Co-staining of microRNAs and their target proteins by miRNA in situ hybridization and immunohistofluorescence on prostate cancer tissue microarrays

• Published in: Laboratory investigation 2019-10, Vol.99 (10), p.1527-1534
• Main Authors: Eckstein, Markus, Sailer, Verena, Nielsen, Boye Schnack, Wittenberg, Thomas, Wiesmann, Veit, Lieb, Verena, Nolte, Elke, Hartmann, Arndt, Kristiansen, Glen, Wernert, Nicolas, Wullich, Bernd, Taubert, Helge, Wach, Sven
• Format: Article
• Language: English
• Subjects: 13/51; 14/32; 631/67/1857; 692/53/2422; 82/80; 96/63; Cytoplasm; Fluorescence; Fluorescent Antibody Technique - methods; Humans; Hybridization; In Situ Hybridization, Fluorescence - methods; Laboratory Medicine; Lymph nodes; Lymphatic system; Male; Medicine; Medicine & Public Health; Metastases; Metastasis; MicroRNAs; MicroRNAs - analysis; MicroRNAs - metabolism; miRNA; Nuclei; Nuclei (cytology); Nucleoli; Pathology; Prostate cancer; Prostatic Neoplasms - chemistry; Prostatic Neoplasms - metabolism; Proteins; Sequestering; Staining; Stromal cells; technical-report; Tissue Array Analysis; Tissues; Tumor cells; Tumors
• ISSN: 0023-6837; 1530-0307
• DOI: 10.1038/s41374-019-0251-8

The co-expression of miRNAs and their target proteins was studied on tissue microarrays from different prostate cancer (PCa) patients. PCa of primary Gleason pattern 4 (GP4), lymph node metastases of GP4, distant metastases, and normal tissue from the transitional and peripheral zones were co-stained by fluorescent miRNA in situ hybridization (miRisH) and protein immunohistofluorescence (IHF). The miRNAs and corresponding target proteins include the pairs miR-145/ERG, miR-143/uPAR, and miR-375/SEC23A. The fluorescence-stained and scanned tissue microarrays (TMAs) were evaluated by experienced uropathologists. The pair miR-145/ERG showed an exclusive staining for miR-145 in the nuclei of stromal cells, both in tumor and normal tissue, and for ERG in the cytoplasm with/without co-expression in the nucleus of tumor cells. The pair miR-143/uPAR revealed a clear distinction between miR-143 in the nuclei of stromal cells and uPAR staining in the cytoplasm of tumor cells. Metastases (lymph node and distant) however, showed tumor cells with cytoplasmic staining for miR-143/uPAR. In normal tissues, beside the nuclei of the stroma cells, gland cells could also express miR-143 and uPAR in the cytoplasm. miR-375 showed particular staining in the nucleoli of GP4 and metastatic samples, suggesting that nucleoli play a special role in sequestering proteins and miRNAs. Combined miRisH/IHF allows for the study of miRNA expression patterns and their target proteins at the single-cell level.