Clonogenic growth of human breast cancer cells co-cultured in direct contact with serum-activated fibroblasts (original) (raw)
Related papers
Multiple Changes Induced by Fibroblasts on Breast Cancer Cells
Connective Tissue Research, 2009
It is now widely recognized that the cross-talk between cancer and stromal cells may play a crucial role in cancer progression. However, little is known about the complex underlying molecular mechanisms that occur within the tumor microenvironment. Fibroblasts are the major stromal cells with multiple roles, especially toward both the extracellular matrix and the neighboring cell population, including neoplastic cells. Consequently, proteomic analyses would provide a wider resource for a better understanding of the potential modulating effects exerted by fibroblasts on cancer cells. In this article we describe the effects of fibroblast stimulation on the breast cancer cell line (8701-BC) proteomics, using a trans-well coculture system. Our results clearly indicate that fibroblasts induce considerable proteomic modulations on 8701-BC, mainly in the cytoskeleton proteins and glycolytic enzymes. Additionally, fibroblast-conditioned medium increased neoplastic cell proliferation and invasion with a concurrent upregulation of the c-Myc oncogene. Collectively these results suggest that fibroblast stimulation may enhance the malignant potential of breast cancer cells in vitro.
International Journal of Molecular Sciences
Breast cancer-associated fibroblasts (BCAFs), the most abundant non-cancer stromal cells of the breast tumor microenvironment (TME), dramatically sustain breast cancer (BC) progression by interacting with BC cells. BCAFs, as well as myofibroblasts, display an up regulation of activation and inflammation markers represented by α-smooth muscle actin (α-SMA) and cyclooxygenase 2 (COX-2). BCAF aggregates have been identified in the peripheral blood of metastatic BC patients. We generated an in vitro stromal model consisting of human primary BCAFs grown as monolayers or 3D cell aggregates, namely spheroids and reverted BCAFs, obtained from BCAF spheroids reverted to 2D cell adhesion growth after 216 h of 3D culture. We firstly evaluated the state of activation and inflammation and the mesenchymal status of the BCAF monolayers, BCAF spheroids and reverted BCAFs. Then, we analyzed the MCF-7 cell viability and migration following treatment with conditioned media from the different BCAF cult...
In Vitro Cellular & Developmental Biology - Animal, 1993
Breast carcinomas commonly contain varying amounts of fibrous stroma and infiltrates of lymphoid cells. Dickson and Lippman (Endocrine Rev., 8,29, 1987) have proposed a model of growth regulation in breast cancer involving interactions between stroma and carcinoma cells. This model is based on results obtained with established cell lines. In an effort to bring experimentation closer to the clinical situation we have used short-term primary cultures from human breast cancer in co-cultures with lymphocytes and fibroblasts. Cultures were established in a chemically defined serum-free medium (CDM3). Cell types were characterized on the basis of live morphology and expression of vimentin and keratin 18. A semi-quantitative system was developed for measuring growth of epithelial cells, thus defining two indices: maximal growth index (GI-max) and growth rate (GR). Moderate-to-good growth was obtained from 34 out of 46 carcinoma samples (74%) and 30 out of 38 parallel samples of non-cancerous tissue (79%). Success in culture was negatively correlated with the amount of hard stroma but unrelated to age of patient or clinical status. Malignant epithelium was clearly identified in 12 out of 34 (35%) carcinoma samples. For the evaluation of responses of epithelial cells in co-cultures, the cultures from each sample were ranked according to GI-max. From 20 co-culture experiments using carcinoma samples, the following results were obtained: the highest GI-max was found in 11 of the co-cuhures with lymphocytes; in six of the co-cultures with fibroblasts; in one case in the control culture without partner cells; and in two experiments there was no difference between controls and co-cultures. The corresponding values for non-cancerous samples were: 5 out of 17, 2/17, 2/17, and 8/17. Control experiments performed without partner cells confirmed that these differences in GI-max between cultures were beyond random variations. Four samples displayed particularly vigorous responses to lymphocytes, and two samples responded extensively to fibroblasts. In four of these six samples cancer ceils proliferated. We conclude that it is feasible to use primary cultures of breast carcinomas for experimentation. Fibroblasts did not have very marked effects on epithelial cell growth, but, contrary to expectation, there was a clear tendency for lymphocytes to stimulate growth.
BMC Cancer, 2010
Introduction Tumour phenotype is regulated in a complex fashion as a result of interactions between malignant cells and the tumour stroma. Fibroblasts are the most abundant and perhaps most active part of the tumour stroma. A better understanding of the changes that occur in fibroblasts in response to the presence of malignant cells may lead to the development of new strategies for cancer treatment. We explored the effects of fibroblasts on the growth and invasion of mammary carcinoma tumour cells in vitro and in vivo. Methods In order to analyse secreted factors that affect invasive abilities of breast cancer cells we co-cultured human mammary fibroblasts (HMF3s) and cancer cells (MCF7S1) in three-dimensional (3D) growth conditions devoid of heterogeneous cell-cell contact. To study the possible influence of fibroblasts on MCF7S1 cancer cell growth in vivo we co-injected HMF3s and MCF7S1 cells in Balb/c nu/nu mice. Results In 3D co-culture both HMF3s and MCF7S1 cells demonstrated e...
Influence of the interaction between nodal fibroblast and breast cancer cells on gene expression
Tumor Biology, 2011
Our aim was to evaluate the interaction between breast cancer cells and nodal fibroblasts, by means of their gene expression profile. Fibroblast primary cultures were established from negative and positive lymph nodes from breast cancer patients and a similar gene expression pattern was identified, following cell culture. Fibroblasts and breast cancer cells (MDA-MB231, MDA-MB435, and MCF7) were cultured alone or co-cultured separated by a porous membrane (which allows passage of soluble factors) for comparison. Each breast cancer lineage exerted a particular effect on fibroblasts viability and transcriptional profile. However, fibroblasts from positive and negative nodes had a parallel transcriptional behavior when co-cultured with a specific breast cancer cell line. The effects of nodal fibroblasts on breast cancer cells were also investigated. MDA MB-231 cells viability and migration were enhanced by the presence of fibroblasts and accordingly, MDA-MB435 and MCF7 cells viability followed a similar pattern. MDA-MB231 gene expression profile, as evaluated by cDNA microarray, was influenced by the fibroblasts presence, and HNMT, COMT, FN3K, and SOD2 were confirmed downregulated in MDA-MB231 co-cultured cells with fibroblasts from both negative and positive nodes, in a new series of RT-PCR assays. In summary, transcriptional changes induced in breast cancer cells by fibroblasts from positive as well as negative nodes are very much alike in a specific lineage. However, fibroblasts effects are distinct in each one of the breast cancer lineages, suggesting that the inter-relationships between stromal and malignant cells are dependent on the intrinsic subtype of the tumor.
BMC Veterinary Research, 2012
Background: It is supposed that fibroblasts present in tumour microenvironment increase cancer invasiveness and its ability to metastasize but the mechanisms have not been clearly defined yet. Thus, the current study was designed to assess changes in gene expression in five various cancer cell lines grown as a co-culture with the carcinoma-associated fibroblasts (CAFs) in vitro. Results: A carcinoma-associated fibroblast cell line was isolated from a canine mammary cancer. Then, a co-culture of cancer cells with the CAFs was established and maintained for 72 hrs. Having sorted the cells, a global gene expression in cancer cells using DNA microarrays was examined. The analysis revealed an up-regulation of 100 genes and a down-regulation of 106 genes in the cancer cells grown as a co-culture with the CAFs in comparison to control conditions. The PANTHER binomial statistics tool was applied to determine statistically over-manifested pathways (p < 0.05). Bulk of the up-regulated genes are involved in the adhesion, the angiogenesis, the epithelialmesenchymal transition (EMT) and generally take part in the developmental processes. These results were further confirmed using real-time qPCR. Moreover, a wound-healing assay and growth characteristics on Matrigel matrix showed that CAFs increase cancer cell migration and matrix invasion.
A FTIR Imaging Characterization of Fibroblasts Stimulated by Various Breast Cancer Cell Lines
PLoS ONE, 2014
It is well known that the microenvironment plays a major role in breast cancer progression. Yet, the mechanism explaining the transition from normal fibroblasts to cancer-stimulated fibroblasts remains to be elucidated. Here we report a FTIR imaging study of the effects of three different breast cancer cell lines on normal fibroblasts in culture. Fibroblast activation process was monitored by FTIR imaging and spectra compared by multivariate statistical analyses. Principal component analysis evidenced that the fibroblasts stimulated by these cancer cell lines grouped together and remained distinctly separated from normal fibroblasts indicating a modified different chemical composition in the cancer-stimulated fibroblasts. Similar changes in fibroblasts were induced by the various breast cancer cell lines belonging to different sub-types. Most significant changes were observed in the region of 2950 and 1230 cm 21 , possibly related to changes in lipids and in the 1230 cm 21 area assigned to phosphate vibrations (nucleotides). Interestingly, the cancer-cell induced changes in the fibroblasts also occurred when there was no possible direct contact between the two cell lines in the co-culture. When contact was possible, the spectral changes were similar, suggesting that soluble factors but not direct cell-cell interactions were responsible for fibroblast activation. Overall, the results indicate that IR imaging could be used in the future for analyzing the microenvironment of breast tumors.
Clinical & Experimental Metastasis, 2011
Fibroblast activation protein-α (FAP) is a cell surface, serine protease of the post-prolyl peptidase family that is expressed in human breast cancer but not in normal tissues. Previously, we showed that FAP expression increased tumor growth rates in a mouse model of human breast cancer. Here the role of the proteolytic activities of FAP in promoting tumor growth, matrix degradation and invasion was investigated. Mammary fat pads of female SCID mice were inoculated with breast cancer cells that express FAP and the mice treated with normal saline or Val-boroPro (talabostat); Glu-boroPro (PT-630); or 1-[[(3-hydroxy-1-adamantyl)amino]acetyl]-2-cyano-(S)-pyrrolidine (LAF-237) that inhibit prolyl peptidases. Other mice were injected with breast cancer cells expressing a catalytically inactive mutant of FAP and did not receive inhibitor treatment. PT-630 and LAF-237 did not slow growth of tumors produced by any of the three cell lines expressing FAP. Talabostat slightly decreased the growth rates of the FAP-expressing tumors but because PT-630 and LAF-237 did not, the growth retardation was likely not related to the inhibition of FAP or the related post-prolyl peptidase dipeptidyl peptidase IV. Breast cancer cells expressing a catalytically inactive mutant of FAP (FAPS624A) also produced tumors that grew rapidly. In vitro studies revealed that cells expressing wild type FAP or FAPS624A degrade extracellular matrix (ECM) more extensively, accumulate higher levels of matrix metalloproteinase-9 (MMP-9) in conditioned medium, are more invasive in type I collagen gels, and have altered signaling compared to control transfectants that do not express FAP and form slow growing tumors. We conclude that the proteolytic activity of FAP participates in matrix degradation, but other functions of the protein stimulate increased tumor growth.