title: Down-Regulation of miR-92 in Breast Epithelial Cells and in Normal but Not Tumour Fibroblasts Contributes to Breast Carcinogenesis.
abstract: BACKGROUND
  MicroRNA (miR) expression is commonly dysregulated in many cancers, including breast. MiR-92 is one of six miRs encoded by the miR-17-92 cluster, one of the best-characterised oncogenic miR clusters. We examined expression of miR-92 in the breast epithelium and stroma during breast cancer progression. We also investigated the role of miR-92 in fibroblasts in vitro and showed that down-regulation in normal fibroblasts enhances the invasion of breast cancer epithelial cells.
  
  
  RESULTS
  We used laser microdissection (LMD) to isolate epithelial cells from matched normal, DCIS and invasive tissue from 9 breast cancer patients and analysed miR-92 expression by qRT-PCR. Expression of ERβ1, a direct miR-92 target, was concurrently analysed for each case by immunohistochemistry. LMD was also used to isolate matched normal (NFs) and cancer-associated fibroblasts (CAFs) from 14 further cases. Effects of miR-92 inhibition in fibroblasts on epithelial cell invasion in vitro was examined using a Matrigel™ assay. miR-92 levels decreased in microdissected epithelial cells during breast cancer progression with highest levels in normal breast epithelium, decreasing in DCIS (p<0.01) and being lowest in invasive breast tissue (p<0.01). This was accompanied by a shift in cell localisation of ERβ1 from nuclear expression in normal breast epithelium to increased cytoplasmic expression during progression to DCIS (p = 0.0078) and invasive breast cancer (p = 0.031). ERβ1 immunoreactivity was also seen in stromal fibroblasts in tissues. Where miR-92 expression was low in microdissected NFs this increased in matched CAFs; a trend also seen in cultured primary fibroblasts. Down-regulation of miR-92 levels in NFs but not CAFs enhanced invasion of both MCF-7 and MDA-MB-231 breast cancer epithelial cells.
  
  
  CONCLUSIONS
  miR-92 is gradually lost in breast epithelial cells during cancer progression correlating with a shift in ERβ1 immunoreactivity from nuclei to the cytoplasm. Our data support a functional role in fibroblasts where modification of miR-92 expression can influence the invasive capacity of breast cancer epithelial cells. However in silico analysis suggests that ERβ1 may not be the most important miR-92 target in breast cancer.
authors.affiliation: Leeds Institute of Cancer and Pathology, University of Leeds, Leeds, United Kingdom.
authors.forename: Laura
authors.initials: L
authors.lastname: Smith
authors.affiliation: Leeds Institute of Cancer and Pathology, University of Leeds, Leeds, United Kingdom.
authors.forename: Euan W
authors.initials: EW
authors.lastname: Baxter
authors.affiliation: Leeds Institute of Cancer and Pathology, University of Leeds, Leeds, United Kingdom.
authors.forename: Philip A
authors.initials: PA
authors.lastname: Chambers
authors.affiliation: Leeds Institute of Cancer and Pathology, University of Leeds, Leeds, United Kingdom.
authors.forename: Caroline A
authors.initials: CA
authors.lastname: Green
authors.affiliation: Leeds Institute of Cancer and Pathology, University of Leeds, Leeds, United Kingdom.
authors.forename: Andrew M
authors.initials: AM
authors.lastname: Hanby
authors.affiliation: Leeds Institute of Biomedical and Clinical Sciences, University of Leeds, Leeds, United Kingdom.
authors.forename: Thomas A
authors.initials: TA
authors.lastname: Hughes
authors.affiliation: Leeds Institute of Cancer and Pathology, University of Leeds, Leeds, United Kingdom.
authors.forename: Claire E
authors.initials: CE
authors.lastname: Nash
authors.affiliation: St James's Institute of Oncology, St James's University Hospital, Leeds, United Kingdom.
authors.forename: Rebecca A
authors.initials: RA
authors.lastname: Millican-Slater
authors.affiliation: Leeds Institute of Cancer and Pathology, University of Leeds, Leeds, United Kingdom.
authors.forename: Lucy F
authors.initials: LF
authors.lastname: Stead
authors.affiliation: Leeds Institute of Cancer and Pathology, University of Leeds, Leeds, United Kingdom.
authors.forename: Eldo T
authors.initials: ET
authors.lastname: Verghese
authors.affiliation: Leeds Institute of Cancer and Pathology, University of Leeds, Leeds, United Kingdom.
authors.forename: Valerie
authors.initials: V
authors.lastname: Speirs
mesh_terms: D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D001943:Breast Neoplasms / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D063646:Carcinogenesis / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D002285:Carcinoma, Intraductal, Noninfiltrating / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D015536:Down-Regulation; D004847:Epithelial Cells / Q000378:metabolism* / Q000473:pathology; D047629:Estrogen Receptor beta / Q000235:genetics / Q000378:metabolism; D005260:Female; D005347:Fibroblasts / Q000378:metabolism* / Q000473:pathology; D020869:Gene Expression Profiling; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D061346:Laser Capture Microdissection; D035683:MicroRNAs / Q000235:genetics / Q000378:metabolism*; D008875:Middle Aged
chemical_list: D047629:Estrogen Receptor beta; C541874:MIRN92 microRNA, human; D035683:MicroRNAs