Gene expression signature of primary breast tumour stromal cells
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Tumour stromal cells secrete proangiogenic and prometastatic factors, such as MMP3 and may support breast cancer progression through promotion of tumour angiogenesis and invasion
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Gene expression signature of primary breast tumour stromal cells
- 1. Gene expression signature of primary breast tumour stromal cells MC Hartmann, RM Dwyer and MJ Kerin Department of Surgery, National University of Ireland, Galway
- 2. Breast cancer microenvironment • The importance of the breast cancer microenvironment in tumour progression is well recognized • Breast tumour stroma consists of various cell types such as immunocytes, pericytes and fibroblasts • Cellular interactions within the breast cancer microenvironment promote tumour growth, invasion, metastasis and resistance to therapy Lorusso et al, Histochem Cell Biol (2008) CAF: carcinoma associated fibroblasts ECM: extracellular matrix TC: tumour cell BMDC: bone marrow derived cell PC: pericyte MSC: mesenchymal stem cell LV: lymphatic vessel
- 3. • Stromal cells are the predominant cell type within the tumour microenvironment • Stromal-epithelial cell cross talk facilitated by paracrine factors and direct cell-cell contact • Tumour stromal cells are actively secreting factors (growth factors, chemokines, cytokines) supporting tumourigenesis Stromal cellTumour cell Tumour stromal-epithelial interaction GF ECM CK The stromal-epithelial interaction in the tumour microenvironment GF : Growth factors ECM : Extracellular matrix CK: Cytokines
- 4. Aim Characterise stromal cells derived from primary breast tumours and potentially elucidate mechanisms through which they exert their actions
- 5. Primary cell culture Stromal cell fraction Breast tumour specimen Finely minced with scalpels Digested overnight Differential centrifugation Epithelial cell fraction Organoid fraction
- 6. Analysis of gene expression Primary stromal cells isolated, cultured and harvested RNA Extraction Targets of interest •Vascular endothelial growth factor (VEGF) • Matrix metalloproteinase 3 (MMP3) • Transforming growth factor beta 1 (TGFβ1) • Transforming growth factor beta receptor 2 (TGFβ R2) •Fibroblast activation protein (FAP) cDNA synthesis Real time quantitative PCR
- 7. MMP3 expression in stromal cells Tan StroTum StroNorm Stro 5 4 3 2 1 Log10relativeMMP3expression MMP3 expr ession in str omal cells N=4 N=24 N=12
- 8. FAP, VEGF, TGFβ1, TGFβR2 expression in stromal cells Tan StroTum StroNorm Stro 20 15 10 5 0 relativeFAPexpression FAP expression in stromal cells Tan StroTum StroNorm Stro 16 14 12 10 8 6 4 2 0 RelativeVEGFexpression VEGF expression in stromal cells Tan stroTum StroNorm stro 12 10 8 6 4 2 0 RelativeTGFb1expression TGFb1 expression in stromal cells Tan StroTum StroNorm Stro 35 30 25 20 15 10 5 0 TGFbR2expressionlevel TGFbR2 expression in stromal cells
- 9. Correlation of TGFβ 1 and TGFβ R2 expression Correlation of TGFβ1 and TGFβ R2 0 2 4 6 8 10 12 14 16 18 Primary stromal cells n=40 RelativeTGFβ1and TGFβR2expressionlevel TGFβ1 TGFβR2 R=0.65 P<0.01
- 10. Correlation of VEGF and MMP3 expression Correlation of VEGF and MMP3 0 2 4 6 8 10 12 14 16 18 Primary stromal cells n=40 RelativelevelofVEGFand MMP3expression MMP3 VEGF R=0.29 P<0.05
- 11. Correlation of FAP and TGFβR2 expression Correlation of FAP and TGFβR2 0 5 10 15 20 25 30 35 Primary stromal cells n=40 RelativelevelofFAPandTGFβR2 FAP TGFBR2 R=0.28 P<0.05
- 12. Summary • Trend towards increased expression of MMP3 in tumour compared to normal stromal cells • Significant positive Pearson correlation found between • TGFβ1 and TGFβR2 • MMP3 and VEGF • FAP and TGFβR2
- 13. Conclusion Tumour stromal cells secrete proangiogenic and prometastatic factors, such as MMP3 and may support breast cancer progression through promotion of tumour angiogenesis and invasion
Editor's Notes
- Fig. 1 Heterotypic cellular interactions in the tumor microenvironment. a Tumor cells orchestrate directly (e.g. though the release of factors) or indirectly (though the induction of tissue hypoxia or appearance of necrosis) the modiWcation of the microenvironment by attracting or activating many non-tumoral cells, including blood and lymphatic endothelial cells and pericytes, carcinoma associated Wbroblast, bone marrow-derived cells, immune and inXammatory cells. Tumor cells can also deposit or modify the extracellular matrix. Most of these stromal modiWcations start early during tumor progression, often at the transition stage from premalignant to malignant lesions. In some cases they may even precede cancer formation, for example in situations of chronic inXammatory conditions. b In turn, tumor microenvironmental events promote tumor progression by stimulating tumor growth and survival, and facilitating invasion and metastasis. Collectively these events will contribute to determine the outcome of tumor progression: tumor growth, tumor dormancy, tumor invasion and metastasis and resistance to therapy. Abbreviations: B B lymphocyte; BMDC bone marrow-derived cells; BV blood vessel; CAF carcinoma associated Wbroblast; EC endothelial cell; ECM extracellular matrix; EMT epithelial to mesenchymal transition; Gr granulocyte; LEC lymphatic endothelial cell; LV lymphatic vessel; Mo monocyte; MSC mesenchymal stem cell; PC pericyte; T T lymphocyte; TAM tumor associated monocyte/macrophage; TC tumor cells