A Study On The Effect Of TRAIL On The Sensitivity Of A Paclitaxel-resistant Human Lung Adenocarcinoma Cell Line To Paclitaxel

OBJECTIVE:Observe whether TNF-related apoptosis inducing ligand (TRAIL) modulates thesensitivity of A549cell’s paclitaxel-resistant sub-strain to paclitaxel, and explore therelated mechanisms.METHODS:1． Cultivate and identify human lung adenocarcinoma paclitaxel-resistantsub-strains: Continuously cultivate the A549cells in low concentrations of paclitaxeland gradually increase the dose to induced the paclitaxel-resistant sub-strain of A549cells, which is called A549/Taxol-R. MTT assay and flow cytometry is used toexamine the growth inhibition rate and apoptosis rate of cells，while MDR1genemRNA expression is detected by RT-PCR.2．Examine how TRAIL and paclitaxel intervention affects the proliferation andapoptosis rate of A549/Taxol-R in vitro: A549/Taxol-R was treated with combined orsingle TRAIL and paclitaxel, then use MTT assay and flow cytometry to examine thegrowth inhibition rate and apoptosis rate of cells.3．Examine how TRAIL and paclitaxel intervention affects the apoptosis relatedgene expression of A549/Taxol-R in vitro: A549/Taxol-R was treated with combinedor single TRAIL and paclitaxel, then the expression of the active cleaved Caspase-3and Bax and Bcl-2mRNA, all of which are apoptosis-related genes, was examined byusing Western Blot and reverse transcription PCR(RT-PCR)RESULTS:1.the paclitaxel resistant strains A549/Taxol-R of human lung adenocarcinomaA549cells was successfully cultivated. Compared to its parental A549cells, thesensitivity of A549/Taxol-R to paclitaxel was significantly reduced, while the MDR1 gene of A549/Taxol-R was highly expressed.2. MTT results showed: comparing to used separately, combined use of TRAILand paclitaxel significantly inhabit the growth rate of A549/Taxol-R cell (P <0.05),the two drugs have a synergistic effect. Flow cytometry results show: combined use ofTRAIL and paclitaxel exceeds both used alone in the apoptosis rate ofA549/Taxol-R(P <0.05)3. Western Blot and RT-PCR showed: TRAIL induced A549/Taxol-R apoptosisvia Caspase-3pathway. Apoptosis-related genes Bax and Bcl-2are involved in theregulation of its apoptosis.CONCLUSION:1. TRAIL can sensitize the A549cell’s paclitaxel-resistant sub-strainA549/Taxol-R to paclitaxel in vitro;2. TRAIL may sensitize the A549cell’s paclitaxel-resistant sub-strainA549/Taxol-R to paclitaxel by promoting apoptosis. The mechanism might involvedin promotion of the expression of Bax, down regulation of the expression of Bcl-2,which raised expression of active cleaved caspase-3.