ABSTRACT
Objective To investigate the different biological characteristics of high-and low-expression of ATP-binding cassette transporter(ABCG2)in human multi-drug resistant nasopharyngeal carcinoma CNE2/DDP cell line(abbreviated as ABCG2High cell and ABCG2Low cell),and to analyze drug-resistant characteristics of the two cell lines.Methods ABCG2High and ABCG2Low cells were isolated by flow cytometry,while their cloning efficiency,cell cycle distribution,and the expression of ABCG2 before and after the 5th passage were examined as well.mRNA expressions of drug-resistant genes,such as ABCG2,Bcl-2,MDR1,MRP and MGMT,were detected by reversed transcriptive polymerase chain reaction(RT-PCR).MTT assay was utilized to detect the light absorbance(A)from day 1 to 14 to determine growth kinetics.Moreover,the drug sensitivity of the two cell lines to fluorouracil,cisplatin,vincristine,carboplatin,epirubicin,daunorubicin,paclitaxel and mitomycin were detected by MTT assay.Results The cloning efficiency of ABCG2High and ABCG2Low cells was 25.24%?1.18% and 16.28%?1.11%,respectively,on day 7.ABCG2High cells in S phase accounted for 41.5%,while the ABCG2Low cells in the G1/M0 phase went up to as high as 63.9%.The expression of ABCG2 in original ABCG2HighCNE2/DDP cells was 98%,while in ABCG2Low CNE2/DDP cells was 2%.However,at the 5th passage,the expression of ABCG2 decreased to 75% in the former cells and increased to 20% in the latter ones.The expression of ABCG2 mRNA in ABCG2High cells was obviously higher than that in ABCG2Low cells,while there was no difference in expression of other drug resistance genes between two cell lines.The growth velocity of ABCG2High cells exceeded that of ABCG2Low cells in early stage,but it slowed down 7 days later.MTT assay showed that IC50 of ABCG2High cells to 8 kinds of anti-tumor drugs were twofold higher than that of ABCG2Low cells,with statistically significant difference between them(P
ABSTRACT
Objective To investigate the mechanisms and effects of Sorafenib on cytotoxic sensitivity of allo-reactive natural killer(Allo-NK) cells against human multi-drug resistant nasopharyngeal carcinoma CNE2/DDP cells which expressing highly ATP-binding cassette superfamily G member 2(ABCG2)(abbr.as ABCG2HighCNE2/DDP cells).Methods ABCG2HighCNE2/DDP and Allo-NK cells were isolated by magnetic bead technique.The target cells were divided into 3 groups: a) treated group(ABCG2HighCNE2/DDP cells incubated with 10 ng/ml sorafenib for 4h);b) untreated group(conventionally cultured ABCG2HighCNE2/DDP cells);and c) control group(conventionally cultured K562 cells).Expression rates of ABCG2 in treated and untreated groups,and of five NKG2D-ligands(MICA,MICB,ULBP1,ULBP2,ULBP3) were evaluated by flow cytometry.The cytotoxic effects of NK cells against different groups of target cells were detected with LDH releasing assay.Results Expression rate of ABCG2 in isolated CNE2/DDP cells was 91.40%?2.32%.The purity of sorted CD3-CD16+CD56+ Allo-NK cells was 90% and higher.The expression rates of NKG2D-ligands(MICA,MICB,ULBP1,ULBP2 and ULBP3) in untreated group were 2.92%?0.33%,4.27%?0.33%,5.80%?0.62%,11.10%?3.15% and 7.75%?1.14%,respectively,which were remarkablely higher than that in treated group(10.38%?1.23%,10.68%?1.26%,11.62%?1.22%,43.24%?4.42% and 11.91%?0.88%,respectively,P