title: Pleiotropic actions of miR-21 highlight the critical role of deregulated stromal microRNAs during colorectal cancer progression.
abstract: The oncogene microRNA-21 (miRNA; miR-21) is overexpressed in most solid organ tumours; however, a recent examination of stage II colorectal cancer (CRC) specimens suggests this may be a stromal phenomenon and not only a feature of cancer cells. In vitro and in vivo studies show that miR-21 has potent pro-metastatic effects in various malignant carcinoma cell lines. The tumour microenvironment has also been identified as a key actor during the metastatic cascade; however to date the significance of deregulated miR-21 expression within the cancer-associated stroma has not been examined. In the present study, a quantitative RT-PCR-based analysis of laser microdissected tissue confirmed that miR-21 expression is associated with a four-fold mean increase in CRC stroma compared with normal tissue. In situ hybridisation using locked nucleic acid probes localised miR-21 expression predominantly to fibroblasts within tumour-associated stroma. To study the molecular and biological impact of deregulated stromal miR-21 in CRC, stable ectopic expression was induced in immortalised fibroblasts. This resulted in upregulated α-smooth muscle actin expression implying miR-21 overexpression is driving the fibroblast-to-myofibroblast transdifferentiation. Conditioned medium from miR-21-overexpressing fibroblasts protected CRC cells from oxaliplatin-induced apoptosis and increased their proliferative capacity. 3D organotypic co-cultures containing fibroblasts and CRC cells revealed that ectopic stromal miR-21 expression was associated with increased epithelial invasiveness. Reversion-inducing cysteine-rich protein with kazal motifs, an inhibitor of matrix-remodelling enzyme MMP2, was significantly downregulated by ectopic miR-21 in established and primary colorectal fibroblasts with a reciprocal rise in MMP2 activity. Inhibition of MMP2 abrogated the invasion-promoting effects of ectopic miR-21. This data, which characterises a novel pro-metastatic mechanism mediated by miR-21 in the CRC stroma, highlights the importance of miRNA deregulation within the tumour microenvironment and identifies a potential application for stromal miRNAs as biomarkers in cancer.
authors.affiliation: Cancer Sciences Division, University of Southampton, Southampton General Hospital, Southampton, UK. m.bullock@soton.ac.uk
authors.forename: M D
authors.initials: MD
authors.lastname: Bullock
authors.forename: K M
authors.initials: KM
authors.lastname: Pickard
authors.forename: B S
authors.initials: BS
authors.lastname: Nielsen
authors.forename: A E
authors.initials: AE
authors.lastname: Sayan
authors.forename: V
authors.initials: V
authors.lastname: Jenei
authors.forename: M
authors.initials: M
authors.lastname: Mellone
authors.forename: R
authors.initials: R
authors.lastname: Mitter
authors.forename: J N
authors.initials: JN
authors.lastname: Primrose
authors.forename: G J
authors.initials: GJ
authors.lastname: Thomas
authors.forename: G K
authors.initials: GK
authors.lastname: Packham
authors.forename: A H
authors.initials: AH
authors.lastname: Mirenzami
authors.forename: A H
authors.initials: AH
authors.lastname: Mirnezami
mesh_terms: D000230:Adenocarcinoma / Q000378:metabolism* / Q000473:pathology; D000970:Antineoplastic Agents / Q000494:pharmacology; D017209:Apoptosis; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D002470:Cell Survival / Q000187:drug effects; D015179:Colorectal Neoplasms / Q000378:metabolism* / Q000473:pathology; D019008:Drug Resistance, Neoplasm; D005347:Fibroblasts / Q000378:metabolism; D058851:GPI-Linked Proteins / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D015972:Gene Expression Regulation, Neoplastic; D058685:Genetic Pleiotropy; D006801:Humans; D020778:Matrix Metalloproteinase 2 / Q000378:metabolism; D035683:MicroRNAs / Q000235:genetics* / Q000378:metabolism; D009361:Neoplasm Invasiveness; D009944:Organoplatinum Compounds / Q000494:pharmacology; D000077150:Oxaliplatin; D034622:RNA Interference; D017154:Stromal Cells / Q000378:metabolism; D059016:Tumor Microenvironment
chemical_list: D000970:Antineoplastic Agents; D058851:GPI-Linked Proteins; C524940:MIRN21 microRNA, human; D035683:MicroRNAs; D009944:Organoplatinum Compounds; C115460:RECK protein, human; D000077150:Oxaliplatin; C522361:MMP2 protein, human; D020778:Matrix Metalloproteinase 2