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Breast stromal fibroblasts from histologically normal surgical margins are pro-carcinogenic.

ABSTRACT: There is evidence that normal breast stromal fibroblasts (NBFs) suppress tumour growth, while cancer-associated fibroblasts (CAFs) promote tumourigenesis through functional interactions with tumour cells. Little is known about the biology and the carcinogenic potential of stromal fibroblasts present in histologically normal surgical margins (TCFs). Therefore, we first undertook gene expression analysis on five CAF/TCF pairs from breast cancer patients and three NBF samples (derived from mammoplasties). This comparative analysis revealed variation in gene expression between these three categories of cells, with a TCF-specific gene expression profile. This variability was higher in TCFs than in their paired CAFs and also NBFs. Cytokine arrays show that TCFs have a specific secretory cytokine profile. In addition, stromal fibroblasts from surgical margins expressed high levels of ?-SMA and SDF-1 and exhibited higher migratory/invasiveness abilities. Indirect co-culture showed that TCF cells enhance the proliferation of non-cancerous mammary epithelial cells and the epithelial-to-mesenchymal transition of breast cancer cells. Moreover, TCF and CAF cells increased the level of PCNA, MMP-2 and the phosphorylated/activated form of Akt in normal breast luminal fibroblasts in a paracrine manner. Furthermore, TCFs were able to promote the formation and growth of humanized orthotopic breast tumours in nude mice. Interestingly, these TCF phenotypes and the extent of their effects were intermediate between those of NBFs and CAFs. Together, these results indicate that stromal fibroblasts located in non-cancerous tissues exhibit a tumour-promoting phenotype, indicating that their presence post-surgery may play important roles in cancer recurrence.

SUBMITTER: Al-Rakan MA

PROVIDER: S-EPMC4284036 | biostudies-literature | 2013 Dec

REPOSITORIES: biostudies-literature

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Breast stromal fibroblasts from histologically normal surgical margins are pro-carcinogenic.

Al-Rakan Maha A MA Colak Dilek D Hendrayani Siti-Faujiah SF Al-Bakheet Albandary A Al-Mohanna Falah H FH Kaya Namik N Al-Malik Osama O Aboussekhra Abdelilah A

The Journal of pathology 20131201 4

There is evidence that normal breast stromal fibroblasts (NBFs) suppress tumour growth, while cancer-associated fibroblasts (CAFs) promote tumourigenesis through functional interactions with tumour cells. Little is known about the biology and the carcinogenic potential of stromal fibroblasts present in histologically normal surgical margins (TCFs). Therefore, we first undertook gene expression analysis on five CAF/TCF pairs from breast cancer patients and three NBF samples (derived from mammopla ...[more]

PMID: 24009142

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Project description:The aim of this study is to provide an extended, retrospective validation of a previously described 4-gene signature (MMP1, COL4A1, P4HA2, and THBS2) predictive of oral carcinoma recurrence. We assessed an independent cohort including 245 histologically normal surgical margins from 62 patients with OSCC. Gene expression was determined using a color-coded probe assay for quantitative gene expression analysis. All margins and tumor blocks were cut (4-5µm tick) and stained with Hematoxylin-Eosin and tissues were subjected to needle microdissection using the stereo microscope Leica EZ4 (Leica Microsystems, Wetzlar, Germany) before RNA extraction, in order to isolate pure tumor or normal cell populations. RNA from FFPE samples was isolated using the RecoverAll Total Nucleic Acid Isolation kit (Ambion/Life Technologies, Carlsbad, CA, USA), following our previously reported protocol with modifications to improve RNA yield [PMID:23835716]. RNA was stored at -80ºC until RNA extraction and gene expression validation⁠. In addition, tumor tissues from the same patients were collected for further analysis (not in scope of the presented study). Probes were designed for the 4-gene signature (MMP1, COL4A1, P4HA2 and THBS2) and contained one capture probe linked to biotin and one reporter probe attached to a color-coded molecular tag for each gene sequence, according to the nCounterTM code-set design. RNA samples were randomized using a numerical ID and were then subjected to NanoString nCounterTM analysis by the Princess Margaret Genomics Centre (https://www.pmgenomics.ca), Toronto, ON, Canada. Patient samples were analyzed in two batches consisting of 125 and 152 margins tissue samples. The detailed protocol for mRNA transcript quantification analysis, including sample preparation, hybridization, detection and scanning followed the manufacturer’s recommendations and was described in [PMID: 21549012]. We used 400 ng of total RNA isolated from FFPE tissues for detection of probe signals. Raw data were analyzed using the nCounterTM digital analyzer software (http://www.nanostring.com/support/ncounter/) and gene expression levels were subjected to further bioinformatic and statistical analyses.

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Breast stromal fibroblasts from histologically normal surgical margins are pro-carcinogenic.

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Breast stromal fibroblasts from histologically normal surgical margins are pro-carcinogenic.

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