miR-125b induces cellular senescence in malignant melanoma.
BACKGROUND
Micro RNAs (miRs) have emerged as key regulators during oncogenesis. They have been found to regulate cell proliferation, differentiation, and apoptosis. Mir-125b has been identified as an oncomir in various forms of tumours, but we have previously proposed that miR-125b is a suppressor of lymph node metastasis in cutaneous malignant melanoma. Our goal was therefore to further examine this theory.


METHODS
We used in-situ-hybridization to visualise miR-125b expression in primary tumours and in lymph node metastasis. Then using a miRVector plasmid containing a miR-125b-1 insert we transfected melanoma cell line Mel-Juso and then investigated the effect of the presence of a stable overexpression of miR-125b on growth by western blotting, flow cytometry and β-galactosidase staining. The tumourogenicity of the transfected cells was tested using a murine model and the tumours were further examined with in-situ-hybridization.


RESULTS
In primary human tumours and in lymph node metastases increased expression of miR-125b was found in single, large tumour cells with abundant cytoplasm. A stable overexpression of miR-125b in human melanoma cell line Mel-Juso resulted in a G0/G1 cell cycle block and emergence of large cells expressing senescence markers: senescence-associated beta-galactosidase, p21, p27 and p53. Mel-Juso cells overexpressing miR-125b were tumourigenic in mice, but the tumours exhibited higher level of cell senescence and decreased expression of proliferation markers, cyclin D1 and Ki67 than the control tumours.


CONCLUSIONS
Our results confirm the theory that miR-125b functions as a tumour supressor in cutaneous malignant melanoma by regulating cellular senescence, which is one of the central mechanisms protecting against the development and progression of malignant melanoma.
Anne Marie
AM
Nyholm
Catharina M
CM
Lerche
Valentina
V
Manfé
Edyta
E
Biskup
Peter
P
Johansen
Niels
N
Morling
Birthe Mørk
BM
Thomsen
Martin
M
Glud
Department of Dermatology, Faculty of Health and Medical Sciences, University of Copenhagen, Bispebjerg Hospital, Copenhagen, Denmark. r.gniadecki@gmail.com.
Robert
R
Gniadecki
D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D000818:Animals; D014408:Biomarkers, Tumor / Q000096:biosynthesis / Q000235:genetics; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D016922:Cellular Senescence; D004195:Disease Models, Animal; D005260:Female; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D008198:Lymph Nodes / Q000473:pathology; D008207:Lymphatic Metastasis; D008297:Male; D008545:Melanoma / Q000235:genetics / Q000378:metabolism* / Q000473:pathology*; D008807:Mice, Inbred BALB C; D008819:Mice, Nude; D035683:MicroRNAs / Q000096:biosynthesis* / Q000235:genetics; D008875:Middle Aged; D012878:Skin Neoplasms; D014162:Transfection; D015854:Up-Regulation; D055815:Young Adult; D000096142:Cutaneous Malignant Melanoma
D014408:Biomarkers, Tumor; C531888:MIRN125 microRNA, human; D035683:MicroRNAs