Differential expression of matrix metalloproteinases in brain- and bone-seeking clones of metastatic MDA-MB-231 breast cancer cells.
Matrix Metalloproteinases (MMPs) play a crucial role in breast cancer metastasis. We examined the mRNA and protein expression of several MMPs in brain- and bone-seeking clones of MDA-MB-231 breast cancer cells, their transcriptional regulation and their functional role in the metastatic process. MMP mRNA expression was examined using real-time reverse transcription polymerase chain reaction. Protein expression was examined using enzyme linked immunosorbent essay (ELISA). The inducibility of mRNA and protein expression was tested with TPA (phorbol 12-myristate 13-acetate; 50 microM); epidermal growth factor and transforming growth factor beta (20 ng/ml both). Migration and invasion assays were performed with the QCM 96-Well Migration/Invasion Assay (8 microm; Chemicon) over 24 h with or without specific MMPs inhibitors (MMP Inhibitor I Mix (5 microM); MMP-2/MMP-9 Inhibitor III (50 microM); EMD Biosciences). We found significantly higher mRNA expression of MMP-1 and -9 in brain-seeking 231-clones in comparison to -bone and -parental cells. In contrast, the mRNA expression of MMP-3 and -14 was comparable in all cells lines examined and MMP-13 expression was lower in both selective metastatic lines. MMP-2 and -8 were not expressed. ELISA revealed a higher amount of total as well as active MMP-1 and -9 in brain-seeking cells. TPA stimulation showed that MMP-1 and -9 transcription was inducible on the mRNA and protein level in 231-parental but not in 231-brain or -bone. 231-brain showed the highest migration and invasive capacity which could be decreased by the application of MMP-1 and/or MMP-9 inhibitor. Our results indicate functional importance of MMP-1 and -9 overexpression in brain metastasis in an in vitro model.
Stark
Andreas M
AM
Department of Neurosurgery, University of Kiel, Schittenhelmstrasse 10, 24105 Kiel, Germany.
Anuszkiewicz
Bartosz
B
Mentlein
Rolf
R
Yoneda
Toshiyuki
T
Mehdorn
H Maximilian
HM
Held-Feindt
Janka
J
D001859:Bone Neoplasms / Q000201:enzymology* / Q000556:secondary; D001932:Brain Neoplasms / Q000201:enzymology* / Q000556:secondary; D001943:Breast Neoplasms / Q000201:enzymology* / Q000473:pathology*; D045744:Cell Line, Tumor; D002999:Clone Cells / Q000201:enzymology / Q000473:pathology; D006801:Humans; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism; D020781:Matrix Metalloproteinase 1 / Q000145:classification / Q000235:genetics / Q000378:metabolism*; D020780:Matrix Metalloproteinase 9 / Q000145:classification / Q000235:genetics / Q000378:metabolism*; D009361:Neoplasm Invasiveness; D009362:Neoplasm Metastasis; D009928:Organ Specificity / Q000502:physiology; D012333:RNA, Messenger / Q000032:analysis; D018709:Statistics, Nonparametric
D007527:Isoenzymes; D012333:RNA, Messenger; D020780:Matrix Metalloproteinase 9; D020781:Matrix Metalloproteinase 1