RESUMO
Acquired resistance to Taxol (TAX) contributes to clinical treatment failure and significantly reduces the survival rate of patients. The present study aimed to explore the effects of exosomal microRNA (miR)-187-5p on TAX resistance in breast cancer cells and its underlying mechanisms. Exosomes were isolated from MCF-7 and TAX-resistant MCF-7/TAX cells, and the miR-187-5p and miR-106a-3p levels of the cells and exosomes were determined using reverse transcription-quantitative polymerase chain reaction (RT-qPCR). Next, MCF-7 cells were treated with TAX for 48 h and either treated with exosomes or transfected with miR-187-5p mimics. Cell viability, apoptosis, migration, invasion and colony formation were determined using Cell Counting Kit-8, flow cytometry, Transwell and colony formation assays, and the expression levels of associated genes and proteins were detected by RT-qPCR and western blotting, respectively. Finally, a dual-luciferase reporter gene assay was performed to confirm the target of miR-187-5p. The results showed that miR-187-5p expression levels increased significantly in TAX-resistant MCF-7 cells and exosomes compared with normal MCF-7 cells and exosomes (P<0.05). However, miR-106a-3p was not detected in the cells or exosomes. Therefore, miR-187-5p was selected for subsequent experiments. A series of cell assays showed that TAX inhibited the viability, migration, invasion and colony formation of MCF-7 cells and promoted their apoptosis; however, these changes were reversed by resistant cell exosomes and miR-187-5p mimics. Additionally, TAX significantly upregulated ABCD2 and downregulated ß-catenin, c-Myc and cyclin D1, whereas resistant exosomes and miR-187-5p mimics reversed the TAX-induced changes in expression. Finally, ABCD2 was confirmed to directly bind with miR-187-5p. It may be concluded that TAX-resistant cell-derived exosomes delivering miR-187-5p may affect the growth of TAX-induced breast cancer cells by targeting ABCD2 and c-Myc/Wnt/ß-catenin signaling.
RESUMO
The anti-inflammatory activity of cirsilineol in in vivo condition was assessed by measuring the relative organ weight, lung dry/wet weight ratio, protein concentration, and infiltration of inflammatory cells in bronchoalveolar lavage fluid. We estimated the myeloperoxidase activity and levels of cytokines, chemokines, and inflammatory markers to analyze the efficacy of cirsilineol against lipopolysaccharide (LPS)-induced lung inflammation. Furthermore, we quantified the gene expression of NFkB/IKK signaling molecules in cirsilineol-treated and untreated acute lung injury mice to confirm the anti-inflammatory property of cirsilineol. The lung histology was assessed with hematoxylin and eosin staining. Apart from in vivo experiments, in vitro tests with LPS-stimulated RAW 264.7 macrophages were also performed. Cell viability assay was performed in the presence and absence of LPS in RAW 264.7 macrophages to determine the cytotoxic effect of cirsilineol against macrophages. Reverse-transcription polymerase chain reaction (RT-PCR) analysis was done to analyze the gene expression of inflammatory markers in LPS-treated RAW 264.7 macrophages to prove that cirsilineol effectively inhibits inflammation in vitro. The results of our study prove that cirsilineol effectively inhibits inflammation in both in vivo and in vitro conditions. RT-PCR analysis results of NFkB/IKK signaling molecules clearly illustrate that cirsilineol inhibited the expression of NFkB/IKK signaling protein and thereby prevented inflammation in in vivo condition, and it is further confirmed with the results of inflammatory protein expression in vitro model. The lung histopathological studies authentically confirm that cirsilineol potentially prevented the mice from LPS-induced lung inflammation.
