| Summary: | 碩士 === 國立臺灣大學 === 動物學研究所 === 92 === Abstract
Lysophosphatidic acid (LPA) and sphingosine 1-phosphate (S1P) are both low molecular weight lysophopholipid (LPLs), which can promote cell proliferation, migration, and invasion via interaction with the endothelial differentiation gene (Edg) family of specific G protein-coupled receptors. Matrix metalloproteinases (MMPs) are a family of zinc-dependent proteolytic enzymes, which are involved in degradation of extracellular matrix, and play critical roles in endothelial cell migration and matrix remodeling during the angiogenesis. Among these MMPs, MMP-2 is an important molecule that is known to trigger the invasion of tumor cells. On the other hand, the nature inhibitors of MMPs, tissue inhibitor of metalloproteinases (TIMPs), are key physiological regulators of MMPs. In our present study, we examine the effects of LPA and S1P on MMPs, especially MMP-2 expression of human endothelial cell. By using RT-PCR, Western blotting, and gelatin zymographic analysis, we showed that LPA and S1P enhanced MMP-2 expression in mRNA, protein levels and also enzymatic activity. The enhancement effects are in a concentration- and time-dependent manner. Next, we evaluated the signal transduction pathway relevant to the MMP-2 expression stimulated by LPLs. Several chemical inhibitors were used including pertussis toxin (Gi blocker), PD98059 (MEK/ERK blocker), U73122 (PLC blocker), LY294002 (PI3 kinase blocker), PDTC (NF-κB blocker), PP2 (Src family kinase blocker), and SKF 96365 (receptor mediate calcium influx blocker). By real-time PCR analysis, the results indicated that these effects are involved in MEK/ERK-, NF-κB-, or calcium influx mediate signaling-dependent pathways. Furthermore, we used chemotaxis chamber to investigate LPL’s effects on cell invasion. Our results showed that endothelial cell invasion was potently increased by LPLs, and the induction can be prevented by preincubation with GM6001, a chemically synthesized MMP inhibitor. Interestingly, we find that S1P also regulated TIMPs expression, including TIMP-2 and TIMP-3. These observations suggest that LPA and S1P may play important roles in endothelial cell invasion by regulating the expression of MMP-2 and may be also MMPs inhibitors.
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