Objective:To investigate the role of macrophages in regulating breast cancer cell migration and its related mechanisms.Methods:Human leukemia monocytic cell line THP-1-secreted exosomes were isolated using multi-step ...Objective:To investigate the role of macrophages in regulating breast cancer cell migration and its related mechanisms.Methods:Human leukemia monocytic cell line THP-1-secreted exosomes were isolated using multi-step ultracentrifugation and verified using nanoparticle tracking analysis.Differentially expressed miRNAs were identified using RNA sequencing.Overexpression of inhibitors of hsa-miR-101-3p in breast cancer MDA-MB-231 cells was performed by infecting their lentiviral constructs.The luciferase reporter assay was used to evaluate the interaction of DLG5 and miR-101.DGL5 expression was detected using qRT-PCR and Western blot analyses.Results:The migration of breast cancer cells was significantly inhibited after addition of exosomes.RNA sequencing results showed that miR-101-3p expression was significantly upregulated.Targetscan analysis predicted that miR-101-3p could target DLG5,and this prediction was verified using the luciferase assay.The addition of the miR-101-3p precursor significantly increased the expression of miR-101-3p,and the mRNA and protein levels of DLG5 were suppressed.In contrast,inhibiting the expression of miR-101-3p increased the mRNA and protein levels of DLG5.Furthermore,the scratch assay showed that inhibiting miR-101-3p could promote the migration of MDA-MB-231 cells.Conclusions:Macrophage exosomes can inhibit the migration of breast cancer cells,and increasing the expression of miR-101-3p to inhibit DLG5 expression may play an important role in this process,which needs further investigation.展开更多
基金supported by the Key Research and Development Program of Hainan Province(ZDYF2020139,ZDYF2018158)the Science and Technology Funding Project of Hainan Province(821MS129).
文摘Objective:To investigate the role of macrophages in regulating breast cancer cell migration and its related mechanisms.Methods:Human leukemia monocytic cell line THP-1-secreted exosomes were isolated using multi-step ultracentrifugation and verified using nanoparticle tracking analysis.Differentially expressed miRNAs were identified using RNA sequencing.Overexpression of inhibitors of hsa-miR-101-3p in breast cancer MDA-MB-231 cells was performed by infecting their lentiviral constructs.The luciferase reporter assay was used to evaluate the interaction of DLG5 and miR-101.DGL5 expression was detected using qRT-PCR and Western blot analyses.Results:The migration of breast cancer cells was significantly inhibited after addition of exosomes.RNA sequencing results showed that miR-101-3p expression was significantly upregulated.Targetscan analysis predicted that miR-101-3p could target DLG5,and this prediction was verified using the luciferase assay.The addition of the miR-101-3p precursor significantly increased the expression of miR-101-3p,and the mRNA and protein levels of DLG5 were suppressed.In contrast,inhibiting the expression of miR-101-3p increased the mRNA and protein levels of DLG5.Furthermore,the scratch assay showed that inhibiting miR-101-3p could promote the migration of MDA-MB-231 cells.Conclusions:Macrophage exosomes can inhibit the migration of breast cancer cells,and increasing the expression of miR-101-3p to inhibit DLG5 expression may play an important role in this process,which needs further investigation.
