Abstract
Membrane type matrix metalloproteinase-1(MT1-MMP) is the identified member of characterized MMPs. Recent researches suggest that MT1-MMP participates in the progress of hematology malignancies, but the underlying mechanisms have not been clearly defined. To explore the effects of MT1-MMP in hematology malignancies, the expression of MT1-MMP as well as MMP-2 and TIMP-2 in 13 malignant hematopoietic cell lines was investigated and its eukaryotic expression vector was constructed and tranfected to HL60 with low expression of MT1-MMP. mRNA expressions of MT1-MMPs, MMP-2 and TIMP-2 in hematopoietic cell lines with different lineages were semi-quantified by RT-PCR. The gelatinolytic activity of MMP-2 was visualized on gelatin zymograms and the expression of MT1-MMP protein was detected by flow cytometry. The eukaryotic expression vector pcDNA3.1/V5-His-TOPO-MT1-MMP was constructed and identified by enzyme digestion, PCR and sequencing, the vector was transfected into HL60 with liposomes and the transfection efficiency was analyzed by RT-PCR, real-time PCR and flow cytometry. HL60/pcDNA3.1 cells were adopted as controls. Cell proliferation was examined by MMT, and expressions of MMP-2, Bcl-2 and Bax were analyzed by real-time PCR. Invasive potential and cell cycle of the transfected cells were studied with transwell plate and flow cytometry. The expression patterns of the MT1-MMP/MMP-2/TIMP-2 were varied in 13 hematopoietic cell lines. No obvious correlation between the origin of the cells and the expression level of the detected genes was observed (P>0.1), while MT1-MMP significantly correlated to MMP-2 (R=0.64, P=0.02). The expression vector of MT1-MMP was successfully constructed. Significantly enhanced proliferative ability was observed in HL60/MT1-MMP than in control (P<0.01). No obvious difference was observed as to the expression of MMP-2, Bcl-2 and Bax between the two groups. At 48h after transfection, significantly increased invasiveness was noted in HL60/MT1-MMP (15.42±2.38%)than in control(5.16±0.78%)(P<0.01). At 24h after transfection, higher ratio of G2 and S phase cells was found in the experiment group than in control (67.5±3.57% vs 57.9±2.52%) (P<0.05), whereas at 48h and 72h after transfection, no obvious difference of cell cycle was observed between the two groups. The association between expression of MT1-MMP and MMP-2 in malignant hematopoietic cell lines may suggest the close relation of their functions. Over-expressing MT1-MMP in leukemic cell lines exhibit enhanced proliferative and invasive potential. No direct relationship can be identified between MMP-2 and enhanced invasiveness. Directive involvement of MT1-MMP may be absent in the regulation of apoptosis induced by the Bcl-2/Bax pathway.
Author notes
Disclosure: No relevant conflicts of interest to declare.
