Project description:To investigate the difference of miRNA expression in exosomes derived from A549 cells and its DDP-resistant cell strain A549/DDP, we have employed miRNA microarray expression to discover the difference expression of miRNAs in exosomes derived from A549 and A549/DDP. We conducted RT-qPCR to examine the expression levels of top differential expressed miRNAs, namely, miR-197-5p, miR-4443, miR-642a-3p, miR-27b-3p and miR-100-5p, confirming low variability between two methods. The A549/DDP was established from A549 in our laboratory, by exposing A549 to gradually increasing DDP concentrations, until the final concentration at 1μg/ml. To avoid the influence of drug to the A549/DDP cells, they were cultured in a drug-free medium for at least two weeks before gene expression analysis. After being incubated for 48-72h, the culture medium of cells was harvested. Exosomes were isolated by ultracentrifugation. And miRNA expression of exosomes derived from A549 and A549/DDP was then analzyed.
Project description:This study aims to identify pathway involvement in the development of cisplatin (cis-diamminedichloroplatinum (II); CDDP) resistance in A549 lung cancer (LC) cells by utilizing advanced bioinformatics software. We developed CDDP-resistant A549 (A549/DDP) cells through prolonged incubation with the drug and performed RNA-seq on RNA extracts to determined differential mRNA and miRNA expression between A549/DDP and A549 cells
Project description:To investigate the difference of miRNA expression in exosomes derived from A549 cells and its DDP-resistant cell strain A549/DDP, we have employed miRNA microarray expression to discover the difference expression of miRNAs in exosomes derived from A549 and A549/DDP. We conducted RT-qPCR to examine the expression levels of top differential expressed miRNAs, namely, miR-197-5p, miR-4443, miR-642a-3p, miR-27b-3p and miR-100-5p, confirming low variability between two methods.
Project description:Combination therapy as an important treatment option for lung cancer has been attracting attention due to the primary and acquired resistance of chemotherapeutic drugs in the clinical application. In the present study, as a new therapy strategy, concomitant treatment with time-restricted feeding (TRF) plus cisplatin (DDP) on lung cancer growth was investigated in DDP-resistant and DDP-sensitive lung cancer cells. We first found that TRF significantly enhanced the drug susceptibility of DDP in DDP-resistant A549 (A549/DDP) cell line, illustrated by reversing the inhibitory concentration 50 (IC50) values of A549/DDP cells to normal level of parental A549 cells. We also found that TRF markedly enhanced DDP inhibition on cell proliferation, migration, as well as promoted apoptosis compared to the DDP-alone group in A549, H460 and A549/DDP cells lines. We further revealed that the synergistic anti-tumor effect of combined DDP and TRF was greater than that of combined DDP and simulated fasting condition (STS), a known anti-tumor cellular medium. Moreover, mRNA sequence analysis from A549/DDP cell line demonstrated the synergistic anti-tumor effect involved in upregulated pathways in p53 signaling pathway and apoptosis. Notably, compared with the DDP-alone group, combination of TRF and DDP robustly upregulated the P53 protein expression without mRNA level change by regulating its stability via promoting protein synthesis and inhibiting degradation, revealed by cycloheximide and MG132 experiments. Collectively, our results suggested that TRF in combination with cisplatin might be an additional novel therapeutic strategy for patients with lung cancer
Project description:To investigate the difference of miRNA expression between lung cancer cell A549 and its DDP-resistant cell strain A549/DDP, we have employed miRNA microarray expression to discover the difference expression of miRNAs of A549 cells and A549/DDP. We conducted RT-qPCR to examine the expression levels of top differential expressed miRNAs, namely, miR-197-5p, miR-4443, miR-642a-3p, miR-27b-3p and miR-100-5p, confirming low variability between two methods. The A549/DDP was established from A549 in our laboratory, by exposing A549 to gradually increasing DDP concentrations, until the final concentration at 1μg/ml. To avoid the influence of drug to the A549/DDP cells, they were cultured in a drug-free medium for at least two weeks before gene expression analysis. miRNA expression of A549 and A549/DDP was then analzyed.
Project description:To investigate the difference of miRNA expression between lung cancer cell A549 and its DDP-resistant cell strain A549/DDP, we have employed miRNA microarray expression to discover the difference expression of miRNAs of A549 cells and A549/DDP. We conducted RT-qPCR to examine the expression levels of top differential expressed miRNAs, namely, miR-197-5p, miR-4443, miR-642a-3p, miR-27b-3p and miR-100-5p, confirming low variability between two methods.
Project description:Continuous exposure to cisplatin can induce drug resistance to limit efficacy, however, the underlying mechanisms correlated to cisplatin resistance are still unclear. Drug-sensitive A549 cells and cisplatin-resistant A549/DDP cells were used to explore the potential metabolic pathways and key targets associated with cisplatin resistance by integrating untargeted metabolomics with transcriptomics. The results of comprehensive analyses showed that 19 metabolites were significantly changed in A549/DDP vs A549 cells, and some pathways had a close relationship with cisplatin resistance, such as the biosynthesis of aminoacyl-tRNA, glycerophospholipid metabolism, and glutathione metabolism. Moreover, transcriptomics analysis showed glutathione metabolism was also obviously affected in A549/DDP, which indicated that glutathione metabolism played an import role in the process of drug resistance. Meanwhile, transcriptomics analysis suggested the four enzymes related to glutathione metabolism - CD13, GPX4, RRM2B, and OPLAH - as potential targets of cisplatin resistance in NSCLC. Further studies identified the over-expressions of these four enzymes in A549/DDP. The elucidation of mechanism and discovery of new potential targets may help us have a better understanding of cisplatin resistance.
Project description:Application of cisplatin (DDP) for treating lung cancer is restricted due to its toxicity and drug resistance. In this study, we aimed to examine whether Jinfukang (JFK), an effective herbal medicine against lung cancer, enhances DDP-induced cytotoxicity in lung cancer cells. Morphologically, we observed JFK increases DDP-induced pro-apoptosis in A549 cells in a synergistic manner. Transcriptome profiling analysis indicated that combination of JFK and DDP regulates genes involved in apoptosis-related signaling pathways. Moreover, we found the combination of JFK and DDP produces synergistic pro-apoptosis effect in other lung cancer cell lines NCI-H1975, NCI-H1650 and NCI-H2228. Particularly, we demonstrated AIFM2 is activated by the combined treatment of JFK and DDP, and partially mediate the synergistic pro-apoptosis effect. Collectively, this study gives the first evidence that activation of AIFM2 contributes to induction of pro-apoptosis by combined treatment with JFK and DDP in human lung cancer cells and provides an insight for its potential clinical application in lung cancer treatment.
Project description:We report the application of specific antibodies and high-throughput sequencing technologies (methylated RNA immunoprecipitation sequencing, MeRIP-seq) for high-throughput profiling of m6A modifications in NSCLC cisplatin resistant cells. We generated maps of m6A modified transcripts in A549 and A549/DDP cells. We find that the m6A level was significantly increased in A549/DDP cells compared to the A549 cells. We show that there was a close correlation between the m6A modification and cisplatin resistance.