal or preneoplastic breast epithelial cells,
respectively.
Contact-dependent Existence with Tumor-derived Breast Fibroblasts
Facilitate Structural and Functional Organization That Is Reminiscent
of the in Vivo Situation.
The results seen in Figs. 1
and 2
demonstrate that benign- or
tumor-derived breast fibroblasts exert inductive branching morphogenic
effects on preneoplastic EIII8 cells that is further enhanced by the
presence of endothelial cells. Histological analysis of H&E-stained
paraffin-embedded EIII8 cocultures revealed the presence of a central
core of fibroblasts from which, and around which, branching epithelial
buds emerge (Figs. 3
and 4)
confirming the data from whole-mount (Fig. 3A)
and
phase-contrast microscopy (Fig. 1, A and B)
.
Further confirmatory biochemical evidence for the localization and
functionality of epithelial, fibroblast, and endothelial components was
obtained by immunochemical analysis of expression of relevant markers:
cytokeratins and muc-1 (epithelial), vimentin (fibroblast), cd31,
Factor VIII, VEGF receptor-2, and
vß3 integrin
(endothelial cells). Results in Fig. 3
show that the majority of the
cytokeratin-positive muc-1-expressing epithelial cells (Fig. 3, F and G)
are organized as buds around the central
vimentin-positive fibroblast core (Fig. 3H)
, whereas a small
fraction of epithelial cells are retained in the stromal core (Fig. 3, F and G)
. This residual epithelial cell
population may represent "fragments of ducts emerging from the
core" or cells that did not participate in the organization process.
The epithelial structures exhibit pronounced branching and increased
potential to invade and degrade the coincident ECM in EIII8 cocultures
containing endothelial cells (Fig. 4A)
. This ability to
degrade the ECM is consistent with the strong reactivity to MMP-9
antibody in the immediate vicinity of epithelial buds in
EIII8–12T-HUVEC tricultures (Fig. 4, E and F)
.
The ability of epithelium to invade and proteolytically degrade the
surrounding ECM requires the presence of endothelial cells because
EIII8 cocultures, containing only benign- or tumor-derived fibroblasts,
lack this property (data not shown). It is interesting to note that the
central fibroblast core in EIII8 cocultures is nonproliferating in
contrast to the high Ki-67 labeling of epithelial buds (Fig. 3E)
, which suggests that the metabolically active yet
nonproliferating fibroblasts are sufficient to serve as a structural
platform for maintenance and support of epithelial survival, growth,
and morphogenesis (Fig. 3)
. An interesting feature observed in the
EIII8 tricultures is the establishment of contact between epithelial,
fibroblast, and endothelial cells that is most suitable for paracrine
interaction. In EIII8–12T-HUVEC tricultures, the cd31-positive,
factor-VIII-expressing endothelial cells are situated in the central
stromal core but predominantly close to the base of the branching
epithelial buds (Fig. 4, A–D)
. These vascular units are
functional because they not only express factor VIII but are also
positive for the endothelial-specific VEGF receptor-2 (Flk-1) and
vß3 integrin, markers
that are indicative of active angiogenesis (Fig. 4, H and I)
. The maintenance of endothelial cell survival and growth
appears to be ensured by large amounts of VEGF expressed and secreted
by EIII8 epithelium because intense immunoreactivity to VEGF antibody
is observed in the epithelial compartment, whereas only focal
immunostaining to VEGF antibody (which is similar to staining patterns
of Flk-1 and vß3
integrin) is observed in the central stromal core (Fig. 4G)
.
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Discussion |
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TopABSTRACTIntroductionMaterials and MethodsResultsDiscussionREFERENCES |
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Our data also show that only breast fibroblasts, but not soft tissue sarcoma-derived malignant mesenchymal cells, normal lung, or embryonal 3T3 fibroblasts, confer morphogenesis-inducing effects. Thus, although the cultures are performed in Matrigel, a reconstituted basement membrane matrix containing a variety of ECM molecules (23) , induction of breast epithelial growth and morphogenesis appears to be mediated by the ECM that is laid down by organ-specific mesenchyme. The ability of normal fibroblasts to convert or revert malignant tumor lines such as basal cell carcinoma of skin (24) and prostatic adenocarcinoma (25) into morphologically benign (24 , 25) or biologically less aggressive cell populations has been demonstrated (25) . Our data show that, although normal reduction mammoplasty fibroblasts can inhibit or retard morphological transformation of both normal MCF10A and preneoplastic EIII8 breast epithelial cells, respectively, tumor-derived breast fibroblasts can override or augment genetic constraints imposed by MCF10A or EIII8 cells, respectively, causing them to undergo ductal-alveolar morphogenesis. This difference in the activities between normal and tumor-derived fibroblasts cannot be accounted for by variations in the ages of cells in culture, because both sets of fibroblasts grew equally well at the passages four to six used for coculture experiments. Although the normal and tumor-derived fibroblasts were derived from donors with mean ages of 30 and 48 years, respectively, deviations in the activities of the fibroblasts cannot be ascribed to differences in age because both normal and tumor-derived fibroblasts exhibited sample-to-sample variations in growth rates that overwhelmed any age-related differences in growth.
Alterations in fibroblasts in the stroma immediately adjacent to transformed epithelial cells have been documented in several tumor systems (26 , 27) . Desmoplasia, a profound stromal response consisting of changes in cellular composition and ECM, is observed in infiltrating ductal carcinoma (28) . Differences in the activity of normal versus tumor-derived fibroblasts may result from variations in the establishment of reciprocal communication between epithelial and fibroblast compartments, which, in turn, could result from differences in molecular and/or cellular mechanisms that are responsible for the production and release of a number of soluble paracrine factors such as FGF(s) (29) , transforming growth factor ß (30) , IGFs (27) , and hepatocyte growth factor (31) , and/or the ability to respond to epithelial-derived signals. Although, potentially, any of the growth factors derived from breast fibroblasts may function as an important regulator of mammary tumor growth, IGFs, and IGF-II in particular, may play a key role in mediating breast tumor growth because IGF-II expression has been reported in the stroma of invasive breast cancers, but not in normal breast (27) , and in the stroma of N-nitrosomethylurea-induced rat mammary tumors (32) . Whereas a principal role for the stroma has often been proposed in mediating steroid action during growth and differentiation in many adult tissues (3) , our study shows that normal fibroblasts have the ability to suppress E2-induced growth of estrogen receptor-positive EIII8 cells (14) , whereas tumor-derived fibroblasts support E2-induced growth of EIII8 cells. Although the addition of 10 nM E2 was required to see a marginal increase in growth over that of control cultures, the E2-induced effect is estrogen receptor-mediated because the addition of the pure estrogen antagonist ICI 182,780 effectively blocked growth in both vehicle-treated and E2-treated cultures. These data suggest that the marginal induction of growth by exogenous estrogen may be attributable to the presence of significant levels of E2 in the control cultures that are probably derived from tumor-derived fibroblasts via aromatase-mediated synthesis of estrogen (16 , 33 , 34) .
An alternate possibility for differences between reduction mammoplasty and tumor-derived fibroblasts may be the presence of genetic abnormalities in the background supportive stroma, i.e., the benign/tumor fibroblasts. Moinfar et al. (9) have recently reported frequent allelic loss in the stroma close to breast cancer, ranging from 10 to 66.5% for ductal carcinoma in situ and from 20 to 75% for invasive ductal cancers, and the absence of loss of heterozygosity either in epithelial or in stromal components in women without any breast disease (9) . Thus, the presence of genetic alterations present in preneoplastic EIII8 cells (which is akin to many benign or early epithelial neoplasms), combined with the presence of a genetically altered stroma (such as that which may be present in benign or malignant tumors of the breast), may underlie the acute inductive responses observed in EIII8–12T or EIII8–38T cocultures. These data are further supported by the absence of similar effects in EIII8-reduction mammoplasty fibroblast cocultures and the occurrence of inductive response in MCF10A-tumor fibroblast cocultures. Stromal-epithelial interaction has a fundamental role in determining normal duct development. Schor et al. (35) have suggested that perturbations in epithelial-mesenchymal interactions caused by the presence of fibroblast subpopulations with "fetal-like" phenotypic properties in breast carcinomas and in histologically normal breast tissue adjacent to a carcinoma may significantly enhance the susceptibility of epithelium to develop cancer. Aberrations in stromal-epithelial interactions resulting from differences in the production and release of growth factors, motility factors, and/or ECM molecules that may potentially ensue from genetic alterations in the stromal compartment may play a vital role in the development and progression of human breast cancer.
It is interesting to note that survival and functionality of endothelial cells are observed only in EIII8 cocultures, because the addition of HUVECs to EIII8–12T or EIII8–38T cocultures resulted in dramatic augmentation of proliferation and branching ductal-alveolar morphogenesis that was accompanied by an increase in invasion and degradation of the coincident ECM and up-regulation of MMP-9. These data are consistent with our previous report (14) that the maintenance of endothelial cell survival and function is promoted by preneoplastic, but not by normal, epithelial cells because the former responds to the presence of endothelial cells with the synthesis and release of the endothelial cell survival factor VEGF (14 , 36 , 37) . These data imply that tumor fibroblasts play an active role as a morphogenic and mitogenic inducer of epithelial cells; however, further enhancement of tumor growth and progression requires the establishment of active angiogenesis.
Our data suggest that therapeutic targeting of stromal cells rather than (or as well as) cancer cells may be beneficial because they appear to have a dominant capacity to modulate and control epithelial morphogenesis and mitogenesis. The inhibition of tumor stroma may result in more efficient tumor regression and treatment by depriving the tumor cells of an essential structural and functional support system. In summary, we have established a novel coculture system that reconstitutes many of the functional interactions between breast epithelial and stromal cells. This assay system will provide us with a unique opportunity to characterize mechanisms regulating breast stromal-epithelial cell interactions.
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ACKNOWLEDGMENTS |
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FOOTNOTES |
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1 Supported by grants from the United States Army
Medical Research and Materiel Command [DAMD17-94-J-4427 and
DAMD17-99-I-9443 (to M. P. V. S) and DAMD17-97-1-7173 to
(R. J. P)] and by NIH Grants CA60881 (to M. P. V. S) and CA22453,
Cancer Center Support Grant (to the Karmanos Cancer Institute). 
2 To whom requests for reprints should be
addressed, at Breast Cancer Program, Karmanos Cancer Institute, 110
East Warren Avenue, Detroit, MI 48201. Phone: (313) 833-0715, extension
2326/2259; Fax: (313) 831-7518; E-mail: shekharm{at}karmanos.org
3 The abbreviations used are: ECM, extracellular
matrix; HUVEC, human umbilical vein endothelial cell; E2,
17ß-estradiol; ATCC, American Type Culture Collection; SFM,
endothelial serum-free basal growth medium; EGF, epidermal growth
factor; bFGF, basic fibroblast growth factor; VEGF, vascular EGF; IGF,
insulin-like growth factor.
Received 7/18/00. Accepted 1/ 3/01.
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 C L Wilson, A H Sims, A Howell, C J Miller, and R B Clarke Effects of oestrogen on gene expression in epithelium and stroma of normal human breast tissue. Endocr. Relat. Cancer, June 1, 2006; 13(2): 617 - 628. [Abstract] [Full Text] [PDF]
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D. E. Costea, K. Kulasekara, E. Neppelberg, A. C. Johannessen, and O. K. Vintermyr Species-Specific Fibroblasts Required for Triggering Invasiveness of Partially Transformed Oral Keratinocytes Am. J. Pathol., June 1, 2006; 168(6): 1889 - 1897. [Abstract] [Full Text] [PDF]
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 K Ishiguro, T Yoshida, H Yagishita, Y Numata, and T Okayasu Epithelial and stromal genetic instability contributes to genesis of colorectal adenomas Gut, May 1, 2006; 55(5): 695 - 702. [Abstract] [Full Text] [PDF]
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M. V. Maffini, J. M. Calabro, A. M. Soto, and C. Sonnenschein Stromal Regulation of Neoplastic Development: Age-Dependent Normalization of Neoplastic Mammary Cells by Mammary Stroma Am. J. Pathol., November 1, 2005; 167(5): 1405 - 1410. [Abstract] [Full Text] [PDF]
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 E Sivridis, A Giatromanolaki, and M I Koukourakis Proliferating fibroblasts at the invading tumour edge of colorectal adenocarcinomas are associated with endogenous markers of hypoxia, acidity, and oxidative stress J. Clin. Pathol., October 1, 2005; 58(10): 1033 - 1038. [Abstract] [Full Text] [PDF]
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 S. Parrinello, J.-P. Coppe, A. Krtolica, and J. Campisi Stromal-epithelial interactions in aging and cancer: senescent fibroblasts alter epithelial cell differentiation J. Cell Sci., February 1, 2005; 118(3): 485 - 496. [Abstract] [Full Text] [PDF]
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 V. Laurent-Matha, S. Maruani-Herrmann, C. Prebois, M. Beaujouin, M. Glondu, A. Noel, M. L. Alvarez-Gonzalez, S. Blacher, P. Coopman, S. Baghdiguian, et al. Catalytically inactive human cathepsin D triggers fibroblast invasive growth J. Cell Biol., January 31, 2005; 168(3): 489 - 499. [Abstract] [Full Text] [PDF]
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K. Fukino, L. Shen, S. Matsumoto, C. D. Morrison, G. L. Mutter, and C. Eng Combined Total Genome Loss of Heterozygosity Scan of Breast Cancer Stroma and Epithelium Reveals Multiplicity of Stromal Targets Cancer Res., October 15, 2004; 64(20): 7231 - 7236. [Abstract] [Full Text] [PDF]
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 J. E. Maglione, E. T. McGoldrick, L. J.T. Young, R. Namba, J. P. Gregg, L. Liu, D. Moghanaki, L. G. Ellies, A. D. Borowsky, R. D. Cardiff, et al. Polyomavirus middle T-induced mammary intraepithelial neoplasia outgrowths: Single origin, divergent evolution, and multiple outcomes Mol. Cancer Ther., August 1, 2004; 3(8): 941 - 953. [Abstract] [Full Text] [PDF]
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M. Elkin, I. Cohen, E. Zcharia, A. Orgel, Z. Guatta-Rangini, T. Peretz, I. Vlodavsky, and H. K. Kleinman Regulation of Heparanase Gene Expression by Estrogen in Breast Cancer Cancer Res., December 15, 2003; 63(24): 8821 - 8826. [Abstract] [Full Text] [PDF]
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J. Walter-Yohrling, X. Cao, M. Callahan, W. Weber, S. Morgenbesser, S. L. Madden, C. Wang, and B. A. Teicher Identification of Genes Expressed in Malignant Cells That Promote Invasion Cancer Res., December 15, 2003; 63(24): 8939 - 8947. [Abstract] [Full Text] [PDF]
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N. Matsumoto, T. Yoshida, and I. Okayasu High Epithelial and Stromal Genetic Instability of Chromosome 17 in Ulcerative Colitis-associated Carcinogenesis Cancer Res., October 1, 2003; 63(19): 6158 - 6161. [Abstract] [Full Text] [PDF]
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 F. Mannello and M. Sebastiani Zymographic Analyses and Measurement of Matrix Metalloproteinase-2 and -9 in Nipple Aspirate Fluids Clin. Chem., September 1, 2003; 49(9): 1546 - 1550. [Full Text] [PDF]
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C. Corsini, P. Mancuso, S. Paul, A. Burlini, G. Martinelli, G. Pruneri, and F. Bertolini Stroma Cells: A Novel Target of Herceptin Activity Clin. Cancer Res., May 1, 2003; 9(5): 1820 - 1825. [Abstract] [Full Text] [PDF]
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H Chauhan, A Abraham, J R A Phillips, J H Pringle, R A Walker, and J L Jones There is more than one kind of myofibroblast: analysis of CD34 expression in benign, in situ, and invasive breast lesions J. Clin. Pathol., April 1, 2003; 56(4): 271 - 276. [Abstract] [Full Text] [PDF]
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 H. Rumpold, K. Mascher, G. Untergasser, E. Plas, M. Hermann, and P. Berger Trans-Differentiation of Prostatic Stromal Cells Leads to Decreased Glycoprotein Hormone {alpha} Production J. Clin. Endocrinol. Metab., November 1, 2002; 87(11): 5297 - 5303. [Abstract] [Full Text] [PDF]
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K. S. Wilson, H. Roberts, R. Leek, A. L. Harris, and J. Geradts Differential Gene Expression Patterns in HER2/neu-Positive and -Negative Breast Cancer Cell Lines and Tissues Am. J. Pathol., October 1, 2002; 161(4): 1171 - 1185. [Abstract] [Full Text] [PDF]
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M. M. Serewko, C. Popa, A. L. Dahler, L. Smith, G. M. Strutton, W. Coman, A. J. Dicker, and N. A. Saunders Alterations in Gene Expression and Activity during Squamous Cell Carcinoma Development Cancer Res., July 1, 2002; 62(13): 3759 - 3765. [Abstract] [Full Text] [PDF]
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 D. Briard, D. Brouty-Boye, B. Azzarone, and C. Jasmin Fibroblasts from Human Spleen Regulate NK Cell Differentiation from Blood CD34+ Progenitors Via Cell Surface IL-15 J. Immunol., May 1, 2002; 168(9): 4326 - 4332. [Abstract] [Full Text] [PDF]
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C. Berking, R. Takemoto, H. Schaider, L. Showe, K. Satyamoorthy, P. Robbins, and M. Herlyn Transforming Growth Factor-{beta}1 Increases Survival of Human Melanoma through Stroma Remodeling Cancer Res., November 1, 2001; 61(22): 8306 - 8316. [Abstract] [Full Text] [PDF]
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