| Objective:The purpose of this study is to investigate the effect of liposomal curcumin on the chemosensitivity of breast cancer cells to Adriamycin via regulating microRNA pathway,and to study its possible mechanism,which could provide a theoretical basis for clinical use of curcumin against human breast cancer chemotherapy resistance.Methods:(1)Liposome curcumin was prepared by liposome encapsulation technology.(2)Fixed dose(6.8,13.58,27?M)of liposomal curcumin was added to the MCF-7/ADR cells at various concentrations of ADR(490,98,19.6,3.92,0.784,0.1568?M)incubating for 48 hours,the inhibition rate and half lethal dose(IC50)of Adriamycin on MCF-7/ADR treated with different concentrations of liposomal curcumin were observed by MTT,and the synergistic effect index(CI)of each group was also calculated.(3)Differentially expressed miRNAs and mRNAs among MCF-7/S,MCF-7/ADR and curcumin-treated MCF-7/ADR were detected using miRNA and mRNA microarray.(4)Candidate genes of dysregulated miRNAs were identified by prediction algorithms based on gene expression profiles.(5)The results of differentially expressed 5 miRNAs(hsa-miR-29b-1-5p,hsa-miR-29b-3p,hsa-miR-6068,hsa-miR-6790-5p and has-miR-4417)and 5 mRNAs(DDIT4,EPAS1,VEGFA,RPS14,DCDC2)in three cell lines were validated by RT-qPCR.(6)Three cell lines modulated by hsa-miR-29b-1-5p were used to further study the mechanism of drug resistance.After cells were transfected with hsa-miR-29b-1-5p mimics or its inhibitors,the effect of miR-29b-1-5p expression on drug resistance of breast cancer cells was observed by MTT.(7)Signaling pathways and target genes involved in Adriamycin resistance were analyzed by networks of KEGG pathways and protein interaction networks(PPI).Statistical analysis was performed using SPSS 22.0.Numerical data are presented as the mean ± SD of at least three independent experiments and the results showed good repeatability.Wilcoxon-Mann-Whitney rank sum test was used to compare the difference between groups.A p-value of<0.05 was considered statistically significant.Results:(1)The combination use of ADR and liposomal curcumin markedly increased inhibition ratio of MCF-7/ADR cells while the IC50 of ADR was significantly decreased and the result indicated that the combination had a synergistic growth-inhibitory effect in MCF-7/ADR cells.(2)The array results showed significant differential expression of miRNA and mRNA among MCF-7/S,MCF-7/ADR and curcumin-treated MCF-7/ADR.There were 67 miRNAs and 323 mRNA(at least two-fold changes)were differentially expressed in three cell lines.In addition,20 target genes(mRNAs)of each dysregulated miRNA not only predicted by prediction algorithms(TargetScan,Pictar,MicroCosm ? miRDB),but also differentially expressed in the microarray were discovered.(3)The expression status of five specific selected miRNAs was validated using RT-qPCR.The results were consistent with the analysis results of miRNA microarray.(4)Compared with negative control,the levels of miR-29b-1-5p expression for MCF-7/S transfected by miR-29b-1-5p mimics were significantly increased,and IC50 of ADR for cells were also significantly increased.In contrast,the levels of miR-29b-1-5p expression for MCF-7/ADR cells transfected by miR-29b-1-5p inhibitors were significantly decreased,and IC50 of ADR for cells were also significantly decreased.However,the levels of miR-29b-1-5p expression for liposomal curcumin treated-MCF-7/ADR transfected by miR-29b-1-5p mimics were significantly increased,and IC50 of ADR for cells were also significantly increased.(5)The results of KEGG and PPI network showed that MAPK,mTOR,PI3K-Akt,AMPK,TNF,Ras signaling pathways and several target genes such as PPARG,RRM2,SRSFland EPAS1,may associate with drug resistance of breast cancer cells to ADR.Conclusions(1)Liposomal curcumin could rescue part of Adriamycin resistance in breast cancer.(2)An altered miRNA expression pattern is involved in acquiring resistance to ADR,miRNA signaling pathways maybe an important molecular pathway through which liposomal curcumin could change the resistance to ADR in breast cancer MCF-7 cells... |
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