The Study Of Lung Cancer Migration And Invasion By Category D Resolvin Of Specialize Pro-resolving Mediators

Objective:To investigate the possible influence of specialize pro-resolving mediators aspirin-triggered resolvin D1 (AT-RvD1) on the metastasis capacity in lung cancer A549 cells, and also the effect of nuclear factor erythrois-2 related factor 2(Nrf2) and pyruvate kinase isozymes M2 (Pkm2). We will provide experimental theory basis for the further explore the metastasis of lung cancer and mechanism of inflammation-related carcinogenesis.Methods:1. The cancer metastasis is very close associated with epithelial-mesenchymai transition (EMT), so we used different concentration of transforming growth factor β1 (TGF-β1) induced EMT process instead of lung cancer metastasis. We use fluorescent probe technique to detect the expression of the reactive oxygen species (ROS) levels, MTT cell proliferation assay kit to detect the proliferation of cell, wound healing to detect the Migration of cell, transwell technique to detect the Invasion of cell, RT-PCR and western blot analysis to detect the expression of epithelial marker E-cadherin, mesenchymal markers vimentin, Nrf2 and Pkm2 by molecular biological technique respectively.2. AT-RvD1 and mammalian target of rapamycin (mTOR) activator were also used to pre-stimulate cells for 30 minutes and investigated the effect on TGF-β1 induced EMT process. We would compare the migration, invasion and proliferative properties and detect the ROS levels and the expression of related gene or proteins.Results:1. TGF-β1-induced EMT in A549 cells enhanced the expression of the mRNA of Nrf2 and mesenchymal markers, such as vimentin, and reduced the expression of Pkm2 and the epithelial marker E-cadherin. The ROS levels were significantly increased by incubation with TGF-β1 and were TGF-β1-concentration dependent. In additon, TGF-β1 clearly enhanced the migration and invasion but did not affect the proliferative properties in A549 cells.2. AT-RvD1 had the ability to inhibit TGF-β1-induced EMT through enhanced the expression of E-cadherin and reduced the expression of vimentin. However, AT-RvD1 clearly reduced the TGF-β1-induced migration and invasion but did not affect the proliferative properties of the TGF-β1-treated A549 cells. Our results also showed that it was associated with the down-regulated Nrf2 and Pkm2 expression.3. Our studies had conclusively shown that the mTOR activator restored the expression of Pkm2, which was suppressed by AT-RvD1, and partly down-regulated the expression of E-cadherin. It also affected the invasiveness but had no effect on the proliferative properties.Conclusions:1. These results suggest that AT-RvD1 inhibited TGF-β1-induced EMT through the inhibition of the migration and invasion, but it had no effect on the proliferative properties.2. AT-RvD1 inhibited TGF-β1-induced EMT via the inhibition of the mTOR pathway by reducing the expression of PKM2, and it was associated with down-regulated Nrf2 expression.